ELSEVIER Journal of Experimental Marine Biology and Ecology,

198 (1996) 269-289

JOURNAL OF EXPERIMENTAL MARINE BIOLOGY AND ECOLOGY

Synchronization of endogenous tidal vertical migration rhythms in laboratory-hatched larvae of the crab Carcinus

maenas

Chaoshu Zeng”, Ernest Naylor

School of Ocean Sciences, University of Wales-Bangor, Menai Bridge, Gwynedd LL59 5EY, UK

Received 30 August 1995; revised 13 December 1995; accepted 20 December 1995

Abstract

Previous studies showed that freshly collected zoea-I larvae of the shore crab Curcirrus maenas

from coastal waters possessed endogenous circatidal vertical migration rhythms. Present experi- ments demonstrate that zoea-1 larvae released by crabs which extruded eggs in the field but were

then kept in laboratory constant conditions since collection, also exhibited endogenous circatidal vertical migration rhythms. Such rhythms were shown by newly hatched larvae from nearly all of 50 females which released larvae in the laboratory, some of which had carried eggs in the laboratory for up to 3-4 months. Thus, since entrainment by tidal variables could not have occurred during embryonic development in these cases, other synchronising factors must set the phase of the larval circatidal rhythms. Present results suggest that the hatching process itself is the

main synchroniser; upward swimming occurred just after hatching and at 12.4 h intervals thereafter. This is consistent with previous observations of peak hatching in Carcinus primarily around nocturnal high tide, with larvae in the field exhibiting tidal vertical migration rhythms of ebb-phased upward swimming associated with dispersal during ebbing tides. The results appear to be the first to characterize a synchroniser for tidal swimming rhythms of planktonic larvae and to provide experimental evidence that the hatching process can serve as the synchroniser for a larval swimming rhythm.

Keywords: Carcinus maenas; Endogenous tidal vertical migration rhythms; Hatching; Larvae; Synchronizer

1. Introduction

Endogenous tidal rhythms of vertical migration in plankton species have been reported so far only in zoea larvae of the crab Rhithropanopeus harrisii (Cronin and

*Corresponding author

0022.0981/96/$15.00 0 1996 Elsevier Science B.V. All rights reserved

P/I SOO22-098 1(96)00004-4

Forward, 1979, 1983), the copepod Eurytemora afjnis (Hough and Naylor, 1992) and megalopae of the fiddler crabs Uca spp. (Tankersley and Forward, 1994), all from estuaries. In estuarine species, endogenous tidal vertical migration rhythms have been regarded typically as the behavioural basis of estuarine retention (Cronin and Forward,

1979, 1982; Cronin, 1982; Hough and Naylor, 1991, 1992) or of estuarine re-invasion mechanisms (De Vries et al., 1994; Tankersley and Forward, 1994), permitting

utilization of vertical variations in velocity and direction of tidal currents. Although the synchronization processes of circatidal rhythms in intertidal animals are

well documented (for reviews, see Palmer, 1974; Enright, 1975; DeCoursey, 1983; Naylor, 1985), synchronization of tidal rhythms in planktonic animals is poorly understood. Most previous studies have been descriptive of endogenous tidal vertical migration rhythms of freshly collected animals (Cronin and Forward, 1979; Hough and

Naylor, 1992; Tankersley and Forward, 1994). The only exception concerns experiments with Rhithropanopeus harrisii (Cronin and Forward, 1983), in which laboratory-hatched

larvae from crabs collected just before larval release were tested. For these larvae, a weak tidal migration rhythm was observed and considered to be entrained by tidal events

in the field during embryonic development (Cronin and Forward, 1983). However, no specific synchroniser has been identified experimentally.

Our previous studies have revealed that zoea-1 larvae of the common shore crab Carcinus maenas (L.), freshly collected from coastal waters, exhibited endogenous tidal vertical migration rhythms of ebb-phased upward swimming which appear to assist

offshore dispersal of the larvae (Zeng and Naylor, 1996). The present paper reports results of experiments with laboratory-hatched zoea-1 from female crabs which extruded

their eggs in the field but were then kept in laboratory constant conditions since collection. We sought to investigate whether such larvae, which had never experienced tidal influences as free-living individuals, exhibit tidal migration rhythms and, if so, how

such rhythms are synchronized.

2. Materials and methods

2. I. Berried females collection and maintenance

During the spring and summer of 1993 and 1994, berried females of the crab Carcinus

maenas were collected by either bottom beam trawling from a small boat at the sandy

beach of Traeth Malynog (53”09’N; 04”19’W), or searching manually under rocks during low tides on a rocky shore beneath the Menai Suspension Bridge (53”13’N; 04”09’W). Both sites are located along the coast of the Menai Strait, North Wales, where freshly collected zoea-1 larvae have been shown to express endogenous circatidal vertical migration rhythms in the laboratory (Zeng and Naylor, 1996). The tides in the Menai Strait are typical semidiurnal tides with a period of approximately 12.4 h (Admiralty Tide Tables, 1995).

After collection, all berried females were immediately labelled on their carapace with the site and date of collection and an assigned number. At the same time a few eggs were detached and the stage of embryo development determined following Boolootian et

C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ecol. 198 (1996) 269-289 271

al. (1959) and Brown and Loveland (1985). The females were then kept communally in

a few big tanks in the laboratory, each with a continuous flow-through of seawater and a

layer of sand as a substrate. The crabs were fed with frozen mackerel or mussels every few days. Water temperatures approximated to those in the field, but aquarium lights were kept on continuously. Crabs with late-stage embryos were removed and put into individual hatching tanks with mesh-protected overflows which retained larvae.

During the two year study period, a total of 250 berried crabs were collected. Larvae hatched from many of these females, which carried eggs ranging from newly extruded to imminent hatching when collected. From among these, the larvae of 50 randomly

selected females were studied for their endogenous migration rhythms. Larval release

occurred from within a few hours to more than 100 days after collection.

2.2. Recording of larval migration rhythm.

In each experiment, between 1000-2000 newly hatched larvae were placed in an actograph system which consisted of a chamber containing seawater and two rows of four infrared transmitters and receivers in an array on each side of the chamber. The chamber was constructed of 6mm Perspex with interior dimensions of 40 cm high, 15

cm wide and 5 cm across. The transmitter/receiver blocks of four, equally spaced along the 15 cm width of the chamber, were arranged in two paired groups facing each other at a distance of 9cm across the narrow section of the chamber. One group of sensors was

placed very close to the water surface, and the other close to the bottom.

During the experiments, an event was recorded each time an infra-red light beam was interrupted by larvae swimming across it. All channels were monitored by a BBC model

B microcomputer and every 15 mins the cumulative sum of beam interruptions in each channel was loaded to a cassette recorder and recorded. A header-tank with a tiny pipe of 1.0 mm diameter provided a very slow clean seawater supply (approx. 150ml/h) into

one end of the chamber, and an outflow pipe was placed at the other end, protected by a mesh panel of 0.2mm aperture to prevent loss of larvae. All experiments were carried out in constant darkness, allowing a l-2 h acclimation period before recording began. Once an experiment started, the whole system was left undisturbed and larvae were not

fed during the experiments. During all experiments except the temperature shock trial. water temperature was maintained at 16% 1°C and salinity varied between 30-34%~.

The records of four channels in the top and the bottom block of sensors were summed and plotted as “swimming activity in top channels” and “swimming activity in bottom channels” against elapsed time respectively. Data plots are of raw data and of periodogram analysis performed as described by Williams and Naylor (1978).

2.3. Experimental rationale

Early experiments showed that newly released larvae from females which had been kept in the laboratory for up to 3-4 months still exhibited clear circatidal vertical migration rhythms (see Section 3). These observations clearly preclude the possibility of entrainment by tidal variables during embryonic development. Moreover, there was no likelihood of entrainment by female circatidal rhythms of locomotor or ventilation

212 C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. I98 (19%) XT%289

activity which would not be apparent after such lengths of time in the laboratory (Naylor, 1985). Also, there could have been no tidal cues of temperature or salinity changes in the tanks in which the females were held, because the laboratory circulation system consisted of two very large storage tanks in series. Experiments were therefore

designed to test whether synchronization was by (1) temperature and/or light/dark

changes when larvae were transferred to the experimental conditions; (2) handling effects or (3) the hatching process. These involved (i) varying the start-times of

experiments in relation to hatching times and the 24 h cycle; (ii) varying the times of introducing batches of larvae into experimental conditions from a single hatch and (iii) varying the temperature in the recording chamber during the course of recording an experiment. In some cases the precise time of hatching was determined by maintaining

ovigerous females which were about to release larvae individually in transparent tanks and continuously monitoring their behaviour with a time-lapse video recorder

(Panasonic model WV-1800 high resolution camera and AG-6720 VHS video recorder) in constant light. In this way, it was possible to obtain larvae known to have been released at particular times of the day.

3. Results

During the hatching seasons of 1993 and 1994 about 50 experiments were carried out using laboratory hatched larvae of Cur&us maenas. In nearly all of these, endogenous circatidal rhythms of vertical migration were apparent (Fig. lA, Fig. 2A and Fig. 3A), even though females had been kept in the laboratory away from the influence of tides for up to 100 days (Fig. 3A). Peaks of swimming activity recurred at approximately 12.4 h

intervals in records from both top and bottom channels, the reciprocal pattern of the records in the two channels indicating clear rhythms of circatidal vertical migration.

Circatidal rhythmicity is confirmed by the periodogram analysis (Fig. lB, Fig. 2B and Fig. 3B), and none of the raw data records showed any indication of circadian modulation of the endogenous circatidal rhythms (Fig. 1 A, Fig. 2A and Fig. 3A). Larval

circatidal vertical migration rhythms appear not to be phased to the starting time of an experiment (Figs. l-8).

The results of an experiment in which two batches of larvae released at the same time from the same female were introduced into experimental conditions at 2 different times,

6 hours apart in tidal antiphase, are illustrated in Fig. 4. The initial 31 h of records are of one group and the subsequent traces are the group introduced 6 h later. The phases of tidal vertical migration rhythms of the two batches of larvae are clearly identical, further suggesting that tidal migration rhythms of laboratory hatched larvae were not phased by either the sudden environmental changes during the transfer or by handling when experiments were set up.

The results of an experiment in which a group of larvae was subjected to a sudden temperature rise of 6°C the maximum extent of a temperature change which larvae might have experienced in experiments such as those illustrated in Figs. 1-3, are illustrated in Fig. 5. At the point when water temperature in the recording chamber was

C. Zeng, E. Naylor If. Exp. Mar. Biol. Ed. 198 (1996) 269-289 273

500

150

250

s E

*

Fig. I. (A) Swimming activity records of laboratory-hatched C. maenaS larvae showing circatidal verttcal

migration rhythms in constant conditions. Upper and lower part of the graph represent records at the surface

and bottom of a vertical chamber. The larvae hatched from an ovigerous crab (precise hatching time was not

determined) kept in laboratory constant conditions without tidal influences for 14 days. The crab was collected

at Traeth Malynog, Menai Strait on 16 June, 1993 with eggs showing clear heartbeat, well formed eyes and

body pigmentation (stage 8 following Boolootian et al. (1959) and Brown and Loveland (1985)). The

experiment started at 20:OOh on 30 June 1993 with an initial number of 1400 newly hatched larvae; 1145

survived when the experiment stopped. (B) Periodogram analysis of the data presented in panel A. Upper and

lower figures represent the periodogram statistic of swimming activity records of the top and bottom channels,

respectively. 95% confidence limits are derived after randomization of the original data (see Williams and

Naylor, 1978).

214 C. Zeng, E. Naylor I J. Exp. Mar. Biol. Ecol. 198 (1996) 269-289

25

2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

25

0 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

Fig. I. (conrinued)

raised from 16°C to 22°C there was a transient effect on the swimming pattern but after a few hours the initial phasing of the swimming rhythm was resumed.

The fact that larval circatidal vertical migration rhythms were not consistently phased to the starting time of an experiment (see Figs. l-8), together with the above results, suggests that environmental changes when transferring larvae and handling them when experiments were set up do not set the phase of tidal migration rhythms of laboratory hatched larvae of C. maenas. Indeed, no matter which treatment was imposed during the present experiments, a consistent phase-relationship was found between larval hatching time and the timing of upward swimming of larvae. In 15 cases in which larval hatching times were precisely established by direct observations or from video records, peaks of

C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. 198 (1996) 269-289 21s

150 ," ,' " ",', "'I ",I' ," ,,,' " ","' ',, " ',,"' ',I' ",',, " "" 0 3 6 0 12 15 IS 2, 24 27 30 33 36 39 42 45 48

0 3 6 9 12 15 18 21 24 27 30 33 36 39 42 45 48

Elapsed time (h)

Fig. 2. (A) Swimming activity records of laboratory-hatched C. ~IXVUS larvae showing circatidal vertical

migration rhythms in constant conditions. The larvae hatched from an ovigerous crab (precise hatching time

was not determined), kept in laboratory constant conditions for 87 days. The crab was collected at Traeth

Malynog on 2 February, 1994 with eggs of approximately I /2 yolk and I /2 transparent embryo with eye

pigment not yet shown (stage 4). The experiment started at l9:OO on I May 1994, with an initial number ot

1200 newly hatched larvae, most larvae survived the entire experiment, (B) Periodogram analysis of the data

presented in A. Symbols and further details as in the legend to Fig. I.

216 C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. IYX (1996) 269-2X9

25

2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 10 19 20 21 22

50 I

Period (h)

10

0 2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

Fig. 2. (conrinued)

abundance at the top of the test chamber consistently occurred soon after every 12.4 h interval from the time of hatching (Fig. 6A, Fig. 7A and Fig. 8A show examples). Again, periodogram analysis confirmed that the rhythms were of circatidal period (Fig. 6B, Fig. 7B and Fig. 8B).

4. Discussion

Present experiments demonstrate that, like freshly collected Curcinus zoea-I from the field (Zeng and Naylor, 1996), larvae hatched from laboratory-held but field-collected

C. Zeng, E. Naylor I J. Exp. Mar. Biol. Ecol. 198 (1996) 269-289 217

r

Fig. 3. (A) Swimming activity records of laboratory-hatched C. marnus larvae showing circatidal vertical

migration rhythms in constant conditions. The larvae hatched from an ovigerous crab (precise hatching time

was not determined) kept in laboratory constant conditions for 100 days. The crab was collected at Traeth

Malynog on I March 1994 and was apparently newly spawned with eggs at the cleavage stage (stage I). The

experiment started at 14:OO h on 9 June 1994, with an initial number of 1200 newly hatched larvae, most

larvae survived the entire experiment. (B) Periodogram analysis of the data presented in A. Symbols and

further details as in the legend to Fig. I.

Elapsed time (h)

278 C. Zag, E. Naylor I .I. Exp. blur. Biol. Ed. 198 (I9961 269-289

I,, “‘,I “““1,‘1”1,’ ” “I’ ““,,‘,‘,f” I,,’ 2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

J 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

Fig. 3. (conrinued)

ovigerous females also displayed synchronised circatidal vertical migration rhythms.

Such synchronised rhythms were expressed despite the fact that the larvae had never experienced tidal conditions as free-living individuals. Moreover, some were hatched from females that had been held in the laboratory in the absence of tides for 3-4 months. It is possible that larvae hatched from females held in the laboratory for only a few days could have been cued by circatidal rhythms of locomotor activity of the female, but overt rhythms of such periodicity would have been lost after several days in the laboratory (Naylor, 1985). Moreover, video recordings of many crabs before hatching failed to reveal rhythmic patterns of egg ventilation behaviour. The question therefore arose as to how such rhythms are synchronized. In these circumstances, it is not reasonable to consider that synchronization occurs by exposure of developing embryos

C. Zeng, E. Naylor J J. Exp. Mar. Biol. Ecol. 198 (1996) 269-289 279

0 4 8 12 16 20 24 28 32 36 40 44 48 52 56 60 64 68 72 76 80 84

Elapsed time (h)

Fig. 4. Swimming activity records of two batches of C. maems larvae from the same release event of a single

crab, with one batch introduced to experimental conditions 6 h later than the other. The larvae hatched at night

on I3 July 1993 and the first 1500 larvae were introduced to the actograph at 14:30 h on 14 July. Both groups

of larvae were maintained under identical conditions in the recording room. The record of the first batch of

larvae terminated after 31 h and, under dim red illumination, the second batch of larvae was switched into the

recording system (indicated by arrows). Vertical broken lines are at 12.4 h intervals from the beginning of the

experiment. Larvae hatched 27 days after collection, from a female collected at Traeth Malynog on 16 June

1993, with eggs of approximately l/2 yolk and l/2 transparent embryo (stage 4).

C. Zen~, E. Nqlor I J. Exp. Mar. Biol. Ecol. 198 (1996) 269-289

0 4 6 12 16 20 24 26 32 36 40 44 46 52 56 60 64 68 72 76 60 84 68 92 96 100

Elapsed time (h)

Fig. 5. Swimming activity records of laboratory-hatched C. maena~ larvae before and after a 6°C temperature

rise. The experiments started at 22:00 h on 18 June at a water temperature of 16°C. After 42 h, the record was

interrupted and, under dim red illumination, water in the test chamber was drained and replaced with seawater

of the same salinity at 22°C (indicated by arrows). Vertical broken lines indicate 12.4 h intervals from the

beginning of the experiment. Second gap on the top channel record indicates temporary malfunction of the

channels. The larvae hatched 20 days after collection, from a female collected at Traeth Malynog on 29 May

1993, which carried eggs with visible trace of brown eye pigment (stage 5).

C. Zeng, E. Naylor I .I. Exp. Mar. Bid. Ed. 198 (1996) 269-289 281

Elapsed time (h)

Fig. 6. (A) Swimming activity records of laboratory-hatched C. rnaenas larvae for which the precise hatchmg

time was known. Arrows indicate 12.4 h intervals from hatching. The female was collected on a rocky shore at

Menai Bridge on 21 April 1994, with eggs of approximately I /2 yolk and I /2 transparent embryo (stage 4).

Hatching occurred around 10:3Oh, 28 May 1994, 37 days after collection. The experiment started at 18:OO h on

28 May with an initial number of 1800 newly released larvae; 1395 survived when the experiment stopped. CB)

Periodogram analysis of the data presented in panel A. Symbols and further details as in the legend to Fig. I.

282 C. Zeng, E. Nuylor I .I. Exp. Mar. Bid. Ed. 198 (1996) 269-289

40

0 2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 16 19 20 21 22

Period (h)

J -2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 16 19 20 21 22

Period (h)

Fig. 6. (continued)

to tidal conditions in the sea which are known to entrain locomotor rhythmicity in this

crab (Naylor and Williams, 1984). Other synchronizing factors should therefore be considered.

Firstly, it was necessary to consider whether the phasing of circatidal swimming rhythms in laboratory-hatched zoeae was a laboratory phenomenon induced by en- vironmental changes when the experiments began. Although temperature shock has been shown to re-initiate or reset tidal locomotor rhythms in adult C. maenas (Naylor, 1963; Williams and Naylor, 1967) and light-dark changes to maintain and reset tidal hatching rhythms in the semi-terrestrial crab Sesarma pictum (Saigusa, 1992) larval circatidal vertical migration rhythms in the present study were not phased to the starting time of an experiment (Figs. l-8). Hence, environmental changes and handling when setting up the

C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. 198 (1996) 269-289 283

0 4 8 12 16 20 24 28 32 36 40 44 48 52 56 60

Elapsed time (h)

Fig 7. (A) Swimming activity records of laboratory-hatched C. marrxas larvae for which the precise hatching time was known. Arrows indicate 12.4 h intervals from hatching. The female was collected at Traeth Malynog

on 23 June 1994, with eggs showing a clear heartbeat, well formed eyes and body pigmentation (stage 8).

Hatching occurred around 01:05 h on I July 1994, 8 days after collection. The experiment started at 0O:OO h on

2 July with an initial number of 1200 newly released larvae of which 989 survived when the experiment

stopped. (B) Periodogram analysis of the data presented in panel A. Symbols and further details as in the legend to Fig. I.

284 C. Zeng, E. Nqdor I J. Exp. Mar. Bid. Ed. I98 (1996) 269-289

40 \ B

0 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

0’,,,~,,‘,,‘,.“,~,‘,~,,,~‘,,1”,1 1”1’,11,‘1,‘11’,,1,” ‘,,l,“l”‘.,‘.i,,‘,“~J 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Period (h)

Fig. 7. (conrimed)

experiments clearly did not set the phase of tidal rhythms in the laboratory-hatched

Carcinus zoeae. Furthermore, the results of experiments specifically designed (Fig. 4 and Fig. 5) to investigate such effects precluded that possibility.

In contrast, larval hatching time showed a consistent phase relationship with the zoeae vertical migration rhythms recorded in the present study, suggesting that it is the hatching process which phases larval tidal migration rhythms. The hatching of the species was characterized by fierce pumping of the abdomen and tearing of the egg plugs by pereiopods, resulting in clouds of larvae being released in a matter of minutes. At this time, the animals stand erect, supported only by a pair of full-stretched pereiopods. This behaviour can be distinguished from egg cleaning and ventilation behaviour (Broekhuysen, 1936) during which the crabs adopt a much lower position,

C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. 198 (1996) 269-289 285

4 8 12 16 20 24 28 32 36 40 44 48 52 56 60 64 68 72

Elapsed time (h)

Fig. 8. (A) Swimming activity records of laboratory hatched C. mcrmas larvae for which the precise hatching

time was known. Arrows indicate 12.4 h intervals from the hatching. The female was collected at Traeth

Malynog on 23 June 1994, with eggs of approximately 2/3 yolk and I /3 transparent embryo (stage 3).

Hatching occurred at 16:45 h, 15 July 1994, 22 days after collection. The experiment started at 22:00 h. 16

July with an initial number of 1200 newly released larvae, most of which survived the entire experiment. (B)

Periodogram analysis of the data presented in panel A. Symbols and further details as in the legend to Fig. I.

286 C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ed. 19X (1996) 269-289

2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 16 19 20 21 :

Period (h)

30, .---7

0 2 3 4 5 6 7 6 9 10 11 12 13 14 15 16 17 16 19 20 21 22

Period (h)

Fig. 8. (conrinurd)

supported by a few pairs of flexed pereiopods, with the egg plug moving gently.

Hatching in C. maenas normally consists of 2-4 main bursts which, in the laboratory, occurred at circadian or circatidal intervals (Zeng and Naylor, submitted). Thus, we were able to predict and witness hatching in several cases. In our experiments in which larval hatching times were established precisely by visual observation and/or video records, results show that upward swimming consistently peaked soon after hatching, and at 12.4 h intervals thereafter. Similarly, freshly collected zoea-1 larvae displayed endogenous circatidal migration rhythms in which peak upward swimming consistently followed the time of expected high tide. In field studies, it has also been demonstrated that first stage zoeae of C. maenas were most abundant at the water surface during ebb tides. As a consequence, upward swimming after the time of high tide by newly released

C. Zeng, E. Naylor I J. Exp. Mar. Bid. Ecol. 198 (1996) 269-289 287

zoeae of C. maenas has been proposed as an offshore dispersal mechanism (Zeng and

Naylor, 1996). We have unpublished data indicating that, like many other crabs

(Forward, 1987), larval release by Curcinus primarily occurs during nocturnal high tides in the field (Zeng and Naylor, submitted). Thus, there is considerable adaptive value for larval tidal migration rhythms to be phased so that upward swimming occurs soon after hatching.

Although hatching is a single event synchroniser which can phase the circatidal vertical migration rhythms in newly released larvae of C. maenas, it remains unclear as to which aspect of the hatching process is involved. Mechanical stimulation has been

shown to be effective in entraining circatidal rhythmicity in other crustaceans (Enright, 1965; Jones and Naylor, 1970) and we have shown that by subjecting detached eggs of the crab to periodic water agitation in the laboratory, tidal rhythms can be entrained in

subsequently hatched larvae (Zeng and Naylor, 1994). Therefore, the characteristic

pumping behaviour observed during the hatching process may be the critically important factor. However, other factors, such as chemical cues released by embryos or the female

at the time of hatching (Forward and Lohmann, 1983; Rittschof et al., 1985) may be also involved.

In the crab Rhithropanopeus harrisii, laboratory-hatched larvae from females col-

lected in an estuary with semidiurnal tides expressed weak circatidal vertical migration rhythms (Cronin and Forward, 1983). However, only larvae hatched within 4 days of collection were used in those studies and most of the hatches were within 24 h of

collection (Cronin and Forward, 1983), leading to the conclusion that entrainment took place in pre-hatching zoeae by tidal changes in the held. Cronin and Forward (1983) recognized that hatching could also act as the synchroniser for larval swimming rhythms

in R. harrisii which are precisely phased to tidal cycles (Forward et al., 1982). However, they considered this unlikely because larvae from crabs in another estuary with aperiodic tides did not always show circatidal rhythms. Clearly, further study of this phenomenon would be worthwhile.

The synchronization process of circatidal rhythms in intertidal animals is well

documented (Palmer, 1974; Enright, 1975; DeCoursey, 1983; Naylor, 1985). For adult C. maenus and other intertidal crustaceans, cyclic changes in turbulence, hydrostatic pressure, salinity, temperature and inundation have been shown to be effective synchronizers (Enright, 1965; Naylor and Atkinson, 1972; Taylor and Naylor, 1977;

Holmstrom and Morgan, 1983; Naylor and Williams, 1984; Bolt and Naylor, 1985; Reid et al.. 1993; Warman and Naylor, 1995). However, for planktonic organisms which remain in the water column, the environmental variables at the land-water interface will

have little influence. Present studies appear to be the first to characterize a synchroniser of tidal swimming rhythms for planktonic larvae and to provide experimental evidence

that the hatching process is the synchroniser of a crustacean larval swimming rhythm. All other vertical swimming rhythms reported so far in marine plankton are of die1 periodicity associated with the day/night cycles.

Finally, it is noteworthy that in the present study, whereas circatidal rhythms displayed by field-collected larvae were always expressed as sharp peaks indicative of closely synchronized swimming (Zeng and Naylor, 1996), laboratory-hatched larvae showed greater variability in synchrony of their rhythmic swimming patterns. Perhaps,

288 C. Zmg, E. Naylor I J. Exp. Mur. Bid. Ed. IYS (1996) 269-289

therefore, some tidal signals are perceived by such planktonic species after their release. In R. harrisii, the rhythms of field-caught larvae were also greatly enhanced in amplitude and coherence when compared with laboratory-hatched larvae. Therefore, Cronin and

Forward (1983) suggested that the entraining stimuli, whether the same or different, are much more effective upon free-living larvae than on developing embryos. The recent discovery of twice tidal rises in the water column of the field of tidal turbulence, on the

ebb and on the flood (Simpson et al., 1995), offers one possibility for further study of entrainment of tidal rhythmicity in an open sea planktonic species.

Acknowledgments

We thank Dr. A. Aagaard for providing periodogram analysis computer program. Thanks are also due to Mrs. S. Rejeki and Mr. B. Roberts for providing some of the ovigerous crabs. This work was carried out during a TC fellowship to CZ and the

research formed a part of his doctoral dissertation.

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