BIOCHEMICAL SOCIETY TRANSACTIONS

900 Exogenous

\

I I I

Endogenous

I

I

q ,gen

1

lipid

Fig. 2. Sources of egg lipid

constant. The increase in triacylglycerol content is represented by the shaded area and accounts for up to 25 % of triacylglycerol fatty acids present during gametogenesis. It is proposed that there is a balance between exogenous (dietary) and endogenous (from mantle glycogen) sources of egg lipid, which depends on the amount of food available during each season. This is shown in Fig. 2. Throughout 1976-1977 the dietary uptake of lipid may have been sufficient or very nearly sufficient to provide all the fatty acids for incorporation into egg triacylglycerol. In the winter of 1977 there may have been less lipid available, and the mantle triacylglycerol, which decreased during November-December, was restored to the normal concentration by an increase in synthesis de novo of fatty acids, particularly palmitoleic acid, from the stored glycogen reserves. In this way the mussel is able to buffer against changes in food availability, and maintain a high concentration of mantle triacylglycerol during the period of gametogenesis. Calzolari, C., Cerrna, E. & Stancher, 8.(1971) La Riuisfa Iral. Sostanze Grasse 48rh, 605-611 Gabbott, P. A. (1975)Proc. Eur. Mar. Biol. Symp. 9th (Barnes, H., ed.), pp. 191-211, Aberdeen University Press, Aberdeen Jeffcoat, R., Brown, P. R., Safford, R. &James, A. T. (1977) Biochern. J . 161,431-437 Jeffcoat, R. & James, A. T. (1977) Lipids 12,469-474 Kanazawa, A,, Teshirna, S. & Tokiwa, S. (1977~)Bull. Jpn. SOC.Sci. Fish.43, 849-856 Kanazawa, A , , Tokiwa, S., Kayarna, M. & Hirata, M. (1977b) Bull. Jpn. Soc. Sci. Fish. 43, 1111-1114

The Contribution of the Pentose Phosphate Cycle to the Central Pathways of Metabolism in the Marine Mussel, Mytifus edufis L. BOGUMIL N. ZABAand J. ISLWYN DAVIES

Department of Biochemistry and Soil Science, University College of North Wales, Bangor, Gwynedd LL57 2UW, Wales, U.K. It is clear that the metabolism of the marine mussel has undergone extensive adaptation to its inter-tidal environment, in which there occur both daily fluctuations in oxygen 1979

901

583rd MEETING, CAMBRIDGE

availability and seasonal changes in the relationship between food supply and energy demand (Gabbott, 1976; De Zwaan, 1977). The occurrence of the pentose phosphate pathway is suggested by the presence of glucose 6-phosphate dehydrogenase and gluconate 6-phosphate dehydrogenase (Rodrigues-Segade et al., 1978)and by the pattern of metabolism of specifically labelled glucose (Bennet & Nakada, 1968). The present paper describes the pentose-phosphate-pathway activity of mussel mantle tissue, which is distinguished from the other tissues mainly in the demands made upon it by the annual gametogenic cycle. Sliced preparations of mantle tissue were incubated at 20°C for 150min in an Oz/C02 (19 :1) atmosphere in Krebs-Ringer bicarbonate buffer (pH7.4) containing various 14C-and 3H-labelled substrates. Its metabolic activity was similar to that of the mantle tissue of the intact animal under aerobic conditions in that the distribution of 14Clabel from [U-14C]glucoseamong the various products formed (in a typical experiment : glycogen, 48%; lipid, 1 %; amino acid, 35%; organic anions, 14%; COz, 5%) was very similar to that in mantle tissue excised after injecting the substrate into the circulatory system of the intact animal and incubating in filtered sea water for 3h. The metabolism of specifically labelled glucose by the mantle slice preparation is shown in Table 1. The proportion of the glucose utilized that is converted to COz is low. This is particularly true for [6-14C]glucose for which l4CO2emerges specifically from the tricarboxylic acid cycle. Since this is also a feature of the utilization of [I-14C]pyruvate, but not of [1,4-14C]succinate,it is tentatively suggested that low pyruvate dehy-

Table 1. Utilization of specifically labelled glucose by mantle tissue of f e d and starved mussels The animals (average length, 6cm; collected January 1979 from Menai Straits, Anglesey) were maintained in an aquarium for 3 weeks with constant-circulating filtered sea-water at ambient temperature. Fed animals were provided with a mixed algal diet at a rate exceeding the maintenance ration. Sliced preparations of mantle tissue (50mg/ml) were incubated (see the text) in media containing dual-labelled glucose ( 5 m ~14C, ; 0.2pCi/ml; 3H, 0.4pCi/ml). 14C02was collected after acidification of the medium, a sample of which was distilled in order to assess 3H20formation. Other products were separated essentially as described by De Zwaan & Van Marrewijk (1973). Each result is the mean of values obtained by analysis of four incubations. Medium glucose utilized (nmol/l50min per lOOmg of tissue) Starved

Fed r

Product 3Hz0 [3H]Glycogen E3H l4COZ [14C]Glycogen 14C-labelled amino acids 14C-labelled organic acids Z14C Z14C/Z3H Pentose phosphate. pathway estimate

Vol. 7

[1-'4C,2-3H] GI ucose 1085 156 1241 34 226 220

[6-14C,2-3H] Glucose 1635 248 1883 4 289 351

33

47

513 0.41

69 1 0.37 10.3

\

[1-'4C,2-3H] Glucose 914 135 1049 33 181 1 60

[6-14C,2-3H] Glucose 1181 182 1363 2 183 294

46

,.

50

420 0.40

529 0.39 /

10.8

902

BIOCHEMICAL SOCIETY TRANSACTIONS

drogenase activity may restrain the operation of the tricarboxylic acid cycle, although this has a considerable oxidative capacity (cf. Addink & Veenhof, 1975; Zaba e t al., 1978). As the formation of 14C02in the tricarboxylic acid cycle decreases, the ease with which the decarboxylation of 6-phosphogluconate in the pentose phosphate pathway can be detected increases. The deviation of the Cl/CBratio from unity is more readily seen in C 0 2 than in other products. Even if the unrealistic assumption is made that all 14C02 released in incubations with [l-'4C]glucose emerges from the pentose phosphate pathway, the implied rate of flux through the pathway remains a very small proportion of total glucose utilization. When calculated using the equation derived by Katz & Wood (1963), the data shown in Table 1 indicate that the flux through the pentose phosphate pathway is 10.5nmol of glucose/l5Oniin per 100mg. By investigating the metabolism of [U-I4C, 2-3H]glucose, differences between tissue preparations in their rates of glucose utilization can be taken into account. The total recovery of 3H (mainly as 3H20and [3H]glycogen) and I4C in the metabolic products appear to diverge at certain periods of the year. It is suggested that the 14C/3Hratio of 0.39 :1 shown in Table 1 may indicate the occurrence of some form of substrate cycling. The precise role of the pentose phosphate pathway remains uncertain. As a source of pentose sugars and NADPH, it is clearly capable of contributing to the synthesis of nucleic acids and lipids required for gametogenesis in male and female mussels respectively. The incorporation of I4C from labelled glucose (exogenous) into lipid is, however, low at all times of the year. Nevertheless, there are indications that the intermediates of glucose metabolism have a considerably lower specific activity than the added substrate, implying that lipogenesis from endogenous sources may be more significant. A proportion of the NADPH generated by the pentose phosphate pathway may be required in regulating the redox balance of the cells and to meet the demands of glutathione-mediated mechanisms, which exist to counter the potential toxicity of the aerobic environment (Fridovich, 1976; cf. Beutler, 1972). We gratefully acknowledge the financial support provided by the Natural Environment Research Council. Addink, A. D. F. & Veenhof, P. R. (1975) in Proc. Eur. Mar. Biol. Symp. 9th (Barnes, H., ed.), pp. 109-1 19, Aberdeen University Press, Aberdeen Bennett, R. & Nakada, H. (1968) Comp. Biochem. Physiul. 24,787-797 Beutler, E. (1972) in ZnheritedBasisof Metabolic Disease. (Stanbury, J. B., Wyngaarden, J. B. & Fredrickson, D. S . , eds.), 3rd edn., pp. 1358-1388, Academic Press, New York De Zwaan, A. (1977) Oceanogr. Mar. Biol. Annu. Rev. 15, 103-187 De Zwaan, A. & Van Marrewijk, W. J. A. (1973) Comp. Biochem. Physiol. B 44,429439 Fridovich, I. (1976)in Free Radicals in Biology(Pryor, W. A., ed.), vol. 1, pp. 239-277, Academic Press, New York Gabbot, P. A. (1976) in Marine Mussels (Bayne, B. L., ed.), pp. 293-335, Cambridge University Press, Cambridge Katz, J. &Wood, H. G . (1963)J. Biol. Chem. 238, 517-523 Rodrigues-Segade, S., Friere, M. & Carrion, A. (1978) Biochem. J . 170,577-582 Zaba, B. N., de Bont, A. M. T. & De Zwaan, A. (1978) Znt. J . Biochem. 9,191-197

The Effects of pp'-DDT [l,l,l-trichloro-2,2-bis(p-chlorophenyl)ethane] on Carbohydrate Metabolism in the Developing Neonatal Rat MICHAEL R. L. STRATFORD* and KEITH SNELL Department of Biochemistry, University o f Surrey, Guildford, Surrey GU2 5 X H , U.K.

The organochlorine insecticide DDT [1,1,l-trichloro-2,2-bis(p-chlorophenyl)ethane] has attracted notoriety with regard to its human toxicity, leading to a partial or total * Present address : Gray Laboratory, Mount Vernon Hospital, Northwood, Middx., U.K. 1979