CA2-TRISPHOSPHATE IS BLOCKED BY THE K+-CHANNEL BLOCKERS APAMIN AND TETRAPENTYLAMMONIUM ION, AND A MONOCLONAL-ANTIBODY, TO A 63 KDA MEMBRANE-PROTEIN - REVERSAL OF BLOCKADE BY K+ IONOPHORES NIGERICIN AND VALINOMYCIN AND PURIFICATION OF THE 63 KDA ANTIBODY-BINDING PROTEIN( RELEASE BY INOSITOL 1,4,5)

Ins(1,4,5)P-3-induced Ca2+ release from platelet membrane vesicles was blocked by apamin, a selective inhibitor of low-conductance Ca2+-acti vated K+ channels, and by tetrapentylammonium ion, and was weakly inhi bited by tetraethylammonium ion. Other K+-channel blockers, i.e. chary bdotoxin, 4-aminopyridine and glybenclamide, were ineffective. A monoc lonal antibody (mAb 213-21) obtained by immunizing mice with the InsP( 3)-sensitive membrane fraction from platelets also blocked Ca2+ releas e by InsP(3) from membrane vesicles obtained from platelets, cerebellu m, aortic smooth muscle, HEL cells and sea-urchin eggs. ATP-dependent Ca2+ uptake and binding of[H-3]InsP(3) to platelet membranes was unaff ected by either K+-channel blockers or mAb 213-21. Blockade of Ca2+ re lease by apamin, tetrapentylammonium and mAb 213-21 was not affected b y the Na+/H+ carrier monensin or the protonophore carbonyl cyanide p-t rifluoromethoxyphenylhydrazone (FCCP), but could be completely reverse d by the K+/H+ ionophore nigericin and partially reversed by the K+ ca rrier valinomycin. The antibody-binding protein (ABP) solubilized from platelets, cerebellum, and smooth:: muscle chromatographed identicall y on gel filtration, anion exchange and heparin-TSK h.p.l.c. ABP was p urified to apparent homogeneity from platelets and aortic smooth muscl e as a 63 kDa protein by immunoaffinity chromatography on mAb 213-21-a garose. These results suggest that optimal Ca2+ release by InsP(3) fro m platelet membrane vesicles may require the tandem function of a K+ c hannel. A counterflow of K+ ions could prevent the build-up of a membr ane potential (inside negative) that would tend to oppose Ca2+ release . The 63 kDa protein may function to regulate K+ permeability that is coupled to the Ca2+ efflux via the InsP(3) receptor.