ATP-dependent calcium uptake into basolateral vesicles from transporting epithelia of intermolt crayfish.

Abstract

ATP-dependent Ca2+ uptake was determined into inside-out basolateral membrane vesicles (BLMV) from intermolt crayfish (Procambarus clarkii) Ca2+-transporting epithelia: gill, hepatopancreas (liver), and antennal gland (kidney). Extravesicular (EV) ATP (5 mM) increased 45Ca2+ uptake (free Ca2+ 5 microM) by fivefold but was abolished by pretreatment with either vanadate or the ionophore A-23187. Addition of A-23187 to Ca2+-loaded vesicles produced 70% efflux. The saturable carrier exhibited a Km for Ca2+ of 0.11-0.27 microM and maximal influx of 20-123 pmol. mg-1. min-1. The Km for ATP was 0.01-0.04 mM. The temperature coefficient ranged from 1.43 to 2.06. EGTA treatment of hepatopancreas and antennal gland vesicles decreased 45Ca2+ uptake by 50-90%; uptake was restorable by calmodulin. However, in gill, 45Ca2+ uptake was unaffected by EGTA treatment and calmodulin decreased uptake in both EGTA-treated and untreated vesicles. Addition of EV Na+ (5 mM) increased ATP-dependent Ca2+ uptake into hepatopancreas and antennal gland BLMV by 60%; in hepatopancreas BLMV, this increase was inhibitable by ouabain. However, ATP-dependent Ca2+ uptake in gill vesicles was Na+ independent. The relative role of each epithelium in whole animal Ca2+ homeostasis has been interpreted based on in vitro characteristics.