CARDIAC FREQUENCY COMPENSATION RESPONSES OF ADULT BLUE CRABS (C~~~~~~C~~~

SAPlDUS RATHBUN~ EXPOSED TO MODERATE TEMPERATURE INCREASES

DENNIS T. BURTON LEONARD B. RICHARDSON and CHARLES J. MOORE*

Academy of Natural Sciences of Philadelphia, Benedict Estuarine Research Laboratory. Benedict, Maryland 20612, U.S.A.

(Received 20 June 1979)

Abstract--l. Cardiac frequency patterns of Callinecres sapidus Rathbun were used to evaluate potential thermal stress after exposure to 5°C increases over a range of acclimation temperatures from 5” to 30°C.

2. An acclimated rate-temperature curve (R-T curve). acute R-T curves of the stabilized rates at the increased tem~ratures and QIo tem~rature coefficients were used to assess the significance of the changes in rate frequency.

3. The acclimated R-T curve showed that blue crabs go through a series of seasonal adaptation types characterized by a plateau of perfect adaptation for both cold and warm adapted organisms. Paradoxi- cal adaptation occurred between the transition from cold to warm acclimation temperatures.

4. The acute R-T curves showed that cardiac frequency was highly responsive to a 5°C increase when the organisms were acclimated to low temperatures.

S. The Qro’s of the acute R-T curves at the warm acclimation temperatures approximated those values derived for the acclimated R-T curve.

6. This suggests that the temperature increase had a negligible effect on the warm adapted crabs, that is. little or no thermal stress occurred.

The ability to predict possible detrimental effects of sudden changes in temperature to poikilotbermic ani- mals is difficult because of the inherent complexity of responses that these organisms exhibit at all levels of biological organization. The evaluation is further complicated by the fact that many other environmen- tal and/or physiologi~l factors influence the re- sponses elicited by a temperature change (Brett, 1970; Fry, 1971; Hutchinson, 1976). When a eurythermal organism is exposed to a gradient of temperatures it survives intermediate exposures, however, the extremes cause death. Responses which favor survival in a changed environment are generally considered to be adaptive. Precht (19%) has described two cate- gories of temperature adaptations. Capacity adapta- tions occur in the mid- or normal temperature range of an organism. Resistance adaptations permit sur- vival at extreme temperatures, however, they are more difficult to quantify and death will ultimately occur (Vernberg & Vernberg, 1975).

This study was designed to determine potential stress effects of moderate sublethal temperature increases on adult blue crabs, Callincctes sapidus Rathbun, in their zone of capacity adaptation. Blue crabs were exposed to temperature increases of 5°C over a 15-min period, which simulated a maximum exposure they could experience if they moved or were entrained into the maximum excess tem~ratu~e iso-

* Present address: State of South Carolina Department of Wildlife and Marine Resources. P.O. Box 12559. Char- leston, SC. 29412. U.S.A.

therm of a modern, low AT power station with a high velocity submerged discharge (Pritchard & Carter, 1965; Shirazi & Davis, 1972). Al1 organisms were held at the elevated temperatures for 24 hr to evaluate possible adverse effects of maximum temperature ele- vation for extended periods of time.

Blue crabs were selected for this study because of their ubiquitous dis~ibution in the Chesapeake Bay and other East and Gulf coast estuaries (Williams, 1974). Studies were conducted at ambient tempera- tures which ranged from 5” to 30°C because of the profound influence seasonal temperatures may play in an organism’s response to increased temperatures (Prosser, 1973). Changes in cardiac frequency were used to assess subtle physiological adjustments to increased temperatures.

MATERIALS AND METHODS

Blue crabs were collected from the Calvert Cliffs area of Chesapeake Bay or the lower Patuxent River by standard collecting procedures. All crabs, with the exception of 5°C and 10°C acclimated animals, were collected at ambient temperatures similar to the experimental temperatures of 5”, 10” 15” 20”. 25” and 30°C. In the cases where seasonal temperatures differed from the experimental temperatures. thermal acclimation was achieved by changing the water temperature l”C/day from ambient to the desired experi- mental temperatures Once the desired temperature was reached, the animals were held at that temperature (&OS”C) for a minimum of 2 weeks before any experimen- tation was initiated.

The organisms were maintained in the laboratory at each acclimation temperature in separate 650 or %Ok.

f .I, P. 65 3R A 259

aquaria which were supplied with a continuous Bow 01 coarsely filtered well-aerated Patuxent River water. The mean salinity of the acclimation and test water throughout the study period was 6.7’:,,, (range. 4.1- 10.8); mean pH 7.8 (range. 7.3 8.3) and dissolved oxygen >5.5 mg./l at all times.

A cyclic photoperiod (incandescent light) of lZL:l?D was maintained for all animais at 5’ I5 C and 16L:8D for organisms at 21) WC during acclimation and experimen- tal study. A 4%min tr~~nsition between light periods was used rather than an abrupt on anti off transition. Crabs were fed regularly with chopped fish. No animal was fed 24 hr preceding a test or during a test. The average weight (_tS.D.) of 60 test organisms which included both males and females in approximately equal numbers was 130.5 (+18.32)g; average carapace width was 13.1 (+_5.13)cm. The organisms were selected to be approximately the same size at all acclimation tempcraturcs in order to make car- diac rate independent of weight (Ansanullah & Newell. 1971)

Experimrntul procedures

Two groups of six animals were studied at each acclima- tion temperature. Individuals in each group were isolated from each other and external stimuli in separate chambers during the course of an experiment. During an experiment, cardiac frequency data from six organisms were recorded simultaneously on a Grxs polygraph (Model 78: C&ass Instrument Co.. Quincy, Mass.) The polygraph was pro- grammed to collect data for 5-min pertods a! 0.5 hr inter- vals for I6 hr before the animals were exposed to a 5 C temperature increase over a 1.5.min period at I700 hr. Car- diac frequency data at the elevated temperature were col- lected at the same time intervals for the next 24 hr. Time and water temperatures were also recorded simultaneously during each measurement period.

The test chambers which housed the animals during the study were supplied by a well-aerated contmuous flow of coarsely filtered water. which entered the chambers at one end and was discharged at the opposite end. The vo]ume of water delivered to the chambers was carefully regulated in order to produce the same test condition during each run. All water delivery lines supplying the six chambers were of equal length and size to ensure that all animals received water at the same temperature at exactly the same time. The temperature increase was achieved by mtroducing water5 C above ambient over a I-min period.

All experimental chambers were just large enough to permit free appendage movement. The terminal segments of the chelipeds were held shut by rubber bands to mini- mize interference during the study. The chambers were constructed of Plexiglas cylinders (9 cm I.D. x 30 cm long) fitted with rubber stoppers at each end. Plexiglas partitions were used in the chambers to keep the animals oriented in the same position throughout the study. A small standpipe was fitted in the top of each chamber to serve as an exit port for electrodes. The flow rate,‘min to the chambers was 250 ml i 1: 7 ml). The Row rate was high enough to ensure that the test organisms did not reduce the dissolved oxygen in the test chamber and influence the physiological re- sponse being measured.

Electrocardiograms (EKG) of each crab were obtained by inserting a small gauge platinum unipolar active elec- trode into the pericardial chamber through a small hole drilled in the carapace (Larimer. 1962). The electrode was placed approx 5.6mm caudad in the mid plane of the pos- terior section of the cervical groove. Each electrode was secured with paraffin or modeling clav and by an elastic band stretched between the lateral spines. No anesthesia was used during electrode placement. The ground and indifferent electrodes were constructed of stainless steel hypodermic needles which were inserted through the stop- pers at each end of the chambers to provide electrical con-

tact with the chamber water. ‘fhc EKG‘s were d~sphjcd on

the polygraph. Cardiac frcqucncy.‘min v.as determined h\ averaging three I-min counts from each 5mtn recording period. The record segments analyzed were selected 10 avoid pause and burst EKG activity (McMahon I% WII- kens. 1977).

The homogeneity of responses between replicates at each ~icclirn~~tion temperature was tested for the pm-cxposurc values by Morrison’s (1967) test for parallel profiles. The hypothesis was rejected at the 5”,, signthcance level for all acclimation groups except crabs at 25 C. Since it signiiicant difference occurred between replicates at 25 C, these data sets could not be pooled and hence were not considered in further analyses, The large differencr m heart rate activity between replicates at ‘5 C is not clear because all experr- mental procedures. with the exception of tempcraturc. wcrc the same as those at the other acclimation temperatures.

Multivariate repeated measurement statistics were used to test the hypothesis that no difference occurred hctwcen mean pre-exposure rate frequency and mean exposure rate frequency for the pooled replicates at each acclimation temperature (Morrison. 1967). All tests taken from Morri- son were performed at the 5”<> sy,nificance level.

The effects of different ~~cclinl;~ti~~n temperat~rcs on rate freytiency of the pre-exposure pooled data Were tested b) Morrison’s (1967) test for parallel protiles. initial analyses shox+ed that the repeated measurements between ali occli- mation groups were not purallcl for some of the groups.

that IS. in some cases the rates decreased from XKH) hr tcr IZOOhr. Therefore. tn the cxxs whcrc this occurred the

variances of the prc-treatment mean\ at each half interval were adjusted for the linear decrease bcforc the cornpart- sons were made between accllmntlon groups at the Cl’,,

significance level.

RESC’LTS

The mean pre-exposure and exposure cardiac rates at each acchmation temperature are summarized in Table 1. Significant increases (P < 0.05) in rate frequency occurred after the temperature increase in blue crabs acclimated to 5 , 10’ and 15-C’. Cardiac frequencies stabilized quickly (within 1 hr) at the higher rates and remained relatively constant throughout the remaining exposure period at the ele- vated temperatures. No significant changes (P > 0.05) occurred in crabs acclimated to 20’ and 30°C.

The effects of seasonal acclimation on rate freyuen- ties were as follows: (1) No significant difference occurred between crabs acclimated to 5’ and 10°C. (2)

Crabs acclimated to 15°C had a signi~cant~y higher frequency than crabs at 5’ and 10°C. (3) No difference occurred between crabs acclimated to 15”. 20. and WC.

DlSCWSiON

Three approaches were used in this study to assess the changes in rate frequency: (1) An acclimated rate-

temperature curve (R--T curve) was used to establish the effects of normal seasonal acclimation tempera- tures on heart rate frequency. The values used to con-

struct the acclimated R--T curve were taken from the me-exposure data at each acclimation temperature. (2) Acute R-T curves were superimposed on the accli- mated R-T curve to indicate the degree of sensitivity

to the increased temperature The stabilized rate fre-

Tab

le

I.

Mea

n pr

e-ex

posu

re

and exposure

heart

rate

s fo

r Cd

hrct

es

supi

dtcs

at

eac

h ac

clim

atio

n te

mpe

ratu

re

Pre

-Ex

po

sure

Rate

Exposure Rate

Significance

Acclimation

(Frequent /Minute)

+

Level Between

Temperature

Mean

(OCl

Pre-Exposure and

(+- S.D.)

Observations

(1 S.D.)

Observations

Exposure Rates

5

33.2(-? 3.58)

329

59.7(+ 4.68)

384

pxo.05

10

31.3(+21*20)

488

62.8(+18.04)

715

p<o.os

15

lOZ.O(t 9.67)

275

126.3(+12.40)

410

p<o.os

20

111.9(t25.83)

419

121.5(+24.39)

638

p>o,o5

-

30

113.9(+25.49)

454

132.7(s22.71)

487

p>o.o5

quency after the temperature increase was used to construct each acute R-T curve. (3) Qlo tempcr~~turc coefficients were calculated for the acclimated R-T curve to establish seasonal compensation patterns. Qlo’s for the acute R~-T curves were used to compare stabilized responses to seasonal compensation pat- terns.

The acclimated R-T curve, acute R--T curves and QIo of each rate change are presented in Fig. 1. If one considers the acclimated R-T curve first, it can be seen that crab heart rates go through a series of seasonal adaptation types characterized by the un- usual occurrence of two plateaus of perfect adapta- tion. One plateau occurred between 5’ and 10°C and another between 15” and 30°C. Although the Qlo on the graph is 0.9 between 5’ and 1O“C and 1.2 between 15” and 20X‘. no statistically sjgnificant difference (P > 0.05) was found between the preexposure rates of each group, hence the Q i o can be expressed as 1 .O or perfect acclimation. Paradoxical adaptation occurred from 10 to IS-C (Q,,, = 10.6).

Cardiac frequencies in crustacea have generally been shown to increase with rising acclimation tem- peratures when measured in the zone of capacity adaptation (Maynard. 1960). The increase is more rapid at low temperatures and tends to decline or level off at higher temperatures much as Bullock (1955) predicted in his review of temperature compen- sation in ~ikiiotherms. Most studies have shown that partial compensation in heart frequency R--T curves may occur in warm adaptated organisms (Ahsanullah & Newell, lY71 : Spaargaren. 1974). deFur & Mangum (1979) have shown that blue crabs have a cardiac frequency Q, 0 of 2.0 between IS and 25’C in contrast to a Qlo of 1.0 in this study between 15’ and 30’ C. Perfect adaptation in decapod heart rate has not been reported until this study. Spaar- garen (1974) has demonstrated, however, that perfect adaptation may occur in “relative” cardiac stroke volume in the shore crab. C’W~~UUS I~WIUS. when measured over a wide range of acclimation tempera- tures under submerged conditions.

i, ‘00 5 __+__&_._&__ ._J5-_..l_J

30 35 TEMPERATURE WI

Fig. 1. Acclimated R-T curve (+ --@). acute R-T curves (O--j and Q,,, temperature coefficients of each for Cal/r-

IIL’Cl(‘S supidtts.

111 contrast to cardiac Ircquenc>. perfect iKiaptat1ol-i in metabolic rate has been found rn se~ral Intertidal species over rather large segments of the acclimated R- T curve (Newell & Northcroft. 1967: New&i & Bayne, 1974; Newell, 1976). Nearly perfect compm- sation in oxygen consumption has been found for juvenile blue crabs between 20’ and 27’c (Leffler. 1972). Laird & Haefner (1976) have suggested that relatively complete tcmpcrature compensation in OX!- gen consumption may also occur in warm acclimated adult blue crabs between 17.5 and 25’ C. Although the correlation between cardiac rate and metabolic rate as a function of temperature is not simple (Ansell. 1973). this study shows that perfect temperature adap- tation does occur in certain components of the car- diac system in blue crabs as it does in whole animal oxygen consumption in other species

The QIo of f in heart rate between 5 and IO C may be related to cold torpor or so called “hiberna- tion”. Blue crabs arc relatively inactive during the winter months in Chesapeake Ba! (Van Engel, 1958; Laird & Haefner, 1976). Qualitative observations of physicial activity and feeding were made in the labor- atory during this study. All animals were quite inac- tive and consumed negligible amounts of food at 5 and 10°C. The oxygen consumptmn of shore crabs has also been shown to be essentially independent of temperature when the animals are starved for approx 3 weeks (Marsden & it/.. 1973). The perfect adaptation of heart rate found between 5 and 1O’C for blue crabs may be an adaptive mecha~iisnl which aids the organisms in coping with the cold winter months when its activity is low and food is less abundant. In contrast, the perfect adaptation plateau between 15 and 30’ indicates that maximum temperature indepen- dence occurs during the months when the animal is most active. One would predict from Bullock’s (1955) work that the Q,(, between the transition of low ac- tivity at cold temperatures to higher activity at warmer temperatures would be fairI? large. This has been documented for cardiac rate in this study and the study by deFur (FL Mangum (1979) for blue crabs (Q,(, = 3.2 between 5 xnd 25 ‘C) ;IS well ~1s for other portunid crabs (A~~~n~~ll~~~~ & Ne\lell. 1971 : Taylor <‘I cti.. 1973).

The acute R T curves for blue crabs showed that cardiac frequency was highly responsive to a 5 C AT when the animals were adapted to low temperatures. The acute.heart rate response from IO to I SC was similar to that of the acclimated rate where ;I large

QI,, was found between the transition from cold to warm conditions. The acute rate from 5 to 10 C shows that no immediate compensation occurs in 24 hr. The acute rate should approach the acclimated rate at l0.C as temperature compensation occurs. It is ditlicult to speculate about the possibility that the initial large increase may be a sign of stress because the correlation between cardiac rate. pump volume and cardiac output is not known at low temperatures.

The Q, c1 values for the stabilized R~ T curves at 15 and ?O’( approximate those values derived for the acclimated R T curve. This suggests that ;I moderate temperature increase for warm adapted crabs has II negligible effect on cardiac frequency. This in turn may be interpreted to mean that little or no thermal stress occurs. Spaargaren Rr Achituv (1977) have made

ERect of temperature on blue crab heart rate 263

similar heart rate observations for several estuarine and marine crustaceans exposed to rapid temperature changes. Some homeostatic compensation in cardiac frequency would probably occur if the animals were acclimated to 20” and 25°C (Presser, 1964). This would bring rates closer to those predicted from the acclimated R-T curve.

Ackrzow(rdyements-The authors wish to thank Philip Abel1 and Stuart Margrey for their technical assistance during the study; Richard Greenstreet for performing the statistical analyses and Virginia Santoro for typing the manuscript. We also thank N. G. Lassahn, Jr and Ehza- beth Bauereis of Baltimore Gas and Eiectric Comnattv. Battimore, M~yland, for their review of the paper and financial support of the study.

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