The Organogenesis and early histogenesis of the bovine ...ventral mesogastrium (future lesser omentum) connected the ventral curvature of the stomach to the liver. Between the stomach

T H E ORGANOGENESIS AND EARLY HISTOGENESIS O F THE BOVINE STOMACH

ELDON D. WARNER Department of Zoology, University of Wisconsin, Milwaukee

TWENTY-EIGHT FIGURES

Early studies on the embryological origin of the ruminant stomach have shown its primordium to be a spindle-shaped enlargement of the digestive tract. This was observed in the sheep by Stoss (1892) and Karl (’15) and in the cow by Zimmerl (’00). More recently, a detailed account of bovine stomach development by Pernkopf (’31) has confirmed these observations. Lewis (’15) pointed out the similarity of the lenticular-shaped stomachs in the young embryos of the sheep, rat, cat and pig. On the basis of his comparative investigations he homologized the rumen, reticulum, omasum and abomasum of the sheep’s stomach respectively with the fundus, corpus, area on the lesser curvature and pars pylorica of the stomachs of the other three forms. The comparative embryology of the mammalian alimentary canal (Kruger, ’29) lends further support to these homologies.

On the other hand, several investigators have concluded that only the abomasum is the ‘‘true stomach” of ruminants. The rumen, reticulum and omasum are considered to be en- largements of the caudal region of the esophagus. According to Bensley (’02) this viewpoint was given impetus by the comparative histological studies of Ellenberger (1884) and Edel- mann (1889). The foundations upon which this concept is based are : (1) the epithelium of the three more cranially lo-

% Supported in part by a grant from the Wisconsin Alumni Research Foundation through the Research Committee of the Graduate School and in part by Badger Breeders Association, Shawano, Wisconsin through the Veterinary Science Depart- ment of the University of Wiseonsin.

33

34 ELDON D. WARNER

cated stomach chambers is of a stratified, non-glandular type like that of the esophagus ; (2) the esophageal groove, extending from the caudal end of the esophagus proper to the omasum along the right interior wall of the reticulum, is interpreted as being a part of the esophagus with the rumen and reticulum as lateral expansions and the omasum as a caudal enlargement; (3) the abomasum alone shows a simple columnar epithelium with typical gastric glands. Favilli (’29) supported this view, holding that the abomasurn can be subdivided into fundus, corpus, pyloric vestibule and pyloric canal all homologous with the corresponding regions of the human stomach. I n Weber’s (‘Die Saugetiere” ( ’28) the rumen and reticulum are repre- sented as crop-like expansions of the esophagus. It is concluded, however, that the omasum was derived from the abomasum and is therefore a part of the true stomach. Hence, the esophageal cardia is believed to be at the junction of the esophageal groove with the omasum.

The idea that the rumen, reticulum and omasum are esophageal dilatations has persisted in the literature in spite of embryological evidence to the contrary. It has been incor- porated into many textbooks of zoology, comparative anatomy and veterinary science including several of recent publication. The present paper is an attempt to evaluate these divergent views by further study of the organogenesis and early histogenesis of the bovine stomach.

MATERIALS AND METHODS

Detailed observations mere carried out on 33 bovine embryos ranging in crown-rump lengths from 9.5 mm to 77 mm. Ten of the smaller specimens were of known ages, having been obtained from University of Wisconsin cattle f o r which in- semination data were available. The remaining material was procured from a Madison, Wisconsin packing plant. The estimated ages of the latter embryos are based on the tables of Winter et al. (’42), Swett et al. (’48) and on comparisons with embryos of known ages in the present studv. Individuals were predominately Guernsey and Holstein with smaller num-

BOVINE STOMACH DEVELOPMENT 35

bers of Hereford and Brown Swiss. No developmental or mor- phological peculiarities ascribable to breed differences were noted. Complete length and age data are given in table 1. Crown-rump measurements mere taken at the time of pro- curement after which the embryos were placed in Mossman's fixative.

TABLE 1

L e n g t h and age data f o r embrglos i n v e s t i g d e d in the present s tudy

CROWN-RUMP ACTUAL OR CROWN-RUMP ACTUAL OR LENGTH ESTIMATED AGE LENGTH ESTIMATED AGE

(mm) 9.5

10.0 11.0 12.0 12.3 12.5 13.0 13.0 14.0 14.0 14.7 16.0 16.7 19.0 19.0 20.0 22.0

(days )

28* 29" 30" 34" 33" 33" 35* 34 36 36" 36* 37 35" 38 38 39 40

(mm) 23.0 23.0 25.0 28.0 28.0 29.0 35.0 35.0 37.0 43.0 50.0 50.0 58.0 61.0 76.0 77.0

( d a y s ) 40 40 41 43 43 44 48 48 49 52 56 56 58 59 64 64

*Actual age.

Seventeen specimens of various ages were processed in toto into paraffin, serially sectioned at 10 p and stained with Harris' liematoxylin and eosin. These slides constituted a series from which studies of developmental changes at all stomach levels were carried out. Paper reconstructions of the gastric epithelium were built up from the serial sections of 9.5 mm, 12.3 mm and 14.7 rnm embryos as an aid in determining the origin of the stomach chambers. Plaster of Paris models were made from these reconstructions.

36 ELDON D. WARNEE

From the remaining I6 embryos the stomachs were removed by dissection under a stereoscopic microscope. They were studied at magnifications of from 10 to 75 diameters to observe gross developmental changes. After gross observation, the stomachs of 5 specimens between 43 and 77 mm were serially sectioned and stained according to the procedure outlined above.

OBSERVATIONS AND DISCUSSION

Origin of the stomach chambers

In the youngest specimens studied a regional differentiation of the digestive tract was well under way. The esophagus was a narrow tube extending from the pharynx to a level just caudal to trhe lung buds. Here the gut expanded rather abruptly into a definite stomach primordium. A model of the gastric epithelium reconstructed from serial sections of a 9.5 mm (28-day) embryo (fig. 1) revealed the shape to be that of a laterally compressed spindle, Both dorsal and ventral curvatures were convex in outline, the former being somewhat more irregular than the latter. Shallow depressions in the lateral walls of the model indicated areas where the gastric mesoderm was thickened, thus bringing about a narrowing of the stomach lumen. These areas of mesodermal thickening and endodermal constriction have been referred to as “Haupt- furchen” by Karl (’15) and “Grenzfurchen” by Pernkopf (’31) because of their important future role in the partition- ing of the developing stomach. Caudally, the gastric primordium tapered more gradually into the intestinal anlage.

Transverse sections of a 10 mm (29-day) embryo showed several features characteristic of early mammalian foregut differentiation. The esophagus was a small endodermal tube imbedded in a thick mass of mesoderm. There was no signifi- cant increase in the size of the esophageal lumen near its junction with the stomach (fig. 2). A characteristic left rotation of the expanded gastric primordium (fig. 3) on its long axis was indicated by the oblique position of the lumen in relation to the mid-sagittal body plane. This rotation approx-


irnated 35 to 40" in the 10 mm embryo. The relatively long dorsal mesogastrium, or future greater omentum, extending well to the left of the mid-line to attach to the dorsal curvature of the stomach, was also involved in the rotation. A shorter ventral mesogastrium (future lesser omentum) connected the ventral curvature of the stomach to the liver. Between the stomach and the right lobe of the liver a vertical slit-like coelo- mic space, the vestibule, was present. An extension of this space toward the left beneath the dorsal mesogastrium marked the beginning of the omental bursa. At the more cranial stomach levels the lumen was slightly expanded both dorsally and ventrally. These expansions appeared to correspond respectively to the fundus and the gastric canal of other embryonic mammalian stomachs as described by Heuser ( '14). Between the expanded areas the lumen was somewhat narrower dne to the thickened mesodermal ridges mentioned above.

The foregoing observations indicate that the bovine stomach at the end of the fourth week of development is essentially like those of other mammalian embryos of comparable age both in structure and in its relatioiiships with neighboring viscera.

During the 5th week a rapid growth of the stomach primordium ensued. This was evident from a size comparison of the 9.5 mm and 12.3 mm (33-day) models. A ventral view of the latter (fig. 4) showed a marked expansion of the fundic region (F) along the anterior part of the greater curvature. Accom- panying this expansion was a deepening of the ventral furrow (V) associated with the ventral mesodermal ridge so that the fundus was more definitely set off from the gastric canal area (G). The dorsal furrow was less conspicuous than the ventral furrow although it was more distinct than at 28 days. Caudal to the gastric canal a slight but definite swelling of the lesser curvature could be distinguished. The tapering caudal portion of the stomach was slightly bent ventrally and toward the right.

The more cephalic sections of the 33-day stomach (fig. 5) revealed a further rotation of the organ, now approximating 90". As a result, the expanded lumen of the fundus lay toward the

38 ELDON D. WARNER

left while the narrowed gastric canal was toward the right. Also associated with the rotation was an increase in length of the dorsal mesogastrium. The dorsal and ventral furrows, occupied by the corresponding mesodermal ridges, formed the boundaries between the fundic and gastric canal regions.

At the mid-stomach level (fig. 6 ) the portion of the lumen lying to the left of the dorsal and ventral furrows was much narrower than that of the fundus with which it was continuous cephalad. On the other hand, the lumen to the right of the furrows and associated cephalad with the gastric canal showed considerable expansion. The more caudal stomach sections were characterized by a gradual disappearance of the ridges and a decrease in the size of the lumen.

By the beginning of the 6th week of development further differentiation of the gastric epithelium madc it possible to distinguish the primordia of the several adult stomach regions. I n the 14.7 mm (36-day) model (fig. 7 ) the groove-like ventral furrow (V) set off more distinctly the areas developing along the greater curvature from those of the lesser curvature. The same was true to a more limited degree of the dorsal furrow. The fundus was now a very prominent enlargement, having expanded cephalad, dorsad and to the left. The position of the fundus and its fate in older cmbryos positively identified it as the primordium of the rumen and the reticulum (AB, fig. 7 ) . The bulb-like swelling of the lesser curvature caudal to the ruminoreticular primordium constituted the major portion of the omasal anlage (C). The tapering caudal stomach region, now showing a more definite bend toward the right, was destined to become the adult abomasum (D). Along the cranial part of the lesser curvature the gastric canal appeared as a cylindrical passage connecting the esophagus with the omasum. This relationship marked it as the developing esophageal groove. Another internal channel, the omasal sulcus (S), could be recognized in the 36-day model. I n the adult, this narrow trough extends along the left wall of the omasum from the reticulum to the abomasum. On the model it was indicated


by a prominence along the greater curvature just caudal to the ruminoreticular primordium.

The relationships of certain stomach regions at 36 clays could be seen more clearly in transverse sections. Cranially, division of the lumen into the large ruminoreticular primordium on the left and the small gastric canal on tlie right was clearly defined (fig. 8). The lips of the latter were made up of the dorsal and ventral mesodermal ridges covered by the endoderm of the corresponding furrows. A further extension of the dorsal mesogastrium toward the left was apparent. At the level of the omasum (fig. 9) the right portion of the lumen was cx- panded to form the major part of that structure. Toward the left the smaller omasal sulcus was bounded by lips that were continuous with the lips of the esophageal groove. I n the region of the abomasum (fig. 10) the lumen narrowed and the dorsal and ventral ridges disappeared.

From the foregoing observations it was evident that the spindle-shaped gastric primordium of the 9.5 mm embryo gave rise to all of the definitive stomach subdivisions. That this primordium is fully homologous with the stomach anlagen of other mammals is further borne out by its characteristic sinis- tral rotation and its association with typical dorsal and ventral mesogastria. It should be emphasized that the esophageal groove, although continuous with the esophagus, was not derived from that organ but from the cranioventral part of the stomach spindle. The esophagus proper in all specimens studied remained a tube of uniformly small diameter with 110 expansions of any kind.

Later nzorphoge.nesis Further development of the stomach to a stage in which it

resembled the adult organ in its general configuration was followed through a series of embryos ranging in crown-rump lengths from 16 mm to 77 mm (estimated ages between 37 and 64 days). Isolated stomachs from representative specimens of this series (figs. 11-16) illustrate the major external changes. Selected transverse sections (figs. 17-22) show certain impor-

40 ELDON D. WARNER

tant internal modifications. In dissecting the younger individuals of this group it was observed that the stomachs had assumed oblique positions in the coelom, lying at angles of from 45 to 60” with the long axis of the body. I n the older specimens this oblique relationship was obscured by expansion and shifting of the stomach chambers. Because of the increasing complexity of later morphogenesis the following dis- cnssion considers separately the changes occurring in each cornpar tment.

In the 16 mm embryo (37 days) the ruminoreticular primordium remained the most prominent external stomach feature, showing further expansion dorsally and toward the left (fig. 11, AB). Subdivision of this primordium into a craniodorsal rumen and a caudoventral reticulum was evident at 20 mm (39 days) (fig. 12). The rumen (A) extended into the left cephalic region of the coelom, lying to the left of the esophagus, ventral to the mesonephros and dorsal to the liver.

Early regional differentiation of the rumen became apparent in the 28 mm embryo (43 days) (fig. 13). The relatively narrow caudal portion, the vestibule (A-3), designated “Vorhof” by Pernkopf (1931) was twisted spirally toward the left. This region communicated caudad with the reticulum (B) and cephalad with a slight prominence, the primordinm of the major portion of the dorsal rumen sac (A-1) . A somewhat larger dorsal protuberance marked the differentiating ventral rumen sac (A-2). The cranial groove of the rumen developed as a transverse furrow across the dorsal surface of the chamber between the vestibule and the ventral rumen sac. Twisting of the vestibule and expansion of the reticulum resulted in the appearance of a distinct ruminoreticular groove along the left wall of the stomach.

Development of the rumen beyond the 43rd day was characterized by marked caudal expansion and migration. During this period the rumen appeared to undergo what had been described by Martin (1899) as a backward rotation of about 90” on a transverse axis through the vestibule. Thus, as the dorsal sac expanded it shifted dorsad, caudad and slightly


toward the right from its original position. Simultaneously, the dorsally situated ventral sac enlarged and mjgrated caudad, ventrad and somewhat toward the left. The dorsal and ventral blind sacs appeared in the 58 mm embryo (58 days) as small dorsal evaginations of the corresponding main rumen sacs. At 77 mm (64 days) (fig. 16, A-4 and A-5) they had expanded considerably and had assumed their definitive caudal positions. The caudal groove of the rumen was evident at 64 days as a transverse notch between the blind sacs. Associated with the caudal expansion of the rumen sacs was a backward and upward movement of the vestibule, its long axis shifting from a horizontal to a nearly vertical inclination. Inspection of older fetal stomachs indicated that the caudal part of the vestibule developed into that area of the adult rumen known as the atrium ventriculi (Sisson and Grossman, '38) while the cranial vestibular region merged with the dorsal sac..

The cranial and caudal rumen grooves were associated respectively with the cranial and caudal pillars, incomplete internal partitions formed by apposition and fusion of the ex- panding rumen walls (figs. 18, 21). As a result of the backward rotation of the rumen, the cranial pillar forming between the ventral sac and the vestibule moved from its original dorsal location successively to caudal and ventral postions. I n the adult stomach the cranial position of this pillar indicates a total end-for-end rotation of nearly 180" during the course of its development. The caudal pillar developing later between the dorsal and ventral blind sacs maintained its early caudal location.

I n the 20 mm embryo the reticulum was apparent as a swelling on the greater curvature of the stomach caudal and ventral to the rumen (fig. 12, B). As it expanded it became more distinctly set off from the rumen and the omasum by ruminoreticular and reticulo-omasal grooves (figs. 14-16). These external furrows were particularly noticeable along the left wall of the stomach. I n the more advanced stages studied, the reticulum showed a gradual shift toward the left and craniad.

42 ELDON D. WARNER

This change in position coincided with the caudal migration of the rumen already described.

Internally, the esophageal groove (figs. 18, 21) in the right reticular wall underwent a marked modification. As it developed, this straight, nearly horizontal channel was transformed into a spirally twisted trough extending vertically from the esophageal cardia to the reticulo-omasal orifice. After torsion the cephalic par t of the groove opened dorsally, the middle portion toward the left and the caudal portion cranially. The lips of the groove, formed from the dorsal and ventral mesodermal ridges of the early embryonic stomach, became more prominent as they developed.

By the end of the 9th week (64 days) the mucosal surface of the reticulum showed little specialization except for faint folds extending laterad from the lips of the esophageal groove. Inspection of several more advanced fetal stomachs revealed that the compartments making up the ‘(honeycomb” did not develop until the middle of the third month.

In embryos between 20 mm and 77 mm the omasum underwent progressive enlargement into a nearly spherical compartment on the lesser curvature of the stomach. As it developed it became more distinctly set off from both the reticulum and the abomasum. Coordinated with the shifts of the rumen and the reticulum, the omasum gradually assumed a position more to the right and anterior in the stomach topography.

Transverse sections of the omasum in the 28 mm embryo revealed the primordia of the laminae omasi or omasal “leaves” as 6 low, longitudinally directed mucosal folds. These are shown in the 35 nim (48-day) stomach (fig. 19). Sub- sequently these folds increased in size, number and complexity until they nearly filled the omasal lumen. In the 77 mm embryo (fig. 22) nearly 50 were present at the level of greatest omasal diameter. At this stage they could be classified as primary, secondary and tertiary in the order of their decreasing height. The omasal sulcus remained a relatively unexpanded groove in the left ornasal wall. Its dorsal and ventral lips were continuous craniad with the corresponding lips of the esophageal


groove. Like the latter they were derived directly from the primary dorsal and ventral ridges of the early stomach.

At the 20 mm stage the abomasum showed differentiation into expanded cranial and narrowed caudal portions (fig. 12) . These formed the primordia respectively of the cardiac and pyloric areas of the adult compartment. As development proceeded the longitudinal abomasal axis was bent increasingly so that the pyloric region extended at first ventrally then toward the right and finally cephalad (figs. 13-16). As a result of this bending, the angular incisure along the lesser curvature between cardiac and pyloric zones became progres- sively more acute. Also in later developmental stages the abomasum was more distinctly set off from the omasum, and cardiac and pyloric zones showed a greater differentiation. The cardiac region in particular increased markedly in both length and diameter. This size increase appeared to fore- shadow the marked growth of the abomasum during the latter two-thirds of the gestation period. According to Becker e t al. ('51) this chamber makes up about 50% of the total stomach volume at the time of parturition.

Folding of the inner abomasal wall was indicated in 19 mm embryos (38 days) by a pair of longitudinally directed dorso- lateral grooves. These were present in both cardiac and pyloric regions. At 35 mm (fig. 20) the left and dorsal portions of the cardiac mucosa were thrown into 7 primary longitudinal folds. Subsequently the entire mucosal surface of the cardiac zone became involved in the folding process with a maximum of 13 primary folds distinguishable at the mid-cardiac level. As development, continued the primary folds increased in prominence and secondary folds appeared at their bases (fig. 22). The pyloric region (fig. 20) developed 4 primary mucosal folds anteriorly and three posteriorly. TJpon these, numerous secondary folds became superimposed later in development. Epithelial pits appeared in the mucosal surface near the angular incisure in the 50 mm embryo and at the 77 mm stage they were widespread in both cardiac and pyloric regions.

44 ELDON D. WARNER

As a result of the above changes the fetal stomach at the end of the 9th week of development displayed most of the morpho- logical features of the adult organ. All of the various parts arose from the spindle-shaped primordium of the early embryo.

Early histogenesis

I n the adult ruminant the stratified, non-glandular epithelium of the rumen, reticulum and omasum is in sharp contrast with the simple columnar, glandular type of the abomasum. This difference forms an important basis for the concept that the three more cranial stomach chambers are esophageal derivatives. It was of interest, therefore, to ascertain when during development epithelial differentiation began and the manner in which it occurred.

In embryos of 19 mm or less, the epithelium of the esophagus was essentially like that of the stomach (figs. 23, 2-10). The cells in both organs were columnar and densely packed, making it difficult to determine in several areas whether true stratification o r pseizdostratification existed. In most regions, however, the epithelium appeared to consist of from two to 4 layers of cells. The nuclei of the basal layer were situated for the most part in the upper portions of the cells. I n the superficial layers on the other hand the nuclei occupied more basal positions. As a result, the nuclei were concentrated in a conspicuous zone in the superficial and middle depths of the epithelium. I n most areas of the stomach the lining was thicker than that of the esophagus, being made up of taller columnar cells but otherwise showing similar characteristics.

Definite histogenetic changes appeared in the esophagus of 23 mm embryos (fig. 24). The most striking general impres- sion obtained was of a more uniform distribution of nuclei throughout the epithelium rather than a concentration in the superficial part of the layer as in younger embryos. Stratifi- cation was more distinct, with cells arranged in from two to 5 layers. The basal cells were columnar in shape with nuclei variable in position, many lying near the basement membrane.


As a result, the nucleus-free basal cytoplasm seen in earlier stages was not in evidence. Cells of the intermediate layers were columnar to polyhedral while those lying next to the lumen were low columnar to cnboidal. This transformation had occurred throughout the length of the esophagus with the exception of the extreme caudal end near its junction with the stomach. I n contrast, the gastric epithelium at 23 mm remained relatively unchanged from the early embryonic condition.

In embryos between 25 mm and 35 mm, certain areas of the stomach showed changes similar to those just described for the esophagus. The floor of the esophageal groove was the first region to be involved. Here, in the 25 mm embryo the epithelium had become distinctly stratified, two to 4 cells in thickness and showed a characteristic dispersal of nuclei. Partial epithelial differentiation had also occurred on the inner surfaces of the lips of the esophageal groove and in an area on the right wall of the omasum just caudad the reticulo-omasal orifice. At 28 mm and 35 mm (fig. 18) these changes were also apparent on the reticular surfaces of the lips of the groove, on the craniodorsal wall of the reticulum and in the caudal portion of the rumen vestibule near its junction with the reticulum. On the floor of the esophageal groove at 35 mm the epithelium had become thicker and the free border showed several irregular cellular projections. I n other regions of the stomach the epithelium remained relatively unchanged.

Distinct epithelial differentiation was apparent in the 43 mm embryo not only in the esophagus and esophageal groove, but in all parts of the rumen and in most of the reticulum as well. Active proliferation of cells had resulted in a marked increase in epithelial height in most areas. This increase was most striking in the rumen and the anterior part of the reticulum (compare figs. 25 and 26). For example, in the dorsal sac of the rumen in the 35 mm embryo, sample measurements of epithelial height ranged from 45 to 80 p whereas at 43 nim the range was from 75 to 175 p. The number of cell layers varied from two to 4 at 35 mm and from 10 to 16 at 43 mm. Through

46 ELDON D. WARNER

a considerable extent of the rumen and reticulum, cordlike masses of cells projected into the lumen, giving the free epithelial surface a very irregular appearance. Intra-epithelial spaces were also abundant in these areas. Increases in epithelial height were not as great in the esophagus and the esophageal groove although the epithelium showed marked differentiation in other respects. The lining oE the lips of the groove was lower than that of the floor, being quite uniformly two cells thick.

I n addition to an increase in thickness the epithelium in the above areas showed a differentiation into superficial and deep zones. The deep or basal zone of the esophagus and the floor of the esophageal groove was composed of from one to three cell layers. In the rumen and in the differentiated portion of the reticulum the same zone was from three to 5 layers in thickness. The basal cells were columnar in shape and fairly uniform in size and arrangement. Their nuclei were round to ovoid and relatively large. The superficial zone was thicker than the deep zone with cell layers ranging from 4 to 7 in the esophagus and esophageal groove floor and from 8 to 12 in the rumen and reticulum. Individual cells were variable in size, but in general, were considerably larger than those of the basal zone. I n shape they were irregularly columnar to polyhedral in the deeper layers and rounded to squamous in the layer adjacent the lumen. Nuclei were small, angular and usually located near the margin of any given cell. These changes possibly foreshadowed the formation of the stratum germinativum and stratum corneum cliaracteristic of the epithelium in the esophagus and the forestomachs of adult ruminants.

The omasal epithelium at the 43 mm stage showed varying degrees of differentiation. On the larger cranial omasal leaves it was quite distinctly stratified and moderately thickened, resembling that of the esophageal groove in the 35 mm embryo. The lining of the smaller caudal leaves on the other hand was relatively low (30 to 50 p) and showed little histogenetic change. The area on the right dorsal wall near the reticulo-


omasal orifice which had undergone some earlier differentiation was now markedly thickened with an irregular free border like that observed in the rumeii and the anterior portion of the reticulum.

Few changes were apparent in the abomasum at 43 mm. In the cardiac region the epithelium was still composed of from two to 4 layers of columnar cells. I n height they ranged from 50 to 75 p. The nuclei were somewhat more scattered than in earlier stages but still occupied mainly the upper portion of the epithelium. Cells were much more densely packed in the pyloric region, but were of about the same height as in the cardiac region. Nuclei here were largely restricted to the superficial portion of the layer.

Beyond the 43 mm stage the stratified epithelium of the esophagus and stomach showed an increasing differentiation into deep and superficial zones. I n the esophagus the deep zone was reduced to one cell layer in thickness in most areas. It was composed of relatively heavily stained columnar cells with round to ovoid nuclei. The superficial zone varied from 4 to 10 layers in height. The cells here were more lightly stained and as at 43 mm were larger than those of the basal zone. They were variable in shape with shrunken, peripheral nuclei. Changes in the esophageal groove paralleled those of the esophagus although the epithelium showed more variations in height. Along the floor of the groove it was thick with an irregular free border, while on the lips it was thinner and more uniform, consisting of single-layered deep and superficial zones. Epithelial height was maximal in the rumen and reticulum. In these chambers the deep zone varied from three to 8 layers in thickness while the superficial zone ranged from 6 to 15 layers. Cells were more croweded here than in the esophagus and esophageal groove. Also, the distinction between deep and superficial zones was less clear cut. I n 50 mm and 58 mm embryos the posterior part of the reticulum showed the characteristic changes seen earlier in the anterior portion. The epithelial cords and intra-epithelial spaces first noted in the rumen and reticulum at 43 mm appeared to reach their

48 ELDON D. WARNER

maximum at 58 mm and then to regress. As a result, the free epithelial borders in these cliambers were more regular a t 77 mm and the height of the epithelium more uniform. The entire omasal surface now showed a differentiation toward the stratified condition although the epithelium in most areas was considerably lower than that of the rumen and reticulum. Along the sides of the developing laminae omasi it became more uniformly two-layered as development proceeded. In height it ranged from 40 to 60 p. The basal cells were low columnar in shape with round nuclei and were characteristically well stained. I n the superficial layer the cells were much taller and were lightly stained. The nuclei were shrnnken and were situated at the extreme inner ends of most of the cells. On the tips of the laminae omasi and also at their bases the epithelium, especially the superficial zone, was considerably thicker than along the sides. The area along the right dorsal wall adjacent to the reticulo-omasal orifice retained its thickness and resembled the lining of the floor of the esophageal groove.

Transformation of the abomasal epithelium toward the simple columnar type was first observed in the 50 mm embryo. This change coincided with the appearance of the pits and secondary folds previously described as forming on the inner abomasal surface. In the cardiac region, from 8 to 10 primary longitudinal folds were present a t 50 mm. Cranially, the epithelium covering these folds was stratified o r pseudostratified as in younger embryos (fig. 27). l\lore caudally, however, near the angular incisure, numerous epithelial pits were developing along with secondary folds at the bases of the primary folds. Nany of the cells lining these areas showed a simple columnar arrangement. Pits and secondary folds were abundant in the pyloric region and some of the epithelium was simple and low columnar to cuboidal in nature. The extreme crowding of cells here made it difficult to determine cell out- lines very readily. I n the 58 mm embryo an increase in pitting and secondary folding was evident in both cardiac and pyloric regions. At 77 mm (fig. 28) the entire cardiac mucosa was strongly pitted and simple columnar epithelium was wide-


spread at all levels. The pyloric epithelium was more com- plexly foldcd than that of the cardiac region and, in general, individual cells were more crowded and somewhat lower. A t this stage of development in both cardiac and pyloric zones the riuclei of the simple columnar cells were situated close to the lumen. This is the reverse of the polarity in the adlilt epithelium where the nnclci are basal in position.

The foregoing histogenetic studies show that the epithelia of both esophagus arid stomach remaincd alike through the early developmental stages up to and including 19 rnm even though tlir two organs had become niorphologically distinct. Epithelial niodifications toward the adult stratified condition were miderway throughout the esophagus by the 23 rnm stage. Shortly thereafter similar changes began in the stomach. These changes were initiated in tlie various regions roughly in the following order : (I) thc esophagcal groove ; (2) portions of thc rumen, reticulum and omasuni adjacent to the esophageal groom; (3) the i*cmainder of the rurnen and the anterior par t of the reticulum; (4 ) the posterior part of the reticnluiii; and (5) the major portion of the omasuni. A t 50 nim the epithelium throughout the esophagus and tlic three cephalic stomach chambers was stratified. This scquence sug- gests a progressive advaiice of stratification from the esophagus along the esophagcal gim~vc~ and into the runien, r e t i c d i m , and omasmi. It does not, l i o ~ ~ e w i - , ofYcr evidcnce of ail esophageal origin of these cliambers sirice they ~ v e r c already morphologically establislied as storriacli derivatives pri 0%- to tlie oiisct of epithelial differentiation.

Ahomasal cliaiiges began later aiid showed no apparent rc- lationsliip to tliosc o c c u r h g in the otlier stomach chambers. EpitheIial pits and areas of simple columnar epithelium a p peared first in the 50 nini embryo near the angular incisure. At 77 nim they were widespread in both cardiac and pyloric 1.egions. Thc formation of pits greatly increased thc area of the abomasal epithelium. This increase in area secnied to be a factor in the transformation of the epithelium f r o m a strati-

50 ELDON D. WARNER

fied or pseudostratified coridi tion to a single-layered arrangement.

The foregoing morpliogenetic and histogenetic observations intlicate that the esophagus does not contribute to the formation of the bovine stomach. The so-called "esophageal" char- acter of the rumen, reticulum and omasurn must therefore be cxplaiiicd in another nianiwr. Clues to the probable explanation are provided hy certain comparative aspects of mammalian gastric morphology ancl histology.

Simple stomachs are found in the majority of marrimals but many nienibors of sewral clift'cr*cnt orders show a variety of gastric sprcializatioiis. Since tliesr have been described and summarizetl in considerable detail by Reiisley ( '02) , TVeber ( '28) arid Plerilr ( '32) only a brief survey will be made here.

Among the marsupials, the kangaroos have elongated, colon- like stomachs with sacculations along the greater curvature. ,4 somewhat similar condition is obserwcl among primates in the laiigur monkey, Sc~mnopitliecus. In the Chii-optera, the stomach of fruit-eating bats has a n enlarged cardiac sac that is further suhdividcd in sonie species into two compartments. In the vainpirc hat, I)csrnodus, this same region is greatly elongated, serving as a 1)lood-storagc organ, The stomach of the edeiitate, Eradypus, lias ail expanded cardiac portion sn1)divideti into three conipartrricnts which lead into a long pointed Hind sac. The pyloric region is also sacculatcd. In several rotlcnts iiicluding hamsters, roles, lemmings and musk- rats the stomacli is made up of a cardiac sac and a pyloric sac sepalbated by a more or less conspicuoixs external furrow. (Yomy)ound stomaelis itre tlic riile among the Sirenia, the cardiac eiilargcmcnt sliowiiig two caecnn-like outpocketings. J3ost Oetacea liavc specialized stomachs divided into 4 or more charri1)ers. The rnacjorbitp of ruminant Artiodactyla have 4- charnhred stomaclis although in members of the Tragulidac tlir omasuni is small o r ak)scnt. The non-ruminnnt pecarries have thrc,c-cliamhcrcd stomachs.

Most of the gastric eulai~genicnts ?just descrilml arc niodifications of the cardiac region, an area that also givcs rise to the

BOVIKE S T O M A C H DEVELOPMENT 51

rumeii and reticulum in the cow and tlic sheep. Since it has been shown that tlie latter chambers are derived embryo- logically from the stomach rather than the esophagus it is reasonablc! to suppose that the cardiac sacs of most other mariimals a re of similar origin. Possible exceptions occur however arrioiig certain Cetacea such as Yhocaeiia and Balaen- optcra. In these forms the first stoniach chamber is directly continuous with the esophagus with no appreciable constriction between the two. The cpitlieliurn of both regions is similar. On tlie basis of these facts and sornc rather limited embryological data, Jungklaus (1898) has concluded that this chamber is derived from the esophagus. This viewpoint has been accepted with reservations by Kerislcy (’02) and Pern- lropf ( ’30) . There is no other substantial support for the concept of an esophageal origin of the cardiac sacs.

On the positive side, Pernkopf (’30) has pointed out homologies in tlie niusclc layers of the sirriple stomachs of most mammals, tlic “ bilocular” stomachs of rodents and the “tri- locular” s tomach of ruminants. This is strong evidence in favor of a gastric origin for the entire niammalian con]- pound stomach.

The histological studies of Etlclrrianii (1889), Oppel (1896) and Kcnslcy ( ’02) show that stratified non-glandular epi- tliclinrri is hy no means restricted to the forestomachs of ruminants. I n the Monotremata the entire simple stomach is lined by s t ra t i f id clpitheliiirn and is devoid of glands. ( ’(1 rtain mc.mhei.s of the 111 a rsupialia, Cliiropte1.a, Eden tat a, Hotlmtia, Perissodactyla a i i t l uon-i~uriiirittnt A rtiodactyla, some with simple stoniaclis and others with corripountl stoniitclis, show vnryiiig t l c g i ~ c ~ ~ of epitlielial stratification arid glantlular modification. It \vonltl he difficult to conceive of all thesc~ c~xaniplcs as being of esophageal origin. Instead, it seems more likely that they reprtlsciit alterations of the stomach lining itself. Thrrc clocs not seem to be a valid I T R S O I ~ nrlly stratified epithcliuni in the stomach must be of extra-g;isti.ic origin.

52 ELDON D. WARNER

Bensley considered the various mammalian gastric pouches and the areas of stratified epithelium in both simple and compound stomachs as evolutionary adaptations of that organ alone to specialized dietary conditions. I n the light of avail- aldc evidence this appears to be a logical conclusion.

SUMMARL'

Alorpliogcnetic arid histogenetic observations of the bovinc stomach v-erc carried out in a series of embryos ranging in c row- rump lerigtli froni 9.5 nim to 77 mm. In the youngest specimens a spindle-shaped gastric enlargernent was present. From this prirnortliurn all of the definitive stomad1 compartments arose. T,ater rniorpliog~nc~sis was followed to a stage in which the adult configuration was attaiiietl. There was no indicatioii of an csopliageal contribution to any par t of the cornpound storriach. Histogcnctic studies showed that the uii- cliff'erciitiatecl epithelia of the esopliagus and the stomach were nearly ideiitical in early stages. Epithelial stixtificatioit I)clg:.aii first in the esopliagus ant1 pi*ogressccl in secyuence to tlic exopliageal g r o o i ~ , to ar t~as of the rnnien, reticulum arid oinasum atljacent lo thc giwovc mid filially to tlie niorc rernote payts of tlicsc c1iarnl)crs. ('hanges in tlie ttl>omasal c.pitliclium to\vartl the simple coluninar condition bcgan relatively late a i i t l a p p a r c d to I)(> unrelated to those occurriiig in tlie other c( )r n pa 1- t 111 en t s . A 11 r ie f c oinpa LYI t ir (1 s LI r' vc y of rnanim aliari stoi-rradi nioiyhology and histology is made with the purpose of cmpliasizing the clcartli of evitlcrice to support the idea of t i n esophng:.c~d origin for aiiy stomacli component.

. .

,2C'liNO~~I,F31)(:I\IE: STS

Sincere tlianks are expressed to Dr. 8. IT. McKutt, Dr. Louise IVipf and Nr. Verlyn I)ary of the Veteriiiary Science Department of tlie 1Tiiivci.si ty of 'Il'iscoiisin f o r providing research space arid technical assistance and f o r facilitating the procuiwnent of embryological material. Thanks are also duo Mr. Tmter Diedrich of tlic Rlilwaukee Public Museum

BOVINE S T O M A C H D E V E L O P M E N T 53

for making the plaster models of embryonic stomachs, Dr. Joseph G. Baier f o r his advice on photographic matters, arid Dr. Harlaiid W. Mossman for his critical appraisal of the present paper.

LITERATURE CITED

BECKER, R. B., P. T. ARNOLD A X D S. P. MARSHALL 1951 Developnient of the

RENSLRY, 1L R. 1902 The cardiac glands of mammals. Am. J. Anat., 2 :

EDELM kNN, R. 1889 Vergleicliend aiiatornische und physiologische Untersuchun- gen uber cine besondere Region der Mageiischleimhaut (Cardiacdrusen- region) hei den Saugrtiercn. Deutsclie Ztschr. f . Thiermed., 15:

l‘:LLENBERGER, w. 1884 Handhuch der vcrglcicheiide~~ IIistologie und I’hysiologie

E’~VILL1, N. 1929 nallo stomaeo (lei ruminanti (tip(i pririiitivo !) allo stornaco dell’uomo. (Contra u ~ i a affcrm,itn e eonfcrniata fi1ogeiiin.i Monitore 2001. I t n l . , 40: 47-61,

GROTE, R. 189G Reitrage z. Entwicklung I). Wiederk:1ueriiiagriis. Zeitschr. f. hinturwiss., 69: 387-479.

HEUSER, C. H. 1914 The form of the stoniac~h in riinrrimxliau rrnhi vos. Anat. Rec., 8 : 130.

JUNGKLAUS, F. 1898 Der JI:igcw (1~1 Cctnccaii. Jciiaisclie Zt’i( 111. f . r\’ntur\v., 3’2: 1-94.

KARL, €I. I915 Dic F!nt~vicklung dcs 3l;rgeus bcim Schnfc. &10rph. Jxlirb., 49: 311-352.

KRVGER. ’Ar. 1929 Der vcrglrichcndc Eiitniclrlungsgeschiciite in1 Dieriste dcr I ihsu~ig c1c.s Homologisici ung5prohlcms an den 1):irni und Gckriisab- srhnitten das Mcnschen und einiger HnuxuBugetierc. Zcitschr. Anat . u. 7’:ntwirklurigb., 90: 458-548.

IIETIS, F. T. 1915 C‘oiiiparntive ernblyologv of the nianinialinn stoni:wh. h : i t . Ree., 9 : 102.

M 4KTIN, 1’. Die Entwicklurig dcs ~ieclerkaiuerm:1geiis nnrl Darmeb. Srhweiz. Arch. Tirrheilk., 31 : 173-214.

€’LXNI\-, IT. 1932 Haiidbnch der niiki oskopischcn hriatoniie dcs Mcnschcn. Ed. by W. ron Mnllcndorf. Springer, Berlin. Sr/Z Vcrdnuungsapparat. Der Mag~n.

OPPEL, A. 1896 Lehrhuch dcr verglcichenden mikroskopischcu Anatonlie der Wirbrlticrc 1. Der Magcn. Fischer, Jena.

Pernkopf, 3:. 1930 Beitrsgr ZUT vergleichendcn Anatonlie des Vertebraten- magcns. 2. Anat. u. Entwicklungs., 91 : 329-390.

PERNKOPP, E. 1931 Die Entwicklung des Vorderdarmes, inbesondere des Magens der Wiedcikauer. Xeitschr. Anat. u. Entwicklungs., 94: 490-622.

STOSS, A. 1890 Vergleichendc ariatomische Untersuehungen ubcr die Eritwick- lung des Verdauungkanales des Wiederkiiuer. Dtsch. Z. Tiermed u. vergl. Path., 16: 96-124.

bovine stoniach during fetal life. Journal of Dairy Science, 34-43.

10.5-156.

165-214.

clcr Hausstugetiere. Barey, Brrliu.

1899

54 ELDOM I). WARNER

SWEET, w. w., c. A. MATHEWS AND M. H. %'OHRMAN 1948 Development O f tlie fetus in thc dairy cow. U.S.U.A. Tecliriical Bulletin #964.

WEBER, M. 1928 Die Saugetiere. 2 AoA. 2 . Fischer, Jena. WINTERS, L. M., W. W. GREEN AND R. F. COXSTOCK 1942 The pre-natal de-

velopment of the bovine. Univ. of Minnesota Ag. Exp. Station Teeli- nical Bullctin #151.

ZIMMERI,, U. 1900 Coiitrihuto a l l s cnuosceriza d e b oiitogenesi dell0 stom:ico del runiin:inti. Rfonitorc Zool. It:il., 11 : 13-29.

PLATES

FXPLAKATION O F PLATES

Abbrevinlioiis riser1 in figures 1 t o 2%:

AB, rumino-reticular primordiurn €3, reticulum A, rumeii C, onlasum

D, aboniasum E, esophagus

B, gastric canal (esophageal groove) R, omasal sulcus V, ventral furrow

1, dorsal rumen sac 2, ventral rumen sac 3, rumen vestibule F, fnndus 4, dorsal blind sac 5, ventral hlind sac

PTIATE 1

EXPLANATION OF FIGIJRES

1 Jlotlcl of tlie gastric cpithrliuin of a 9.5 111111 c.nibryo, left ventral riew. x 3 2 . l'raiis\rrse wetion through thc IMSC of tliv esopliagus of :t 10 iiiin embryo. X 41.

' ~ i i i i i s v ~ r s e bec'tioii tliiough the htoiiiacli of :i 11) n11ii viiibrj o at the level of i t s grcantrst t l ian~rter . Iruiiien of the fundus upper lrft , lomc~ii of gastric c:iii:il lower right. X 41.

hlodel of thv gastr ic epithelium of a 12.3 min t w h i yo, lef t ventral viev. X 3 2 .

Traiixvei se section through tlic fu11tlos nirtl gasti ic cannl of a 12.3 niin e m b q o. Fundus toward thr lef t , pb t r i c c:rii:rl toii:iid thc right.

2

.3

4

.j

X 41.

6 Trniisverse sectioii through the xtoiiiatli of a 12.3 iiim r m b r p candad the funidus :md gnstlic canal. x 11.

i Model of tllc g T I C c~pitlirliuii~ of :i 14.7 nim e i~ ibqo , l c t t vriitrnl view. x 3 2 .

8 Traiihvrrw wetioii tlitougli tlic, riiiiiiiiO-iftiCulfli piiiiiortliurtl ( lef t ) and csoplingtvl g i o 0 w ( l i g h t ) of a 1 4 . i 111111 cmhlyo. x 41.

9 'Fr:ili s w t i o ~ tliiougli tlrc or11 I siilriis (lc,ft\ niid omasiiiri (right) of 21 14.7 111111 elllbr\.l). x 41.

tion tlirougli the :rho~iiabiiiii of :I 14.7 riiiii eriibryo. X 41.

56

BOVINE STOMACH DEVELOPMENT FLDON D. WAENER

PLATE I

57

PLATE 2

ESPLBNATION O F FIGURES

(The stomachs in figures 11 to 16 were dissected from fixed bovine embryos and photographed in 80% nlcohol from the left sidr.)

11

1 2

13

14

15

16

Stomach from :t 16 intii embryo. Note the conspicuons rumino-reticdar primordium. x 8. Stomach from a 20 nim embryo. The rumiiio-reticular priniordium is dif- ferentiatiiig into a runien and reticulum. The otiiasuni and abomawni are also apparciit. X 8.

Rtoniacli from a 28 min embryo. The rumen shows the beginning of differentiation into dorsal sac, ventral sac and vestibule.

Stomach from x 50 uim embryo. The runicn has expaiided and shiftcd exudad in relation to the other conipartnieiits. The aboniasuin is distinctl? dif - fereiitiated into cardiac and pyloric regions. x 5. Stomach f rom a 61 in in enibryo. The ruinen shows further caudal expansion and thc dorsal blind sac is apparent. The other chambers are more distinctly tliffrrentiatcd. x 3. Stoni:rcli from a 7 7 niiii cmbryo. The expanded runieii show5 distiiict dorsal and ventral blind mcs. The other chanibcrs s I i o \ ~ further differentiation and eqxiiisiion. x 5.

X 8.

58

BOVINE STOMACH DEVELOPRIEXT ELDON D. WARNER

PLATE 2

PLATE 3

ESt'LANATION OF FIGURES

1 7 Transverse scctioii through tlie ventral sac, rc%stibule aiid csopliagus of a 35 mni enibrgo. The cpithelinm of tlica csopliagus is differeiitiatiiig while that of the rumen is of tlic early enil)rgonic tgpc.. X 24.

18 Transverse srction through the wstihiile, rcticuluni and csoplingenl groove of a 35 inni c.nibryo. Notc tlir differeiiti:~ting epitlieliuin in t h e csophngenl groove and the proximal vestibular Irgion. X 24.

19 Transverse wct ioi i through the oiii:isun~ of a 35 inin emliryo sliowiiig the primary oniasnl lcnvcs :ind on1

'l'r:rlisvcrse scc?tioli tlirough the :11~oiii:1su111 siioviiig tlie cardiac region (upper l e f t ) and tlir pploric, region (lower riglit 1 . Note the priiiiary loiigit,udiiial folds. x 24. Tr:iiisvc~se section through tlic runic~i , rcticrriuiii aiid esop11age:il groove of :I .58 mrii embryo. Tliickeiicyl, stratifietl epitlicliuni is :rpi)tireiit iii all of the regions. x 11.

20

21

se sectiorl through tlie oiii:isurii (riglit) a i d ahomnsmii ( left) of a i7 nim crubrgo. Notc tlie prim:iry, secondary aiid tcrtinry leaves in the oiiiilsurn xiid the primary x r i d sccuiid:iry folds in the abomnsuiii. x 11.

60

BOVINE STOXACH DEVELOPMEXT BLUOX I). \VARNER

PLATE 3

23 Tralisvcrsc~ section tlii ougli tlie (~soph:igus of i i 1 9 iiini embryo sliowing tlie typical uiidiffereiitiatcd cyitlieliuni. x 260.

section through tllc esop1i;ig.u~ of a 23 nini embryo slioning the X 260. increased epithelial strat1fic:itioii and dispersal of nuclei.

Tr:rnb~rrsc~ sertioii through tlie dors:il iiinien sac of a 35 nini embryo showing the undiffereiitiated epitliclitiin. M, basement nicrnbrane. x 260. Trunsverse srctioii througli tlir dorsal riinirn sac of a 58 niin embryo. h'ote the striking epithelial prolifcration acc*oirip:i~iicd by ce1lul:rr projections into the lumen and the nppwr:iiir.r of intra cyitliehal vacuoles. M, basement meml,ra11e. x 260. Transverse section through the cnitlinc iegioii of tEic abomastmi of a 50 111111 tmhryo. Tlie epitlic~lium slion \ littlr ch:iIigc f r o m the carly embryonic condition. x 260. Trmsvcrsr srction throiigli tlic. c,iitli:ic. legion of thcx :il~oiiinsu~n of a 7 7 inm enil)r.~o \lion nig (,pi tlielinl pits aiid iliff c~reiitinting siniple coliiiiinar epithelium. x 260.

2.5

26

27

28

62

BOVINE S T O M S C H DEVELOPMENT I L D O N D. WAENER

PLATE 4

63

Documents

The Organogenesis and early histogenesis of the bovine ...ventral mesogastrium (future lesser omentum) connected the ventral curvature of the stomach to the liver. Between the stomach