Functional evidence for the presence of a carbonic anhydrase repressor in the eyestalk of the euryhaline green crabCarcinus maenas

Abstract

Carbonic anhydrase (CA) activity and relative expression of CA mRNA were measured in the gills of the euryhaline green crab Carcinus maenas in response to eyestalk ablation (ESA), injection of eyestalk extract and exposure to low salinity. For crabs acclimated to 32 p.p.t. salinity, ESA alone resulted in an increase in both CA activity and relative mRNA expression in the posterior, ion-transporting gills, but not in the anterior, respiratory gills. The ESA-stimulated increase in CA activity was abolished by injections of extracts of eyestalks taken from crabs acclimated to 32 p.p.t. salinity. Transfer of intact crabs from 32 to 10 p.p.t. salinity for 7 days resulted in an eightfold increase in CA activity and a sixfold increase in mRNA expression in posterior gills. ESA potentiated the normal low salinity-mediated CA induction by 23%. Daily injections of eyestalk extract reduced low salinity-stimulated CA induction by nearly 50% in intact crabs and by almost 75% in eyestalk ablated crabs. A 4-day transfer to 10 p.p.t. salinity also caused significant increases in both CA activity and mRNA expression in posterior gills, and ESA resulted in a 32% increase in the normal degree of CA induction. Daily injections of eyestalk extracts reduced CA induction in a dose-dependent manner over the 4-day time course. When CA induction was reduced by 66%, hemolymph osmotic regulation was also disrupted. These results are functional evidence for the presence of a CA repressor in the major endocrine complex of the crab, the eyestalk. This compound appears to function in keeping CA expression at low, baseline levels in crabs at high salinity. Exposure to low salinity removes the effects of the putative repressor, allowing CA expression, and thus CA activity, to increase.