Comparative morphology of the gallbladder and biliary tract in vertebrates: Variation in structure, homology in function and gallstones

Comparative Morphology of the Gallbladder and Biliary Tractin Vertebrates: Variation in Structure, Homology in Functionand GallstonesCARLA K. OLDHAM-OTT1,2 AND JACQUES GILLOTEAUX*,2,3

1Department of Biological Sciences, Kent State University, Kent, Ohio 442422Summa Health System Foundation, Akron, Ohio 443043Laboratoire Oceanologique Arago de l’Universite Pierre et Marie Curie, Banyuls 66650, France

KEY WORDS gallbladder; biliary tract; vertebrates; ultrastructure; surface epithelium

ABSTRACT A review of investigations on the morphology of the gallbladder and biliary tract infish, reptiles, amphibians, birds, and mammals was performed. Scanning electron microscopy,transmission electron microscopy, and light microscopy observations by the authors were alsoincluded. Variations in the presence or absence of a gallbladder, surface epithelium of thegallbladder, and differences in the morphology of the biliary tract in vertebrates were reported.Many differences were diet-related. Despite some dissimilarities observed, analogous functioning ofthe biliary system was accomplished by its various components, with the biliary ducts performingthe function of the gallbladder when this organ was absent. In addition, the occurrence of peculiarparasitism and gallstones among some cases of vertebrates, including humans, was presented.Microsc. Res. Tech. 38:571–597, 1997. r 1997 Wiley-Liss, Inc.

INTRODUCTIONIn those animals which possess a gallbladder, the

function is essentially the same in all species: tofunction as an accessory organ of the digestive tract instoring bile, concentrating the bile by removing waterand exchanging electrolytes, actively secreting, modify-ing, and circulating the bile through the enterohepaticsystem, and affecting the digestive uptake of lipid-soluble compounds. Thus, the gallbladder functions asa reservoir and mechanical pump (Carey and Duane,1994), among its other functions. The size of thegallbladder, the amount of bile produced, and theextent to which the bile is concentrated varies widelyamong species (Gorham and Ivy, 1938). Bile is concen-trated only one to two times in cattle, four to ten timesin the dog, and eight to ten times in the human andhamster (Gabe, 1973; Gorham and Ivy, 1938). Variousother functions are also performed by the epithelialcells of the gallbladder, including the secretion andexcretion of substances such as hormones and xenobiot-ics. In addition, the production and secretion of aprotective mucus film at the cell surface of the gallblad-der and of several parts of the biliary tract is aremainder of the mucus-producing epithelial surface ofthe endodermal foregut lining.

In general, variability in gallbladder morphology isdependent mainly upon diet. Carnivores (except whales)invariably possess a gallbladder. The gallbladder mayor may not be present in omnivores and herbivores(Gorham and Ivy, 1938). In animals which eat onlyperiodically, as well as in humans, the bile receivedfrom the liver becomes more concentrated and is storedfor longer time periods than is the bile in vertebratesthat eat frequently, such as guinea pigs (Schoenfield,1977). Some animals, such as pigeons, rats, and deer,which eat almost continuously, have no gallbladder; the

constant flow of bile from the liver to the intestine isadequate for their digestive needs (Gorham and Ivy,1938; Bellairs, 1970). Diet is not the only factor in-volved in the differences seen in gallbladder morphol-ogy. Physical constraints imposed by the form of theanimal also play a part. A snake’s internal organs are,by necessity, more elongated than those of other ani-mals. The presence of a gallbladder appears to be aprimitive trait. It is found in most fish and all adultreptiles and amphibians and has been well conserved inmammals, for the most part. If the gallbladder isabsent, its functions are compensated for by differentparts of the biliary tract (Dorst, 1973). Table 1 illus-trates the presence or absence of a gallbladder through-out the vertebrates.

This study will attempt to summarize the data thathas been obtained in studies of the gallbladder andbiliary tract of vertebrates: fishes (Classes Agnatha,Chondrichthyes, and Osteichthyes), amphibians (ClassAmphibia), reptiles (Class Reptilia), birds (Class Aves),and mammals (Class Mammalia). A brief discussion ofthe parasites infesting the gallbladder in the lowergroups will also be included. The authors have alsoincluded the results obtained from scanning electronmicroscopy (SEM) and some transmission electron mi-croscopy (TEM) of the gallbladders and biliary tracts ofmembers of these vertebrate groups.

MATERIALS AND METHODSFishes (Torpedo marmorata Risso, Triglosporum las-

toviza Brunnich, Boops boops, and several specimens of

Contract grant sponsor: Summa Health System Foundation.*Correspondence to Dr. J. Gilloteaux, Summa Health System Foundation, 41

Arch Street, Suite 506, Akron, Ohio 44304, USA.Received 10 July 1995; accepted in revised form 25 July 1995

MICROSCOPY RESEARCH AND TECHNIQUE 38:571–597 (1997)

r 1997 WILEY-LISS, INC.

TABLE 1. Presence (1) and absence (2) of gallbladder or unreported existence (?) throughout Zoology Systematic with specific remarks ( )

ChordataPhylum Urochordatae or Tunicates ................................................................................................................. 2Phylum Cephalochordatae or Acrania

Fam. Branchiostomidae Branchiostoma sp. ......................................................................................... 2Asymmetron sp. ............................................................................................ 2

Phylum VertebrataSuperclass AgnathaClass Cyclostomata

Order Petromyzonta Petromyzon sp. .................................................................................................... 1 (larval stage only)Order Myxinoidea Myxine sp. ............................................................................................................ 1

Class Chondrichthyes (All) ............................................................................................................. 1Subclass Selachii

Order ElasmobranchiiiSubclass Bradyodontia

Order HolocephalaClass Osteichthyes (All) ............................................................................................................. 1

Subclass ActinopterygiiSuperOrder ChondrosteiSuperOrder HolosteiSuperOrder Teleostei

Subclass BrachiopterygiiSubclass CrossopterygiiSubclass Dipnoi

Class Amphibia (All) ............................................................................................................. 1Subclass Labyrinthodontii

SuperOrder AnouresSubclass Urodelomorphii

SuperOrder UrodeliSuperOrder Gymnophioni

Class Reptilia (All) ............................................................................................................. 1Subclass Anapsidei

Order CheloniiSubclass Diapsidei

Order SquamataOrder Crocodilii

Class Aves.Subclass Ratitii

Order Struthioniformi .............................................................................................................................. 2Order Apterygiformi ................................................................................................................................. 1

Subclass CarinatiiOrder Colymbiformi .................................................................................................................................. 1Order Alciformi ......................................................................................................................................... 1Order Procellariiformi .............................................................................................................................. 1Order Pelecaniformi ................................................................................................................................. 1Order Ardeiformi ...................................................................................................................................... 1Order Anseriformi ..................................................................................................................................... 1Order Lariformi ........................................................................................................................................ 1Order Charadriiformi ............................................................................................................................... 1

except Erolia alpinaOrder Ralliformi or Gruiformi .................................................................................................................. 1

except Truix tankiOrder Tinamiformi ................................................................................................................................... 1Order Galliformi ....................................................................................................................................... 1Order Columbiformi (most species) ......................................................................................................... 2

except Ptilinopus insolitusCoryphoenas crassirostrisCarpophaga sp.Lopholaemus sp.Ptilonopus sp.

Order Falconiformi ................................................................................................................................... 1Order Strigiformi ...................................................................................................................................... 1Order Psittaciformi ................................................................................................................................... 2

except Cacatua sp.Calopsitta novae-hollandiae

Order Cuculiformi ..................................................................................................................................... 2Order Piciformi ......................................................................................................................................... 1

except Picumnus squamulatusOrder Caprimulgiformi ............................................................................................................................ 1

except some of Chordeiles sp.Order Apodiformi ...................................................................................................................................... 1Order Coraciadiformi ............................................................................................................................... 2

except Halcyon smyrnensisOrder Passeriformi ................................................................................................................................... 1 (most species)

except:Fam. Pittidae ..................................................................................................................................... 2 (Pitta oatesi)Fam. Dicruridae ................................................................................................................................ 2 (Chaptea aena)Fam. Paridae ..................................................................................................................................... 2 (Aegithaliscus annamensis)Fam. Timeliidae ................................................................................................................................. 1

except Mixornis sp.Fam. Bombycillidae ........................................................................................................................... 2 (Bombycilla garrula)Fam. Nectariidae ............................................................................................................................... 2 (all)

Order Sphenisciformi ............................................................................................................................... 1

TABLE 1. (continued)

Class MammaliaSubclass Prototheria

Order Monotrema ..................................................................................................................................... 1 (not Ornithorhychus)Subclass Theria

Order Metatheria or Marsupialia ............................................................................................................ 1Order Polyprotodonta ............................................................................................................................... ?Order Paucituberculata ............................................................................................................................ ?Order Diprotodontia ................................................................................................................................. ?

Subclass Eutheria or PlacentaliaOrder Zalambdodonta .............................................................................................................................. ?Order Insectivora ...................................................................................................................................... 1Order Macroscelidea ................................................................................................................................. ?Order Tupaioidea ...................................................................................................................................... 1Order Chiroptera ...................................................................................................................................... 1Order Dermoptera .................................................................................................................................... 1Order Xenarthra ....................................................................................................................................... 1Order Cingulata ........................................................................................................................................ 1Order Tardigrada ...................................................................................................................................... 1

except Bradypus sp.Order Pholidota ........................................................................................................................................ 1Order Rodentia ......................................................................................................................................... 1

Sub Order ProtogomorphaFam. Aplodontidae ............................................................................................................................. 2

Sub Order SciuromorphaFam. Sciuridae ................................................................................................................................... 1Fam. Ctenodactylidae ....................................................................................................................... 1Fam. Castoridae ................................................................................................................................ 1Fam. Heteromyidae ........................................................................................................................... 2Fam. Geomyidae ................................................................................................................................ 2

Sub Order Theridomorpha .................................................................................................................... ?Sub Order Gliromorpha ........................................................................................................................ ?Sub Order Myomorpha ......................................................................................................................... 1

Fam. Muriodeae ................................................................................................................................. 1Fam. Cricetidae ................................................................................................................................. 1Fam. Muridae .................................................................................................................................... 1

except Rattus and Apodemus sp.Sub Order Caviomorpha

Fam. Erethizontidae .......................................................................................................................... 2Fam. Octodontidae ............................................................................................................................ 2Fam. Echimyidae ............................................................................................................................... 1Fam. Caviidae .................................................................................................................................... 1

Sub Order HystricomorphaFam. Hystricidae ............................................................................................................................... 2Fam. Thryonomyidae ........................................................................................................................ ?Fam. Petromuridae ........................................................................................................................... 2Fam. Bathyergidae ............................................................................................................................ ?Fam. Pedetidae .................................................................................................................................. 2

Order CarnivoraSub Order Fissipedia ............................................................................................................................ 1

except Nandinia sp.Sub Order Pinnipedia ........................................................................................................................... 1

Order Lagomorpha ................................................................................................................................... 1Order Tubulidentata ................................................................................................................................. 1 (Orycteropus afer)Order Perissodactyla ................................................................................................................................ 2Order Sirenia ............................................................................................................................................ 1

except Rhytina sp.Order Proboscidea .................................................................................................................................... 2Order Hyracoidea ..................................................................................................................................... 1 (variable development)Order Artiodactyla

Sub Order SuiformesFam. Hippopotamidae ....................................................................................................................... 1 (not constant)Fam. Suidea ....................................................................................................................................... 1

except Dicotylinae sp.Sub Order Tylopoda .............................................................................................................................. 2Sub Order Ruminantia

Fam. Tragulidae ................................................................................................................................ 1Fam. Moschidae ................................................................................................................................. 1Fam. Cervidae ................................................................................................................................... 2Fam. Giraffidae .................................................................................................................................. 1 (not constant)Fam. Antilocapridae .......................................................................................................................... 1Fam. Bovidae ..................................................................................................................................... 1

except Cephalophinae sp.Order Cetacea ........................................................................................................................................... 2Order Primates ......................................................................................................................................... 1

This table was made by using Grasse (1958–1983), Grasse and Devillers (1965), Remane et al. (1976), and Grzimek (1972–1975) systematic zoology texts and containsonly animal groups for which records of gallbladder were made or found. All fossil groups were omitted. In addition, Gorham and Ivy (1938) was used to complete thetable.

573VERTEBRATE BILIARY TRACT AND GALLSTONES

Uranoscopus sp.), along with another set of 12 species(to be studied later), were captured outside the BanyulsBay (France; Mediterranean Gulf of the Lion), bynet trailing between 50–60 m of depth along a NNW-SSE line facing Cape Bear to Cape de l’Abeille (at theedge of the National Biological Marine Reserve). Follow-ing capture, the fishes were kept and transported inrunning sea water tanks until they were placed in thelaboratory aquaria where, after decapitation, they weredissected, the gallbladder and liver were excised andfixed for 1.5 h duration in 3% buffered glutaraldehyde(0.1 M sodium cacodylate) diluted in seawater (diluted1:3 [v:v] by distilled water). Following this step, wash-ing and postfixation with 2% OsO4 aqueous solutionfollowed; tissues were then washed in buffer containing10% sucrose and then dehydrated by graded alcohols.Most of the tissues were then prepared for scanningmicroscopy (SEM) via critical-point drying followed bycoating the samples with 15–20-nm-thick gold layer.Other samples were prepared for transmission electronmicroscopy (TEM) by embedding in Polybed 812 epoxyresin (Polysciences, Warrington, PA) and ultrathin sec-tions were cut for observation in the transmissionelectron microscope. Amphibians such as the sala-mander (Ambystoma laterale) and toad (Bufo america-nus) were captured in two wet areas of home backyardin Kent, OH. Armadillo lizards, originating in SouthAfrica (Cordylus cataphractus), were obtained from alocal pet store. Tree frogs (Hyla sp.) and anoles (Anoliscarolinensis [not illustrated here]) were collected in abackyard and swampy ground near the TangipahoaRiver in Louisiana. Three-month-old female Syrianhamsters (Mesocricetus auratus Waterhouse) were pur-chased from BioBreeders (Cambridge, MA) and wereused for other studies (Gilloteaux et al., 1993c) andhuman samples were obtained from cholecystectomiesand fixed in 3.5% buffered glutaraldehyde (Gilloteauxet al., 1989). All samples from live amphibians orreptiles were obtained after euthanasia or were ob-tained already fixed by formaldehyde and preserved in3.2% buffered (0.1 M cacodylate buffer) glutaraldehydefor 2 h at room temperature and then were kept at 4°Cuntil microscopic dissections took place. Samples werethen prepared for TEM observation by epoxy embed-ding as described above and in a previous report(Gilloteaux et al., 1992, 1995).

FISHFish constitute a diversified group divided into three

classes: Class Agnatha or Cyclostomata, Class Chon-drichthyes, and Class Osteichthyes (Winchester andJaques, 1981). The diet of fish is extremely varied. Bothherbivory and carnivory are practiced and the type offeeding habit or niche adaptation is reflected by thespecialization and diversity of digestive system morphol-ogy (including the liver, biliary tracts, and gallbladder).One can distinguish between planctonophagy (Syn-gnathidae), predatory or carnivorous feeding (manyspecies), madrepore-eating (Scaridae), insect-eaters(Toxotes), bottom-feeders (Mugilidae, Mullidae), andherbivorous feeders.

In general, the fish livers are not as definitely ar-ranged in cords as in the mammalian liver (Bluntschli,1904). Portal triads are seldom observed in fish. Theliver is a more or less tubular gland in lower groups

(Biagianti-Risbourg, 1990; Elias and Bengelsdorf, 1952;Gonzalez et al., 1993; Hampton et al., 1988). The biliaryductules or cholangioles, which may be very long,contain numerous actin filaments surrounding the lu-men (Figure 10, and Gilloteaux et al., 1995). Thegallbladder is occasionally absent in fish (Hyrtl, 1868;Mentzer, 1929a) but these finding may indicate congeni-tal anomalies and were not verified elsewhere. Thelayers found in mammalian gallbladders can also beseen in fish and in all vertebrates. A surface epithelium,lamina propria, fibromuscular layer, connective tissueor subserosal layer, and serosal layer are all present.

Class Agnatha: Jawless FishThe most primitive class of fish is the jawless fish. In

the liver of the larval sea lamprey, Petromyzon mari-nus, hepatocytes are found in a tubular pattern aroundthe bile canaliculi (Peek et al., 1979). The gallbladder isintrahepatic. It lies within the anterior portion of theliver and the gallbladder cells are separated from theliver cells by blood vessels, connective tissue, andmuscle fibers (Youson and Sidon, 1978). ‘‘Leaky’’ zonulaoccludentes of the bile canaliculi form a bile-bloodbarrier (Youson et al., 1987). The bile ducts are formedof cells with lateral folds and numerous microvilli onthe apical surface, forming a brush border that nearlyfills the lumen. The gallbladder epithelial cells are verylike those of the bile ducts but have a less-developedbrush border. The gallbladder cells contain glycogen,analogous to that found in some mammals. Fewermitochondria and narrower intercellular spaces thanare found in mammals are present, indicating thattransport may be reduced (Sidon et al., 1980).

When the larval sea lamprey metamorphoses into anadult, the intrahepatic gallbladder and bile ducts arelost. In human, biliary atresia is a fatal disease ofunknown etiology, which affects infants and it is diffi-cult to reproduce with an animal model. This primitivefish could be used as a model for the human condition(Youson and Sidon, 1978). The livers of adult lampreys,possess cells similar to those of other vertebrates anddo not show marked changes due to bile stasis (Yousonet al., 1985). The vascular channels surrounding thebile ducts become separated from the ducts by thickfibrous tissue (Yamamoto et al., 1986). Extrahepaticcommon bile duct cells dedifferentiate and in time giverise to the adult pancreas (Elliott and Youson, 1993;Youson et al., 1987). Tight junctions between the paren-chymal cells of the adult liver are lost after biliaryatresia. However, gap junctions increase, thus allowingfor greater cell contact (Youson et al., 1987).

The biliary tree of the larval brook lamprey, Lampe-tra lamottenii Le Sueur, is organized much like that ofthe sea lamprey. A large spherical-to-ovoid nucleus isfound in the basal portion and numerous mucus gran-ules in the apical region of each bile duct cell. Anoccasional cilium was observed at the apical surface ofthe intrahepatic bile duct cells (Eng and Youson, 1992).Interesting studies done by Sorensen and colleagues (Liet al., 1995) indicate that lampreys appear to find theirspawning stream by following the scent of a unique bilesalt, petromyzonol sulfate, only made when they arelarvae. This compound could be used to fool adults intofinding unsuitable spawning grounds and thus prevent

574 C.K. OLDHAM-OTT AND J. GILLOTEAUX

growth population of this parasitic fish consuming alarge number of the Great Lakes’ native fishes.

Class Chondrichthyes: Cartilaginous FishIn sharks, the liver is horse-shoe shaped and has

equal-sized right and left lobes. The gallbladder iselongated and is usually embedded in the right lobe.Occasionally, it is totally surrounded by hepatic tissues(Bertin, 1958; Gilloteaux et al., 1995).

The electric ray, Torpedo marmorata Risso, possessesa gallbladder composed of the characteristic layerspreviously mentioned, with a thickness of up to 500 µm.This fish and the others observed in our laboratory arebottom feeders, collected from the Mediterranean Sea.T. marmorata has a typical epithelium with numerousshallow folds, as shown by SEM (Fig. 1A) and bulgingcell apices of different sizes and general aspects. Usinglight microscopy of 1-µm-thick sections (Fig. 1B), thecells are easily differentiated from one another bybasolateral spaces attached to the basal lamina and arelinked to each other by junctional complexes. The basalportion of the cells contains numerous mitochondriaand a few wandering cells may be observed. The oval,elongated nuclei, containing at least two nucleolarbodies, are found at the same level in the lower half ofthe cells. Junctional complexes, reaching approxi-mately 2 µm into the cell, are observed between thewell-delineated cells when the epithelium is observedwith TEM (Fig. 1C). The bulging apices of the cells andcytoplasmic material within the gallbladder lumenwere also observed in Figure 1B and C. Abundantmitochondria are located throughout the cell but aremore abundant in the supranuclear region; elongateddense bodies are also apparent. The bulging apices ofthese epithelial cells (Fig. 1D) are observed with SEMto be covered by short, fine microvilli interspersed withminute bulging excrescences. These apical excres-cences may be stages in the secretion of mucous-containing vesicles (Gilloteaux et al., 1995). The spheri-cal or conical apices seem to form complementarycontours secreting a layer of mucus. The cell apices,viewed with TEM (Fig. 1E), appear to be filled withglycogen and other materials, but discrete mucus gran-ules are not observed. As the mucus granules reach theapex, they fuse with the material there and merge withit, making them difficult to discern within the apicalmass. The cell seems to segregate this apical masswhich is then liberated into the lumen.

Class Osteichthyes: Bony FishThe bony fish are perhaps the most studied of the

groups of fishes, especially the Superorder Teleostei. Agreat variability in liver morphology is present. In thelungfishes, Neoceratodus has a gallbladder that isassociated with the anterior lobe of the liver (Blunt-schli, 1904). The ray-finned fishes have a liver with one,two, or three or more lobes. The liver is like that ofhigher vertebrates in that the biliary canaliculi areformed by junctional complexes of the plasma mem-branes of the hepatocytes (Hampton et al., 1988).Melano-macrophage centers, found in the parenchymaof many fish livers, may perform a variety of activities,among them storage of waste products such as lipofus-cin, storage of metals such as hemosiderin, and theaccumulation of melanin (Biagianti-Risbourg, 1990;

Gonzalez et al., 1993). The bile duct may be short andpossess a muscular sphincter, as in Cottus gobio (West-ern, 1969).

The gallbladder of teleosts, if present, may be spheri-cal, ovoid, elongated, or more varied. The gallbladder ofC. gobio is relatively large and spherical when full,while that of Parenophrys bubalis is more elongate andhighly elastic. Both of these fish are strict carnivores(Western, 1969). The gallbladder epithelial cells of thetench have been observed to contain glycogen (Vieh-berger, 1982). Peroxisomes have been described in thegallbladder epithelial cells of the stickleback, Gasteros-teus aculeatus (Ruiter et al., 1988). The injection ofradioactively labelled thyroid hormones in the brooktrout (Salvelinus fontinalis) resulted in biliary excre-tion of these compounds, as has been seen in severalmammals (Eales et al., 1971).

The rainbow trout, Salmo gairdneri, has a gallblad-der epithelium composed of cells covered with numer-ous microvilli, forming a authentic brush border. Inother species, the microvilli are less regularly arranged.The epithelial cells in fish range from high columnar tolow columnar to cuboidal, have microvilli in greater orlesser amounts, and also have a glycocalyx. In the carp(Cyprinus carpio L.) the epithelial cells are very tall(75–125 µm). Three populations of mitochondria arepresent in the epithelial cells of the carp: supranuclear,basal, and circumnuclear. According to Bader (1965),b-cytomembrane whorls are present in the infranuclearregion. It is possible that these serve as a reserve ofendoplasmic reticulum. Regeneration of gallbladdercells in this fish arises from small, undifferentiatedcells attached to the basal lamina, which grow to reachthe height of the other cells.

The gallbladder of Triglosporum lastoviza Brunnich,when viewed with light microscopy, demonstrates tall,columnar epithelial cells (Fig. 2A). The cells are sonarrow and elongated that the epithelium gives theappearance of pseudostratification. The bulging apiceshave a pale appearance, suggesting the presence ofmucus glycoproteinaceous secretory releases. The en-tire gallbladder wall has been previously described(Gilloteaux et al., 1995). TEM displays the extensivejunctional complexes between cells bulging and decapi-tating their apical excrescences in the lumen (Fig. 2B).The secreted material in the lumen contains no organ-elles, but within the electron dense material, one mayobserve the material mixed with mucus granules (about1 µm in diameter) still bounded by membranes. Some ofthe apical excrescences display electron-lucent mucoussecretory granules and mitochondria (Fig. 2B).

Mucinous secretions are widespread in fish. Numer-ous small granules have been observed in the epithelialcells of the lungfish, Protopterus, usually near a well-developed Golgi apparatus; these granules were alsoobserved in cystic duct epithelial cells in this fish(Higashi et al., 1986). Madrid and collaborators (1989)have shown that gallbladder of the sea bream has aPAS positive luminal surface, their observations indicat-ing the presence of mucins, sialomucins, and glycogens.The gallbladder of Boops boops, a Mediterranean school-ing fish, gives evidence of the abundant mucus secre-tion which may be present. This fish’s gallbladder cellsurfaces, observed in our laboratory using SEM, beginthe secretory cycle with the formation of bulging epithe-


lial apices, which then are discharged, and comminglewith the mucus secretions of nearby cells, forming afilm of mucus (Fig. 3A–D). Following discharge, the cellsurfaces ‘‘heal,’’ forming a uniform surface outlined bymarked junctional complexes (Fig. 3E). The cycle be-gins anew as the central portion of some cells againshow evidence of the beginnings of secretion (i.e.,bulging apices) following healing and regrowth of apicalaspects (Fig. 3F).

Uranoscopus sp. have a gallbladder with a very thickwall (Gilloteaux et al., 1995) and a most complex

mucosal surface. The epithelial cells, observed usinglight microscopy (Fig. 4A), have spherical nuclei in thelower half of the cell. Toluidine blue staining displaysgranules up to 1.5 µm in diameter. The supranuclearregion is usually empty of granules, as is the upper 3–5µm of the apical zone. Extended apical processes reach-ing 10 µm have been noted. TEM shows that most of thegranules are moderately electron dense (Fig. 4B) andappear to be made of mixed lipids, often containingcurved to circular electron lucent, paracrystalline inclu-sions resembling those found in human cholesterolosis,

Fig. 1. Torpedo marmorata gallbladder epithelium. A: Shallowconvoluted folds of the mucosal surface display apical bulgings ofvariable shape and size. Bar 5 10 µm. B: LM of epoxy embeddedspecimen, 1 µm-thick, Toluidine blue stained section showing some ofthe pleomorphic apical excrescences displayed in A. Arrows indicatemigrating cells. L: lumen. Bar 5 10 µm. C: TEM view of supranuclearepithelial regions rich in mitochondria an a few lysosomal bodies.Notice the lack or paucity in organelles in the apical excrescences.

Small arrows depict long, well-defined junctional complexes. Bar 5 5µm. D: Detail SEM View of narrow and wide apical excrescences.Bar 5 1 µm. E: TEM view of similar field of view observed in D. Thebulging apices almost only contain a fine granular cytoplasm segre-gated from the main cell body at the level of the junctional complex(arrows) where electron lucent mucus inclusions are scattered amidstsmall rounded mitochondria and residual bodies. Bar 5 1 µm.


although in fish these structures are enclosed withinlipoid droplets and are not found within the cytoplasmas in humans. In the upper region of the cytoplasm,delineated by thin microfilaments (actin), electron lu-cent, round mucus droplets were observed (Fig. 4C).Mucus granules are also located between the lipiddroplets and a population of small (0.2 to 0.5 µm indiameter), electron densely contrasted and pear-shaped, elongated mucous secretory granules are ob-served forming an apical, 1.0–1.5-µm-thick layer (Fig.4B). Abundant mitochondria with pale matrices anddense bodies are also present. The cells contain exten-sive junctional complexes and clavate apices. Microvillisometimes surround the bulging, clavate apices. Adetailed view of the elongated, electron-densely con-trasted (acidic) mucous granules is visible in Figure 4D.SEM confirms the existence of a complex apical surface.Conical, clavate, and spherical excrescences may bepresent, surrounded by long microvilli, some of whichmay have been damaged by fixation procedures. Open-ings on the apical surface appear to be remnants ofrecent apical discharge (Fig. 4E).

The quasi-spherical gallbladder of Scorpena scrofa L.appears indented after fixation as a result of a commonattachment of pancreatic mesentery (Fig. 5A). Thegallbladder wall is thin and pancreatic tissue is associ-ated with the wall, as may also be seen in Figure 5B.The gallbladder epithelium is composed of low cylindri-cal to cuboidal cells (Fig. 5B, inset). The fibromuscular

layer is narrow (Fig. 5C) and the cytoplasm appearscondensed, perhaps due to fixation or to the presence ofthe probable parasites pictured here, which will bediscussed in a later section of this review. The epithelialcells can be observed discharging single or groupedmucus granules (Fig. 5D) enmeshed in a fine, granularmaterial. Tall microvilli coated with an abundant glyco-calyx are also present. A detailed description of thegallbladder morphology of S. scrofa will be found else-where (Gilloteaux and Gilloteaux, manuscript in prepa-ration).

AMPHIBIANSAmphibians are usually classified into three orders:

Order Gymnophiona or Apoda (caecilians), Order Cau-data or Urodela (salamanders, newts, sirens, mudpup-pies), and Order Anura or Salientia (frogs and toads)(Grasse and Devillers, 1965; Grzimek, 1972–1975; Re-mane et al., 1976; Winchester and Jaques, 1981). Allknown adult members of this group are carnivorous.The diet of amphibian larvae, however, can be varied.Carnivores, omnivores, and herbivores are all repre-sented by larval stages (Burggren and Just, 1992). Mostanurans possess a bilobate liver, while in the OrderCaudata, the liver is elongated. Three genera of tad-poles have been found to have a trilobed liver (Nodzen-ski et al., 1989). In caecilians, which are limbless, theliver is greatly elongated. The hepatocytes of some frogsappear to be arranged in rows or hepatic cords, similar

Fig. 2. Triglosporum lastoviza gallbladder epithelium. A: LM viewof epoxy embedded specimen, Toluidine blue stained 1-µm thicksection. Tall cylindrical epithelium is illustrated with nuclei located atdifferent height, and apical excrescences (viewed as stained lilac andblue by LM) are suggestive of a glycoproteinaceous content. Bar 5 10µm. B: TEM view of several adjacent apices of the epithelium of A,demonstrating the heterogenous content of the tall, pencil-like excres-

cences containing fine cytoplasmic granulations mixed with a largenumber of mucus granules (m). Large arrows indicate exocytosis ofmucus while apices are themselves in the process of being severedfrom the epithelial cells. Small arrows indicate junctional complexes.S: base of mucin layer in the lumen still containing mucus granulesdelineated by their membrane boundary. Bar 5 1 µm.


to those in mammals, although no portal triads arefound (Azanza et al., 1989).

As shown in Table 1, the gallbladder is present in allamphibians. In most amphibians, the gallbladder isattached to the liver (Duellman and Trueb, 1986). Insome tadpoles, it appears as a large transparent sacbetween two of the three lobes of the liver (Nodzenski etal., 1989). The gallbladder of Triturus pyrrhogaster(Caudata) has an epithelium comprised of simple, highcolumnar cells, usually uninucleated with from 1–3nucleoli. The mitochondria of these cells are found

mostly between the nucleus and the free cell surface,but this organelle’s spatial distribution varies with thecell’s functional activity. Glycogen may be present,usually in the basal portion of the cell. Lipid dropletsare present in the epithelium and lamina propria.Secretory vesicles may be seen; apocrine secretion alsooccurs, probably of a mucus-like material (Togari andOkada, 1954).

The gallbladder of the frog is similar. Mitochondriaare found in the basal zone, a brush border is present,and some cells with bulging apices projecting into the

Fig. 3. Boops boops gallbladder epithelium. A–D: Apical excres-cence (*) from adjacent cells forming by confluence a thick mucus (m)film (in C) above the epithelium. D: Extruding apices where curvedarrows indicate cell junctions. Bars 5 5 µm. E: Coalescent apicaldischarges and mucus are shown at the surface of empty-like, apical

regions of epithelial cells only demarcated by their remaining junc-tional complexes (arrow). Bar 5 1 µm. F: Apical surface of epithelialcells after ‘‘healing’’ arrows indicate junctional complex outlining acoat of short blunted microvilli. Bar 5 1 µm.


Fig. 4. Uranoscopus gallbladder epithelium. A: Toluidine bluestained, 1-µm-thick epoxy section of sample demonstrating the granu-lar content of the supranuclear cytoplasm. Extended apical processescan be detected (arrow) in the lumen (L). Loosely arranged smoothmuscle fibers (f) in the lamina propria and a large diameter bloodvessel can be seen. Bar 5 5 µm. B: TEM view of the apices of twoadjacent cells. It is possible to contrast large diameter lipid-likeinclusions containing a curved or circular electron lucent crystalloidinclusion (small arrows) surrounded by mitochondria. Small electron-dense apical granules located all along the conical apical edgescontaining small (in D) electron dense and lucent mucus granules. L:

lumen. Bar 5 1 µm. C: Detailed TEM view of adjacent cell apicesshowing the complex subcellular architecture of these cells. Longmicrovilli are surrounding the clavate apex (large arrow) reaching thelumen (L). li: lipoid granules with crystalloid (arrow); m: mucusgranules. Curved arrows indicate an example of elongated junctionalcomplex. Bar 5 1 µm. D: Detailed view of pear-shaped, electrondensely contrasted apical mucus granules (small arrows). Open arrowindicates tall microvilli. Bar 5 1 µm. E: SEM view of apical surfacesshowing a few long microvilli (open arrow). An exocytotic site (crater-like) can be detected (*) among conical to clavate apices. Bar 5 1 µm.


Fig. 5. Scorpena scrofa gallbladder epithelium and parasites. A:Overall view of the gallbladder. One-micron-thick section, Toluidineblue stained. P (small arrows): mixed exocrine and endocrine pancre-atic tissue normally adhering to the gallbladder. Bar 5 50 µm. B:Transverse 1-µm-thick section, Toluidine blue stained, of the gallblad-der wall, showing the entire wall histology and associated pancreatictissue richly vascularized. Notice four subepithelial parasites. Inset:Example of encysted parasite (arrow). Bars 5 10 µm. C: TEM view of

the gallbladder wall and a parasite. f: smooth muscle fibers of the thinmuscular tunics; n: nucleus. Bar 5 5 µm. D: TEM view of a section of aparasite. m: mucus exocytosis; n: nucleus. Open arrow indicates one ofthe phospholipidic whorls or micelle present in the cytoplasm. A smallarrow indicates one of the micelles entangled among the parasiteprotoplasmic extensions ‘‘crawling’’ between the short epithelial micro-villi. Bar 5 1 µm. E: Detailed TEM view of phospholipidic whorlsamong the mucin covering the epithelial surfaces. Bar 5 200 nm.


lumen, suggesting apocrine secretion, are present(Grzycki and Nowakowski, 1967). The epithelium ofRana esculenta L. is that of a typical amphibian,although it is more cuboidal than cylindrical. Somepreliminary studies (Bader, 1965) of the metamorpho-sis of the larval to the adult stage indicate morphologi-cal changes occurring: as the epithelium degenerates,there is an invasion of blood cells (leukocytes andmacrophages) followed by regeneration. Further ultra-structural studies are needed to elucidate the changesthat occur in this metamorphosis. The diet of a tadpoleis herbivorous while that of an adult is omnivorous orcarnivorous; it is probable that the change in diet isreflected in morphological differences.

Tadpoles of Rana perezi have a gallbladder epithe-lium composed of simple cuboidal or low cylindricalcells. With the application of the carbamate insecticideZZ-Aphox, this epithelium will consist of flat cells untilthe 4th day of development, but the cells will recover tonormal height by the 14th day; it is suggested that bilemay be an important route for the excretion of lipid-based xenobiotics (Honrubia et al., 1993).

PAS-positive granules in the cytoplasm of the Natter-jack toad gallbladder epithelium have been found.These granules appear in both the cytoplasm andluminal surface in the lake frog (Rana perezi). Lectinhistochemistry indicates that the residues are sialicacid in the frog, but appear to be GlcNAc in the toad(Madrid et al., 1989). Both have been identified in thehuman gallbladder. Monoamine oxidase activity hasbeen observed in the epithelium of the frog (Grzyckiand Nowakowski, 1967). Melanin granules, often lo-cated near blood vessels and nerves, have been found inboth the liver and the gallbladder of Rana ridibunda(Azanza et al., 1989).

In the course of studies related to the comprehensionof fluid and ion transport across the gallbladder wall,many researchers have identified tight junctions in thegallbladder epithelium of Necturus (mudpuppy) (Bent-zel et al., 1980, 1987; Hill and Shachar-Hill, 1993;Loeschke and Bentzel, 1994; Palant et al., 1983;Shachar-Hill and Hill, 1993). Recent freeze-fracture ofthis tissue has revealed the tight junctions to beinterconnecting strands encircling each cell, thus sepa-rating the apical from the basolateral membranes.Occasional strands have been seen in the zonula ad-herens (Bentzel et al., 1987). A decrease in strandnumber, and thus resistance, has been observed withthe removal of calcium (Palant et al., 1983). Celldimensions and fluid transport were also investigatedby Rostgaard and Frederiksen (1981) and volume regu-latory decrease has also been shown to be dependent oncalmodulin and a microfilament network (Foskett andSpring, 1985). This confirms the work of Bentzel et al.(1980) showing that an application of phalloidin orcytochalasin B increases resistance. A naturally occur-ring protein, protamine, increases transepithelial resis-tance, causing the tight junction domain to expandvertically; this is reversible with heparin (Bentzel et al.,1987). Necturus gallbladder has been studied exten-sively as a model of transepithelial pathway (Larsonand Spring, 1984; Reuss, 1979, 1983; Reuss et al., 1980)and this fluid transport, epithelial cells, and intercellu-lar spaces were also studied in Rana esculenta gall-bladder.

Preliminary investigations in our laboratory usingSEM have provided descriptions of the gallbladders ofthree amphibians, furnishing an overall view of theorgan that is similar in all three specimens.

The epithelial cells of the salamander (Ambystomalaterale) gallbladder show a fairly smooth surface, withlittle mucus and only shallow undulations present (Fig.6A). There are no deep folds as are found in reptiles ormammals; the general appearance is more like that ofthe fish gallbladder. Numerous microvilli, approxi-mately 1 µm in length, are apparent on the bulgingapical cells (Fig. 6B) and their delicate microvillar coatsappear almost as willow inflorescence.

The tree frog, Hyla sp., demonstrates a shallowlyfolded epithelial surface similar to the salamander (Fig.7A). The microvilli are more clumped than in thesalamander, probably as a result of increased mucussecretion. Scattered bulging apices are covered by tallerand delicate microvilli (Fig. 7B), up to 5 µm in length.These microvilli have an appearance similar to that ofthe tuft cells seen in the mouse (Luciano, 1972; Lucianoand Reale, 1969, 1990; Nevalainen, 1977), although wehave not yet performed TEM studies to confirm thisobservation. Based on this peculiar morphological as-pect, it is possible that these tall microvilli possess apossible mechano- or chemo-sensory function.

Bufo americanus (common toad) exhibits a shallowlyfolded epithelium similar to the other two amphibiansobserved in our laboratory. There are, however, epithe-lial cells with significant apical bulging present (Fig.8A), which produce the mucus material seen in Figure8F. This mucus is retained on the epithelium afterfixation, producing the clumped microvilli seen in Fig-ure 8B. Outlines of individuals cells and microvilli areblurred as a result of this secretion. There are areas inthe toad gallbladder devoid of mucoid material (Fig.8C). These areas are very similar to the salamandergallbladder, with the cell boundaries more well definedby the evenly distributed microvilli. A large field of viewof the gallbladder epithelium demonstrated the varyinglarge and small aggregates of microvilli present (Fig.8D). Deposits resembling microcalculi were found be-tween the cell boundaries (Fig. 8E). Mucus-like mate-rial is occasionally observed secreted in globs or blobs of4–5 µm in width before it is spread over the surface ofthe epithelium. A detailed view of the clumped micro-villi shows some of them to be tall (ranging from 1.5 to2.5 µm) and curved at the dilated tip (Fig. 8G). A TEMstudy will be necessary to further elucidate the distribu-tion of mucus and the glycocalyx.

REPTILESClass Reptilia is comprised of the following orders:

Order Testidunata or Chelonia (turtles and tortoises),Order Rhynchocephalia (tuatara), Order Crocodilia (ga-vials, alligators, caimans, and crocodiles), and OrderSquamata or Lepidosauria (lizards and snakes) (Re-mane et al., 1976; Winchester and Jaques, 1981). Asshown in Table 1, all reptiles studied possess a gallblad-der (Guibe, 1970; Remane et al., 1976). Although bothherbivory and carnivory are found in reptiles, many eatinfrequently; the long time interval between their largemeals requires a sudden need for a large amount of bile(Bellairs, 1970). In addition, the diet of the carnivorousreptiles has a high lipid content. The gallbladder and


biliary tract of reptiles has been visualized by means ofgross dissection, light microscopy, some immunocyto-chemical techniques, and ultrasonography (Frye, 1984;Madrid et al., 1989; Penninck et al., 1991), but littleultrastructural research has been done at the TEM andSEM level.

The shape of the liver of reptiles varies with the bodyshape. It is elongated and unlobed in snakes and otherelongate reptiles, has a deep indentation delineating akind of lobation in lizards (Lacertilae), and is clearlydivided into two main lobes in Chelonians and Crocodil-ians (Guibe, 1970). The liver may contain large amountsof melanin pigment. The liver is not separated into thedistinct lobules observed in higher vertebrates, al-though the classic liver lobule has been seen in some

species. In some reptiles, the hepatocytes are arrangedin branching cords, the width of two cells, with the bilecanaliculi present in connective tissue septa-like areasbetween the cords (Elias and Bengelsdorf, 1952; Frye,1984). The gallbladder is usually positioned close to theliver, although in snakes and some lizards, it may befound to be shifted backward and connected to the liverby a long cystic duct (Bellairs, 1970; Frye, 1984). Thegallbladder of the desert tortoise (Xerobates agassizi) isfound in the right lobe of the liver. It has an oval shape,as is found in mammals (Penninck et al., 1991).

Modifications of the duct system may occur. In Chryse-mys, the hepatic duct directly joins the gallbladder,with the bile being evacuated from the gallbladder by acystic duct which joins the duodenum directly behind

Fig. 6. A,B: SEM views of Ambystoma laterale gallbladder epithelium. Shallow epithelial surfacedemonstrating numerous delicate and long microvilli. Bars 5 10 µm in A, 1 µm in B.

Fig. 7. A,B: SEM views of Hyla sp. gallbladder epithelial surface: SEM aspects. Shallowly foldedepithelial surface depicting small (small arrow) to tall (open arrow) clumps of extended microvilli, whichappear like the apices of tuft cells. Bars 5 10 µm in A, 1 µm in B.


the main pancreatic duct (Guibe, 1970). A classic com-mon bile duct draining the cystic duct and the hepaticduct has been observed in some reptiles, namely Ki-

nosternon (Guibe, 1970), but numerous variations maybe present. The liver may be linked directly to theduodenum via a hepatico-interior canal network at the

Fig. 8. Bufo americanus gallbladder epithelium surface: SEMaspects. A: Typical shallow folds of the epithelial surface depict a smallnumber of bulging apices with small (arrows) and large (curved arrow)and deposits. Bar 5 100 µm. B, D, G: Epithelial surfaces demonstrat-ing numerous microvilli clumped by mucus, creating tepee-shapedmicrostructures. The outlines of individual cells (in D) and microvilli

(arrows in G) are blurred by the retention of the mucus. Bars 5 1 µm.C: Area of the surface epithelium devoid of mucus. Bar 5 1 µm. E, F:Surface of epithelium with blobs (in F) and microcrystal depositssimilar to those found in the mammalian gallbladder (in E). Bars 51 µm.


surface of the gallbladder as in Varans (van Dijk, 1935).In the Crocodilians, there is a canal draining thegallbladder, which traverses the pancreas and attachesto the intestine (Guibe, 1970).

The gallbladder of most reptiles demonstrates asimple or pseudostratified columnar epithelium, alamina propria, a fine layer of smooth muscle, andmucin glands or goblet cells. In some reptiles, themicrovilli on the epithelial cells are short and wide. Thegallbladder of the gopher tortoise, Gopherus agassizi,possesses a simple columnar epithelium, lamina pro-pria, fibromuscular tunic, a layer of perimuscular con-nective tissue, and a stroma (Frye, 1984). PAS positivegranules in the cytoplasm and luminal surface ofepithelial cells of the ladder snake and moorish geckosuggest the presence of mucins, sialomucins, and glyco-gen. Lectin histochemistry in the snake suggests thatthe residues on the luminal surface are possibly GlcNAc(N-acetylglucosamine) rather than sialic acid (Madridet al., 1989). Yamada (1960) described a mode ofapocrine mucus secretion for Clemmys japonica (sicChlemys), which resembles a process found in mamma-lians.

Recent observations by the authors (not illustrated)demonstrate that the gallbladder epithelium of theanole is fairly smooth and only shallow, axial longitudi-nal folds are detected. The epithelial cells are of equalsize with numerous long microvilli obscuring the out-lines of the cells. Calculi were found, the largest beingapproximately 20 by 50 µm, resembling the smallgallstones described in humans (Gilloteaux et al., manu-script in preparation) and the Syrian hamster (Gillote-aux et al., 1993a,b; Karkare et al., 1995; Karkare andGilloteaux, 1995).

The gallbladder of the armadillo lizard (Cordyluscataphractus) shows between 8 to 13 longitudinal foldsbut the epithelial surface is functionally more complex,illustrating morphological differences varying with theregion being observed and the phase of the secretorycycle. In Figure 9A, the tall columnar epithelium isobserved to be covered by typical 1-µm-tall microvilli.Interspersed among these microvilli are extremely tallmicrovilli (up to 3.5 µm), collapsed here after fixation,and hence following the landscape of the epithelialsurface. Some of the cell apices demonstrate intensebulging, with only a small number of the long microvilliobservable. The smaller microvilli are no long observed,being obscured by a heavy covering extruded mucus. Inregions with a moderate covering of extruded mucus(Fig. 9C), the epithelium has a more shallow or flatappearance and the cell outline is defined by themicrovilli. Single, extremely tall microvilli are againobserved. Tightly packed clumps of microvilli in dis-persed cell apices (Fig. 9D, E) resemble the tuft cellsseen in the mouse (Luciano and Reale, 1969, 1990,1997) and proposed to be present in the amphibian ofthe genus Bufo (Fig. 7A, B). A higher magnification ofan example of apical surface of one of these cells may beobserved in Figure 9E. An occasional depression (Fig.9F) in the epithelial surface indicates the possibleconclusion of the secretory cycle or remnants of aspecial cell damaged by abrasion. The microvilli areblunted, only 200–300 nm long.

PARASITES OF THE GALLBLADDER IN FISH,REPTILES, AND AMPHIBIANS

Fish, reptiles, and amphibians are hosts to numerousparasites. The relationship may be parasitic or commen-salistic; the animal may be an intermediate or defini-tive host. Parasites may infect the blood, muscle, brain,gut, liver, kidney, heart, lung, spleen, testis, and othertissues. The gallbladder is a target organ for manyparasites. Many of these parasites are host-specific, butsome, such as the myxosporidians (multicellular organ-isms), may be more tissue-specific and show little hostspecificity (Frank, 1984; Poisson, 1953), at least inreptiles and amphibians. A complete account of themany parasites of fish, reptiles, and amphibians is notpossible in this space; a short summary is presentedhere.

One species of the myxosporidian genus Leptothecaparasitizes fish of the genera Scopeaena and Trygon.Probable Leptotheca spores have been found among theepithelial cells of S. scrofa (Fig. 5B, inset), with otherpossible life stages of this parasite found in the gallblad-der lumen in the same fish, as shown at low power TEMin Figure 5C. Higher power TEM shows that theparasites appear either uninucleate or binucleate, de-pending on the plane of sectioning. A ring of condensedmitochondria surround the nucleus (Fig. 5D) and othermitochondria were observed in the cytoplasm alongwith numerous small electron dense granules (the sizeof ribosomes) and glycogen. Whorled, onion-like bodies,as small as 50 nm and as large as 1 µm, appear bothinside and surrounding the parasite. Figure 5B–Dsuggests the appearance of a crawling protozoan, feed-ing on the whorled bodies found on the gallbladderapical surface. These whorled bodies (Fig. 5E) arepostulated to be phospholipids or micelles. Other cyto-plasmic structures are unclear. This fish was fixed infield conditions and it is possible that the parasitesrequire special fixation techniques. The parasites wereunexpected additions to our observations.

Another species of Leptotheca may be found in thegallbladder of Zeus faber. Sphaeromyxa, Zschokella,Chloromyxum, and Mixidium are other myxosporid-ians found in the gallbladders of fish. One myxosporid-ian, Ceratomyxa, is found in the gallbladder of therabbit fish; this parasite is itself parasitized by themicrosporidian Nosema certomyxae (Diamant and Pa-perna, 1989).

Parasitic nematods have also been found in thegallbladders of Uranoscopus (cited in Gilloteaux et al.,1995). The nematode Truttaedacnitis stelmiodes infectsthe bile ducts of the larval brook lamprey (Lampetralamottenii). The intrahepatic bile ducts are dilated andthe apical cell surfaces ranged from those with appear-ances similar to uninfected fish to surfaces with short-ened microvilli, dome-shaped apices, flattened apices,and many with cytoplasmic extrusions (Eng and You-son, 1992).

Many types of protozoan parasites are found inreptiles. The flagellate Hexamita has been found in thebile ducts of the liver and the gallbladder, but its majorpathological effect is on the kidneys. This parasite hasalso been found in amphibians. Eimeria, another typeof protozoan parasite of reptiles and amphibians, in-vades the host by the uptake of spores when the animal


feeds. E. bitis, found in North American snakes, causedthe snake’s gallbladder epithelium to be stripped andthe bile to become viscid. E. cascubeli in the gallbladderand extrahepatic ducts of rattlesnakes caused a prolif-eration of connective tissue and erosion and enlarge-ment of the liver. The young are much more affected bythis parasite; among chameleons in captivity, there wasa high mortality rate for the infected young, while theinfected mother remained apparently healthy (Frank,

1984). Another eimerid species, Tyzzeria chalcides,infects the ocellated skink; eimerids have also beenfound in lizards in Asia (Telford, 1992). Various lifestages of this parasite inhabit the epithelial cells of thebile ducts, the epithelial cells of the gallbladder, and thegallbladder lumen (Probert et al., 1988).

Other types of protozoan parasites are also foundamong the reptilia. Paradistomum geckonum has beenseen in the gallbladder and bile ducts of lizards from

Fig. 9. Armadillo lizard (Cordylus cataphractus) gallbladder sur-face epithelium: SEM aspects. A, B: Surface epithelium demonstrat-ing short and long microvilli, free (in A) or agglomerated by theremaining mucus (in B). A small number of excrescences (*) suggest aprocess of mucus secretory activity similar to that of mammals. Bars 55 µm in A, 1 µm in B. C: Epithelial region without secretory events,depicting an overall short microvillar apical coating. A single, ex-

tremely tall microvillus per apex can also be detected. Bar 5 1 µm. D,E: Tightly packed, tall microvilli (open arrow) of a small number ofcells resembling the tuft cells described in the mouse by Luciano andReale (1969, 1990, and 1997). Bars 5 1 µm. F: Example of depressionin the surface epithelium, showing apex of a cell concluding a secretorycycle or repairing from abrasion of microvilli. Bar 5 1 µm.


Indonesia (Kennedy et al., 1987). Myxosporidians arealso found in reptiles, but seem to be more common inamphibians. They are usually introduced into the hostby the uptake of infected water. Pathogenicity is usu-ally low; the greater the infection, the greater thepathological destruction observed. Leptotheca, Myxobo-lus, Myxosoma, Hemneguya, and Myxidium are allfound in amphibians. Dual infection is also possible;spores of both Myxidium and Hemneguya were found inthe gallbladder of Pipa; the gallbladder had a gelati-nous texture (Frank, 1984). Myxidium lesminteri wasobserved in the gallbladders of three southern Africananurans (Delvinquier et al., 1992). Myxidium immer-sum is a gallbladder parasite of the African cane toad(Delvinquier, 1986; Delvinquier and Freeland, 1988).

In addition to these aforementioned parasites, Nema-thelminth worms can also be found in the biliary tractof specific fishes; an example is Gordiacea worms,which have been observed to occupy some specimens ofUranoscopus gallbladder. In one specimen, we encoun-ter three of these nemathelminths having an average of35 mm in length.

BIRDSThe class Aves forms a diversified group with many

orders (see Table 1). The liver in domestic birds is atubular, bilobed gland (Nickel et al., 1977). The largebile ducts of the penguin demonstrate a columnarepithelium and many tubulo-alveolar glands (Andrewsand Andrews, 1979), although glands are not present inthe large bile ducts and glands of domestic birds (Nickelet al., 1977). The presence or absence of a gallbladder isextremely variable in birds, although it is usually foundin the carnivores of this group (Gorham and Ivy, 1938;Mentzer, 1929a). As shown in Table 1, the gallbladder isabsent in the Struthioniformes, one Charadriiforme(Erolia alpina), one Ralliforme (Truix tanki), most ofthe Columbiformes, most of the Psittaciformes, onegenus (Chordeiles) of the Caprimulgiformes, most of theCoraciadiformes, and a few species of the Passerifor-mes. In the Massai ostrich (Struthio camelus massaicusNeuman), which does not have a gallbladder, fourhepatic ducts join together to form a common hepaticduct, which empties into the duodenum (Mentzer,1929b). The gallbladder in domestic birds may bepear-shaped or tubular and is partially fused to theright lobe of the liver (Nickel et al., 1977). The gallblad-der of the marabou stork (Leptoptilos crumeniferusLesson) is small, with a very short cystic duct (Mentzer,1929b). The gallbladder epithelium of the chick is welldeveloped by the 15th day of embryonic development,demonstrating an epithelium with PAS-positive colum-nar cells and a lamina propria (Nesci and Palermo,1970), structurally similar to adult chickens and mam-mals. It is interesting to note that neutral mucopolysac-charides appear early (14th day of incubation) but arefollowed by sulfated secretory material (15th day ofincubation) in the invaginated epithelial surfaces(Gheri et al., 1988).

MAMMALSThe orders of Class Mammalia are many and diverse

(see Table 1). Carnivory, omnivory, and herbivory areall present. The gallbladder and biliary tracts of mam-

mals are among the most studied of all vertebrategroups.

The classic liver lobule may be observed in mammals.It is especially well demonstrated in the pig, where thelobules are totally surrounded by connective tissue(Stinson and Calhoun, 1976). Unusual cells known asductular hepatocytes have been observed in man andanimals and seem characteristic of a disease state.These cells have an appearance intermediate betweenhepatocytes and intrahepatic bile duct epithelial cells(Sirica, 1995). Developmental studies are difficult tofind in the literature, at least studies concerning itsultrastructure. Needle biopsies are providing new in-sight about the presence of a hepatocyte growth factor(Strain et al., 1995). Contributions of Wood (1965) and,especially, Koga (1971) dealt with morphogenesis ofintrahepatic bile ducts in the human fetus.

Comparative ultrastructure of the terminal branchesof the biliary tree has been examined in the dog, rabbit,rats, and human by Steiner and Carruthers (1961). Athree-dimensional fine structure of the biliary tract(from bile canaliculi to bile ducts) including its vascularsupply has been shown by SEM (Yamamoto et al.,1990). In this journal, Nakanuma et al. (1997) andOhtani and others (1997) have also illustrated theorganization of the blood and lymphatic microvascula-ture of the gallbladder in the guinea pig.

The bile canaliculi in humans have been described byRouiller (1956) but have been well characterized at theultrastructural level later (Rouiller and Jezequel, 1963;Matter et al., 1969). Many aspects relevant to the cellbiology and pathology of the biliary epithelia have beensurveyed in Alpini et al. (1994) and Sirica (1995). Therole of the cytoskeleton, especially actin (Phillips et al.,1982; Watanabe et al., 1983, 1985), has been the objectof attention, especially in regard to its impact onnormal bile flow, permeability of hepatocytes, and itspathologic state in terms of cholestasis (Phillips, 1994;Steiner and Carruthers, 1962). Micrographs obtainedfrom Dr. P. Novikoff comparing an intact and a patho-logic (dilated) bile canalicular networks of rat viewed byconfocal microscopy can illustrate the difference inactin patterns (Fig. 10A vs. B).

The bile ductules of humans, connecting the bilecanaliculi with the intrahepatic bile ducts, containmicrovilli and a single cilium on the ductular cells,possibly having a role in bile flow (Itoshima et al.,1977). Ciliated cells or oligocilia have also been ob-served in the bile ductules of rats, hamsters (Gilloteauxet al., manuscript in preparation), and Yorkshire pigs(Gilroy et al., 1995). In the pig, solitary and oligociliatedcells are found among the epithelial microvilli. Al-though the classic 9 1 2 axonemal structure was notpresent, it was postulated that the cilia served a motileas well as a sensory function (Gilroy et al., 1995). In therat, a bile ductular plexus has been observed, possiblyfunctioning in storage and modification of the bile, asthis mammal does not have a gallbladder (Sirica, 1995;Yamamoto and Phillips, 1984; Yamamoto et al., 1985).

The biliary epithelia biology and development wassurveyed by Alpini et al., (1994) and Hayward (1965),and Koga (1971) has studied some of the developmentalultrastructural aspects. The bile ducts of man andanimals begin as a single layer of cuboidal cells, pro-gressing to a simple columnar epithelium in the larger


ducts (Sirica, 1995). Man, the dog, and the cat havefewer and smaller bile ducts than are found in the rat,guinea pig, and rabbit, with many tubulo-alveolarglands being present in the large ducts, and peribiliaryglands are also found in the intrahepatic tree (Ishida etal., 1989). Most of the bile is secreted into the bilecanaliculi in these groups, while in the rat, guinea pig,and rabbit, which have few to no tubulo-alveolar glands,it is postulated that much bile is secreted by small bileduct cells (Andrews and Andrews, 1979). In man, thelarge intrahepatic bile ducts have extensive sidebranches and pouches in one plane. The pig has pouchesbut no side branches, while the dog and guinea pig haveneither pouches nor side branches (Yamamoto et al.,1985). The intrahepatic duct in mammals occasionallycontains goblet cells; these are especially conspicuousin the guinea pig (Gabe, 1973). The intrahepatic bileduct epithelial cells of the rat have many microvilli,lysosomes, many vesicles near the apical surface, coatedpits and coated vesicles on both the luminal andbasolateral plasma membranes (probable indicators ofreceptor-mediated endocytosis), tight junctions, and anoccasional centriole in the apical cytoplasm (Ishii et al.,1989). The lamina propria of the extrahepatic biliarytract in the rat demonstrates a rich network of bloodvessels under the epithelium, possibly having a role in

reabsorption of water and solutes from the bile, since,as stated previously, this mammal does not have agallbladder (Gaudio et al., 1993). The extrahepatic bileduct of rodents and humans has a weak muscularis, butthe muscularis is quite thick and continuous in rumi-nants (Gabe, 1973). The bile duct of the cow is short andenters the duodenum. The bile ducts of the pig, dog, andcat also enter the duodenum. The bile ducts of goatsand sheep join the pancreatic duct and form a commonbile duct into the duodenum (Stinson and Calhoun,1976).

The hepatic duct and common bile duct in mammalsshow a mucosa, lamina propria, fibromuscular layer,and serosa, as are found in the gallbladder also. Themuscular layer of the ducts is very thin in carnivores,thickest in bovine genera, and discontinuous in somemammals. The hepatic duct in the horse carries uncon-centrated bile directly to the duodenum. The cystic ductin humans has been extensively described by Gillote-aux et al. (1997). Its absence in some humans has beenreviewed by Mentzer (1929b). The aardvark (Oryctero-pus) has been found to have two cystic ducts that fusebefore joining the hepatic duct (Mentzer 1929a). Thecommon bile duct in the rat demonstrates a columnarepithelium consisting of many light cells with micro-villi, numerous vesicles, micropinocytotic images, abun-

Fig. 10. A, B: Bile canaliculi (arrows) from normal rat liver (A) and altered rat liver (B) are delineatedby staining for actin using FITC-labeled phalloidin and imaged by confocal microscopy. Bar 5 10 µm.Reproduced from P.M. Novikoff and M. Cammer, Dept. of Pathology and Image Analysis Facility, AlbertEinstein College of Medicine.


dant microfilaments, junctional complexes, few organ-elles, minimal PAS staining, and small dense granules.Fewer dark cells with no microvilli, and larger lyso-somal-like dark granules were also observed. Electronlucent apical protrusions were also present (Leeson andMelax, 1969; Yamada, 1969). The common bile duct ofthe mouse and hamster, however, demonstrate fewermicrovilli, few indications of micropinocytosis, manyorganelles, fewer microfilaments, abundant secretorygranules, and strong PAS staining, possibly signifying aprotection function from concentrated bile flowing fromthe gallbladder, a function unnecessary in the rat,which does not have a gallbladder (Yamada, 1969).

The gallbladder is absent in some mammals (Table1). Various other gallbladder anomalies have beenreviewed by Mentzer (1929a), including the congenitalabsence in some humans and duplication in cats,giraffe, aardvarks, and humans.

In those mammals that possess a gallbladder, theposition may vary, as may the size and the route bywhich the bile is secreted into the duodenum (Mentzer,1929b). The gallbladder is found embedded completelyin the liver in marsupials and in a few human cases(Mentzer, 1929a). The gallbladder epithelium is formedby tall, columnar cells with numerous microvilli. Darkand light cells are found in the dog (Stinson andCalhoun, 1976) and in the ground squirrel (Pemsingh etal., 1987). Glands and goblet cells may occasionally beobserved between epithelial cells and are particularlywell demonstrated in large ruminants (Stinson andCalhoun, 1976). Histology and basic histochemistry ofmucus production and secretion were reviewed in carni-vores by Seeliger (1938). The mouse gallbladder wasstudied by Meenen and Schiebler (1978), and Laitio andTerho (1975) have studied polysaccharides in the hu-man gallbladder wall and bile.

The hedgehog (Erinaceus europaeus) has a gallblad-der epithelial surface similar to that of other mammals.During hibernation, however, the hedgehog gallbladderepithelium becomes modified. Microvilli are more sparseand irregular. The mitochondria contain electron-densegranules. The basal portion of the cells narrows and asmaller amount of glycogen is observed. The Golgivesicles are very large, indicating unreleased secretoryproducts, and the endoplasmic reticulum is reduced(Allara, 1966).

Examination of the Syrian hamster (Mesocricetusauratus Waterhouse) gallbladder in SEM (Fig. 11A)demonstrates the typical layers consisting of surfaceepithelium, lamina propria, fibromuscular layer, connec-tive tissue or subserosal layer, and serosal layer. TEMand SEM detailed views and descriptions of the surfaceepithelium have been published elsewhere (Gilloteauxet al., 1992, 1993a,b). In Figure 11A, a cross-section wasobtained by scalpel cut; it is possible to show the organwall and detect several blood vessels, i.e., arteries and alarge space delineating a venous return. In the sameview, the mucosal surface shows an artificial spaceresulting from this hand-sectioning. The surface epithe-lium shows typical hexagonal-like epithelial cell out-lines with the apical covering by numerous microvilli,and normal functional pleomorphism, according to therelative amount of mucins coating the microvillar sur-faces and bulging apices, are displayed in Figure 11B–E.Characteristically, the microvilli in the central apical

regions appear the shortest (0.5–0.6 µm), while thoselocated near the cell borders are taller (0.7–0.8 µm). Asmall number of microvilli attaining a height of approxi-mately 1.0–1.2 µm are visible (Fig. 11B–D). In Figure11D, an oblique view of the cell surfaces shows blobs ofmucin located in the grooves between the cells andassociated with the tallest microvilli. When finely coatedby mucus, the microvillar tips can be detected anddisplay a comparable aspect to what was observed inthe amphibians (Fig. 11E). Following induction of chole-lithiasis, and preceding this condition, anionic secre-tory mucus granules are formed, accumulated andsecreted through the apical surfaces as mucus (Gillote-aux et al., 1992, 1993a,b; Karkare et al., 1995; Karkareand Gilloteaux, 1995). The guinea pig (Cavia sp.)gallbladder epithelial cells also produce glycoprotein-aceous-rich (anionic) mucus characterized by Thiery’smethod (Inferrera and Ferlazzo, 1974). Sulfated muco-polysaccharides were also detected in the gallbladderepithelium of the guinea pig and the ram (Tusques etal., 1964).

The human gallbladder reveals an extensively foldedsurface (Pfuhl, 1932), with both primary and secondaryfolds (Fig. 12A) and although the histology and histo-chemistry of mucus have been studied long ago (Gomp-per, 1951; Pfuhl, 1932), it is only with the advent ofelectron microscopy that a more detailed investigationhas been made possible (Bader, 1962, 1965; Evett et al.,1964; Ferner, 1949; Fox, 1972; Laitio and Nevalainen,1972; Wolf-Heidegger et al., 1965). Gallstones disruptthis surface, obliterating these folds. Small bilestonesleave spaces between major folds (Fig. 12B). The epithe-lium of surgical samples reveals the presence of bulgingcells throughout the epithelium (Fig. 12C) and a pleo-morphic surface, with small and large cells present(Fig. 12D). Erosion of the apical surface is also appar-ent, an indication of secretory processes or the abrasivepresence of calculi. Mucus secretion is indicated by thebulging apices of the cells, devoid of microvilli (Fig.12E). Microvilli near the apex are more blunted thanthe elongated microvilli near the cell periphery; smallmucous blebs are present on the periphery. TEM of asimilar field of view shows the large accumulation andfusion of mucus granules (Fig. 12F) near the apex andthe loss of the apical microvilli. High magnificationSEM (Fig. 13A) presents the apices with a delicatecoating of microvilli and a rich glycocalyx as contrastedwith cells undergoing various stages of the secretoryprocess (Fig. 13B). At the conclusion of the secretoryprocess, the apical cells present a crater-like appear-ance (Fig. 13C). Some granular mucus remains and afilm of mucus still coats the microvilli.

TEM of human cholecystic gallbladder epithelialcells (Fig. 14) discloses the tall (15–25 µm in height)columnar cells attached to the basal lamina. Pale anddark cells are attached by junctional complexes in theapical region and coated by 0.2–0.5-µm-long microvillicoated with a glycocalyx, which protrude into thelumen. Large nuclei with a single large nucleolusoccupy the basal portion of the cells. Basolateral exten-sions interdigitate with those of nearby cells (Gillote-aux et al., 1989).

If the gallbladder is absent, its functions are compen-sated for by other parts of the biliary tract. In theProboscidians (elephants), the cystic duct is large and


long and has an internal structure similar to that of thegallbladder of other mammals. Although Cetaceans donot have gallbladder, the biliary canal of Tursiops isdilated to form a large reservoir (Dorst, 1973).

The gallbladder musculature has been restricted tostudies of the dog (MacPherson et al., 1984, 1986) andguinea pig gallbladders (Cai and Gabella, 1983). Hu-man gallbladder musculature was initiated by recon-struction techniques (Scanga et al., 1984) and shows anoverall spiralling of the muscle layer along the longitu-dinal axis of the organ and there are suggestions for

‘‘finely braided muscle bundles’’ making ‘‘reconstructiontedious’’ (MacPherson et al., 1986). This original effortappears to have not helped these investigators andothers to decipher a complex histostructure.

GALLSTONESGallstones, common in humans, are uncommon in

primates, but have been found in the gallbladder ofmarmosets and tamarins, correlated with septation ofthe gallbladder, although one species demonstratedseptation without the presence of calculi (Pissinatti et

Fig. 11. SEM aspects of Syrian hamster (Mesocricetus auratus)gallbladder. A: Highly folded surface epithelium of gallbladder wallcross-sectioned by scalpel dissection depicting all the tunics or layers.b: blood vessel; L: lumen. Open arrow: artefactual space created bydetachment of epithelium from its basement membrane. Bar 5 10 µm.B: Apical surface illustrating the bulging apices covered by numerousshort microvilli and showing a small number of tall microvilli in the

groove delineating adjacent cells. Bar 5 5 µm. C: Detail view ofadjacent cell apices illustrating the rich microvillar coat free of mucus.Bar 5 5 µm. D: Profile of apical surfaces with mucus remainingassociated with the cell boundaries and the tall microvilli, havecreated minute blebs. Bar 5 1 µm. E: Enlarged view of microvillarapices. Scale is 1 µm.


Fig. 12. SEM and TEM views of human gallbladder surface. A:Highly folded surface displaying eroded areas (arrows) as a result ofadhering or gallstones rubbing against the mucosal folded surface.Bar 5 1 mm. B: Primary folds created by the nesting of stones (*).Secondary folds can be detected in the deepest areas. Bar 5 100 µm. C:General view of surface of ‘‘normal’’ mucosal aspect in a cholelithiaticspecimen (notice stands of mucins, small arrows). Bar 5 10 µm. D:General view of epithelium depicting pleomorphism in a cholelithiatic

specimen, which presents adenomatous changes. Notice decapitatedapices following mucus releases. Bar 5 10 µm. E: Bulging apex (*)before mucus release. Notice small mucinous blebs. Bar 5 1 µm. F:TEM view of apical region of a cell similar to the one depicted in Eillustrating the accumulation of apical mucus granules (m), loss ofapical microvilli (arrows) and adjacent endosomal and lysosomalbodies. Bar 5 1 µm.


al., 1992). The presence of gallstones in other primateswas reported in Papio sp., in Aotus, in Callithrixjacchus. Also experimental induction of cholelithiasiswas studied in Saimiri and Pongo (surveyed in Pissi-natti et al., 1992).

Gallstones have affected man since antiquity. Asstated by Schoenfield et al. (1989), gallstones are amajor health problem in the United States and otherWestern countries. Between 16 and 20 million Ameri-cans have gallstones, and about 800,000 to one millionnew cases are discovered each year. In the UnitedStates alone, about 10–20% of the adult populationharbors gallstones (Robbins and Cotran, 1979; Shafferand Small, 1976) and this is not different from thevalues reported in an earlier Canadian study by Fisher(1979) and for the town of Malmo, in Sweden (Lind-

strom, 1971). Their prevalence progressively increasesto reach a peak in the sixth and seventh decades.Women are affected far more than men in ratios of 2:1(Australian) and 4:1 (Bantu and in most developedWestern countries). In addition, a significant and in-creased prevalence of gallstones and related diseases(diabetes, obesity) can be found among the AmericanIndians (e.g., Sampliner et al., 1970), such as the Pima(Comess et al., 1967; Grundy et al., 1972; Hesse, 1947;Lam, 1954) and Chippewa (Thistle and Schoenfield,1970). However, it has been reported that the Masaitribe of Africa does not have gallstones (Biss et al.,1971). In addition to geographic prevalence, there aredeterminants of gallstones prevalence such as age,gender, social class, genetic factors, and clinical associa-tions (pancreatitis, carcinoma, cirrhosis, hemolytic states,

Fig. 13. SEM views of human gallbladder. A: High magnificationexemplifying the numerous apical microvilli of each cell apex finelycoated by the glycocalyx. This rich coat produces a blurry effect of theirtips. Bar 5 1 µm. B: Stages of apical discharge, including an apical

decapitation (*). Bar 5 5 µm. C: Following the secretory process, thelarge amount of remaining mucus (small arrow) clumps the cellsurface and creates a crater-like appearance (open arrows) of the cellapices. Bar 5 10 µm.


chronic ileal diseases, and obesity). Other probableassociations also mentioned by Shaffer and Small (1976)include diet, atherosclerosis, hyperlipidemia, diabetesmellitus, peptic ulcer disease, and other disorders.

Multiple defects are involved in the production ofthese calculi. It appears from the recent reviews pro-vided by Moody et al. (1989), Schoenfield and others(1989), Carey (1993), and Carey and Dwane (1994), aswell as experimental induction of gallstones in animalmodels, that supersaturation of cholesterol, nucleation(mucins, calcium salts, cell debris, etc.), bile defects,and gallbladder hypomotility can result in gallstoneformation. Mucus seems a quite common factor suscep-tible to forming or facilitating a gallstone nucleus (Dotyet al., 1961; LaMont, 1982; LaMont et al., 1983; Lee,1990), and even though cholesterol in gallbladder biledecreased significantly during human weight reduc-tion, both gallbladder mucin and Ca21 concentration

increased with gallstone formation (Shiffman et al.,1992). In the case of pigment stones, hemolytic anemia(Bates and Brown, 1952; Jordan, 1957) provides anexcessive amount of bile pigments and, hence, pigmentstones. In order to study the origin of these stones, amouse model has been investigated (Trotman et al.,1982). Pigment gallstones have been studied exten-sively in Japan because of their high incidence (Suzuki,1966; Trotman, 1991). Calcium bilirubinate salts arecommonly found associated with Escherichia coli andAscaris infections (Maki, 1966; Trotman, 1991). If gall-stones are not caused by bacterial infection, commonbile duct infections can be associated with calculi andcholangitis (Scott, 1971). Recently, molecular geneticshas demonstrated bacterial colonization of gallstones(Swidsinski et al., 1995) and shown that mixed stoneshave the highest incidence of bacterial content. Ultra-structural investigations conducted in our laboratory

Fig. 14. General aspect of human cholecystic gallbladder surface epithelium. Columnar clear cells (C)and dark (D) cells contain mucus secretory granules (m) in their apical zones; some granules containelectron dense cores (large open arrows). d: basolateral digitations; bm: basement membrane. Bar 5 2 µm.Reproduced from Gilloteaux et al. (1989) with permission of the publisher.


on a small number of calculi have shown the presence ofalternate layers of cholesterol and bile pigments withbacteria. Only a small proportion (1:20) of humangallstones examined contained such ultrastructuralevidence (Gilloteaux and Kelly, manuscript in prepara-tion).

Recent developments in medical care have probablycreated other conditions for gallstone formation andhave attracted interest. The mode of induction of thesecalculi is unknown, but they appear to be on the rise inpatients following organ transplantation (Hudson etal., 1992; Lowell et al., 1993; Steck et al., 1991) orconditions of immunosuppression by cyclosporine (Speset al., 1990), in postchemotherapy treatment (Mah-moud et al., 1991), as well as immunoglobulin defi-ciency (Diaz-Buxo et al., 1975).

The morphology and histochemistry of gallstoneshave been studied by Wallraff and Dietrich (1957), andthe stones’ composition varies according to ethnic ori-gin. In Caucasians, for example, mixed stones with highcholesterol and esters of cholesterol content are predomi-nant, while pigment stones are more common in Asiansand the Bantus (Robbins et Cotran, 1979). Crystallo-graphic studies (x-ray diffraction: Juniper and Woolf,1956; x-ray powder method: Sutor and Wooley, 1971)have demonstrated several forms of cholesterol andradially oriented crystals with respect to the nucleus(Bills and Lewis, 1975). Bile acid composition of gall-stones from man was also studied by Schoenfield et al.(1966). The development and growth pattern of solitaryand multiple cholesterol gallbladder stones has beenstudied in an informative paper of Wolpers and Hof-mann (1993). Calcium salts have been observed in thenucleus (Bills and Lewis, 1975; Cowie et al., 1973) aswell as in concentric layers suggesting the sequentialgrowth of the gallstone (Kaufman et al., 1990; Gillo-teaux et al., 1997). Ca-binding proteins have beendetected in bile and gallstones (Kestell et al., 1992) andmay be an important nucleator of cholesterol andbilirubinate in bile (Donovan et al., 1994). A cholesterolcrystallization promoter (Abei et al., 1993) as well asinhibitor factors have been claimed (Ohya et al., 1993).Moreover, in addition to cholesterol, a major componentof hard gallstones, calcium carbonate is also present inthe forms of calcite, aragonite, and/or vaterite (Billsand Lewis, 1975). Other salts, such as calcium bilirubi-nate and palmitate, are also present (Cowie et al., 1973;Gilloteaux et al., 1991; Kaufman et al., 1990). Inaddition to a radiating, scaffolding matrix containingalso Ca phosphates (Sutor and Wooley, 1974), otherpeculiar macromolecular co-aggregates of peculiar ste-rols have been detected, e.g., sapogenins, in the form ofdiosgenin salts (Gilloteaux et al., manuscript in prepa-ration; Troxler, 1986) as well as, surprisingly, siliciumsalts. The nuclei of gallstones are known to be rich inproteinaceous compounds (e.g., immunoglobulins: Af-dhal et al., 1990; Delacroix, 1985; Delacroix et al., 1982;Dive, 1970; Dive and Heremans, 1974; Dive et al., 1974;Harvey et al., 1991; Lipsett et al., 1994) and scanningmicroscopic aspects of cores of cholesterol, mixed andpigment stones have been reported (Gilloteaux et al.,1991).

Animal models such as the mouse (Caldwell et al.,1965; Hayward, 1962a; Laitio and Terho, 1975; Lee,1980; Lee and Scott, 1982; Luciano, 1972; Luciano et

al., 1973; Trotman et al., 1982; Wahlin, 1976, 1977;Ziegler et al., 1982), gerbil (Wahlin, 1979) groundsquirrel (MacPherson et al., 1987; Pemsingh et al.,1988), guinea pig (Brotschi et al., 1984; Geleff and Bock,1984; Hayward, 1962b; Wahlin and Schiebler, 1975;etc.), prairie dog (Haley-Russell et al., 1989; LaMont etal., 1984; Moody et al., 1989), rabbit (Borgman andHaselden, 1969; Lee et al., 1986), and Syrian hamster(Christensen and Dam, 1954; Dam, 1971; Gilloteaux etal., 1992, 1993a,b; Karkare and Gilloteaux, 1995;Karkare et al., 1995) have been used in the study ofgallstone formation. Animal models of calcium choleli-thiasis were reviewed by Hofmann (1984). Hemolyticmodels have also been successfully developed in themouse (Trotman et al., 1982). In the case of the Syrianhamster, it is possible that, besides a metabolic defectassociated with lowering the bile acid pool, a specific‘‘female protein’’ (Coe, 1982) can be induced by femalesex steroids (Coe, 1977; Coe et al., 1981) and couldparticipate as one of the nucleating participants, alongwith mucins and cell debris, to the complex organicmatrix of the gallstones.

SUMMARYThe gallbladder and biliary tract of the vertebrate

groups discussed here have some morphological differ-ences, but a basic similarity persists. The liver in manygroups shows variable degrees of lobulation and doesnot normally demonstrate the cords or plates of hepato-cytes found in mammals, although this feature ispresent in a few species. The lengthy biliary ductulesfound in some fish may be associated with the tubulararchitecture of the fish liver; the mammalian liver isusually correlated with short ductules. Although dataon the biliary ductules of snakes was not found, it ispossible that they too possess long biliary ductules. Thegallbladder in these groups may or may not be contigu-ous with the liver. The gallbladder of the reptiles mostresembles that of mammals, but in all cases encoun-tered, fish display the most diversity. Gallstones can befound not only in humans, but in other mammals and,perhaps, in lower vertebrates. Gallbladders of fishescan be hosts of specific biliary tract parasites.

REFERENCESAbei, M., Kawzak, P., Nuutinen, H., Langnas, A., Svanvik, and

Holzbach, R.T. (1993) Isolation and characterization of a cholesterolcrystallization promoter from human bile. Gastroenterology, 104:539–548.

Afdhal, N.H., Offner, G.D., Murray, F.E., Troxler, R.F., and Smith, B.F.(1990) Isolation and characterization of peptides from the proteincore of bovine gallbladder mucin. Gastroenterology, 98:1633–1641.

Allara, E. (1966) Struttura e ultrastruttura della cistifellea del riccio(Erinaceus europaeus) durante l’ibernazione. Boll. Soc. Ital. Biol.Sperim., 42:936–937.

Alpini, G., Phillips, J.O., and LaRusso, N.F. (1994) The biology ofbiliary epithelia. In: The Liver: Biology and Pathobiology, 3rd. ed.I.M. Arias, J.L. Boyer, N. Fausto, W.B. Jakoby, D.A. Schacter, andD.A. Shafritz, eds. Raven Press, Ltd., New York, pp. 623–653.

Andrews, C.J.H., and Andrews, W.H.H. (1979) The relation betweenstructure and function of bile ducts in man, some laboratory animalsand the Adelie penguin. Q. J. Exp. Physiol., 64:61–67.

Azanza, M.J., Aisa, J., Junquera, C., and Castiella, T. (1989) Theautonomic innervation of the liver and gallbladder of Rana ridi-bunda. Histol. Histopathol., 4:405–410.

Bader, G. (1962) Die submikroskopische Struktur des Gallenblasenepi-thels. II. Mitteilung: Das Epithel der Steingallblase des Menschen.Frankf. Z. Pathol. 74:501–511.

Bader, G. (1965) Die submikroskopische Struktur des Gallenblasenepi-


thels und seiner Regeneration I. Mitteilung: Karpfen (Cyprinuscarpio L.) und Frosch (Rana esculenta L.). Frankf. Z. Pathol.77:92–107.

Bates, G.C., and Brown, C.H. (1952) Incidence of gallbladder diseasein chronic hemolytic anemia (spherocytosis). Gastroenterology, 21:104–109.

Bellairs, A. (1970) The Life of Reptiles. Vol. 1. Universe Books, NewYork, pp. 262–272.

Bentzel, C.J., Hainau, B., Ho, S., Hui, S.W., Edel, A., Anagnostopoulos,T., and Benedetti, E.L. (1980) Cytoplasmic regulation of tight-junction permeability: Effect of plant cytokinins. Am. J. Physiol.,239:C75–C89.

Bentzel, C.J., Fromm, M., Palant, C.E., and Hegel, U. (1987) Prot-amine alters structure and conductance of Necturus gallbladdertight junctions without major electrical effects on the apical cellmembrane. J. Membr. Biol., 95:9–20.

Bertin, L. (1958) Appareil digestif. In: Traite de Zoologie, Tome XIIIFasc. 2, Agnates et Poissons. Masson & Cie, Paris, pp. 1288–1302,1710–1754.

Biagianti-Risbourg, S. (1990) Contribution a l’etude du foie de juve-niles de muges (Teleosteens, Mugilidae) contamines experimentale-ment par l’atrazine (S-triazine herbicide): approche ultrastructuraleet metabolique: Interet en ecotoxicologie. Thesis, Universite dePerpignan, France.

Bills, P.M., and Lewis, D. (1975) A structural study of gallstones. Gut,16:630–637.

Biss, K., Ho, K.-J., Mikkelson, B., Lewis, L., and Taylor, B. (1971)Some unique biologic characteristics of the Masai of East Africa. N.Engl. J. Med., 284:694–699.

Bluntschli, H. (1904) Der feinere Bau der Leber von Ceratodus,zuglich ein Beitrag zur vergleichenden Histologie der Fishleber.Jena Denkschr., 4:335–375.

Borgman, R.F., and Haselden, F.H. (1969) Cholelithiasis in rabbits:Effects of bile constituents and hormones on dissolution of gall-stones. Am. J. Vet. Res., 30:107–112.

Brotschi, E.A., LaMorte, W.W., and L.F. Williams (1984) Effect ofdietary cholesterol and indomethacin on cholelithiasis and gallblad-der motility in guinea pig. Dig. Dis. Sci., 29:1050–1056.

Burggren, W.W., and Just, J.J. (1992) Developmental changes inphysiological systems. In: Environmental Physiology of the Amphib-ians. M.E. Feder and W.W. Burggren, eds. The University of ChicagoPress, Chicago, pp. 467–530.

Cai, W., and Gabella, G. (1983) Innervation of the gall bladder andbiliary pathways in the guinea pigs. J. Anat. (Lond.), 136:97–109.

Caldwell, F.T., Levitsky, K., and B. Rosenberg (1965) Dietary produc-tion and dissolution of cholesterol gallstones in the mouse. Am. J.Physiol., 209:473–478.

Carey, M.C. (1993) Pathogenesis of gallstones. Am. J. Surg., 165:410–419.

Carey, M.C., and Duane, W.C. (1994) Enterohepatic circulation. In:The Liver: Biology and Pathobiology, 3rd. ed. I.M. Arias, J.L. Boyer,N. Fausto, W.B. Jakoby, D.A. Schacter, and D.A. Shafritz, eds.Raven Press, Ltd., New York, pp. 719–767.

Christensen, F., and Dam, H. (1954) Alimentary production of gall-stones in hamsters. Acta Physiol. Scand., 31:75–82.

Coe, J.E. (1977) A sex-limited serum protein of Syrian hamsters:Definition of female protein and regulation by testosterone. Proc.Natl. Acad. Sci. U.S.A., 74:730–733.

Coe, J.E. (1982) Female protein of the Syrian hamster: A homolog ofC-reactive protein. Ann. NY Acad. Sci., 389:299–307.

Coe, J.E., Margossian, S.S., Slayter, H.S., and J.A. Sogn (1981)Hamster female protein. A new pentraxin structurally and function-ally similar to C-reactive protein and amyloid P component. J. Exp.Med., 153:977–991.

Comess, I.J., Benett, P.H., and Burch, T.A. (1967) Clinical gallbladderdisease in Pima Indians: Its high prevalence in contrast to Framing-ham, Massachusetts, N. Engl. J. Med., 277:894–898.

Cowie, A.G.A., Sutor, D.J., Woolery, S.E., and Clark, C.G. (1973) Thecalcium palmitate gall-stone. Br. J. Surg., 60:16–18.

Dam, H. (1971) Determinants of cholesterol cholelithiasis in man andanimals. Am. J. Med., 51:596–613.

Delacroix, J.P. (1985) The human immunoglobulin: A system, itsvascular compartment. Thesis, Louvain, Fac. Medicine, EuropeanMedical Press, Brugge, pp. 50–71.

Delacroix, J.P., Hodgson, H.J.F., McPherson, A., Dive, C., and Vaer-man, J.P. (1982) Selective transport of polymeric IgA in bile.Quantitative relationships of monomeric and polymeric IgA, IgM,and other proteins in serum, bile and saliva. J. Clin. Invest.,70:230–241.

Delvinquier, B.L.J. (1986) Myxidium immersum (Protozoa: Myxo-

sporea) of the can toad, Bufo marinus, in Australian Anura, with asynopsis of the genus in amphibians. Aust. J. Zool., 34:843–854.

Delvinquier, B.L.J., and Freeland, W.J. (1988) Protozoan parasites ofthe cane toad, Bufo marinus, in Australia. Aust. J. Zool., 36:301–316.

Delvinquier, B.L.J., Markus, M.B., and Passmore, N.I. (1992) Myx-idium lesminteri, new species (Myxosporea: Myxidiidae) from thegallbladder of three southern African Anura. Sys. Parasitol., 23:25–30.

Diamant, A., and Paperna, I. (1989) Cytopathology of Ceratomyxa sp.(Myxosporea) hyperparasitized with the microsporidian Nosemaceratomyxae. Dis. Aquat. Org., 6:75–79.

Diaz-Buxo, J.A., Hermans, P.E., and L.R. Elveback (1975) Prevalenceof cholelithiasis in idiopathic late-onset immunoglobulin deficiency.Ann. Intern. Med., 82:213–214.

Dive, C. (1970) Les proteines de la bile. Leur composition et leurorigine. Thesis Louvain, Arscia Publ., Brussels, pp. 105–169.

Dive, C., and Heremans, J.F. (1974) Nature and origin of the protyeinsin bile. I. A comparative analysis of serum and bile proteins in man.Eur. J. Clin. Invest., 4:235–239.

Dive, C., Nadalini, R.A., Vaerman, J.P., and Heremans, J.F. (1974)Origin and nature of the proteins of bile. II. Comparative analysis ofserum, hepatic lymph and bile proteins in the dog. Eur. J. Clin.Invest., 4:241–246.

Donovan, J.M., Leonard, M.R., Batta, A.K., and Carey, M.C. (1994)Calcium affinity for biliary aggregates in model biles: Complemen-tary importance of bile saklts and lecithin. Gastroenterology, 107:831–846.

Dorst, J. (1973) Appareil digestif et annexes. In: Traite de Zoologie,Tome XVI, Mammiferes, Fasc. 5A, Vol. 1, Masson & Cie, Paris, pp.250–382.

Doty, J.E., Pitt, H.I., Kuchenbecker, S.L., Portel-Fink, V., and Berter,L.D.W. (1961) Role of gallbladder mucus in the pathogenesis ofcholesterol gallstones. Am. J. Surg., 145:54–61.

Duellman, W.E., and Trueb, L. (1986) Biology of Amphibians. TheJohns Hopkins University Press, Baltimore, pp. 408–411.

Eales, J.G., Welsh, L.A., and Chan, H.H. (1971) Biliary excretion of3,5,38-Triiodo-L-thyronine-125I by the brook trout, Salvelinus fontina-lis (Mitchill). Gen. Comp. Endocrinol., 16:169–175.

Elias, H., and Bengelsdorf, H. (1952) The structure of the liver ofvertebrates. Acta Anat., 14:297–337.

Elliott, W.M., and Youson, J.H. (1993) Development of the adultendocrine pancreas during metamorphosis in the sea lamprey,Petromyzon marinus L. II. Electron microscopy and immunocyto-chemistry. Anat. Rec., 237:271–290.

Eng, F., and Youson, J.H. (1992) Morphology of the bile ducts of thebrook lamprey, Lampetra lamottenii (Le Sueur) and during infectionwith the nematode, Truttaedacnitis stelmiodes (Vessichelli, 1910)(Nematoda: Cucullanidae). Anat. Rec., 234:201–214.

Evett, R.D., J.A. Higgins, and A.L. Brown Jr. (1964) The fine structureof normal mucosa in human gallbladder. Gastroenterology, 47:49–60.

Ferner, H. (1949) Ueber das Epithel der menschlichen Gallenblase. Z.Zellforsch. 34:503–513.

Fisher, M.M. (1979) Perspectives on gallstones. In: Gallstones. M.M.Fisher, C.A. Goresky, E.A. Shaffer, and S.M. Strasberg, eds. PlenumPress, New York, pp. 1–17.

Foskett, J.K., and Spring, K.R. (1985) Involvement of calcium andcytoskeleton in gallbladder epithelial cell volume regulation. Am. J.Physiol., 248:C27–C36.

Fox, H. (1972) Ultrastructure of the human gallbladder epithelium incholelithiasis and chronic cholecystitis. J. Pathol., 108:157–164.

Frank, W. (1984) Non-hemoparasitic protozoans. In: Diseases ofAmphibians and Reptiles. G.L. Hoff, F.L. Frye, and E.R. Jacobson,eds. Plenum Press, New York, pp. 259–384.

Frye, F.L. (1984) Euthanasia, Necroscopy Techniques, and Compara-tive Histology of Reptiles. In: Diseases of Amphibians and Reptiles.G.L. Hoff, F.L. Frye, and E.R. Jacobson, eds. Plenum Press, NewYork, pp. 703–756.

Gabe, M. (1973) Anatomie microscopique de l’appareil digestif desmammiferes. In: Traite de Zoologie, Tome XVI, Fasc. 5 A, Vol. 1,Masson & Cie, Paris, pp. 383–483.

Gaudio, E., Onori, P., Marinozzi, P., and Marinozzi, G. (1993) Microcir-culation of the extrahepatic biliary tree: A scanning electron micros-copy study of corrosion casts. J. Anat., 182:37–44.

Geleff, S., and Bock, P. (1984) Lectin binding of surface epithelia andconcomitant glands of gallbladder and biliary ducts in the guineapig. Histochemistry, 81:543–549.

Gheri, G., Gheri Bryk, S., and Orlandini, G.E. (1988) Histochemistryof mucosubstances in the gallbladder epithelium of the chickembryo. Histochemistry, 88:519–524.


Gilloteaux, J., Pomerants, B., and Kelly, T.R. (1989) Human gallblad-der mucosa ultrastructure: Evidence of intraepithelial nerve struc-tures. Am. J. Anat., 184:323–335.

Gilloteaux, J., Hawkins, W.S., and T.R. Kelly (1991) Scanning electronmicroscopic study of human gallstone cores. J. Scan. Microsc.,13(Suppl. I):114–115.

Gilloteaux, J., Karkare, S., Ko, W., and Kelly, T.R. (1992) Female sexsteroid induced epithelial changes in the gallbladder of the ovariec-tomized Syrian hamster. Tissue & Cell., 24:869–878.

Gilloteaux, J., Kosek, E., and Kelly, T.R. (1993a) Sex steroid inductionof gallstones in the male Syrian hamster. J. Submicrosc. Cytol.Pathol., 25:157–172.

Gilloteaux, J., Kosek, E., and Kelly, T.R. (1993b) Epithelial surfacechanges and induction of gallstones in the male Syrian hamstergallbladder as a result of two-month sex steroid treatment. J.Submicrosc. Cytol. Pathol., 25:519–533.

Gilloteaux, J., Karkare, S., and Kelly, T.R. (1993c) Apical excrescencesin the gallbladder epithelium of the female Syrian hamster inresponse to medroxyprogesterone. Anat. Rec., 236:479–489.

Gilloteaux, J., Hawkins, W.S., Gilloteaux, L.C., and Kelly, T.R. (1996)Ultrastructural aspects of human cystic duct epithelium as a resultof cholelithiasis and cholesterolosis. Microsc. Res. Tech., 38:643–659.

Gilroy, C., Singh, A., and Shahidi, E. (1995) Cilia in the porcine bileductule: Motile or sensory? Histol. Histopathol., 10:301–304.

Gompper, H. (1951) Ueber das schleimartige Sekret der Gallenblase.Z. Zellforsch., 57:280–303.

Gonzalez, G., Crespo, S., and Brusle, J. (1993) Histo-cytological studyof the liver of the cabrilla sea bass, Serranus cabrilla (Teleostei,Serranidae), an available model for marine fish experimental stud-ies. J. Fish Biol., 43:363–373.

Gorham, F.W., and Ivy, A.C. (1938) General function of the gall bladderfrom the evolutionary standpoint. Field Mus. Nat. Hist., Zool. Ser.,22:159–213.

Grasse, P.-P., and Devillers, Ch. (1965) Zoologie II. Vertebres. Massonet Cie publ., Paris.

Grundy, S.M., Duane, W.C., Adler, R.D., Aron, J.M., and Metzger, A.L.(1972) Mechanisms of lithogenic bile formation in American Indianwomen with cholesterol gallstones. J. Clin. Invest., 51:3026–3043.

Grzimek, B. (1972–1975) Grzimek’s Animal Life Encyclopedia. VanNostrand Reinhold Co. Pub. New York, Vol. 4 (Fish) to Vol. 13(Mammals).

Grzycki, S., and Nowakowski, A. (1967) The histotopochemical obser-vation of monoamine oxidase (MAO) in the gall bladder epitheliumof the frog and mouse. Z. Mikr. Anat. Forsch., 77:611–616.

Guibe, J. (1970) L’appareil digestif. In: Traite de Zoologie. P.-P. Grasse,ed. Masson & Cie, Paris, Tome XIII, Agnates et Poissons, pp.521–548.

Haley-Russell, D., Husband, K.J., and Moody, F.G. (1989) Morphologyof the prairie dog gallbladder: Normal characteristics and changesduring early lithogenesis. Am. J. Anat., 186:133–143.

Hampton, J.A., Lantz, R.C., Goldblatte, P.J., Lauren, D.J., and Hinton,D.E. (1988) Functional units in rainbow trout (Salmo gairdneri,Richardson) liver. II. The biliary system. Anat. Rec., 221:619–634.

Harvey, P.R., Upadhya, G.A., and Strasberg, S.M. (1991) Immunoglobu-lins as nucleating proteins in the gallbladder bile of patients withcholesterol gallstones. J. Biol. Chem., 166:13996–14003.

Hayward, A.F. (1962a) Aspects of the fine structure of the gall bladderepithelium of the mouse. J. Anat. (Lond.) 96:227–236.

Hayward, A.F. (1962b) Electron microscopic observations on absorp-tion in the epithelium of the guinea pig gall bladder. Z. Zellforsch.,56:197–202.

Hayward, A.F. (1965) The fine structure of the gallbladder epitheliumof the sheep. Z. Zellforsch., 65:331–339.

Hesse, F.G. (1947) Incidence of cholecystitis and other diseases amongPima Indians of Southern Arizona. J.A.M.A., 170:1789–1790.

Higashi, K., Sasa, S., Gomi, T., Kimura, A., Tsuchiya, H., andHashimoto, T. (1986) Morphological study of the lungish (Protop-terus sp.) gallbladder. J. Med. Soc. Toho Univ., 33:237–246.

Hill, A.E., and Shachar-Hill, B. (1993) A mechanism for isotonic fluidflow through the tight junctions of Necturus gallbladder epithelium.J. Membr. Biol., 136:253–262.

Hofmann,A.F. (1984)Animal models of calcium cholelithiasis. Hepatol-ogy, 4:209S–211S.

Honrubia, M.P., Paz Herraez, M., and Alvarez, R. (1993) The carba-mate insecticide ZZ-Aphox induced structural changes of gills, liver,gall-bladder, heart and notochord of Rana perezi tadpoles. Arch.Environ. Contamin. Toxicol., 25:184–191.

Hudson, H.M., Hakaim, A.G., and Birkett, D.H. (1992) Laparoscopiccholecystectomy in a renal transplant recipient. Surg. Endosc.,6:193–194.

Hyrtl, C.J. (1868) Uber Ampulla am Ductus cysticus der Fisches.Denkschr. K. Akad. Wiss. Wien (Math. Naturwiss.), 28:185–190.

Inferrera, C., and Ferlazzo, A. (1974) Cytochemical observations in theelectron microscope of the gall bladder epithelium. J. Submicrosc.Cytol., 6:261–274.

Ishida, F., Terada, T., and Nakanuma, Y. (1989) Histologic andscanning electron microscopic observations of intrahepatic peribili-ary glands in normal human livers. Lab. Invest., 60:260–265.

Ishii, M. Vroman, B., and LaRusso, N. (1989) Isolation and morpho-logic characterization of bile duct epithelial cells from normal ratliver. Gastroenterology, 97:1236–1247.

Itoshima, T., Yoshino, K., Yamamoto, K., Ohta, W., Kubota, M., Ukida,M., Ito, T., Hirakawa, W., Munetomo, F., and Shimada, Y. (1977)Scanning electron microscopy of the bile ductule. Gastroenterol.Jpn., 12:476–482.

Jordan, R.A. (1957) Cholelithiasis in sickle cell disease. Gastroenterol-ogy, 33:952–958.

Juniper, K., and Woolf, W.E. (1956) Biliary tract studies. 1. X-raydiffraction analysis; correlation with occurrence of microcrystals inbile. Am. J. Med., 20:383–391.

Karkare, S., and Gilloteaux, J. (1995) Gallstones induced by sexsteroids in the female Syrian hamster: Duration effects. J. Submi-crosc. Cytol. Pathol., 27:53–74.

Karkare, S., Kelly, T.R., and Gilloteaux, J. (1995) Morphologicalaspects of female Syrian hamster gallbladder induced by one-monthsex steroid treatment. J. Submicrosc. Cytol. Pathol., 27:35–52.

Kaufman, H.S., Littlemoe, K.D., Magnuson, T.H., Frasca, P., and Pitt,H.A. (1990) Backscattered electron imaging and windowless energydispersive X-ray microanalysis: A new technique for gallstoneanalysis. Scanning Microsc., 4:853–862.

Kennedy, M.J., Killick, L.M., and Beverley-Burton, M. (1987) Theprevalence of Paradistomum geckonum, Mesocoelium sociale, andPostorchigenes ovatus (Digenea) in lizards (Sauria) from Indonesia.Can. J. Zool., 65:1291–1294.

Kestell, M.F., Sekijima, J., Lee, S.P., Park, H.Z., Long, M., and Kaler,E.W. (1992) A calcium-binding protein in bile and gallstones.Hepatology, 19:1315–1321.

Koga, A. (1971) Morphogenesis of intrahepatic bile ducts of the humanfetus. Z. Anat. Entwicklungsgesch., 135:156–184.

Laitio, M., and Nevalainen, T. (1972) Scanning and transmissionelectron microscope observations on human gallbladder epithelium.I. Adult structure. Z. Anat. Entwicklungsgesch., 136:319–325.

Laitio, T., and Terho, M. (1975) Polysaccharides of metaplastic mousegallbladder mucosa and carcinoma of the gallbladder. Lab. Invest.,32:183–189.

Lam, R.C. (1954) Gallbladder diseases among the American Indians.Lancet, 74:305–309.

LaMont, J.T. (1982) Gallbladder mucin glycoprotein hypersecretion inexperimental cholelithiasis: Role of mucin gel in nucleation ofcholesterol gallstones. Adv. Exp. Med. Biol., 144:231–234.

LaMont, J.T., Ventola, A.S., Trotman, B.W., and Soloway, R.D. (1983)Mucin glycoprotein content of human pigment gallstones. Hepatol-ogy, 3:377–382.

LaMont, J.T., Smith, B.F., and Moore, J.R.L. (1984) Role of gallbladdermucin in pathophysiology of gallstones. Hepatology, 4:51S–56S.

Larson, M., and Spring, K.R. (1984) Volume regulation by Necturusgallbladder: Basolateral KCl exit. J. Membr. Biol., 81:219–232.

Lee, S.P. (1980) The mechanism of mucus secretion by the gallbladderepithelium. Br. J. Exp. Pathol., 61:117–119.

Lee, S.P. (1990) Role of mucus in sludge and stone formation. In:Current Topics in Gastroenterology and Hepatology. Georg ThiemeVerlag, Stuttgart, pp. 403–414.

Lee, S.P., and Scott, A.J. (1982) The evolution of morphologicalchanges in the gallbladder before stone formation in mice fed acholesterol-cholic acid diet. Am. J. Pathol., 108:1–8.

Lee, S.P., Tasman-Jones, C., and Carlisle, V.F. (1986) Oleic acid-induced cholelithiasis in rabbits. Am. J. Pathol., 124:18–24.

Leeson, T.S., and Melax, H. (1969) Fine structure of the common bileduct in adult rats. Can. J. Zool., 47:33–35.

Li, W., Sorensen, P.W., and Gallaher, D.D. (1995) The olfactory systemof migratory adult sea lamprey (Petromyzon marinus) is specificallyand acutely sensitive to unique bile acids released by conspecificlarvae. J. Gen. Physiol., 105:569–587.

Lindstrom, C.G. (1971) The frequency of gallstone-disease in the townof Malmo. Acta Path. Microbiol. Scand., Sect A., 79:311–312.

Lipsett, P.A., Hildreth, J., Kaufman, H.S., Littlemoe, K.D., and Pitt,H.A. (1994) Human gallstones contain pronucleating nonmucinglycoproteins that are immunoglobulins. Ann. Surg., 219:25–33.

Loeschke, K., and Bentzel, C.J. (1994) Osmotic water flow pathwaysacross Necturus gallbladder: Role of the tight junction. Am. J.Physiol., 266:G722–G730.


Lowell, J.A., Stratta, R.J., Taylor, R.J., Bynon, J.S., Larsen, J.L., andNelson, N.L. (1993) Cholelithiasis in pancreas and kidney trans-plant recipients with diabetes. Surgery, 114:858–863.

Luciano, L. (1972) Die Feinstruktur der Gallenblase und der Gallen-gange. I. Das Epithel der Gallenblase der Maus. Z. Zellforsch.,135:87–102.

Luciano, L., and Reale, E. (1969) A new cell type (‘‘brush cell’’) in thegall bladder epithelium of the mouse. J. Submicrosc. Cytol., 1:43–52.

Luciano, L., and Reale, E. (1990) Brush cells of the mouse gallbladder.A correlative light- and electron-microscopical study. Cell TissueRes., 262:339–349.

Luciano, L., and Reale, E. (1997) The presence of brush cells in themouse gallbladder. Microsc. Res. Tech., 38:598–608.

Luciano, L., Wermbter, G., and Reale, E. (1973) Die Feinstruktur derGallenblase und der Gallengange. III. Beobachtungen an gefriergeat-zen Preparaten der Gallenblase der Maus unter besonderer Beruck-sichtigung der dichten Korper und des Verbindunskomplexes. Cyto-biologie, 7:76–88.

MacPherson, B.R., Scott, G.W., Chansouria, J.P.N., and Fisher, A.W.F.(1984) The muscle layer of the canine gallbladder and cystic duct.Acta Anat., 120:117–122.

MacPherson, B.R., Yiu, V., Lee, W., and Scott, G.W. (1986) A scanningelectron-microscopic study of the muscle layer of the canine gallblad-der. Acta Anat., 127:59–64.

MacPherson, B.R., Pemsingh, R.S., and Scott, G.W. (1987) Experimen-tal cholelithiasis in the ground squirrel. Lab. Invest., 56:138–145.

Madrid, J.F., Ballesta, J., Galera, T., Castells, M.T., and Perez-Tomas,R. (1989) Histochemistry of glycoconjugates in the gallbladderepithelium of ten animal species. Histochemistry, 91:437–443.

Mahmoud, H., Schell, M., and Pui, C.H. (1991) Cholelithiasis aftertreatment for childhood cancer. Cancer, 67:1439–1442.

Maki, T. (1966) Pathogenesis of calcium bilirubinate gallstone: Role ofE. coli glucuronidase and coagulation by inorganic ions, polyelectro-lytes and agitation. Ann. Surg., 164:90–100, 91:437–443.

Matter, A., Orci, L., and Rouiller, C. (1969) A study on the permeabilitybarriers between Disse’s space and the bile canaliculus. J. Ultra-struct. Res., (Suppl) 11:1–71.

Meenen, N.M., and Schiebler, T.H. (1978) Zur Entwicklung desGallenblasenepithels der Maus. Anat. Anz., 144:407–428.

Mentzer, S.H. (1929a) Anomalous bile ducts in man based on a study ofcomparative anatomy. J.A.M.A., 93:1273–1277.

Mentzer, S.H. (1929b) Comparative anatomy of the biliary system.Calif. West. Med., 30:315–321.

Moody, F.G., Haley-Russel, D., Li, Y.F., Husband, K.J., Weisbrodt,N.W., and Dewey, R.B. (1989) The effect of lithogenic bile ongallbladder epithelium. Ann. Surg., 210:406–416.

Nakanuma, Y., Hoso, M., Terada, T., Sanzen, T., and Sasaki, M. (1997)Microstructure and development of the normal and pathologicbiliary tract in humans, including blood supply. Microsc. Res. Tech.,38:552–570.

Nesci, E., and Palermo, D. (1970) Osservazioni sul differenziamentodell’epitelio di rivestimento della colecisti durante lo sviluppo delGallus dom. Boll.-Soc. Ital. Biol. Sperim., 46:264–265.

Nevalainen, T.J. (1977) Ultrastructural characteristics of tuft cells inmouse gallbladder epithelium. Acta Anat., 98:210–220.

Nickel, R., Schummer, A., Seiferle, E., Siller, W.G., and Wight, P.A.L.(1977) Anatomy of the Domestic Birds, Verlag Paul Parey, Berlin,pp. 57–60.

Nodzenski, E., Waserssug, R.J., and Inger, R.F. (1989) Developmentaldifferences in visceral morphology of megophryine pelobatid tad-poles in relation to their body form and mode of life. Biol. J. LinneanSoc., 38:369–388.

Ohtani, O., Lee, M.-H., Wang, Q.-X., and Uchino, S. (1997) Theorganization of the blood and lymphatic microvasculature of thegallbladder in the guinea pig: A scanning electron microscopic study.Microsc. Res. Tech., 38:660–666.

Ohya, T., Swarzendrube, J., Busch, N., Gresky, S., Chandler, K.,Takabayashi, A., Igimi, H., Egami, K., and Holzbach, R.T. (1993) Ahuman glycoprotein that inhibits formation and growth of choles-terol crystals: Purification and characterization. Gastroenterology,104:527–538.

Palant, C.E., Duffey, M.E., Mookerjee, B.K., and Bentzel, C.J. (1983)Ca21 regulation of tight-junction permeability and structure inNecturus gallbladder. Am. J. Physiol., 245:C203–C212.

Peek, W.D., Sidon, E.W., Youson, J.H., and Fisher, M.M. (1979) Finestructure of the liver in the larval lamprey, Petromyzon marinus L.;hepatocytes and sinusoids. Am. J. Anat., 156:231–250.

Pemsingh, R.S., MacPherson, B.R., and Scott, G.W. (1987) Ultrastruc-tural observations on the gallbladder of Richardson’s ground squir-rel (Spermophilus richardsonii). Acta Anat., 129:179–181.

Pemsingh, R.S., MacPherson, B.R., and Scott, G.W. (1988) Character-

ization of lipid accumulation in the gallbladder mucosa of theground squirrel fed a lithogenic diet. J. Pathol., 154:173–180.

Penninck, D.G., Stewart, J.S., Paul-Murphy, J., and Pion, P. (1991)Ultrasonography of the California desert tortoise (Xerobates agas-sizi): Anatomy and application. Vet. Radiol., 32:112–116.

Pfuhl, W. (1932) Die Leber, die Gallenblase und die extrahepatischenGallengange. Mollendorffs Hand. Micro. Anat., 5:235–462.

Phillips, M.J. (1994) Biology and pathobiology of actin in the liver. In:The Liver: Biology and Pathobiology, 3rd. ed. I.M. Arias, J.L. Boyer,N. Fausto, W.B. Jakoby, D.A. Schachter and D.A. Shafritz, eds.Raven Press Ltd., New York, pp. 19–32.

Phillips, M.J., Oshio, C., Miyairi, M., Katz, H., and Smith, C.R. (1982)A study of bile canalicular contractions in isolated hepatocytes:Methods and time series analysis. Hepatology, 2:763–768.

Pissinatti, A., Batista da Cruz, J., Dias do Nascimento, M., do Rocha eSilva, R., and Coimbra-Filho, A.F. (1992) Spontaneous gallstones inmarmosets and tamarins. Folia Primatol., 59:44–50.

Poisson, R. (1953) Sous-Embranchement des Cnidosporidies. In: Traitede Zoologie, Protozoaires, Rhizopodes, Actinopodes, Sporozoaires,Cnidoporides. P.-P. Grasse, ed. Masson & Cie, Paris, Tome 1, pp.1008–1088.

Probert, A.J., Roberts, A.H., and Wilson, I.B. (1988) Tyzzeria chalcides,new species from the ocellated skink, Chalcides ocellatus. J. Proto-zool., 35:211–213.

Remane, A., V. Starch, and Welsch, U. (1976) Systematische Zoologie.Stamme des Tierreichs. Gustav Fisher Verlag, Stuttgart.

Reuss, L. (1979) Electrical properties of the cellular transepithelialpathway in Necturus gallbladder: III. Ionic permeability of thebasolateral cell membrane. J. Membr. Biol., 47:239–259.

Reuss, L. (1983) Basolateral KCl co-transport in a NaCl-absorbingepithelium. Nature (London), 305:723–726.

Reuss, L., Weinman, S.A., and Grady, T.O. (1980) Intracellular K1

activity and its relation to basolateral membrane ion transport inNecturus gallbladder epithelium. J. Gen. Physiol., 76:33–52.

Robbins, S.C., and Cotran, R.S. (1979) Pathologic Basis of Disease. 2nded. W.B. Saunders, Philadelphia, pp. 1071–1080.

Rostgaard, J., and Frederiksen, O. (1981) Fluid transport and dimen-sions of epithelial cells and intercellular spaces in frog gallbladder.Cell Tissue Res., 215:223–247.

Rouiller, C. (1956) Les canalicules biliaires. Acta Anat., 26:94–109.Rouiller, C., and Jezequel, A.M. (1963) Electron microscopy of the liver.

In: The Liver, vol. 1. C. Rouiller, ed. Academic Press, New York, pp.195–264.

Ruiter, A.J.H., Veenhuis, M., and Wendelaar Bonga, S.E. (1988)Peroxisomes in the intestinal and gallbladder epithelial cells of thestickleback, Gasterosteus aculeatus L. (Teleostei). Cell Tissue Res.,251:685–689.

Sampliner, R.E., Benett, P.H., Comess, L.J., Rose, F.A., and Burch,T.A. (1970) Gallbladder disease in Pima indians. N. Engl. J. Med.,283:1358–1364.

Scanga, D.G., Scott, C.J., MacPherson, B.R., Clanachan, A.S., andScott, G.W. (1984) The structural organization and contractility ofhuman gallbladder muscle. Dig. Dis. Sci., 29:363–378.

Schoenfield, L.J. (1977) Diseases of the Gallbladder and BiliarySystem. John Wiley & Sons, New York, pp. 1–80.

Schoenfield, L.J., Sjovall, J., and Sjovall, K. (1966) Bile acid composi-tion of gallstones from man. J. Lab. Clin. Med., 68:186–194.

Schoenfield, L.J., Carey, M.C., Marks, J.W., and Thistle, J.T. (1989)Gallstones: An update. Am. J. Gastroenterol., 84:999–1007.

Scott, A.J. (1971) Bacteria and disease of the biliary tract. Gut,12:487–492.

Seelinger, M. (1938) Ueber den Bau des Gallengangsystems beidenCarnivoren (Hund und Katze) mit besonderer Berucksichtigung derSchleimbildung und des Glykogen gehaltes. Z. Zellforsch. 26:578–602.

Shachar-Hill, B., and Hill, A.E. (1993) Convective fluid flow throughthe paracellular system of Necturus gallbladder epithelium asrevealed by dextran probes. J. Physiol. (Lond.), 468:463–486.

Shaffer, E.A., and Small, D.M. (1976) Gallstone disease: Pathogenesisand management. Current Prob. Surg., 13:1–72.

Shiffman, M.L., Sugerman, H.J., Kellum, J.M., and Moore, E.W. (1992)Changes in gallbladder bile composition following gallstone forma-tion and weight reduction. Gastroenterology, 103:214–221.

Sidon, E.W., Peek, W.D., Youson, J.H., and Fisher, M.M. (1980) Finestructure of the liver in the larval lamprey, Petromyzon marinus L.;bile ducts and gall bladder. J. Anat., 131:501–519.

Sirica, A.E. (1995) Ductular hepatocytes. Histol. Histopathol., 10:433–456.

Spes, C.H., Angermann, C.E., Beyer, R.W., Schreiner, J., Lehnert, P.,Kemkes, B.M., and Theisen, K. (1990) Increased incidence of


cholelithiasis in heart transplant recipients receiving cyclosporinetherapy. Heart Transplant., 9:404–407.

Steck, T.B., Costanzo-Nordin, M.R., and Kehavarzian, A. (1991)Prevalence and management of cholelithiasis in heart transplantpatients. J. Heart Lung Transplant., 10:1029–1032.

Steiner, J.W., and Carruthers, J.S. (1961) Studies on the fine structureof the terminal branches of the biliary tree. Am. J. Pathol., 38:639–649.

Steiner, J.W., and Carruthers, J.S. (1962) Experimental extrahepaticbiliary obstruction. Some aspects of the fine structural changes ofbile ductules and pre-ductules (ducts of Hering). Am. J. Pathol.,40:253–270.

Stinson, A.W., and Calhoun, M.L. (1976) Digestive system. In: Text-book of Veterinary Histology. H. Dellmann and E.M. Brown, eds. Leaand Febiger, Philadelphia, pp. 205–265.

Strain, A.J., Wallace, L., Joplin, R., Daikuhara, Y., Ishii, T., Kelly, D.A.,and Neuberger, J.N. (1995) Characterization of biliary epithelialcells isolated from needle biopsies of human liver in the presence ofhepatocyte growth factor. Am. J. Pathol., 146:537–545.

Sutor, D.J., and Wooley, S.E. (1971) A statistical survey of thecomposition of gallstones in eight countries. Gut, 12:55–64.

Sutor, D.J., and Wooley, S.E. (1974) The organic matrix of gallstones.Gut, 15:487–491.

Suzuki, N. (1966) On a bilirubin-metal complex compounds in relationto black pigment of gallstones. Tohoku J. Exp. Med., 90:195–205.

Swidsinski, A., Ludwig, W., Pahlig, H., and Priem, F. (1995) Moleculargenetic evidence of bacterial colonization of cholesterol gallstones.Gastroenterology, 108:860–864.

Telford, S.R., Jr. (1992) An eimeriid species (Apicomplexa: Eimeriidae)that parasitises the gallbladder and bile-duct of three species ofTakydromus (Sauria: Lacertidae) in eastern and southeastern Asia.Syst. Parasitol., 23:203–208.

Thistle, J.L., and Schoenfield, L.J. (1970) Lithogenic bile among youngIndian women, a high risk group for cholelithiasis. Gastroenterol-ogy, 58:1000.

Togari, C., and Okada, T. (1954) Cytological observations on theepithelium of the gallbladder in the urodele amphibian (Trituruspyrrhogaster). Okajimas Folia Anat. Jpn., 26:327–333.

Trotman, B.W. (1991) Pigment gallstone disease. Gastroenterol. Clin.Am., 20:111–126.

Trotman, B.W., Bongiovanni, M.B., Kahn, M.J., and S.E. Bernstein(1982) A morphological study of the liver and the gallbladder inhemolysis-induced gallstone disease in mice. Hepatology, 2:863–869.

Troxler, R.F. (1986) Bile pigments in plants. In: Bile Pigments andJaundice. J.D. Ostrow, ed. Marcel Dekker, Inc., New York, pp.649–688.

Tusques, J., R. Senelar, Y. Ginguene, and Vaugien, M. (1964) Etudehistochimique sur les vesicules biliaires du mouton et de cobaye:mucopolysaccharides. Ann. Histochem., 9:269–274.

Dijk, A.W. van. (1935) Ausfuhrgange der Leber und des Pankreas beiVaranus komodoensis und einigen anderen Reptilien. Anat. Anz.Jena., 80:347–355.

Viehberger, G. (1982) Apical surface of the epithelial cells in thegallbladder of the rainbow trout and the tench. Cell Tissue Res.,224:449–454.

Wahlin, T. (1976) Effects of lithogenic diets on mouse gallbladderepithelium. Virchows Arch. [B], 22:273–286.

Wahlin, T. (1977) Synthesis of glycoproteins in the Golgi complex of the

mouse gallbladder epithelium during fasting, refeeding, and gall-stone formation. Histochemistry, 51:133–140.

Wahlin, T. (1979) Histochemical analysis of mucosubstances andcytochemical studies of the Golgi region and the secretory granulesof the normal gerbil gallbladder principal cells. Acta Anat., 103:468–476.

Wahlin, T., and Schiebler, T.H. (1975) Zur Entwicklung des Gallenbla-senepithels des Meerschweinchens. II. Elektronenmikroskopischeund enzymhistochemische Untersuchungen. Histochemistry, 44:253–275.

Walraff, J., and Dietrich, K.F. (1957) Zur Morphologie und Histoche-mie der Steingallenblase des Menschen. Z. Zellforsch., 46:155–231.

Watanabe, S., Miyairi, M., Oshio, C., Smith, C.R., and Phillips, M.J.(1983) Phalloidin alters bile canalicular contractility in primarymonolayer cultures of rat liver. Gastroenterology, 85:245–253.

Watanabe, S., Brown, C.R., and Phillips, M.J. (1985) Coordination ofthe contractile activity of bile canaliculi. Lab. Invest., 53:275–279.

Western, J.R.H. (1969) Studies on the diet, feeding mechanism andalimentary tract in two closely related Teleosts, the freshwaterCottus gobbio L. and the marine Parenophrys bubalis Euphrasen.Acta Zool., 50:185–205.

Winchester, A.M., and Jaques, H.E. (1981) How to Know the LivingThings. pp. 138–149. W.C. Brown Company, Publishers, Iowa City:Iowa, pp. 138–149.

Wolf-Heidegger, G., Staubli, W., and Hess, R. (1965) Zur Ultrastrukturund Histochemie der Gallenblasenschleimhaut des Menschen undder Katze. Acta Anat., 62:606–618.

Wolpers, C., and Hofmann, A.F. (1993) Solitary versus multiplecholesterol gallbladder stones. Clin. Invest. 71:423–434.

Wood, R.L. (1965) An electron microscope study of developing bilecanaliculi in the rat. Anat. Rec., 151:507–530.

Yamada, K. (1960) A unique mode of apocrine secretion in the gallbladder epithelium of Chlemys japonica with a note on the histo-chemical quality of the secretory material. Okajimas Folia Anat.Jpn., 35:47–76.

Yamada, K. (1969) Fine structure of rodent common bile duct epithe-lium. J. Anat., 105:511–523.

Yamamoto, K., and Phillips, M.J. (1984) A hitherto unrecognized bileductular plexus in normal rat liver. Hepatology, 4:381–385.

Yamamoto, K., Fisher, M.M., and Phillips, M.J. (1985) Hilar biliaryplexus in human liver. Lab. Invest., 52:103–106.

Yamamoto, K., Sargent, P.A., Fisher, M.M., and Youson, J.H. (1986)Periductal fibrosis and lipocytes (fat-storing cells or Ito cells) duringbiliary atresia in the lamprey. Hepatology, 6:54–59.

Yamamoto, K., Itoshima, T., Tsuji, T., and Murakami, T. (1990)Three-dimensional fine structure of the biliary tract: Scanningelectron microscopy of biliary casts. J. Electron Microsc. Tech.,14:208–217.

Youson, J.H., and Sidon, E.W. (1978) Lamprey biliary atresia: Firstmodel system for the human condition? Experientia, 34:1084–1086.

Youson, J.H., Yamamoto, K., and Shivers, R.R. (1985) Nonparenchy-mal liver cells in a vertebrate without bile ducts. Anat. Embryol.,172:89–96.

Youson, J.H., Ellis, L.C., Ogilvie, D., and Shivers, R.R. (1987) Gapjunctions and zonulae occludentes of hepatocytes during biliaryatresia in the lamprey. Tissue & Cell., 19:531–548.

Ziegler, U., Palme, G., and Marker, H.J. (1982) Morphological alter-ations in epithelial cells of the mouse gallbladder 30 hours aftertreatment with a lithogenic diet. Pathol. Res. Pract., 174:116–130.


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Comparative morphology of the gallbladder and biliary tract in vertebrates: Variation in structure, homology in function and gallstones