Pergamon Comp. Biochem. Physiol. Vol. IOU, Nos 213, pp. 337-341, 1994

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Comparative Nutrition Papers

Seasonal variation of the carbohydrate and lipid metabolism in a land pulmonate gastropod, Megalobulimus oblongus

Roselis S. M. Da Silva and Denise Maria Zancan Departamento de Fisiologia, Centro de Endocrinologia Experimental, Instituto de BiociCncias, Universidade Federal do Rio Grande do Sul, 90.050-170 Porto Alegre-RS-Brazil

Seasonal variations of carbohydrate and lipid metabolism in Megafobuhus oblongus were investigated. Haemolymph glucose levels were higher in summer than in other seasons. The glycogen content of the hepatopancreas, muscle and mantle was constant throughout the year, except in winter when lower values were found. Hepatopancreas total lipid increased in spring, whereas gonad total lipid increased in winter and spring. Glycogen metabolism seems to constitute the source of energy substrate during winter while lipid metabolism may be related to the annual reproductive cycle.

Key words: Megalobulimus oblongus; Seasonal variation.

Comp. Biochem. Physiol. 108A, 337-341, 1994.

Introduction

In their habitat, snails are exposed to a great number of environmental variables such as temperature, humidity, photoperi- odicity and a reduced availability of energy substrate, that induce behavioral and meta- bolic changes.

In many molluscs, the glycogen storage- mobilization cycle is directly and inversely related to the annual reproductive cycle: glycogen is stored during periods of abun- dant food supply and is mobilized to fuel gametogenesis in winter and early spring (de Zwaan and Zandee, 1972; Gabbott, 1975, 1983). Pulmonate snails show a clear seasonality in their glycogen reserves. The

Correspondence fo : Roselis Silveira Martins da Silva, Departamento de Fisiologia, Instituto de Biocih- cias UFRGS, Rua Sannento Leite, 500, 90.050- 170 Port0 Alegre-RS-Brazil.

Received 15 July 1993; accepted 17 September 1993.

terrestrial gastropod Helix pomatia showed a marked deposition of glycogen in tissues during the autumn months preceding hiber- nation. The high level of glycogen found in the tissues declined during hibernation, and also, albeit somewhat less clearly, during the period of egg-laying (Joosse, 1988).

In the natural habitat of the freshwater pulmonate Lymnaea stagnalis, glycogen levels are low during the reproductive sea- son, in the summer, and increase during autumn, when oviposition stops, but food consumption continues (Scheerboom and van Elk, 1978). Hemminga et al. (1985) have demonstrated that the photoperiod might be the key factor in inducing the above mentioned seasonal changes in glyco- gen storage in Lymnaea.

In order to obtain more information on the control of the storage-mobilization

337

338 Rosclis S. M. Da Silva and Denise Maria Zancan

cycle of energy reserves in the terrestrial pulmonate snail Megalobulimus oblongus, in this paper we investigate the effects of seasonal variation on the energy metab- olism and on the weight of albumen gland and gonad.

Materials and Methods Experimental animals

Adult Megalobulimus oblongus (Pul- monata, Stylomatophora), snails weighing 45-70 g were collected at Charqueadas, Rio Grande do Sul, a subtropical region in Brazil. The animals were kept at ambient temperature in terraria containing a sand layer, flushed with tap water, and were fed fresh lettuce daily. The mean ambient tem- perature was 24°C in summer, 17°C in fall, 16°C in winter and 20°C in spring. The animals were used 15 days after capture. The snails were kept in the laboratory on the day of the experiment. The experiments were performed throughout the year; five snails were used each month.

Fig. 1. Seasonal variation in haemolymph glucose concentration in Megalobulimus oblongus. Data are given as means f SEM. Number of snails in paren-

theses.

again, reaching significantly higher levels (P < 0.01) in summer as compared to winter.

The haemolymph samples were taken by heart ventricle puncture. Gonad, albumen gland and samples of hepatopancreas, an- terior mantle and muscle (diaphragm) were dissected, weighed and immediately pro- cessed.

The results in Fig. 2 show the glycogen concentration in the muscle and mantle. Throughout the year, a certain constancy can be observed in glycogen levels in the two tissues, except in winter when lower values were found (P < 0.01) as compared to mean levels in the other three seasons.

Chemical analyses

Haemolymph glucose concentration was determined by the enzymatic oxidase method. Hepatopancreas, mantle and muscle glycogen was extracted essentially according to van Handel (1965) and deter- mined as glucose after acid hydrolysis. Tis- sue lipids were determined gravimetrically after extraction by the procedure of Folch et al. (1957).

Statistical analyses were performed by 8 analyses of variance followed by multiple 14 2

range testing; P < 0.05 was taken as the criterion of significance.

1

ReSUlts

Figure 1 shows that in summer, high haemolymph glucose levels are found, which decrease (P < 0.01) during autumn and winter, when the lowest concentrations occur. However, in the spring, the haemolymph glucose concentration rises

Fig. 2. Seasonal variation in glycogen content in the mantle and muscle in Meg~obulimus oblongus. Data are given as means f WM. Number of snails in . .

parentnews.

Seasonal variation metabolism in a gastropod mollusc 339

q sumtaor q wiator R Poll Rslh

(26) (14) -r

Fig. 3. Seasonal variation in glycogen content in the hepatopancreas in Megalobulimus oblongus. Data are given as means f SEM. Number of snails in paren-

theses.

Seasonal variations in the glycogen con- tent of hepatopancreas were detected (Fig. 3). The lowest (P < 0.01) values are found during winter; in the spring they increase 32% as compared to winter, reach- ing 63% and 80% increases in summer and autumn, respectively.

Figure 4 shows the data regarding the total lipid concentration in hepatopancreas

10

9

8

7

6

5

4

Heptiopulcreu (18) T

(8)

Fig. 4. Seasonal variation in total lipid content in the hepatopancreas and gonads in Megalobulimus oblongus. Data are given as means f SEM. Number

of snails in parentheses.

Table 1. Seasonal variation in the wet weight of hepatopancreas and gonads in Megalobulimus

oblongus

Albuman gland Gonads (mg of wet weight)

Fall 456 f 140 460*40 (23) (12)

Winter 244f73 215 f 32 (17) (11)

Spring 2517 + 358 441*35 (18) (13)

Summer 667 f 166 460 & 49 (16) (16)

Data are given as means If: SEM. Number of snails in parentheses.

and gonads. The total lipid values in hepa- topancreas increase 2-fold in spring as com- pared to those in the other three seasons. Winter and spring induce a 2-3-fold in- crease in the gonadal total lipid concen- tration when compared to that in summer and autumn.

Table 1 shows the wet weight of the albumen gland and gonad. The wet weight of the albumen gland increases lo-fold (P < 0.001) in spring when compared to that in winter. In the summer and autumn the wet weight of albumen gland decreases 4-6-fold (P < O.Ol), when compared to that in spring. The wet weight of the gonad shows a certain constancy throughout the year, except in winter, when the weight decreases 2-fold as compared to that in the other seasons.

Discussion

In the course of the year, the haemo- lymph glucose concentrations of the field Megdobulimus exhibited seasonal vari- ations. The highest values were found during summer, decreasing during autumn and winter; in spring a 30% increase was found in relation to winter, indicating a tendency to increase confirmed in summer (Fig. 1). These data agree with the results of Marques and Pereira (1970), working with the same species and Scheerboom and van Elk (1978) in Lymnuea stugnulis. However, in Megulobulimus the order of magnitude of variation found between the maximum (summer) and minimum (winter) values of haemolymph glucose levels were lower than in Lymnueu (Scheerboom and van Elk, 1978).

340 Roselis S. M. Da Silva and Denise Maria Zancan

Few variations in glycogen contents in mantle and muscle were found during the seasons, except in winter, when a rather sharp drop in these levels was found (Fig. 2). The glycogen concentration in hepatopancreas was higher in summer and autumn and was markedly reduced during winter. In spring, the mean glycogen con- centration rose 32% compared to that in winter, indicating a tendency to increase confirmed in summer and autumn (Fig. 3). This slower increase in glycogen levels in hepatopancreas was accompanied by 93% increase in total lipid concentration in the digestive gland (Fig. 4). In the winter months, M. oblongus stay in their shell buried in the earth and food consumption decreases, as in other gastropod molluscs (Joosse, 1988). The glycogen concentration in tissues seems to constitute the major source of energy during hibernation, since the total lipid levels in the hepatopancreas remain constant during autumn and winter time (Fig. 4). However, the stabilization of haemolymph sugar levels during winter time, and the fact that appreciable levels of hepatopancreas and mantle glycogen were found, and the muscle glycogen levels were utilized, seem to indicate that lactate de- rived from muscle polysaccharides might be an important source of carbon chains for gluconeogenesis in this mollusc. Moreover, the stabilization of haemolymph glucose levels during winter may be due to the reduced utilization of carbohydrate as a consequence of a decrease in metabolic rate (Marques and Pereira, 1970).

Despite the fact that, in Megdobulimus, the foot muscle glycogen did not show any consistent change during the year (Marques and Pereira, 1970), in the present study a parallel change in the glycogen contents of the mantle and diaphragm muscle was found (Fig. 2). However, in contrast to the reserves in the mantle, glycogen in muscle cells is presumably only an intracellular energy reservoir.

The total lipid contents in ovotestis and hepatopancreas rose considerably during late winter and spring (Fig. 4). At the same time the ovotestis began the female gameto- genesis; the total lipid levels were maximal when the ovotestes were fully gravid, and markedly decreased after the liberation of gametes (Horn and Zancan, 1992). These

data agree with the results of Nanaware and Varute (1976), working in Semperuh macu - lata, another Stylomatophora. These authors suggest that lipids may be con- verted into sugars and used as energy during gametogenesis. The total lipid levels in the ovotestis can also be related to its hormonal activity, since many authors have demonstrated the synthetic capacity for, or the presence of, steroids in gastropod go- nadal tissue (de Jong-Brink et al., 198 1; Lehoux and Williams, 1971; Takeda, 1979). Also the total lipid in ovotestis can be used as a nutritive substance for the eggs, whereas in Lymnuea and Bithyniu large lipid droplets are found in the follicle cells and oocytes during the vitellogenic phase (Dohmen, 1983).

The haemolymph glucose concentration will be the result of the glucose influx via the gut on the one hand, and of the glucose utilizing processes of the tissues on the other. Adult specimens show an annual cycle of oviposition correlated with the end of spring and the beginning of the summer (Horn and Zancan, 1992). This maturation of the female reproductive system can be demonstrated by determining the wet weight of the albumen gland (Table 1). Therefore, the most likely explanation for the low levels in haemolymph glucose and hepatopancreas glycogen in spring is an increased utilization of glucose by the syn- thetic processes (galactogen, glycogen and lipid synthesis) in the female organs. Veld- huijzen and Dogterom (1975) concluded that, in Lymnaea, the galactogen and glyco- gen syntheses in the female organs were very important for the dynamics of the haemolymph glucose concentration.

The annual reproductive cycle in Megdobulimus shows that male gametoge- nesis occurs twice a year, during spring and autumn. Sperm appeared in the hermaphrodite duct in January (summer) and in July (winter), while female gameto- genesis takes place only during the spring. As the sperm was voided from the gonad into the hermaphrodite duct, the decrease in gonadal wet weight in winter (Table 1) was reflected histologically by the shrunken acini (Horn and Zancan, 1992).

In Megdobulimus oblongus, under field conditions, the hepatopancreas, mantle and diaphragm muscle glycogen seem to

Seasonal variation metaboli km in a gastropod mollusc 341

Joosse J. (1985) Effects of photoperiod and tem- perature on the glycogen stores in the mantle and the head-foot muscles of the freshwater pulmonate snail Lymnaea stagnalis. Comp. Biochem. Physiol. NIB, 139-143.

Horn A. C. M. and Zancan D. M. (1992) Ciclo reprodutivo do gartropodo pulmonado Megalobu- limus oblongus. In Proceeding of VII Reunirio Anual da Federa@ de Sociedades de Biologia Experimen - tal p. 241. Caxambti, MG, Brasil.

Joosse J. (1988) The hormones of molluscs. In Invert - ebrate Endocrinology. Endocrinology of Selected Invertebrate Types (Edited by Laufer H. and Downer R. G. H.), Vol. 2, pp. 89-140.

Lehoux J. G. and Williams E. E. (1971) Metabolism of progesterone by gonadal tissue of Littorina littorea (L.) (Prosobranchia, Gastropoda). J. En- doer. 51, 411-412.

Marques M. and Pereira S. (1970) Seasonal vari- ations in blood glucose and glycogen levels of some tissues of Strophocheilus oblongus. Rev. Brasil. Biol. 30, 4348.

Nanaware S. G. and Varute A. T. (1976) Biochemical studies on the reproductive organs of a land pul- monate, Semperula maculata (Tompleton, 1858; Semper, 1885) during seasonal breeding-aestiva- tion cycle: I. Biochemical seasonal variation in proteins and lipids. The Veliger 19, 96106.

Scheerboom J. E. M. and van Elk R. (1978) Field observations on the seasonal variations in the natural diet and the haemolymph-glucose concen- tration of the pond snail Lymnaea stagnalis (L.). Proc. Kon. Ned. Akad. Wet. 81C, 365376.

Takeda N. (1979) Induction of egg-laying by steroid hormones in slugs. Comp. Biochem. Physiol. 62A, 273-278.

van Handel E. (1965) Estimation of glycogen in small amount soft tissue. Analyt. Biochem. 11, 256-265.

Veldhuijzen J. P. and Dogterom, G. E.(1975) Incor- poration of “C-glucose in the pond snail Lymnaea stagnalis as affected by starvation. Neth. J. Zool. 25, 247-260.

constitute the largest source of energy sub- strate during winter, when the animal is less active, whereas the hepatopancreas and gonad lipids can be closely related to the annual oogenesis cycle.

Acknowledgements-This study was supported in part by grants from conv&io FINEP/UFRGS (No. 66.91. 0509.00), from CNPq and FAPERGS.

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