Effect Of Cadmium On The Activltes Of Glutamate Dehydrogemase, Alanine And Aspartate Aminotransferase In Fresh Water Snails, Helisoma Duryi And Lymnea Natalensis

Presented at the Sida,SAREC Water Project Conference in December 2000.,Snails are known to accumulate metal ion pollutants in their tissues this being attributed to induction of metal binding proteins (Petering and Fowler. 1986). A consequence of the presence of pollutants in water inhabited by aquatic organisms may be the induction of enzymes required to metabolise or degrade the pollutants in such organisms. Other enzymes may also be induced in response to toxic effects of these pollutants on metabolic pathways in which these enzymes are involved. We are investigating the potential use of key enzymes of amino acid metabolism as markers of pollution due to metal ions in fresh water snails. Helisoma dllryi and Lnnl1ea natalensis. Experimental snails were drawn from concrete breeding tanks where they were regularly fed on lettuce. The snails were exposed for 96 hours to 0.0 1. 0.1 and I ppm concentrations of cadmium ion as a chloride salt. After exposure. snails were shelled excluding any dead snails. The tissue was homogenised and centrifuged at 500 x g for 10 minutes at 4PC to pellet nuclei and unbroken cells. The post nuclear supernatant was centrifuged at 10 000 x g for 10 minutes at 4uC. After suspension of the resulting pellet in buffer. both the pellet fraction ("mitochondria'" fraction) and the supernatant (cytosolic fraction) were aliquoted and stored at -82uC. The samples \vere assayed for the activities of glutamate dehydrogenase. and aspartate and alanine aminotransferases. The concentration of cadmium in breeding waters and in shells and tissues (not homogenised) was also determined. The concentration of cadmium in tissues rose with increasing concentTation the metal ion in breeding waters. In the absence of Cd added i.e. 0.03 and 0.02 1lg./ml Cd concentration in the breeding waters of Helisoma and Ll'lIlI1t!a respectively. the concentration of Cd was 0.08Ilg/g. in Helisvma and Ln7lnea tissues. This concentration rose to 0.47 and 0.37 Ilg/g. at I ppm added Cd for Helisoma and L.1'ml1ea respectively. showing 5.9 and 4.6 fold increases over initial concentration. Cadmium was also found to accumulate in shells of the two snail species. In general the activities of glutamate dehydrogenase (GDH). aspartate and alanine aminotransferases (AST and ALT) increased with concentration of cadmium but then decreased at Ippm added metal for GDH and AST. and at 0.1 ppm for ALT. In both snails consistent changes in GDH activity were seen in the homogenate and 10 000 x g pellet. At 0.1 ppm added cadmium the increased activity of GDH in the 10 000 x g pellet was 2.5 and 3.6 fold over initial activities for Helisol11a and Ll'Il1l1ea respectively. AST however showed consistent changes in the homogenates and 10 000 x g supernatants for both snails. At 0.1 ppm added cadmium the increased activity of AST in the 10 000 x g sup ematant was 3.4 and 1.8 over initial activities for Helisol11o and (1'IIlI1ea respectively. ALT also showed a similar pattern of activity change in the homogenate and 10 000 x g supernatant although decrease in activity staned much earlier at 0.1 ppm added metal. In the 10 000 x g pellets ALT activity progressively declined froin the initial values to reach 3-t?'0 and 43% of these values at I ppm added Cd for Helisol11a and (Hl/l1ea respectively. Since alanine and aspanate aminotransferases are known to be dually localised in the mitochondria and cy10so1 in a number of species. a possibility that enzyme could have redistributed due to organelle damage is unlikely in view of the low initial activities in the pellets. Funher the increase in enzyme activit. with metal ion concentration also occurred in the homogenates. The increases in enzyme activities were therefore likely to be due to induction of enzymes. Homogenate enzyme activities as well as those of pellet GDH and ALT. and supernatant AST could be sensitive indicators of Cd pollution,Sida,SAREC Water Projec