CALCIUM-RELEASE FROM INTRACELLULAR STORES EVOKED BY EXTRACELLULAR ATPIN A XENOPUS RENAL EPITHELIAL-CELL LINE

1. The signal transduction mechanism mediating extracellular adenosine 5'-triphosphate (ATP)-induced calcium release in a renal epithelial c ell line (A6) was investigated using the whole-cell voltage-clamp tech nique and fura-2 fluorescence measurement. 2. ATP (10 mu M) activated calcium-dependent non-selective cation channels in cells held under vo ltage clamp. 3. Guanosine 5'-O-(2-thiodiphosphate) (GDP beta X; 0.1-1. 0 mM) in the pipette inhibited the ATP-activated calcium-dependent cur rents. With guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; 0.1-1.0 mM) in the pipette, the currents were spontaneously elicited without a pplication of ATP. Pretreatment with pertussis toxin (PTX) affected ne ither the ATP-activated currents nor the increase in intracellular fre e calcium concentration ([Cd2+](i)) evoked by ATP. 4. Intracellular ap plication of neomycin or heparin inhibited the ATP-activated currents. Inositol 1,4,5-trisphosphate (IP3; 0.1-100 mu M) in the internal solu tion produced currents similar to those due to ATP activation. 5. Thes e results suggest that a PTX-insensitive guanosine 5'-triphosphate (GT P)-binding regulatory protein (G protein) is involved in extracellular ATP-induced phosphoinositide turnover and subsequent calcium release from IP3-sensitive stores, which subsequently activates the calcium-de pendent channels in AB cells.