Carbonic anhydrase in branchial tissues of osmoregulating shore crabs, Carcinus maenas

Abstract

AbstractActivities of carbonic anhydrase (CA) were determined in various organs of shore crabs, Carcinus maenas, acclimated to salinities of 10, 20, 30, 40, and 50‰. In addition, some properties of the branchial enzyme (effects of detergents, storage conditions, and specific inhibitors) were tested. Subcellular distribution of gill CA was analyzed by means of centrifugation procedures. In hemolymph no CA activity could be detected. In all other organs except gills activities were low and insensitive to salinity change. High activities of CA were found in the gills. Activities in posterior gills were—in comparison with anterior gills—more than twofold increased. Acclimation of crabs to reduced ambient salinities resulted in significant increases in CA only in posterior gills. In the gill enzyme specific activities and stability during storage could be increased by means of the detergent Triton X‐100 used at 1% (v/v) in the homogenizing medium. Sulfonamide inhibitors reduced CA activities effectively. Inhibition constants Ki of the gill enzyme were 7.0 × 10−9 M for acetazolamide and 3.1 × 10−8 M for methazolamide. Measurements of CA activities in supernatants and pellets obtained by differential centrifugation and in fractions obtained by centrifugation of 7,500g supernatants in a sucrose density gradient indicated that the major portion of CA activity in the gill (ca. 75%) was bound to cellular particulates including plasma membranes and ca. 25% occurred in a soluble form in the cytoplasm. Both the bound and the soluble forms of CA were increased in crabs acclimated to low salinity, suggesting that CA in crab gills, besides its known role in CO2 excretion and acid‐base balance, functions in osmoregulatory processes.