Directory UMM :Data Elmu:jurnal:J-a:Journal of Experimental Marine Biology and Ecology:Vol247.Issue1.Apr2000:

(1)

L

Journal of Experimental Marine Biology and Ecology 247 (2000) 29–49

www.elsevier.nl / locate / jembe

Effect of diet and temperature upon muscle metabolic

capacities and biochemical composition of gonad and muscle

in Argopecten purpuratus Lamarck 1819

a ,_* b a a

Gloria Martınez , Katherina Brokordt , Cristian Aguilera , Viterbo Soto ,

Helga Guderley

Departamento de Biologıa Marina, Universidad Catolica del Norte, Casilla 117, Coquimbo, Chile

´ ´ ´

Departement de Biologie, Universite Laval, Quebec, P.Q., Canada G1K 7P4

Received 2 August 1999; received in revised form 1 November 1999; accepted 15 December 1999

Abstract

Recently spawned Argopecten purpuratus broodstock were conditioned at two temperatures and fed three different diets (microalgae, microalgae mixed with lipids and microalgae mixed with carbohydrates) to examine changes in the biochemical composition of gonad and muscle as well as muscle metabolic capacities. During one experiment, scallops were fed at 3% of their dry mass per day whereas during a second experiment, they were fed at 6% of their dry mass per day. During both experiments, total gonadal levels of lipids and protein increased markedly during con-ditioning with the two mixed diets at 168C. These increases were less pronounced at 208C. Carbohydrate gonadal levels only increased during the second experiment at both temperatures and with the three diets. Of the major biochemical components of the adductor muscle, carbohydrate levels changed most during conditioning. Whereas muscle protein levels increased slightly with gonadal maturation, carbohydrate levels dropped considerably. Despite the marked drop in the levels of glycolytic substrates, only the activities of octopine dehydrogenase in the adductor muscle of the scallops conditioned at 168C consistently decreased. Muscle levels of glycogen phosphorylase were higher in mature than in recently spawned (control) scallops, suggesting a role in the transfer of glucose equivalents from the adductor muscle to other tissues.

Keywords: Argopecten purpuratus; Diet; Temperature; Gonads; Muscle; Metabolic capacity; Composition

*Corresponding author. Tel.: 156-51-209-793; fax: 156-51-209-812. ´

E-mail address: [email protected] (G. Martınez)

(2)

1. Introduction

In most animals, but particularly in broadcast spawners, gonadal maturation is a costly process requiring considerable investment of material and energy. When food availability is high, the cost of gonadal maturation can be covered entirely by absorbed materials and energy, but when food availability is low, gonadal maturation can require mobilization of macromolecular components from other tissues for their use for gametogenesis. Scallops are broadcast spawners in which the mature gonad can account for more than 20% of the body mass. Inverse cycles of the biochemical components in the adductor muscle and gonad occur during the reproductive cycle of various scallop species, supporting the interpretation that materials are mobilized from the muscle to support gonadal maturation. Such cycles have been observed for Chlamys opercularis (Taylor and Venn, 1979), Chlamys varia (Shafee, 1981), Argopecten irradians concentricus (Barber and Blake, 1981), Argopecten irradians irradians (Epp et al., 1988),

Placopec-´

ten magellanicus (Couturier and Newkirk, 1991), Argopecten purpuratus (Martınez,

1991; Martınez and Mettifogo, 1998), Pecten maximus (Pazos et al., 1997) and Euvola (Pecten) ziczac (Brea, 1986; Boadas et al., 1997). Whereas glycogen and protein are the principal macromolecules which are mobilized from muscle (Taylor and Venn, 1979; Barber and Blake, 1981, 1985; Pazos et al., 1997), lipid levels in the adductor muscle may also decline during gonadal maturation (Epp et al., 1988). Clearly, beyond its obvious contractile role, the adductor muscle fulfills an important role as a site of reserve deposition in scallops.

When muscle macromolecules are mobilized to support the metabolic requirements of other tissues, both the levels of energetic reserves, such as glycogen and lipid, and those of enzymes or contractile proteins can be affected. For example, during their non-feeding spawning migration, sockeye salmon, Oncorhynchus nerka, first metabolize their lipid deposits in muscle and then break down muscle enzymes and sarcoplasmic proteins (Mommsen et al., 1980). During starvation, cod, Gadus morhua, first mobilise hepatic lipid, then hepatic and muscle glycogen and finally muscle proteins (Black and Love, 1986). During the nutritional stress caused by winter conditions, mussels experience a high energy loss from body tissues and proteins accounts for 75% of this loss (Gabbott and Bayne, 1973). During gametogenesis in Argopecten irradians irradians, digestive gland lipid reserves are first mobilized followed by muscle glycogen and finally protein reserves (Barber and Blake, 1981). The sparing of muscle proteins during starvation likely reflects the fact that proteins play more roles than simply serving as an energetic reserve. Effectively, the mobilisation of muscle proteins during starvation or reproduc-tive maturation decreases muscle metabolic capacities (Guderley et al., 1994, 1996), particularly the glycolytic capacity of white muscle. Studies of relative utilization of dietary protein for energy and biosynthesis by the mussel Mytilus edulis have demonstrated a continuous conservation of amino-N over amino-C to satisfy bio-synthetic demands (Kreeger et al., 1995, 1996).

The extensive mobilization of muscle reserves during gametogenesis in scallops suggests that muscle metabolic capacities change during gonadal maturation. However, little is known of the impact of gonadal maturation upon the metabolic capacities of the adductor muscle. The oxidative capacities and respiratory control ratios of mitochondria

(3)

isolated from the adductor muscle of Euvola (Pecten) ziczac during their first spawning were lower than in those isolated at other periods, apparently due to the low food availability and high temperatures in the preceding period (Boadas et al., 1997). When gonadal maturation is induced in recently spawned scallops, muscle metabolic capacities may first rise and then fall if muscle proteins are mobilized for gonadal maturation (Pazos et al., 1997). If muscle enzymes are simply used to provide energy and amino acids, all enzymes should decrease in parallel during protein mobilization. On the other hand, enzymes which facilitate reserve mobilization or which are restricted to specific organelles may be spared. For example, as glycogen phosphorylase liberates glucose moieties from glycogen, its levels may be maintained.

Reinitiation of gonadal maturation following spawning is of particular interest for the aquaculture industry, which depends upon a ready availability of high quality larvae. The extent and quality of this gonadal recuperation depend upon the available food, with certain diets leading to faster gonadal maturation, greater fertilization success and

improved larval survival than others (Martınez et al., 2000). Thus, in Argopecten

purpuratus, gonadal maturation is faster at 168C than at 208C and the best larval survival is obtained with a microalgal diet supplemented with a lipid emulsion. During conditioning of A. purpuratus, considerable time is required for gonadal maturation,

even with appropriate diets and temperatures (Martınez et al., 2000). Thus, recently spawned individuals are likely to replenish tissue reserves before undertaking gonadal maturation. The levels of biochemical components in muscle would increase before any improvement in gonadal status is apparent and then decrease during gonadal maturation, particularly if food availability is insufficient to cover the requirements of gonadal maturation. Under this scenario, muscle metabolic capacities may first rise and then fall during broodstock conditioning, although enzymes which play a role in reserve mobilisation may be spared.

To examine these possibilities, we evaluated the biochemical composition of muscle and gonad and muscle metabolic capacities in the scallop, A. purpuratus, during conditioning of recently spawned individuals at 16 and 208C using three diets, one consisting of microalgae, another of microalgae supplemented with a lipid emulsion and the last of microalgae supplemented with carbohydrate. We measured levels of carbohydrates, lipids and proteins in muscle and gonad. Since the scallops conditioned at 168C showed more complete gonadal maturation than those at 208C and as we wished to ascertain the impact of mobilization of muscle reserves in support of gonadal maturation upon muscle metabolic capacities, we determined the activities of glycogen phosphor-ylase (GP), pyruvate kinase (PK), octopine dehydrogenase (ODH) and citrate synthase (CS) only in the muscles of scallops conditioned at 168C. GP is responsible for the liberation of glucosyl units from glycogen, PK catalyses an ATP generating reaction in glycolysis, whereas ODH catalyses the reductive condensation of pyruvate and arginine, thereby regenerating NAD. CS is a component of the Krebs cycle and serves as a marker of mitochondrial abundance. If these enzymes show the same variation during mobilization of muscle reserves, it would indicate non-selective protein breakdown. Specific changes, in which certain activities were maintained whereas others decline, would indicate targetted protein mobilization. We predicted that the levels of the glycolytic enzymes, PK and ODH, would decrease whereas GP would be spared during

(4)

mobilization of muscle reserves, given its role in liberating glucose equivalents from glycogen. We reasoned that the localization of CS in the mitochondrial matrix would protect it during reserve mobilization.

2. Materials and methods

2.1. Feeding and sampling of scallops

Scallops (experiment 1: shell height of 80–90 mm; experiment 2: shell height of 70–80 mm) just obtained in mature stage from Culture Centers in Tongoy Bay (308S) were induced to spawn by increasing the temperature and adding excess microalgae consecutive years. The spent animals were distributed among nine tanks (34 animals per tank) at 168C and nine tanks at 208C. Three groups at each temperature were fed with a mixture of microalgae (50% of Isochrysis galbana150% of Chaetoceros gracilis), three other groups received 70% of the same mixture of algae plus 30% carbohydrates (commercial potato starch), and the other tanks received 70% microalgae and 30% of a lipid emulsion (provided by Artemia Reference Center in Belgium, corresponded to the EmDHA whose composition is described in Caers et al., 1999). The gross biochemical composition of these diets is given in Table 1. More details about diets and their absorption efficiency are described in Navarro et al. (2000). The daily food ration, supplied continuously by a peristaltic pump, amounted to 3% of the animals’ dry mass during the first experiment and to 6% during the second experiment. Both experiments were run from January to March in consecutive years (1998 and 1999).

Before beginning conditioning, five spent scallops were sampled to evaluate their biochemical status (this sample was called ‘control’). Periodically, similar analyses were performed for three scallops from each experimental treatment (each animal from a different tank) to follow the course of gonadal recovery and the levels of the biochemical components. For this, the scallops were sorted into three groups according to their visually determined degree of maturation and one scallop from the largest group in each tank was sampled. The duration of the experiments was determined according to the progress of gonadal ripening; when about 40% of the scallops appeared mature in any of the tanks, the entire experiment was ended to avoid spontaneous spawning. When the first experiment (3% ration) was ended, the gonadal indices in the groups with the best performance approached 10% whereas in the second experiment (6% ration), the

gonadal indices of the best groups were greater than 13% (Martınez et al., 2000).

Table 1

Biochemical composition of the three experimental diets used for conditioning A. purpuratus

Proteins Lipids Carbohydrates

Microalgae 212.99611.02 75.0566.69 62.9564.16

Microalgae1lipids 191.9568.45 108.8762.92 69.8267.07 Microalgae1carbohydrates 200.00615.43 80.00616.11 215.00626.26

a 21

(5)

2.2. Biochemical and enzymatic analyses

The scallops were sacrificed and their gonads and muscles were weighed before freezing and storage in liquid nitrogen for subsequent biochemical determinations. The muscle samples used for enzymatic measurements were transported on dry ice to

Universite Laval where they were stored at 2708C before analysis. The levels of proteins, total carbohydrates and lipids in muscle and gonad were analysed following the

methods of Martınez (1991). Muscle levels were presented as concentrations (mg g dry mass), whereas gonadal components were presented as total gonadal contents (for the female and male portions respectively) given the marked changes in gonadal size and no significant changes in muscle size during conditioning (gonadal size increase was more than 300%)

For measurements of PK and ODH, portions of the adductor muscle were homogen-ized in 50 mM imidazole–HCl, 2 mM Na -EDTA, 5 mM EGTA, 1 mM dithiothreitol2

and 0.1% Triton X-100, pH 6.6. For GP and CS measurements, samples of adductor muscle were homogenized in this buffer at pH 7.2 supplemented with 150 mM KCl. The pH of all solutions was adjusted at room temperature. Homogenisation of the muscle samples (1:10 m / v) was carried out on ice using a Polytron (Brinkmann, Rexdale, Ontario) at 50% maximal speed for three, 30-s periods separated by 1-min rest periods. Spectrophotometric measurements were carried out using a Beckman DU-640 UV-Vis spectrophotometer. The cuvette temperature was controlled at 168C by a Haake G8 refrigerating circulator. Measurements were made at 340 nm for PK, ODH and GP following the changes in absorbance of NAD(P)H, whereas measurements of CS followed the absorbance of 5,59dithio-bis(2-nitro)benzoic acid (DTNB) at 412 nm. Enzyme activities were measured using the following conditions:

Pyruvate kinase: 50 mM imidazole–HCl, 13 mM MgSO , 100 mM KCl, 5 mM ADP,4

0.2 mM NADH, pH 6.6, 5 U lactate dehydrogenase and 5 mM phosphoenolpyruvate (omitted for the control).

Octopine dehydrogenase: 50 mM imidazole–HCl, 2 mM Na EDTA, 5 mM EGTA, 12

mM K-cyanide, 0.2 mM NADH, 6 mM Na-pyruvate and 6 mM arginine (omitted for the control). The control activity, which represents that of lactate dehydrogenase, was virtually nil.

Glycogen phosphorylase: 50 mM imidazole–HCl, 80 mM KH PO , 5 mM Mg-2 4

acetate, 2.5 mM Na EDTA, 0.8 mM AMP, 0.5 mM cyclic AMP, 0.6 mM NADP, 0.0042

mM glucose-1,6-diphosphate, 2 U glucose-6-phosphate dehydrogenase, 2.5 U

phosphog-21

lucomutase, 10 mg ml glycogen, pH 7.5.

Citrate synthase: 75 mM Tris–HCl, 0.25 mM DTNB, 0.35 mM acetyl CoA, 0.05 mM oxaloacetate, pH 8.0. Oxaloacetate was omitted for the control.

The mmolar extinction coefficients were 6.22 and 13.6 for NAD(P)H and DTNB, respectively. Enzyme activities are expressed as international units (mmol of substrate

21 21

converted to product min )3g wet mass. 2.3. Calculations and statistical comparisons

(6)

concentration of the component in a sample of the female and male gonad, and then extrapolated to the total content of that section by assuming that the gonad was equally divided into male and female sections (sections never differed by more than 10% and histological analysis showed a clear separation between them). The effect of diet and duration of conditioning upon the levels of biochemical components in gonad and muscle, upon enzyme levels in muscle and upon muscle mass and % water content was examined using two-way ANOVAs which considered diet and duration of conditioning as the two factors (Systat). When no interactions were observed between the factors, one-way ANOVAs considering only the effect of duration of conditioning were run for each diet. When significant effects of duration of conditioning were observed, Tukey a posteriori tests were used to identify significant differences (P,0.05). For each experiment, the data obtained under the two thermal regimes were analysed separately.

3. Results

3.1. Biochemical composition of muscle

When recently spawned Argopecten purpuratus, maintained at 16 or 208C, were supplied with the three experimental diets at 3 or 6% of their dry mass per day, carbohydrate levels in the adductor muscle showed the most pronounced fluctuations. During experiment I significant initial increases were followed by marked decreases in all treatments (Table 2, Tukey a posteriori tests, P,0.05) (Fig. 1). Initial carbohydrate levels were higher in the second experiment than in the first (Student’s test, P,0.05). Presumably this is why muscle carbohydrates did not rise at the start of the experiment II.

During conditioning at 3%, muscle protein levels showed slight statistical differences between diets at both temperatures (Table 2, Fig. 2); slight increases were detected at the end of conditioning of scallops fed with microalgae plus starch or only microalgae at 168C. At the end of conditioning at 208C, muscle protein levels declined in scallops conditioned with the mixture microalgae–carbohydrate. In contrast to experiment I, muscle protein levels of animals fed at 6% of their dry mass per day rose at the start of conditioning, but did not increase further nor fall below initial values in scallops conditioned at either temperature.

In the first experiment, lipid levels in the adductor muscle were higher, particularly on day 32, in the scallops fed microalgae supplemented with lipids or starch than in the muscle of scallops fed only microalgae (P,0.05) (Fig. 3). Initial muscle lipid levels were higher (Student’s test, P,0.05) in the scallops conditioned at the higher level of food (experiment 2) and declined gradually during their conditioning at 168C; at 208C, except with enriched-lipid diet, there was an initial fall (on day 10) then initial values were recovered, followed by a gradual decline during the final maturation of the gonad. The changes in the biochemical components in muscle during conditioning were not accompanied by shifts in water content nor wet mass (ANOVA, P.0.05).

(7)

Table 2

Two-factor analysis of variance for adductor muscle biochemical components of Argopecten purpuratus

conditioned at 16 and 208C with three diets

df F

168C 208C

Experiment I

Carbohydrates

Diet 2 3.484* 1.112 ns

Days of conditioning 4 35.689*** 46.080***

Interaction 8 7.834*** 17.481***

Proteins

Diet 2 2.588 ns 1.147 ns

Days of conditioning 4 9.181*** 7.151***

Interaction 8 1.654 ns 1.270 ns

Lipids

Diet 2 1.353 ns 2.146 ns

Days of conditioning 4 6.924*** 8.641***

Interaction 8 0.568 ns 2.272 ns

Experiment II

Carbohydrates

Diet 2 10.867*** 6.782**

Days of conditioning 6 19.103*** 17.385***

Interaction 12 2.631** 4.024***

Proteins

Diet 2 2.893 ns 1.328 ns

Days of conditioning 6 9.368*** 14.823***

Interaction 12 0.460 ns 1.272 ns

Lipids

Diet 2 5.878** 7.241**

Days of conditioning 6 30.513*** 22.909***

Interaction 12 3.510** 3.130**

Diets and time of conditioning are the main effects considered. ns, not significant; * 0.01,P,0.05; ** 0.001,P,0.01; *** P,0.001.

3.2. Biochemical composition of gonad 3.2.1. Experiment I

Conditioning of the scallops at 168C, at 3% of their dry body mass per day, led to marked fluctuations of the total content of protein and lipid in the female portion of the gonad (Figs. 4 and 5). These changes were most pronounced for scallops fed microalgae supplemented with carbohydrates or lipids. Lipid contents increased at the end of conditioning relative to levels during the remainder of the experiment (two-way ANOVA; P,0.05). Carbohydrate contents of the female gonad fluctuated little during conditioning at 168C (Fig. 6), but final values were lower during conditioning in the lipid-supplemented diet (P,0.05). Carbohydrate contents of the male portion of the gonad were low and did not change during conditioning. Lipid levels rose slightly towards the end of conditioning (P,0.05) with lipid-supplemented diet.

(8)

Fig. 1. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the concentrations of carbohydrates in

adductor muscle (mg g dry mass). (A) Scallops fed with microalgae supplemented with starch, (B) scallops fed with microalgae supplemented with a lipid emulsion; (C) scallops fed only with microalgae. Values represent the mean6S.E. (N53). Different letters mean significantly different values (Tukey’s P,0.05); a single letter means significantly different from the rest of the values of the same treatment and values without letter mean not different from any of the values of the same treatment.

(9)

Fig. 2. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the concentration of protein in adductor

muscle (mg g dry mass). (A) Scallops fed with microalgae supplemented with starch, (B) scallops fed with microalgae supplemented with a lipid emulsion; (C) scallops fed only with microalgae. Values represent the mean6S.E. (N53). Different letters mean significantly different values (Tukey’s P,0.05); a single letter means significantly different from the rest of the values of the same treatment and values without letter mean not different from any of the values of the same treatment.

(10)

Fig. 3. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the concentration of lipids in adductor

(11)

Fig. 4. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the total content of protein in each

gonadal portion (mg gonad ). (A) Scallops fed with microalgae supplemented with starch; (B) scallops fed with microalgae supplemented with a lipid emulsion; (C) scallops fed only with microalgae. Values represent the mean6S.E. (N53). Different letters mean significantly different values (Tukey’s P,0.05); a single letter means significantly different from the rest of the values of the same treatment and values without letter mean not different from any of the values of the same treatment. Upper-case letters for male gonadal portion and lower-case letters for female gonadal portion.

(12)

Fig. 5. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the total content of lipids in each

(13)

Fig. 6. Effect of conditioning recently spent Argopecten purpuratus at 16 and 208C with three diets at 3 (experiment I) and 6% (experiment II) of their body mass per day, on the total content of carbohydrate in each

(14)

changes in gonadal contents of carbohydrates and proteins. Diet had a significant effect on the protein content of the female gonad (Fig. 4), as well as on the lipid and carbohydrate content of the male gonad (two-way ANOVA; P,0.05) (Figs. 5 and 6). The fluctuations of the levels of biochemical components led the duration of con-ditioning to significantly affect the contents of proteins and carbohydrates in the female gonad and on all of the components in the male gonad. Inspection of the overall changes in the gonadal contents of biochemical components indicates that gonadal levels of lipids and proteins rise during final maturation, whereas increases in proteins which are not accompanied by increases in lipids may occur well before gonadal maturation. 3.2.2. Experiment II

The protein content of the female gonad markedly increased at the start of conditioning with all diets and at both temperatures (Fig. 4). Subsequent fluctuations of protein contents occurred under all experimental conditions, but were more pronounced at 168C. The slight decrease at 70 days was likely to due to a partial spawning. Protein contents of the female gonad were affected by diet and duration of conditioning at 168C and only by the duration of conditioning at 208C (Table 3, two-way ANOVA; P,0.05). At 168C, lipid contents increased considerably during the last 20 days of the experiment. Lipid and carbohydrate contents of the female gonad were affected by duration of conditioning and diet at 168C (Table 3, Figs. 5 and 6). For each diet, the scallops fed at 208C tended to have lower gonadal protein contents than those fed at 168C. The increase of lipid contents towards the end of conditioning was less pronounced at 20 than at 168C.

Protein contents of the male gonad rose considerably at the start of conditioning, and oscillated before showing their final increase in the scallops conditioned at 168C with the two supplemented diets (Fig. 4). Again these oscillations may reflect partial spawnings. The carbohydrate contents of the male gonad were affected by both duration of conditioning and diet at both temperatures (Table 3, two-way ANOVA; P,0.05). At 16 and 208C, lipid and protein contents were only affected by duration of conditioning (Table 3). At 208C, a rise in protein contents was noted only for scallops conditioned with the microalgae supplemented with carbohydrates. In contrast to the scallops conditioned in the first experiment, the lipid contents of the male gonad changed little during the final stages of conditioning.

3.3. Muscle enzyme levels

Muscle enzyme levels were expressed as U g wet mass since the water content of the muscles did not change during conditioning at 168C (ANOVA, F3,6850.332,

P.0.05). A two-factor ANOVA with time of conditioning and diet as factors indicated that time of conditioning significantly affected GP levels (P,0.001), but that diet did not. No interaction was found between time and diet. Both diet (P50.044) and conditioning time (P50.009) significantly affected PK levels. Throughout conditioning, scallops fed microalgae alone maintained lower levels of PK than those fed microalgae with lipids, whereas those fed microalgae supplemented with carbohydrates showed decreases followed by an increase at the end of the experiment (Fig. 7). ODH levels

(15)

Table 3

Two-factor analysis of variance for gonadal biochemical components of Argopecten purpuratus conditioned at

16 and 208C with three diets

df F F

Female portion Male portion

168C 208C 168C 208C

Experiment I

Carbohydrates

Diet 2 7.156** 7.010** 1.097 ns 4.940*

Days of conditioning 4 5.256** 8.222*** 3.846* 10.184***

Interaction 8 6.521*** 1.520 ns 1.465 ns 2.107 ns

Proteins

Diet 2 0.001 ns 4.182* 0.371ns 1.083 ns

Days of conditioning 4 7.872*** 10.960*** 3.054* 4.667**

Interaction 8 1.060 ns 1.145 ns 0.633 ns 1.619 ns

Lipids

Diet 2 3.065 ns 0.859 ns 0.681 ns 4.350*

Days of conditioning 4 6.211*** 1.167 ns 8.256*** 5.051**

Interaction 8 1.486 ns 3.915** 0.909 ns 1.315 ns

Experiment II

Carbohydrates

Diet 2 4.290* 2.631 ns 15.209*** 6.591**

Days of conditioning 6 7.636*** 12.925*** 34.483*** 6.620***

Interaction 12 1.164 ns 1.131 ns 3.216** 1.784 ns

Proteins

Diet 2 3.691* 0.674 ns 6.838** 0.676 ns

Days of conditioning 6 10.963*** 10.674*** 21.007*** 7.034***

Interaction 12 2.292* 1.024 ns 0.959 ns 1.607 ns

Lipids

Diet 2 8.079*** 2.731 ns 2.382 ns 0.355 ns

Days of conditioning 6 8.410*** 6.346*** 16.022*** 9.452***

Interaction 12 1.743 ns 0.680 ns 1.219 ns 1.004 ns

Diets and time of conditioning are the main effects considered. ns, not significant; * 0.01,P,0.05; ** 0.001,P,0.01; *** P,0.001.

were significantly reduced after 40 days of conditioning (P50.003), but the effect of the diet was not significant (P50.478). Time of conditioning (P50.096) and diet (P5

0.126) did not significantly affect the activities of CS.

4. Discussion

When recently spawned Argopecten purpuratus were conditioned at 16 and 208C at 3 and 6% of their dry mass per day, diet and temperature consistently modified the responses of the biochemical components of the gonad. At both feeding levels, the accumulation of biochemical components in the male and female sections of the gonad was more complete at 16 than at 208C. At 208C, extensive increases in gonadal proteins and lipids only occurred when scallops were fed supplemented microalgae. While

(16)

Fig. 7. Effect of conditioning recently spent Argopecten purpuratus at 168C with three experimental diets at 3% of their body mass per day, on the activities of glycogen phosphorylase, pyruvate kinase, octopine

dehydrogenase and citrate synthase in the adductor muscle. Activities are presented as U g wet mass. Each value represent the means6S.E. (N56). The asterisk over a set of columns indicates that the activities at that sampling time were significantly different from those obtained in the initial sampling (two-factor ANOVA, followed by Tukey’s test, P,0.05).

(17)

gonadal accumulation of biochemical components was more extensive in the scallops fed at 6% of their dry mass per day, similar responses to the experimental diets and temperatures were obtained during the two experiments. At the higher temperature, the microalgal diet must be deficient in some components since even higher feeding levels (6% respect to 3% of the body mass) did not allow sufficient accumulation of biochemical components in the gonad. Significantly lower values of scope for growth were found for the microalgal diet (Navarro et al., 2000). We chose this higher temperature, which is above the range experienced by this population of A. purpuratus to evaluate whether gonadal maturation is accelerated by this fairly small increase in temperature. Instead, our results show that this higher temperature impedes maturation

(Martınez et al., 2000).

In both experiments, muscle carbohydrate levels decreased during accumulation of protein and lipid in the gonad. According to the status of the control scallops, conditioning led to initial increases in muscle carbohydrate levels (experiment I) or to initial maintenance of high levels (experiment II). Subsequently, muscle carbohydrate levels declined gradually, particularly towards the end of the experiment when gonadal protein and lipid contents rose markedly. The scallops fed at 6% of their mass per day depleted muscle carbohydrates to a lesser extent than those fed at 3% in muscle, maintaining 3–5-fold higher values. Muscle carbohydrate reserves are likely mobilised to provide energy or precursors for lipid or protein synthesis in the gonad. In Pecten

maximus, gonadal growth in periods of poor food quality has been shown to coincide with a fall of reserves in muscle (Pazos et al., 1997). Measurements of muscle glycogen levels and ratios of oxygen consumption to nitrogen excretion (O / N ratios) during gametogenesis in Argopecten irradians concentricus (Barber and Blake, 1985), indicate that muscle carbohydrate is converted to lipid which is stored in developing ova.

In agreement with our predictions, the activities of glycogen phosphorylase in the adductor muscle did not decrease during conditioning, but increased during final gonadal maturation. Glycogen phosphorylase liberates glucose moieties from glycogen and would be important for transfer of carbohydrate reserves from muscle to other tissues, particularly if glucosidase activities are in the range typical of muscle (Fournier and Guderley, 1993). After phosphorylase produces glucose-1-phosphate, it can then be converted to glucose-6-phosphate and liberated as glucose after dephosphorylation by glucose-6-phosphatase. The levels of these enzymes should also be maintained during mobilization of muscle carbohydrate.

Reserve mobilization from muscle was accompanied by decreases in the levels of ODH, fluctuations in those of PK and no changes in CS levels. As protein levels did not decrease in the scallops conditioned at 168C, a generalized depletion of muscle proteins cannot explain these changes. ODH, one of the enzymes which maintains redox balance during anaerobic glycolysis (Fields, 1988), decreased during gonadal maturation in scallops fed the three diets. ODH activities follow carbohydrate levels, suggesting their co-regulation. The activities of PK only decreased in scallops fed microalgae, whereas those fed the supplemented diets either showed no changes (lipids) or decreases followed by increases (carbohydrates). PK has a major role in glycolytic regulation in marine invertebrates (Munday et al., 1980; Gaitanaki et al., 1990), being subject to an inhibitory phosphorylation (Holwerda et al., 1981, 1983; Michaelidis et al., 1988;

(18)

Whitwam and Storey, 1990). This regulatory role may require its protection during mobilization of muscle reserves. Interestingly, PK levels were maintained best in the scallops conditioned with the most adequate diet (microalgae supplemented with lipids). The lack of change in CS activity may be due to its mitochondrial localization. Changes in nutritional status bring concomitant modifications of muscle metabolic capacities in numerous fish species. Ration level affects the activity of lactate dehydrogenase in white muscle of sablefish (Anoplopoma fimbria) (Sullivan and Somero, 1983). Activities of several glycolytic enzymes are positively correlated with growth rates in saithe (Pollachius virens) after 2 weeks under differing feeding conditions (Mathers et al., 1992) and in cod (Gadus morhua) after 6–16 weeks exposure to different feeding conditions (Pelletier et al., 1993, 1994; Dutil et al., 1998). Mitochondrial enzymes change less during shifts in growth rates than glycolytic enzymes (Blier et al., 1997).

The impact of changes in reproductive status on muscle metabolic capacities in ectotherms has not been systematically examined, but androgen stimulation of chum salmon (Oncorhynchus keta) decreases the levels of sarcoplasmic proteins (Ando et al., 1986), as does the spawning migration of sockeye salmon (O. nerka) (Mommsen et al., 1980). Since salmon feed little during reproductive maturation, the requirements of gonadal production are largely covered by reserves in the organism. White muscle components thus become prime candidates for deposition in the gonad. Changes in protein levels should have a more direct impact upon the metabolic capacities of muscle than the depletion of carbohydrate reserves.

In scallops, a central role of carbohydrate reserves of adductor muscle in gonadal production has been shown for Chlamys varia (Shafee, 1981), Argopecten concentricus (Barber and Blake, 1981), Argopecten irradians irradians (Epp et al., 1988),

Placopec-ten magellanicus (Couturier and Newkirk, 1991) as well as for ArgopecPlacopec-ten purpuratus

´ ´

(Martınez, 1991; Martınez and Mettifogo, 1998). Interestingly, this role remains even when food availability was quite high as in our second experiment. Presumably, at even higher food levels, the requirement for the depletion of carbohydrate reserves in muscle could disappear. This argument is supported by the maintenance of higher muscle carbohydrate levels in scallops fed at 6% rather than 3% of their dry mass. Muscle protein levels declined during conditioning of adult Argopecten purpuratus when food availability was relatively low (3%) and temperatures high (208C). In Euvola (Pecten)

ziczac, oxidative capacities of mitochondria isolated from the adductor muscle of

scallops which had spawned after a period of high temperatures and low food availability (May) decreased relative to those of mitochondria isolated in more favorable periods (Boadas et al., 1997). Protein levels in the adductor muscle of Euvola were lower in May than in the other periods.

Although muscle protein levels remained stable or increased during conditioning of

Argopecten purpuratus at 168C, muscle metabolic capacities decreased. While this may reflect the greater sensitivity of enzymatic measurements, this pattern is also suggestive of modulation of muscle enzyme levels in accordance with their specific roles. The enzyme that decreased most during reproductive conditioning was ODH, a terminal enzyme of anaerobic glycolysis in molluscs. In gastropods, fish, frogs and mammals, glycolytic enzymes in muscle can be citosolic or attached to structural macromolecules

(19)

(Clarke et al., 1984; Plaxton and Storey, 1986; Lowery et al., 1987; Brooks and Storey, 1991; Guderley et al., 1989; Huber and Guderley, 1993). Enzyme binding is enhanced when glycolytic rates increase, such as during burst exercise. Hence, when glycogen levels decrease, the binding sites for glycolytic enzymes may decrease, accelerating the turnover of these enzymes.

In conclusion, during conditioning of adult Argopecten purpuratus, mobilization of muscle carbohydrate reserves appears to play an important role as it consistently accompanied gonadal maturation. Nonetheless, the depletion of muscle carbohydrate reserves was less extensive when scallops were fed with a higher ration level. Changes in muscle metabolic capacities occurred despite the maintenance of muscle protein levels. The enhanced glycogen phosphorylase activities in muscle towards the end of conditioning may facilitate this carbohydrate mobilization. The decrease in ODH levels during conditioning may reflect their co-regulation with glycogen levels. Finally, conditioning Argopecten purpuratus at 208C did not accelerate gonadal maturation, if anything the increased metabolic demands brought by this higher temperature con-sistently decreased the accumulation of biochemical components in the gonad and led to depletion of muscle reserves, particularly at the lower ration level.

Acknowledgements

´ This study was supported by a grant from the Programa Acuicultura y Biotecnologıa Marina, 1 (97), FONDAP, Chile (Sub-programa Invertebrados). Enzymatic studies were partially supported by a grant to Helga Guderley from NSERC of Canada. [SS]

References

Ando, S., Hatano, M., Zama, K., 1986. Protein degradation and protease activity of chum salmon (Oncorhynchus keta) muscle during spawning migration. Fish Physiol. Biochem. 1, 17–26.

Barber, B., Blake, N., 1981. Energy storage and utilization in relation to gametogenesis in Argopecten

irradians concentricus (Gay). J. Exp. Mar. Biol. Ecol. 52, 121–134.

Barber, B., Blake, N., 1985. Substrate catabolism related to reproduction in the bay scallop Argopecten

irradians concentricus, as determined by O / N and RQ physiological indexes. Mar. Biol. 87, 13–18.

Black, D., Love, R.M., 1986. The sequential mobilization and restoration of energy reserves in tissues of Atlantic cod during starvation and refeeding. J. Comp. Physiol. B. 156, 469–479.

Blier, P.U., Pelletier, D., Dutil, J.D., 1997. Does aerobic capacity set a limit upon fish growth rate? Rev. Fish Sci. 5, 323–340.

Boadas, M.A., Nusetti, O., Mundarain, F., Lodeiros, C., Guderley, H., 1997. Seasonal variation in the properties of muscle mitochondria from the tropical scallop Euvola (Pecten) ziczac. Mar. Biol. 28, 247–255.

´ ´ ´

Brea, J., 1986. Variaciones energeticas estacionales en la composicion quımica de Pecten ziczac (Linne 1758)

´ ´

en relacion con el metabolismo energetico, reproductivo y crecimiento. Thesis, Biology Department, ´

Universidad de Oriente, Cumana, Venezuela.

Brooks, S.P.J., Storey, K.B., 1991. Studies on the regulation of enzyme binding during anoxia in isolated tissues of Busycon canaliculatum. J. Exp. Biol. 156, 193–204.

(20)

Caers, M., Coutteau, P., Cure, K., Morales, V., Gajardo, G., Sorgeloos, P., 1999. The Chilean scallop

Argopecten purpuratus (Lamarck, 1819). II. Manipulation of the fatty acid composition and lipid content of

the eggs via lipid supplementation of the broodstock diet. Comp. Biochem. Physiol. B. 123, 97–103. Clarke, F.M., Stephan, P., Huxham, G., Hamilton, D., Morton, D.J., 1984. Metabolic dependence of glycolytic

enzyme binding in rat and sheep heart. Eur. J. Biochem. 138, 643–649.

Couturier, C.Y., Newkirk, G.F., 1991. Biochemical and gametogenetic cycles in scallops, Placopecten

magellanicus (Gmelin 1791) held in suspension culture. In: Shumway, S.E., Sandiger, P.A. (Eds.), An

International Compendium of Scallop Biology and Culture, World Aquaculture Workshops, Vol. 1, World Aquaculture Society, Baton Rouge, LA, pp. 107–117.

Dutil, J.D., Lambert, Y., Guderley, H., Blier, P.U., Pelletier, D., Desroches, M., 1998. Nucleic acids and enzymes in Atlantic cod (Gadus morhua) differing in condition and growth rate trajectories. Can. J. Fish. Aquat. Sci. 55, 788–795.

Epp, J., Bricelj, V., Malouf, R., 1988. Seasonal partitioning and utilization of energy reserves in two age classes of the bay scallop Argopecten irradians irradians. J. Exp. Mar. Biol. Ecol. 121, 113–136.

Fields, J.H.A., 1988. Fermentative adaptations to the lack of oxygen. Can. J. Zool. 66, 1036–1040. Fournier, P.A., Guderley, H., 1993. Muscle: the predominant glucose-producing organ in the leopard frog

during exercise. Am. J. Physiol. 264 (Regul. Integr. Comp. Physiol. 33), R239–R243.

Gabbott, P.A., Bayne, B.L., 1973. Biochemical effects of temperature and nutritive stress on Mytilus edulis L. J. Mar. Biol. Assoc. UK 53, 269–286.

Gaitanaki, C., Papadopoulos, A., Beis, I., 1990. Time-course of covalent modification of pyruvate kinase during anaerobiosis in the mantle muscle and the hepatopancreas of the limpet Patella caerulea (L). J. Comp. Physiol. B. 160, 529–535.

Guderley, H., Jean, C., Blouin, M., 1989. The effect of fatigue on the binding of glycolytic enzymes in the isolated gastrocnemius muscle of Rana pipiens. Biochim. Biophys. Acta 977, 87–90.

Guderley, H., Lavoie, B.A., Dubois, N., 1994. The influence of age, temperature and growth rate in determining muscle metabolic capacities and tissue masses in the stickleback. Fish Physiol. Biochem. 13, 419–431.

Guderley, H., Dutil, J.D., Pelletier, D., 1996. The physiological status of Atlantic cod, Gadus morhua, in the wild and the laboratory: estimates of growth rates under field conditions. Can. J. Fish Aquat. Sci. 53, 550–557.

Holwerda, D.A., Kruitwagen, E.C.J., De Bont, A.M.T.H., 1981. Regulation of pyruvate kinase and phosphoenol-pyruvate carboxykinase activity during anaerobiosis in Mytilus edulis (L.). Mol. Physiol. 1, 165–171.

Holwerda, D.A., Veenhof, P.R., Van Heugten, H.A.A., Zandee, D.I., 1983. Modification of mussel pyruvate kinase during anaerobiosis and after temperature acclimation. Mol. Physiol. 3, 225–234.

Huber, M., Guderley, H., 1993. The effect of thermal acclimation and exercise upon the binding of glycolytic enzymes in muscle of the goldfish (Carassius auratus). J. Exp. Biol. 175, 195–209.

Kreeger, D.A., Hawkins, A.J.S., Bayne, B.L., Lowe, D.M., 1995. Seasonal variation in the relative utilization of dietary protein for energy and biosynthesis by the mussel Mytilus edulis. Mar. Ecol. Prog. Ser. 126, 177–184.

Kreeger, D.A., Hawkins, A.J.S., Bayne, B.L., 1996. Use of dual-labeled microcapsules to discern the physiological fates of assimilated carbohydrates, protein carbon, and protein nitrogen in suspension-feeding organisms. Limnol. Oceanogr. 41, 208–215.

Lowery, M.S., Roberts, S.J., Somero, G.N., 1987. Effects of starvation on the activities and localization of glycolytic enzymes in the white muscle of the barred sand bass Paralabrax nebulifer. Physiol. Zool. 60, 538–549.

Martınez, G., 1991. Seasonal variation in biochemical composition of three size classes of the Chilean scallop,

Argopecten purpuratus Lamarck 1819. Veliger 34, 335–343.

Martınez, G., Mettifogo, L., 1998. Mobilization of energy from adductor muscle for gametogenesis of the scallop, Argopecten purpuratus Lamarck. J. Shellfish Res. 17, 113–116.

Martınez, G., Aguilera, C., Mettifogo, L., 2000. Interactive effects of diet and temperature on reproductive conditioning of Argopecten purpuratus broodstock Lamarck. Aquaculture 183, 149–159.

Mathers, E.M., Houlihan, D.F., Cunningham, M.J., 1992. Nucleic acids concentrations and enzyme activities as correlates of growth rate in the saithe, Pollachius virens: growth rate estimates of open sea fish. Mar. Biol. 112, 363–369.

(21)

Michaelidis, B., Gaitanaki, C., Beis, I., 1988. Modification of pyruvate kinase from the foot muscle of Patella

caerulea (L.) during anaerobiosis. J. Exp. Zool. 248, 264–271.

Mommsen, T.P., French, C.J., Hochachka, P.W., 1980. Sites and patterns of protein and amino acid utilization during the spawning migration of salmon. Can. J. Zool. 58, 1785–1799.

Munday, K.A., Giles, I.G., Poat, P.C., 1980. Review of the comparative biochemistry of pyruvate kinase. Comp. Biochem. Physiol. B 67, 403–411.

Navarro, J.M., Leiva, G.E., Martınez, G., Aguilera, C., 2000. Interactive effects of diet and temperature on the scope for growth of the scallop Argopecten purpuratus during reproductive conditioning. J. Exp. Mar. Biol. Ecol. (in press).

Pazos, A.J., Roman, G., Acosta, C.P., Abad, M., Sanchez, J.L., 1997. Seasonal changes in condition and biochemical composition of the scallop Pecten maximus L. from suspended culture in the Ria de Arousa (Galicia, N.W. Spain) in relation to environmental conditions. J. Exp. Mar. Biol. Ecol. 211, 169–193. Pelletier, D., Guderley, H., Dutil, J.D., 1993. Effect of growth rate, temperature and season on glycolytic

enzyme activities in white muscle of cod Gadusmorhua. J. Exp. Zool. 265, 477–487.

Pelletier, D., Dutil, J.D., Blier, P.U., Guderley, H., 1994. Relation between growth rate and metabolic organisation of white muscle, liver and digestive tract in cod, Gadus morhua. J. Comp. Physiol. B 164, 179–190.

Plaxton, W.C., Storey, K.B., 1986. Glycolytic enzyme binding and metabolic control in anaerobiosis. J. Comp. Physiol. B156, 635–640.

Shafee, M.S., 1981. Seasonal changes in the biochemical composition and calorific content of the black scallop (Chlamys varia (L.)) from Lanceoc Bay of Brest. Oceanol. Acta 4, 331–341.

Sullivan, K.M., Somero, G.N., 1983. Size- and diet-related variations in enzymatic activity and tissue composition in the sablefish Anoplopoma fimbria. Biol. Bull. (Woods Hole) 164, 315–326.

Taylor, A.C., Venn, T.J., 1979. Seasonal variation in weight and biochemical composition of the tissues of the queen scallop, Chlamys opercularis, from the Clyde Sea Area. J. Mar Biol. Assoc. UK 59, 605–621. Whitwam, R.E., Storey, K.B., 1990. Organ specific analysis of the time course of covalent modification of

(1)

(2)

gonadal accumulation of biochemical components was more extensive in the scallops

fed at 6% of their dry mass per day, similar responses to the experimental diets and

temperatures were obtained during the two experiments. At the higher temperature, the

microalgal diet must be deficient in some components since even higher feeding levels

(6% respect to 3% of the body mass) did not allow sufficient accumulation of

biochemical components in the gonad. Significantly lower values of scope for growth

were found for the microalgal diet (Navarro et al., 2000). We chose this higher

temperature, which is above the range experienced by this population of A. purpuratus

to evaluate whether gonadal maturation is accelerated by this fairly small increase in

temperature. Instead, our results show that this higher temperature impedes maturation

´

(Martınez et al., 2000).

In both experiments, muscle carbohydrate levels decreased during accumulation of

protein and lipid in the gonad. According to the status of the control scallops,

conditioning led to initial increases in muscle carbohydrate levels (experiment I) or to

initial maintenance of high levels (experiment II). Subsequently, muscle carbohydrate

levels declined gradually, particularly towards the end of the experiment when gonadal

protein and lipid contents rose markedly. The scallops fed at 6% of their mass per day

depleted muscle carbohydrates to a lesser extent than those fed at 3% in muscle,

maintaining 3–5-fold higher values. Muscle carbohydrate reserves are likely mobilised

to provide energy or precursors for lipid or protein synthesis in the gonad. In Pecten

maximus

, gonadal growth in periods of poor food quality has been shown to coincide

with a fall of reserves in muscle (Pazos et al., 1997). Measurements of muscle glycogen

levels and ratios of oxygen consumption to nitrogen excretion (O / N ratios) during

gametogenesis in Argopecten irradians concentricus (Barber and Blake, 1985), indicate

that muscle carbohydrate is converted to lipid which is stored in developing ova.

In agreement with our predictions, the activities of glycogen phosphorylase in the

adductor muscle did not decrease during conditioning, but increased during final gonadal

maturation. Glycogen phosphorylase liberates glucose moieties from glycogen and

would be important for transfer of carbohydrate reserves from muscle to other tissues,

particularly if glucosidase activities are in the range typical of muscle (Fournier and

Guderley, 1993). After phosphorylase produces glucose-1-phosphate, it can then be

converted to glucose-6-phosphate and liberated as glucose after dephosphorylation by

glucose-6-phosphatase. The levels of these enzymes should also be maintained during

mobilization of muscle carbohydrate.

Reserve mobilization from muscle was accompanied by decreases in the levels of

ODH, fluctuations in those of PK and no changes in CS levels. As protein levels did not

decrease in the scallops conditioned at 16

8 C, a generalized depletion of muscle proteins

cannot explain these changes. ODH, one of the enzymes which maintains redox balance

during anaerobic glycolysis (Fields, 1988), decreased during gonadal maturation in

scallops fed the three diets. ODH activities follow carbohydrate levels, suggesting their

co-regulation. The activities of PK only decreased in scallops fed microalgae, whereas

those fed the supplemented diets either showed no changes (lipids) or decreases

followed by increases (carbohydrates). PK has a major role in glycolytic regulation in

marine invertebrates (Munday et al., 1980; Gaitanaki et al., 1990), being subject to an

inhibitory phosphorylation (Holwerda et al., 1981, 1983; Michaelidis et al., 1988;

(3)

Whitwam and Storey, 1990). This regulatory role may require its protection during

mobilization of muscle reserves. Interestingly, PK levels were maintained best in the

scallops conditioned with the most adequate diet (microalgae supplemented with lipids).

The lack of change in CS activity may be due to its mitochondrial localization.

Changes in nutritional status bring concomitant modifications of muscle metabolic

capacities in numerous fish species. Ration level affects the activity of lactate

dehydrogenase in white muscle of sablefish (Anoplopoma fimbria) (Sullivan and

Somero, 1983). Activities of several glycolytic enzymes are positively correlated with

growth rates in saithe (Pollachius virens) after 2 weeks under differing feeding

conditions (Mathers et al., 1992) and in cod (Gadus morhua) after 6–16 weeks exposure

to different feeding conditions (Pelletier et al., 1993, 1994; Dutil et al., 1998).

Mitochondrial enzymes change less during shifts in growth rates than glycolytic

enzymes (Blier et al., 1997).

The impact of changes in reproductive status on muscle metabolic capacities in

ectotherms has not been systematically examined, but androgen stimulation of chum

salmon (Oncorhynchus keta) decreases the levels of sarcoplasmic proteins (Ando et al.,

1986), as does the spawning migration of sockeye salmon (O. nerka) (Mommsen et al.,

1980). Since salmon feed little during reproductive maturation, the requirements of

gonadal production are largely covered by reserves in the organism. White muscle

components thus become prime candidates for deposition in the gonad. Changes in

protein levels should have a more direct impact upon the metabolic capacities of muscle

than the depletion of carbohydrate reserves.

In scallops, a central role of carbohydrate reserves of adductor muscle in gonadal

production has been shown for Chlamys varia (Shafee, 1981), Argopecten concentricus

(Barber and Blake, 1981), Argopecten irradians irradians (Epp et al., 1988),

Placopec-ten magellanicus (Couturier and Newkirk, 1991) as well as for ArgopecPlacopec-ten purpuratus

´

(Martınez, 1991; Martınez and Mettifogo, 1998). Interestingly, this role remains even

when food availability was quite high as in our second experiment. Presumably, at even

higher food levels, the requirement for the depletion of carbohydrate reserves in muscle

could disappear. This argument is supported by the maintenance of higher muscle

carbohydrate levels in scallops fed at 6% rather than 3% of their dry mass. Muscle

protein levels declined during conditioning of adult Argopecten purpuratus when food

availability was relatively low (3%) and temperatures high (20

8 C). In Euvola (Pecten)

ziczac, oxidative capacities of mitochondria isolated from the adductor muscle of

scallops which had spawned after a period of high temperatures and low food

availability (May) decreased relative to those of mitochondria isolated in more favorable

periods (Boadas et al., 1997). Protein levels in the adductor muscle of Euvola were

lower in May than in the other periods.

Although muscle protein levels remained stable or increased during conditioning of

Argopecten purpuratus at 16

8 C, muscle metabolic capacities decreased. While this may

reflect the greater sensitivity of enzymatic measurements, this pattern is also suggestive

of modulation of muscle enzyme levels in accordance with their specific roles. The

enzyme that decreased most during reproductive conditioning was ODH, a terminal

enzyme of anaerobic glycolysis in molluscs. In gastropods, fish, frogs and mammals,

glycolytic enzymes in muscle can be citosolic or attached to structural macromolecules

(4)

(Clarke et al., 1984; Plaxton and Storey, 1986; Lowery et al., 1987; Brooks and Storey,

1991; Guderley et al., 1989; Huber and Guderley, 1993). Enzyme binding is enhanced

when glycolytic rates increase, such as during burst exercise. Hence, when glycogen

levels decrease, the binding sites for glycolytic enzymes may decrease, accelerating the

turnover of these enzymes.

In conclusion, during conditioning of adult Argopecten purpuratus, mobilization of

muscle carbohydrate reserves appears to play an important role as it consistently

accompanied gonadal maturation. Nonetheless, the depletion of muscle carbohydrate

reserves was less extensive when scallops were fed with a higher ration level. Changes

in muscle metabolic capacities occurred despite the maintenance of muscle protein

levels. The enhanced glycogen phosphorylase activities in muscle towards the end of

conditioning may facilitate this carbohydrate mobilization. The decrease in ODH levels

during conditioning may reflect their co-regulation with glycogen levels. Finally,

conditioning Argopecten purpuratus at 20

8 C did not accelerate gonadal maturation, if

anything the increased metabolic demands brought by this higher temperature

con-sistently decreased the accumulation of biochemical components in the gonad and led to

depletion of muscle reserves, particularly at the lower ration level.

Acknowledgements

´

This study was supported by a grant from the Programa Acuicultura y Biotecnologıa

Marina, 1 (97), FONDAP, Chile (Sub-programa Invertebrados). Enzymatic studies were

partially supported by a grant to Helga Guderley from NSERC of Canada.

[SS]

References