A procedure was developed for the preparation of inside-out vesicles from plasma membranes isolated from the branchial epithelium of the green shore crab Carcinus maenas (L.). Procedures normally applied to fish branchial epithelium required the introduction of an additional hypotonic shock to obtain a preparation containing 22% inside-out vesicles, 33% right-side-out vesicles and 45% leaky membrane fragments. In such membrane preparations, the first direct evidence for uphill (against a [Ca2+] gradient) ATP-dependent and Na(+)-gradient-dependent Ca2+ transport in crustacean gills was found. The affinity for Ca2+ of the ATP-driven Ca2+ transporter was 149 nmol l-1 and that of the Na+/Ca2+ exchanger was 1.78 mumol l-1; the Vmax values were 1.73 and 9.88 nmol min-1 mg-1 protein respectively. The relative importance of these carriers for Ca2+ transport in the branchial epithelium of the crab is evaluated on the basis of their calcium kinetics.

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