亚硝态氮胁迫对日本蟳消化酶活力及同工酶表达的影响

亚硝态氮是养殖水体中常见的胁迫因子.为探讨亚硝态氮对养殖蟹类的毒性作用,采用生物酶测定及聚丙烯酰胺凝胶电泳的方法,研究了水体中不同浓度(0.15,0.3,2,5,10,20mg/L)亚硝态氮胁迫下日本蟳肝胰腺消化酶活力的变化,以及高浓度(20mg/L)亚硝态氮胁迫对日本蟳同工酶表达的影响.结果表明:低浓度(0.15,0.3mg/L)的亚硝态氮胁迫对碱性蛋白酶,酸性蛋白酶,脂肪酶和淀粉酶活力均产生一定的诱导效应,且在处理7d时,除淀粉酶活力略低于对照组外,其他3种消化酶活力仍保持在较高的水平.较高浓度(2,5,10mg/L)亚硝态氮的短期胁迫(0.5~1d)会诱导酸性蛋白酶,碱性蛋白酶和脂肪酶活力的迅速升高,但随即快速下降.高浓度(20mg/L)的亚硝态氮胁迫对蛋白酶和淀粉酶活力表现出明显的抑制效应.处理7d时,胁迫浓度2mg/L及以上的实验组各消化酶活力与剂量浓度间均呈显著负相关关系.20mg/L亚硝态氮胁迫下,除肌肉中新增两条酶带(MDH-2和MDH-4)外,日本蟳鳃,肝胰腺,胃,心脏,卵,精子等组织中的α-淀粉酶(α-AMY)同工酶,乳酸脱氢酶(LDH)同工酶,苹果酸脱氢酶(MDH)同工酶和过氧化物酶(POD)同工酶均出现活性减弱或酶带数量减少现象.结果显示高浓度亚硝态氮胁迫对日本蟳消化酶活力和同工酶表达均表现出明显的抑制作用.

The effect of nitrite stress on digestive enzyme activities and isozyme expression of marine crab,Charybdis japonica

Nitrite is one of the most common stress factors in aquaculture water. In this study, the digestive enzyme activities in the hepatopancreas of Charybdis japonica under different concentrations of nitrite stress (0.15, 0.3, 2, 5, 10, 20mg/L) were measured by using bioenzymatic determination, and meanwhile the high concentration of nitrite stress (20mg/L) on the expression of tissue isozymes were also examined with polyacrylamid gel electrophoresis in order to explore the toxic mechanism of nitrite on aquatic crabs. The results showed that the activities of akaline protease, acid protease, lipase and amylase were all induced to some degree after exposure to lower concentrations of nitrite (0.15, 0.3mg/L). On day 7after exposure, all enzyme activities were still kept at high levels except the amylase activity was slightly lower than the control group. The activities of hepatopancreatic akaline protease, acid protease and lipase first increased rapidly, then following a dramatic decrease after exposure to moderate nitrite stress (2, 5, 10mg/L) for short time (0.5~1d). But higher concentration of nitrite stress (20mg/L), resulted in a significant inhibition of hepatopancreatic protease and amylase activities. On day 7after exposure, digestive enzyme activities showed a significantly negative correlation with increasing nitrite concentrations after the stress was over 2mg/L. The expression of α-amylase (α-AMY), lactate dehydrogenase (LDH), malic dehydrogenase (MDH) and peroxidase (POD) isozymes in gill, hepatopancreas, stomach, heart, ovum and sperm was significantly affected by high concentration of nitrite stress (20mg/L): isozyme activities declined or their expression bands reduced except that the muscle was induced to express MDH-2and MDH-4. The results indicated that high concentration of nitrite had a significant adverse effect on the digestive enzyme activities and the expression of tissue isozymes of C. japonica.