SHAFER, T.H.*; MONTOYA, L.M.; HOFFMAN, G.G.: Developmental and Environmental Expression of the Gene for a Na/K/2Cl Cotransporter in the Blue Crab (Callinectes sapidus)

Juvenile blue crabs (Callinectes sapidus) inhabit low salinity environments (5 ppt or less) in the tidal rivers and creeks of southeastern NC. Larvae generally hatch from ovigerous females in sea water (about 35 ppt). It is possible that an ontogenetic change in the capacity to osmoregulate dictates the developmental stage at which recruitment to estuaries occurs. Reverse transcriptase-polymerase chain reaction (RT-PCR) was employed to examine the expression of genes potentially coding for parts of the osmoregulatory mechanism. A V-type proton pump is expressed at all embryonic and larval stages examined. It is also expressed at high levels in both the anterior (gas exchange) and posterior (ion transporting) gills of adult blue crabs. Expression of the V-ATPase is equally high in the hepatopancreas. For these reasons, this gene does not appear to be specifically induced as a requirement for osmoregulation. On the other hand, the mRNA for a Na/K/2Cl cotransporter shows interesting tissue-related, stage-related, and salinity-related expression. It is present in both gill types but is much more strongly expressed in the posterior gills. It is absent in the hepatopancreas. The cotransporter mRNA is present at high levels in all embryonic stages from gastrula to hatching. However, the gene is not expressed in megalopae at 35 ppt salinity. At this stage it is induced by gradually decreasing the salinity to 5 ppt. This suggests that the cotransporter is part of the osmoregulatory mechanism and that it can be activated as needed. Zoea stages are being cultured to determine when the cotransporter product is no longer constitutively present (as in embryos) and when it becomes inducible by hyposaline conditions (as in megalopae).