Photoperiodic Regulatio onf Gametogenesi isn Sea Stars, with … · 2018-10-14 · Photoperiodic Regulatio onf Gametogenesi isn Sea Stars, with ... bu, t not in the sea star Patiria

AMER. ZOOL., 26:417-431 (1986)

Photoperiodic Regulation of Gametogenesis in Sea Stars, withEvidence for an Annual Calendar Independent of Fixed Daylength1'1

JOHN S. PEARSE, DOUGLAS J. EERNISSE,3 VICKI B. PEARSE,AND KATHERINE A. BEAUCHAMP

Institute of Marine Sciences and Biology Board of Studies, University of California,Santa Cruz, California 95064

SYNOPSIS. Gametogenesis and gonadal growth in the west coast sea star Pisaster ochraceusnormally begins in the fall and leads to large gonads full of gametes in the spring, whenspawning occurs. The timing of gametogenesis can be shifted simply by maintaining theanimals on a seasonally changing photoperiodic regime out of phase with ambient. Whenthey are kept on a spring-summer photoperiodic regime during the fall and winter, game-togenesis proceeds 6 mo ahead of schedule in the following spring and summer. Game-togenesis can be shifted out of phase even when the eyespots are removed. Short daylengthsthat normally occur during the fall and winter are not required for gametogenesis toproceed, nor are even the long daylengths of spring and summer that precede the initiationof gametogenesis in the fall. The temporal program is insensitive to fixed daylengths (LD15:9, 13:11, 12:12, 9:15) and appears to involve an endogenous calendar.

Shifting the photoperiodic regime 6 mo out of phase also leads to a shift of the game-togenic temporal program in the sea stars Leptasterias sp. (a brooder) and Asterias vulgaris(from the New England coast), but not in the sea star Patiria miniata. Gametogenic timingalso can be switched in the sea urchin Strongylocentrotus purpuratus but the mechanism ofthe photoperiodic response is fundamentally different; gametogenesis requires short day-lengths; continues indefinitely under a repeated short day, fall-winter photoperiod regime,and apparently does not involve an endogenous calendar. As photoperiodic responses areinvestigated further in these and other marine invertebrates, the models developed pri-marily from studies on terrestrial organisms may need to be extensively modified oradditional new models required.

INTRODUCTION

The seasonal production of gametes bysea urchins, sea stars, and other marineanimals has been a familiar fact of life forwell over a century, particularly to cell anddevelopmental biologists who havedepended on the gametes of these animalsfor research material. Until recently, how-ever, little attention has been paid to theenvironmental and physiological mecha-nisms that control this seasonality, or tothe ecological and evolutionary mecha-nisms that maintain it. Orton's (1920) clas-sic paper pointed to seasonal changes ofsea temperature as the major environmen-tal factor determining the timing of repro-

1 From the Symposium on Photoperiodism in theMarine Environment presented at the Annual Meetingof the American Society of Zoologists, 27-30 Decem-ber 1984, at Denver, Colorado.

* We dedicate this paper to Professor Arthur C.Giese on his 80th birthday, 19 December 1984.

5 Current address: Dr. Douglas J. Eernisse, FridayHarbor Laboratories, University of Washington, Fri-day Harbor, WA 98250.

ductive activity of marine animals. Suffi-cient correlative evidence accumulatedover the following decades for Thorson(1946) to propose that Orton's suggestiontake on the status of a general rule("Orton's Rule"); subsequent experimen-tal work showed that changes in sea tem-perature can modify reproductive timingin many marine species, especially in theNorth Atlantic (reviewed by Giese andPearse, 1974). Nevertheless, seasonallychanging sea temperatures are not likelyto be very influential in controlling the tim-ing of reproduction in species that live inareas where sea temperature undergoes lit-tle seasonal change {e.g., the west coast ofmuch of North America). Moreover, evenin areas with substantial seasonal changeof sea temperature, the sequence of eventsinvolved in gamete production often seemstoo precisely synchronized from place toplace and year to year to be determinedsolely by changing sea temperatures.

The importance of seasonally changingphotoperiods in synchronizing reproduc-tive activities of terrestrial plants and ani-

417

418 J. S. PEARSE ET AL.

mals has been recognized for nearly 50years, and a large and cumbersome liter-ature has accumulated on various ramifi-cations of the system. However, despiteGiese's (1959) suggestion that photoperi-odism may be important in the sea as well,and work by Richard (1971) signalling suchimportance, its role has only recently beenrecognized and more thoroughly exam-ined in both marine plants and animals (seepapers in this symposium).

Sea urchins and sea stars recently havejoined the ranks of animals known to havephotoperiodic responses. When reared ina photoperiodic regime of seasonallychanging daylengths that is 6 mo out ofphase with ambient {i.e., the longest day ofthe year is 21 December), individuals ofthe west coast sea urchin Strongylocentrotuspurpuratus both grow and produce gametes6 mo out of phase with those in the fieldand those reared in a seasonally changingphotoperiod regime in phase with that inthe field (Pearse et al., 1986). Gametogenicactivity is maintained, apparently contin-uously, when animals are kept on repeatedshort-day fall-winter photoperiod regimes,but suppressed when they are kept onrepeated long-day spring-summer photo-period regimes. The animals thus appearto be typical "short-day" organisms withrespect to gametogenesis, and there is noevidence that they possess an endogenousrhythm or calendar controlling growth orgametogenesis.

Photoperiodic control of growth of stor-age organs (pyloric ceca) and gonads alsohas been demonstrated in several speciesof sea stars, including Pisaster ochraceus byPearse and Eernisse (1982) and Asterias vul-garis by Pearse and Walker (1986), bothspecies with planktotrophic larvae, andLeptasterias sp. by Pearse and Beauchamp(1986), a brooding species without larvae.Both P. ochraceus and Leptasterias sp. occuron the west coast of North America wheretemperature fluctuations are moderate, butA. vulgaris occurs on the northeast coast ofNorth America where seasonal tempera-tures range over nearly 20°C. These obser-vations suggest that photoperiodic controlof gametogenesis is widespread in shallow-water sea stars. However, we found no evi-

dence of photoperiodic control of game-togenesis in another common west coastsea star, Patiria miniata (K. K. Davis and J.S. Pearse, unpublished observation).

Continuing work in our laboratory hasrevealed that the photoperiodic responsein Pisaster ochraceus is unlike that in Stron-gylocentrotus purpuratus. Individuals of P.ochraceus appear to be unresponsive to theshort days of fall and winter, but rathercan be caused to produce gametes out ofphase by exposure to the long days of springand summer. Although they thus seem tobe "long-day" animals, they are unrespon-sive to fixed daylengths, either short, neu-tral, or long, nor are long daylengths ofspring and summer necessary for game-togenesis to proceed in the fall. There isevidence in these animals for an endoge-nous calendar, which is perhaps set veryearly in life and which underlies the tem-poral program of gametogenesis. Thissymposium paper summarizes some of ourwork on P. ochraceus that is aimed at char-acterizing the nature of the photoperiodicresponse in these animals, and places it inperspective with current ideas about pho-toperiodic phenomena.

MATERIALS AND METHODS

Our basic experimental protocol is givenin Pearse and Eernisse (1982) and Pearseet al. (1986). Individuals of the sea starPisaster ochraceus, common intertidally onthe Pacific coast of North America, werecollected from an intertidal mussel bed inSanta Cruz, California. These animals weremaintained in tanks with running seawaterunder identical conditions of food (unlim-ited supply of mussels upon which all fednearly continuously), of sea temperature(ambient; ranging from 10—16°C in winterand spring to 12-18CC in summer and fall,see Pearse and Beauchamp, 1986), and ofdensity but under contrasting photoperi-odic regimes. In one light-tight room anastronomic time switch (R. W. Cramer &Co., Type SY Model SOL) turned the flu-orescent lights (G.E. F40D Daylight) on andoff in phase with local sunrise and sunset;the animals in this room were under an"ambient" or "in-phase" photoperiodicregime. In the adjacent room the astro-

PHOTOPERIODISM IN SEA STARS 419

nomic time switch was set to turn the flu-orescent lights on and ofF6 mo out of phasewith local sunrise and sunset so that thelongest day of the year was on 21 Decem-ber and the shortest day was 21 June; ani-mals in this room were under an "out-of-phase" photoperiodic regime. Behavior,feeding, growth, and gametogenic activitywere monitored in animals maintained inthe two rooms. Subsamples of 2 to 7 ani-mals were taken on a quarterly to semi-annual schedule, gonadal indexes deter-mined (gonadal index = gonadal wetweight x 100/total animal wet weight),and the gonads prepared and analyzed his-tologically. As found in earlier studies (e.g.,Giese, 1959), there was no statistical dif-ference between the gonadal indexes ofmales and females taken on any given date;the values therefore were combined forplotting and analyses.

In the first experiment, lasting fromDecember 1978 to August 1980, animalswere simply maintained in separate roomsunder one or the other of the contrastingphotoperiodic regimes; this experimentdemonstrated photoperiodic control ofgametogenesis in these animals as reportedby Pearse and Eernisse (1982). In the sec-ond experiment, using 60 animals and last-ing from March 1980 to December 1981,individuals were transferred from one ofthe light-tight photoperiod rooms to theother at 6 mo intervals; some experiencedrepeated spring-summer regimes ("longday") while others experienced repeatedfall-winter regimes ("short day"). In thethird set of experiments, lasting fromDecember 1981 to August 1983, 192 indi-viduals were maintained 16 each in 12 plas-tic laundry sinks (50 x 50 x 35 cm) cov-ered with light-tight wooden boxes, eachequipped with an overhead 24-inch flu-orescent light (G.E. F24D Daylight). Forsome of the boxes, the astronomic timerswere set to produce seasonally changingphotoperiods so that there were in-phase(ambient), 3-mo out-of-phase, and 6-moout-of-phase regimes. For others, 24-hourtimers (Intermatic, Models D - l l l andD-811) were set to produce fixed dailyhours of light (L) and dark (D)—LD 15:9,13:11, 12:12, and 9:15. In addition, to

examine the possibility that an "hourglass"model of photoperiodic control is involved(see Saunders, 1982), long days were inter-rupted by 1 and 5 hours of darkness everyday (i.e., LDLD 8:1:7:8 and 8:5:7:4). Finally,animals in one box were initially exposedto a fixed photoperiod of LD 15:9 for onemonth, then to LD 9:15 for the remainderof the experiment.

In the experiments described above, theanimals were killed upon dissection, so thelength of an experiment was determinedin part by the initial number of animalsmaintained (12 or more per treatment), thefrequency of subsampling, and the numberof animals in the subsamples. In order toinclude both males and females in the sub-samples, animals were sexed by examininggonadal tissue withdrawn with a hypoder-mic needle and syringe. In several addi-tional experiments small numbers of juve-niles were reared and maintained for fourto five years (the "entrainment experi-ments" presented at the end of the Resultssection); these individuals were biopsiedsemiannually, but not killed. Initially, a sin-gle gonad was removed from each animalthrough a small slit cut in the base of a ray,weighed to estimate the gonadal index(assuming all gonads had similar weights),and then fixed for histological analysis.However, often the wound did not heal,and the slit remained as an opening intothe coelom. Such animals appeared col-lapsed and stressed, and they died withinweeks to months. Later biopsies were doneby cutting off one whole ray with a razorblade and removing the gonads from thatray. After ray-removal, the animals healedwell, and the regenerating ray bud reacheda centimeter or more in length within about6 mo. The effect of regeneration on game-togenesis is unknown; it probably does notchange gametogenic timing, but maydepress gonadal growth (Harrold andPearse, 1980).

RESULTS

Switching experiments: Repeated"winters" and "summers"

Animals collected in March, when theycontained large gonads full of ripe gametes,all spawned in the laboratory in June


S O N D J F M A M J J A S O N D ' J F M A M J J A S O N D '1980 1981

Field Lab-FIG. 1. Changes in the gonadal indexes (males and females combined) of Pisaster ochraceus maintained ondifferent regimes of seasonally changing photoperiods. Black areas represent periods when daylengths wereless than 12 hr; photoperiods ranged from 9 hr at winter solstice to 15 hr at summer solstice as indicated atthe bottom of each panel. Sixty animals were brought into the laboratory in March 1980 and divided equallybetween photoperiod regimes that were in phase (lower two panels) and out of phase (upper two panels); halfof each group was switched to the alternate photoperiodic regime in September 1980. Sample sizes for eachdissection date were: September 1979 and March 1980, 10 animals (field samples); September 1980, 5 animals


regardless of photoperiodic regimes, andall had very small gonads in September (Fig.1). Moreover, all underwent a gameto-genic cycle between September and March,regardless of photoperiod regime, includ-ing those transferred from an in-phase toan out-of-phase regime ("2 summers"),those maintained out-of-phase ("2 win-ters"), and those transferred from an out-of-phase to an in-phase regime ("3 win-ters"). During the following year, thosereturned to in-phase conditions after beingmaintained for 6 mo under out-of-phaseconditions ("3 winters") followed the in-phase temporal program. In contrast, thosemaintained under the out-of-phase regime,beginning either in March ("2 winters") orin September after being held on the in-phase regime for 6 mo ("2 summers"),underwent a second gametogenic cycle inthe spring and had large gonads filled withfull-grown oocytes or sperms in August1981. These animals were seen spawningin December, 6 mo out of phase with thein-phase animals, and had small undevel-oped gonads upon the final December dis-section.

These experiments show that the game-togenic cycle (both in ovaries and testes) inthese animals can proceed 1) whether ornot it is preceded by the long days of springand summer, and 2) whether or not it isaccompanied by the short days of fall andwinter. Moreover, as found earlier (Pearseand Eernisse, 1982), when the animals weremaintained under long days of spring andsummer during the fall and winter theyinitiated a gametogenic cycle 6 mo early.

Phase-shifting experiments: 3-mo and6-rno out-of-phase regimes

Although Pearse and Eernisse (1982) haddemonstrated that the gametogenic pro-gram in Pisaster ochraceus could be shifted6 mo out of phase by placing the animalsunder a photoperiod regime 6 mo out of

phase, we needed also to determine theflexibility of the program. Could it be resetto any phase? After animals had been main-tained for 20 mo on a photoperiod regime3 mo out of phase with ambient, theirgonadal size and oogenic condition (esti-mated by analysis of oocyte sizes) wereintermediate between those of animalsmaintained on in-phase and 6-mo out-of-phase regimes (Fig. 2). The shift, and theintermediate status of the 3-mo out-of-phase animals, was evident within a year,in December, when those 6 mo out of phasewere seen spawning; the 3-mo out-of-phaseanimals were seen spawning in Februaryand April, and those under the in-phaseregime spawned in May, June, and July.

It should be noted that for unknown rea-sons gonadal growth was suppressed dur-ing the first spring in animals maintainedunder both 3-mo and 6-mo out-of-phaseregimes, although all the animals spawnedbetween May and July (Fig. 2). A similar,but more severe suppression (the animalsdid not spawn the first year) was seen inanimals maintained under an interruptedfixed photoperiod (LDLD 8:1:7:8).

The role of the terminal eyespots

Many sea stars, including Pisaster ochra-ceus, have an ocellus at the tip of each ray.Variation in light received by these ocellimodifies sea star behavior (Yoshida andOhtsuki, 1966). Moreover, transmissionelectronmicroscopic studies have demon-strated a nightly rhythm of membraneregeneration in the ocellar cells (Branden-burger and Eakin, 1980). Such a circadianrhythm could provide a mechanism formeasuring seasonally changing daylength(Pittendrigh, 1981a).

We tested the possibility that the eye-spots serve as the receptors for the pho-toperiodic response by surgically removingthe eyes and a small portion of tissue aroundthem (several mm2, including the terminal

per treatment (in phase and out of phase); March, August, and December 1981, 4 animals per treatment.(The remaining two provided the authors with an excellent dinner.) Arrows indicate when long spring-summer daylengths apparently influence the initiation of gametogenic cycles; dotted lines suggest course ofoverlapping gametogenic cycles.


^Jp=spawned 6-mo.Out-of-Phase

J F M A M J J

1 9 8 2

A S O N D J F M A M J J A S O N D

1 9 8 3

FIG. 2. Changes in gonadal indexes (solid lines) and size distributions of oocytes (shaded polygons) of Pisasterochraceus maintained under seasonally changing photoperiods with different phases. Animals in 3-mo out-of-phase treatment were initially shifted ahead from mid-December photoperiod to that equivalent of mid-March; those in 6-mo out-of-phase treatment were shifted to equivalent of mid-June. Initial sample size 10animals, thereafter field samples ranged from 5 to 10 animals, and laboratory samples ranged from 2 to 6animals; final laboratory sample sizes were 5 to 6 animals per treatment. Field animals presumably spawnedbetween February and June each year.

tube feet). Treated animals then weremaintained under both in-phase and 6-moout-of-phase photoperiodic regimes.Regeneration of the eyespots is rapid (seePenn and Alexander, 1980); we found smallregenerated ocelli within 3 to 4 weeks afterremoval. Consequently, we removed theeyespots every 3 weeks. The behavior ofthe surgically treated animals was affected;they rarely moved around the tanks andtheir feeding rates were less than half thoseof untreated animals.

Despite the obvious adverse effects ofeyespot removal, the gametogenic cycle wasvirtually identical between treated anduntreated animals maintained under thein-phase regime (Fig. 3). However, thosewith the eyespots removed were never seento spawn, and the gametes remaining inthe gonads of the August subsample oftreated animals were disintegrating relicts.Perhaps the eyespots have a role in spawn-ing (but see below).

The gametogenic cycle also was very


6 mo. Out-of-Phase,Eyes Removed

I IIn-Phase, Eyes Removed

F M A M J J A S O N D1982 I 9 8 3

FIG. 3. Changes in gonadal indexes (solid lines) and size distributions of oocytes (shaded polygons) ofPisasterochraceus maintained under seasonally changing photoperiods that were in phase and 6 mo out of phase. Theeyes (ocelli) at the tips of the rays were removed surgically every 3 to 4 weeks from one set of animalsmaintained under each photoperiod regime. "Relicts" refers to oocytes that were disintegrating, as seen inhistological preparations. Sample sizes for the April 1983 dissections were 7 animals each for the eye-removaltreatments and 4 animals each for the eye-intact treatments.

similar between treated and untreated ani-mals maintained under the 6-mo out-of-phase regime; in both sets of animals it hadshifted out of phase (Fig. 3). This resultsuggests that the eyespots are not the pho-toreceptors responsible for mediating thephotoperiodic response of gametogenesis.Moreover, although the treated, out-of-phase animals did not spawn the first year,a few were seen spawning in January oftheir second year, showing that spawningcan occur even in the absence of the ocelli.

Fixed daylengths

The gametogenic cycles were virtuallyindistinguishable among animals main-tained under fixed daylengths (Fig. 4),including long days (LD 15:9, and 13:11not shown), neutral days (LD 12:12), andshort days (LD 9:15); these cycles wereessentially the same as those in animalsmaintained under the in-phase photope-riodic regime. Only two differences werenoted. 1) Animals under long-day and neu-


150—1 *=spawned

100-

D 'J F M A M J1982 1983

FIG. 4. Changes in gonadal indexes (solid lines) and size distributions of oocytes (shaded polygons) of Pisasterochraceus maintained under fixed daylengths in contrast with those in animals maintained under seasonallychanging in-phase photoperiods. Final sample sizes for the August 1983 dissections were 6 animals for theLD 15:9, 9:15, and in-phase treatments, and 3 animals for the LD 12:12 treatment.

tral regimes spawned one to two monthsbefore short-day animals, again indicatinga possible role of light in spawning. 2)Vitellogenic oocytes, more than 50 /j,m indiameter, were found in August of bothyears only in the ovaries of long-day ani-mals (both LD 15:9 and 13:11). This obser-vation suggests that unseasonal gameto-genic activity is stimulated by longdaylengths, but is subsequently suppressedby other factors.

The gametogenic cycles of animalsmaintained on fixed but interrupted long-

day regimes also were similar to those ofin-phase animals (Fig. 5). However, as withthe animals on fixed long-day regimes, ani-mals on fixed "interrupted" regimes hadvitellogenic oocytes above 50 pm in diam-eter in August of both years. Moreover,in the animals maintained on the LDLD8:1:7:8 regime, the gametogenic cyclealready in progress at the start of theexperiment was severely suppressed,spawning was not seen the first year, andspawning was several months later the sec-ond year than in other long-day, neutral-


150-n

100—

8L:5Dt7L:4D

J F M A M J J

1982

A S 0 N D J F M A M J J A S O N D

1983

0

FIG. 5. Changes in gonadal indexes (solid lines) and size distributions of oocytes (shaded polygons) ofPisasterochraceus maintained under interrupted, fixed, long daylengths in contrast with those in animals maintainedunder seasonally changing in-phase photoperiods. Final sample sizes for the August 1983 dissections rangedfrom 5 to 7 animals.

day, and in-phase animals. It is not clearhow to interpret these findings.

Never was a complete out-of-phasegametogenic cycle produced by placing in-phase animals into any of the continuousfixed-daylength regimes. However, onesuch cycle appeared to result when animalswere placed under long days (LD 15:9) forone month in December-January, thentransferred to and maintained under a con-tinuous short-day regime (LD 9:15) (Fig.6). In this case large vitellogenic and full-grown oocytes were found in the Augustsample of the first year and spawningoccurred in December, as in the animalsheld 6 mo out of phase. However, the ani-mals also had large gonads full of gametesin April of the second year and their gonadswere small and their ovaries containedmainly very small oocytes in August of thesecond year, as in the animals held in phase.

The brief period of unseasonal long daysseems to have stimulated a single, extra,out-of-phase gametogenic cycle.

Entrainment experiments

The above experiments, that using neu-tral daylengths, in particular, indicate thatthe gametogenic cycle in Pisaster ochraceusis at least partly independent of photope-riod. There are at least two possible expla-nations. In the absence of photoperiod cues,the animals could 1) follow changes in someother environmental variable, such as insea temperature or food quality, or 2) fol-low changes in an endogenous calendar,analogous to the self-sustaining oscillatorsin circadian clocks (Pittendrigh, 19816). Ifan endogenous calendar is present, it mightbe possible to initiate or set it out of phase.Evidence for such a calendar would be thefinding that animals set out of phase would


6-mo. Out-of- Phase

£=1

150-1

100-

5 0 -

0

i

* = spawned

\

*

1

-15

- 1 0

- 5

l5L:9Dl s t mo(L) | - l5

J F M A M J J A S O N D ' J F M A M J J A S O N D

1982 1983FIG. 6. Changes in gonadal indexes (solid lines) and size distributions of oocytes (shaded polygons) of Pisasterochraceus maintained for one mo under fixed long daylengths (LD 15:9), then for the remainder of theexperiment under fixed short daylengths (LD 9:15), in contrast with those in animals maintained underseasonally changing photoperiods that were in phase and 6 mo out of phase with ambient photoperiods. Finalsample sizes for the August 1983 dissection ranged from 5 to 6 animals.

remain out of phase in the absence of pho-toperiodic cues {i.e., when maintained atLD 12:12), even when kept under identicalconditions of sea temperature and foodsupply as in-phase animals.

We made two experimental attempts toentrain animals out of phase and then testtheir ability to maintain an out-of-phasecycle under ambient sea temperatures andconstant food supplies. Juveniles between5 and 20 g wet weight were used in bothof these experiments in an attempt to avoidalready entrained cycles. Gonads do notbegin growth in animals less than 50-75g, and gametes have not been found inanimals less than 100 g (J. S. Pearse,unpublished data).

In one experiment 12 juveniles were col-lected in March 1981 and divided equallybetween the in-phase and out-of-phase

rooms. Because of their small sizes theywere maintained in plastic salad collandersand fed mussels less than 1 cm in length.After one year, when they had grown to amean wet weight of 124 g, they were trans-ferred to plastic dish pans, and the two panswere placed side-by-side in the out-of-phaseroom. Subsequently, every September andevery March, both pans of animals weretransferred to the alternate rooms so thatthe animals experienced repeated fall-win-ter photoperiod regimes, but not the longdays of spring and summer that can resetthe gametogenic cycle (Fig. 7, top).

The animals were first biopsied in March1983, after they had been held for 2 yearsand had reached a mean wet weight of 480g; the mean gonadal index (calculated bymultiplying the wet weight of one gonadby 10) of the in-phase entrained animals


In-Phose —" Winters" 6-mo. Out-of-Phase —" Winters"

Collect BiopsyJ - I I 1 1

80 8> 82 83 84

(6)

M A M J J A S O N D1983

F M A M J J A1984

M A M J J A S O N D1983

F M A M J1984

J A

FIG. 7. Changes in gonadal indexes of Pisaster ochraceus reared for one year under seasonally changingphotoperiods that were in phase or 6 mo out of phase with ambient photoperiods, then maintained underrepeated fall-winter photoperiods with daylengths less than 12 hr. Numbers in parentheses give number ofanimals surviving and sampled by biopsy on each date; all surviving animals were biopsied on each successivesampling date. Note temporal scales in top and bottom figures do not correspond.

was significantly (P < 0.05) larger than thatof the out-of-phase entrained animals (Fig.7, bottom); however, that of the latter waslarger than would be expected, and thegonads of both sets of animals containednumerous oocytes of various sizes or sper-matocytes and sperms, and they were indis-tinguishable histologically. In September1983, the gonadal index of the out-of-phaseentrained animals was significantly largerthan that of the in-phase entrained ani-mals. Moreover, unlike the gonads of thein-phase entrained animals, those of theout-of-phase animals were full of full-grownoocytes or sperms. Subsequent biopsies inMarch and August 1984, however, showedno differences between the two sets of ani-mals, and both sets followed the in-phasecourse of gametogenesis.

In the other entrainment experiment, 5juveniles were collected in March 1980 andreared in the out-of-phase room for threeyears. Three other juveniles were rearedin the in-phase room beginning in March1981. All these animals were biopsied inMarch 1983 and then placed under a neu-tral photoperiod (LD 12:12), after which

they were biopsied semiannually (Fig. 8).From March 1983 to March 1984, the twosets of animals remained 6 mo out of phasewith each other; the in-phase entrainedanimals had large gonads full of gametesin March of 1983 and 1984, and very smallgonads with very few gametes in Septem-ber 1983; the out-of-phase entrained ani-mals had large gonads full of gametes onlyin September 1983. However, by August1984, the remaining animals, all of whichwere entrained to be out of phase, hadgonads that were in every way like thoseof in-phase animals. Mortality of the biop-sied animals was high between March andAugust 1984 when three of the six animalsin the experiment died, perhaps as a resultof the repeated biopsies. Whether the smallsize and undeveloped state of the gonadsin these animals were due to their revert-ing to in-phase conditions or to stress,therefore, is unclear.

These entrainment experiments wereintriguing yet inconclusive. In both exper-iments, the gametogenic cycle in animalsentrained out of phase remained at leastpartly out of phase for the first year, indi-


InPhase— 12 L: 12 D

Collect Biopsy

6 mo. Out-of-Phase— I2UI2D

Collect Biopsy

1 1 1 1

79 80 ' 81 ' 82 ' 83 ' 84 7 9 r 6 O 8 1 ^ 8 2 8 3 8 4

a

10-

% 5Hoco

, (3)

(2)

M A M J J A S O N D J F M A

1 9 8 3 1 9 8 4

M A M J J A S O N D J F M A M J J A

1 9 8 3 1 9 8 4

FIG. 8. Changes in gonadal indexes of Pisaster ochraceus reared for two to three years under seasonallychanging photoperiods that were in phase or 6 mo out of phase with ambient photoperiods, then maintainedunder a fixed daylength of LD 12:12. Numbers in parentheses give number of animals surviving and sampledby biopsy on each date. Note temporal scales in top and bottom figures do not correspond.

eating that the persistence of the cycle inthe absence of photoperiodic clues is notdue to the animals following some otherseasonally changing environmental factor(e.g., slight seasonal changes in sea tem-perature). On the other hand, after abouta year, the gametogenic cycles in animalsentrained out of phase shifted into phasewith in-phase animals. This suggests thatthe endogenous calendar, if present, wasalready set very early in life and is resistantto all but temporary entrainment.

DISCUSSION

The photoperiodic control system thatwe describe for gametogenesis in Pisasterochraceus appears to be fundamentally dif-ferent from that known to date for othersystems. Even photoperiodism in the seaurchin Strongylocentrotus purpuratus seemsfundamentally different from that in P.ochraceus (Pearse etal, 1986), although bothspecies share the same body plan (echino-derm) and the same habitat (rocky shallow-water of western North America). More-over, both spawn gametes at about the sametime (winter and spring) and share similarcircannual rhythms (Halberg et al., 1986).In S. purpuratus gametogenesis appears to

require short daylengths of fall and winterto proceed, while in P. ochraceus gameto-genesis is nearly or completely insensitiveto short daylengths. Long daylengths ofspring and summer apparently suppressgametogenesis in S. purpuratus, but lead tothe initiation of a gametogenic cycle in thefollowing 3 months in P. ochraceus. Finally,while there are experimental indications ofan endogenous calendar underlying thegametogenic cycle of P. ochraceus, similarexperiments with repeated "summers" and"winters" provide no evidence for such acalendar in S. purpuratus. However, exper-iments have not been done with 5. purpur-atus using fixed daylengths and we cannotyet say with any certainty that an endog-enous calendar is not present.

Endogenous circannual rhythms or cal-endars are known to occur in a wide varietyof animals (Saunders, 1982; Halberg^ al.,1983), and the presence of such a calendarunder photoperiodic control in sea starswould not be unusual in itself. For exam-ple, Schwab (1971) showed photoperiodiccontrol of testicular activity in starlings byobserving that the testes remained largeand spermatogenic when the animals wereheld at daylengths less than 11 hr, but theywere always small and quiescent when held


at daylengths more than 13 hr. He furtherconcluded that there was an endogenouscircannual rhythm because the annual tes-ticular cycle continued when the birds wereheld on a photoperiod of LD 12:12 (butsee Hamner, 1971). In contrast, we foundthat the circannual gametogenic rhythm inPisaster ochraceus continued unmodified atLD 15:9, 13:11, and 9:15, as well as at12:12; the gametogenic rhythm could bephase-shifted only when the seasonal pho-toperiod rhythm was shifted, either 3 moor 6 mo out of phase.

The gametogenic cycle in Pisaster ochra-ceus seems similar to the circannual rhythmof ovarian growth in the catfish Hetero-pneustesfossilis; that rhythm persists for overa year at constant temperature and contin-uous light, continuous darkness, LD 14:10,12:12, and 9:15 (Sundararaj et al., 1973,1982). However, no attempt has yet beenmade to alter the rhythm by using season-ally changing daylengths as done with P.ochraceus, and there is no evidence of pho-toperiodic control of the rhythm. Suchmanipulation, indicative of photoperiodiccontrol, has been done with the cycle ofantler shedding and regeneration in deer(Goss, 1969a, b, 1980; Goss et al, 1974).Like the gametogenic cycle in P. ochraceus,the antler growth cycle can be shifted 6 moout of phase simply by shifting the seasonallight cycle 6 mo out of phase. Moreover,the antler shedding cycle is maintainedunder fixed photoperiods of LD 8:16,13:11, 16:8, and 24:0. On the other hand,unlike the gametogenic cycle in P. ochra-ceus, the antler shedding cycle is obliter-ated at fixed daylengths between LD 12:12and 13:11 (Goss, 19696) and this is due tothe 12-hr duration of the light or darkperiods (Goss, 1984). Goss et al. (1974)found that they could increase or decreasethe frequency of the antler growth cycleby changing the frequency of the changinglight cycles; it would be valuable to deter-mine whether the frequency of the game-togenic cycle in P. ochraceus could be sim-ilarly modified.

Olive and Garwood (1983) showed thatwhen individuals of the polychete Nereisdiversicolor are maintained under constanttemperatures and "static" daylength

regimes, they become gravid at differenttimes separated by about 240 days. Theseobservations were interpreted as evidencefor a long-term endogenous rhythm thatis "gated" and allows maturation to pro-ceed at particular times after birth. Envi-ronmental factors such as light and tem-perature would act mainly to modify thematuration time before or after the gate ispassed. Our finding that the gametogeniccycle of Pisaster ochraceus remains unal-tered at different fixed photoperiods is evi-dence for the presence of such a long-termendogenous rhythm or calendar. More-over, because the gametogenic cycle wasevident in animals reared from juvenilesthat were held under a 6-mo out-of-phasephotoperiodic regime for one year (Fig. 7),the rhythm apparently is set early in life.The presence of such an innate rhythmshould be tested by rearing animals result-ing from spawnings 6 mo apart. However,Olive and Garwood's model does notexplain how the gametogenic cycle can beshifted as much as 6 mo out of phase simplyby shifting the phase relationship of theannual photoperiodic cycle.

We were only able to shift the gameto-genic cycle of Pisaster ochraceus out of phaseby using an out-of-phase photoperiodicregime of seasonally changing daylengths;continuous exposure to either long or shortdaylengths had little or no effect on theoverall expression of the cycle. Still, itwould be premature to conclude that crit-ical daylengths are not involved in regu-lating the photoperiodic response in P.ochraceus. Several species of neuropteraninsects, for example, must experience achange from a critical short daylength toa critical long daylength to avert or ter-minate diapause (Tauber and Tauber,1982). Our seasonal switching experiments(Fig. 1) indicate that long daylengths ofspring and summer lead to the initiationof gametogenesis 6 mo out of phase withambient, and critical long daylengths maybe involved. In support of this suggestionwe succeeded in initiating an out-of-phasegametogenic cycle by exposing animals toLD 15:9 for one month in midwinter andthen maintaining them under LD 9:15(although the animals later reverted to an


in-phase cycle) (Fig. 6). Finally, femalesexposed to continuous light regimes of LD15:9 or 13:11 beginning in December con-tained vitellogenic oocytes the followingAugust (Fig. 4), indicating a precocious ini-tiation of gametogenesis. It is tempting topropose that some critical long daylengthcan initiate gametogenesis out of phase,and that long daylength needs to be fol-lowed by some critical short daylength forgametogenesis to continue. The problemwith this proposal, however, is our findingthat the gametogenic cycle continues inphase even when the spring-summer longdaylengths were repeated (2 "summers,"Fig. 1), or when the animals are maintainedcontinuously on LD 15:9 or 13:11 (Fig. 4).

Currently there are two main modelsproposed to explain photoperiodic phe-nomena (Saunders, 1982). The "hour-glass" model proposes that some criticalsubstance is produced or degraded over acritical period of light or dark, while the"circadian periodicity" or "Bunning'shypothesis" model proposes that a circa-dian oscillator measures daylength. Pitten-drigh (1981a) reviewed the evidence forthese models and concluded that most orall cases of photoperiodism can beexplained by the latter model. This con-clusion was reinforced by Pittendrigh et al.(1984), while Veerman and Vaz Nunes(1984) developed an "hour-glass time-oscillator counter" model that accountedremarkably well for photoperiodic induc-tion of diapause in spider mites. If the pho-toperiodic response of the gametogeniccycle in Pisaster ochraceus is indeed insen-sitive to fixed daylengths, it (and the antlergrowth cycle) cannot be readily accountedfor by either of these models. Rather, ourexperiments suggest that a new or muchmodified existing model is needed thatincorporates changing daylengths and/orseveral different critical daylengths. AsGoss et al. (1974) concluded 10 years ago:"In our present state of ignorance, we canonly continue to gather data on thisintriguing problem and hope that some daythe bewildering profusion of facts may fallinto place." Strange as it seems, sea starsappear to have joined deer in displaying aperplexing form of photoperiodism.

Because sea stars and deer are so differentin so many ways, this form of photoperi-odism may be widespread and merits care-ful attention.

ACKNOWLEDGMENTS

We thank Betsy Steele for coordinatingthe daily monitoring schedule of our ani-mals, and work-study students Lisa Borok,Ted Cranford, Kathy Embrey, JenniferHolder, Genine Scelfo, and Denise Scra-maglia for faithfully checking them overthe years; Gary McDonald for constructingthe light-tight boxes; Diana Custer, Rich-ard Goss, Franz Halberg, and Nancy Mar-cus for suggestions on the manuscript; andWilliam Doyle, Director of the Institute ofMarine Sciences, University of California,Santa Cruz, for encouragement and finan-cial support.

REFERENCES

Brandenburger, J. L. and R. M. Eakin. 1980. Cyto-chemical localization of acid phosphatase in ocelliof the sea star Patiria miniata during recycling ofphotoreceptoral membranes. J. Exp. Zool. 138:127-140.

Giese, A. C. 1959. Reproductive cycles of some westcoast invertebrates. In R. B. Withrow (ed.), Pho-toperiodism and related phenomena in plants and ani-mals, pp. 625-638. Amer. Assoc. AdvancementSci., Washington, D.C.

Giese, A. C. and J. S. Pearse. 1974. Introduction:General principles. In A. C. Giese and J. S. Pearse(eds.), Reproduction of marine invertebrates, Vol. 1,pp. 1-49. Academic Press, New York.

Goss, R. J. 1969a. Photoperiodic control of antlercycles in deer. I. Phase shift and frequencychanges. J. Exp. Zool. 170:311-324.

Goss, R. J. 1969ft. Photoperiodic control of antlercycles in deer. II. Alterations in amplitude. J.Exp. Zool. 171:223-234.

Goss, R. J. 1980. Photoperiodic control of antlercycles in deer. V. Reversed seasons. J. Exp. Zool.211:101-105.

Goss, R. J. 1984. Photoperiodic control of antlercycles in deer. VI. Circannual rhythms on alteredday lengths. J. Exp. Zool. 230:265-271.

Goss, R. J., C. E. Dinsmore, L. N. Grimes, and J. K.Rosen. 1974. Expression and suppression of thecircannual antler growth cycle in deer. In E. T.Pengelley (ed.), Circannual clocks, pp. 393-422.Academic Press, New York.

Halberg, F., M. Lagoguey, and A. Reinberg. 1983.Human circannual rhythms over a broad spec-trum of physiological processes. Interntl. J.Chronobiol. 8:225-268.

Halberg, F., K. Shankaraiah, A. C. Giese, and F. Hal-berg. 1986. The chronobiology of marine inver-tebrates: Methods of analysis. In A. C. Giese and


J. S. Pearse (eds.), Reproduction of marine inverte-brates, Vol. 9. Blackwell Scientific/Boxwood, PaloAlto. (In press)

Hamner, W. M. 1971. On seeking an alternative tothe endogenous reproductive rhythm hypothesisin birds. In M. Menaker (ed.), Biochronometry, pp.448-462. Nat. Acad. Sci., Washington, D.C.

Harrold, C. and J. S. Pearse. 1980. Allocation ofpyloric caecum reserves in fed and starved seastars, Pisastergiganteus (Stimpson): Somatic main-tenance comes before reproduction. J. Exp. Mar.Biol. Ecol. 48:169-183.

Olive, P. J. W. and P. R. Garwood. 1983. The impor-tance of long term endogenous rhythms in themaintenance of reproductive cycles of marineinvertebrates: A reappraisal. Int. J. Invert.Reprod. 6:339-347.

Orton, J. H. 1920. Sea temperatures, breeding anddistribution in marine animals. J. Mar. Biol. Ass.,U.K. 12:339-366.

Pearse, J. S. and K. A. Beauchamp. 1986. Photo-periodic regulation of feeding and reproductionin a brooding sea star from central California.Interntl. J. Invert. Reprod. Develop. (In press)

Pearse, J. S. and D.J. Eernisse. 1982. Photoperiodicregulation of gametogenesis and gonadal growthin the sea star Pisaster ochraceus. Mar. Biol. 67:121-125.

Pearse, J. S., V. B. Pearse, and K. K. Davis. 1986.Photoperiodic regulation of gametogenesis andgrowth in the sea urchin Strongylocentrotus pur-puratus. J. Exp. Zool. 237:107-118.

Pearse.J.S. and C.W. Walker. 1986. Photoperiodicregulation of gametogenesis in a North Atlanticsea star, Asterias vulgaris. Interntl. J. Invert.Reprod. Develop. 9:71-77.

Penn, P. E. and C. G. Alexander. 1980. Fine struc-ture of the optic cushion in the asteroid Nepanthiabelcheri. Mar. Biol. 58:251-256.

Pittendrigh, C. S. 1981a. Circadian organization andthe photoperiodic phenomena. In B. K. and D.E. Follett (eds.), Biological clocks in seasonal repro-ductive cycles, pp. 1—35. John Wright, Bristol.

Pittendrigh, C. S. 1981 A. Circadian systems: Entrain-ment. In Handbook of behavioral neurobiology 7:95-124. Plenum Press, New York.

Pittendrigh, C. S., J. Elliot, and T. Takamura. 1984.The circadian component in photoperiodicinduction. In Photoperiodic regulation of insect andmolluscan hormones. Ciba Found. Symp. 104:26—41. Pitman, London.

Richard, A. 1971. Action qualitative de la lumieredans le determinisme du cycle sexuel chez cepha-lopode Sepia officinalis L. C. R. Hebd. Seanc. Acad.Sci., Paris (Ser. D) 272:106-109.

Saunders, D. S. 1982. Photoperiodism in animals andplants. In J. Brady (ed.), Biological timekeeping, pp.65-82. Cambridge University Press, Cambridge,U.K.

Schwab, R. G. 1971. Circannian testicular periodicityin the European starling in the absence of pho-toperiodic change. In M. Menaker (ed.), Biochron-ometry, pp. 428-445. Nat. Acad. Sci., Washing-ton, D.C.

Sundararaj, B. I., S. Vasal, and F. Halberg. 1973.Circannual rhythmic ovarian recrudescence inthe catfish, Heteropneustes fossilis. Interntl. J.Chronobiol. 1:362-363.

Sundararaj, B. I., S. Vasal, and F. Halberg. 1982.Circannual rhythmic ovarian recrudescence in thecatfish Heteropneustes fossilis (Bloch). In R. Taka-hashi, F. Halberg, and C. A. Walker (eds.), Towardchronopharmacology, pp. 319-337. Pergamon Press,New York.

Tauber, C. A. and M.J. Tauber. 1982. Evolution ofseasonal adaptations and life history traits inChrysopa: response to diverse selective pressures.In H. Dingle and J. P. Hegmann, Jr. (eds.), Evo-lution of genetics and life histories, pp. 51-72.Springer-Verlag, New York.

Thorson, G. 1946. Reproduction and larval devel-opment of Danish marine bottom invertebrates.Medd. Komm. Danmarks Fiskeri-og Havunders.,Serie:Plankton 4:1-523.

Veerman, A. and M. Vaz Nunes. 1984. Photoperiodreception in spider mites: Photoreceptor, clockand counter. In Photoperiodic regulation of insectand molluscan hormones. Ciba Found. Symp. 104:48-59. Pitman, London.

Yoshida, M. and H. Ohtsuki. 1966. Compound ocel-lus of a starfish: its function. Science 153:197—198.

Documents

Photoperiodic Regulatio onf Gametogenesi isn Sea Stars, with … · 2018-10-14 · Photoperiodic Regulatio onf Gametogenesi isn Sea Stars, with ... bu, t not in the sea star Patiria