Inositol 1,4,5-trisphosphate mediates adrenaline activation of K+ conductance in mouse peritoneal macrophages

In mouse peritoneal macrophages, alpha 1-adrenoceptor stimulation evokes a Ca(2+)-dependent K+ current [Io(Adr)][Hara et al. (1991) Pflügers Arch 419:371-379]. The roles of D-myo-inositol 1,4,5-trisphosphate (InsP3) and a GTP-binding protein (G protein) in Io(Adr) were investigated with tight-seal whole-cell recordings and fura-2 fluorescence measurements. Intracellular injection of InsP3 (5-50 microM) evoked transient outward currents [Io(InsP3)] with or without damped oscillations in membrane currents at -40 mV. Dialysis with 0.2 mM guanosine 5'-[3-thio]triphosphate (GTP[gamma S], a poorly hydrolysable GTP analogue) at -40 mV activated oscillatory outward currents or a slowly developing steady current on which such oscillations were superimposed after a delay of 10-90 s. Io(InsP3) and the GTP[gamma S]-induced current [Io(GTP[gamma S])] were accompanied by an increase in conductance. Reversal potentials of both responses closely depended on the extracellular K+ concentration. Fura-2 measurements revealed that Io(InsP3) and Io(GTP[gamma S]) result from a rise in intracellular free Ca2+ concentration ([Ca2+]i). Removal of extracellular Ca2+ did not abolish Io(InsP3) and Io(GTP[gamma S]). Both were blocked by bath-applied charybdotoxin. Intracellular D-myo-inositol 1,3,4,5-tetrakisphosphate (InsP4, 50 microM) did not evoke any responses, whereas D-myo-inositol 2,4,5-trisphosphate [InsP3(2,4,5), 20 microM] elicited an outward current at -40 mV. Io(InsP3) was completely blocked by prior dialysis with the InsP3 receptor antagonist heparin (5 mg/ml). Inclusion of guanosine 5'-[2-thiol] diphosphate (GDP[beta S], 2 mM) or heparin (5 mg/ml) together with GTP[gamma S] in the patch pipette solution completely blocked Io(GTP[gamma S]). These results indicate that intracellular injection of InsP3 or GTP[gamma S] mimic Io(Adr).(ABSTRACT TRUNCATED AT 250 WORDS)