Regulation of the cerebellar inositol 1,4,5-trisphosphate receptor by univalent cations

In the present Study we investigated the effects of K+ and other univalent cations on [H-3]InsP(3) {[H-3]Ins(1,4,5)P-3} binding to sheep cerebellar microsomes. In equilibrium binding experiments performed over 4 s at pH 7.1 and 20 degreesC, the addition of K+ to the binding medium decreased the affinity and increased the total number Of bindin g sites for InsP(3) in a close-dependent manner. At low InsP(3) concentration (0.5 nM) these effects resulted in a biphasic dose-response curve, with maximal binding at about 75 mM K+. In contrast. the dose-response curve calculated for InsP(3) at the physiological concentration of 5 muM, was linear up to 200 mM K+. Univalent inorganic cations stimulated [H-3]InsP(3) binding to various extents, with the following descending order of efficiency at 75 mM: Cs+ approximate to Rb+ approximate to K+ > Na+ > Li+. The effect of K+ on InsP(3)R affinity was rapidly reversed upon cation removal. We were therefore also able to demonstrate that K+ increased B-max (maximal specific binding) by pre-treating microsomes with K+ before measuring [H-3]InsP(3) binding in the absence of that cation. The increase in B-max was reversible, but this reversal occurred less rapidly than the change in affinity. These results are consistent with a process by which K+ reversibly converted very low-affinity sites into sites with higher affinity, making them detectable in competitive binding experiments. They suggest that interconversion between these two affinity states constitutes the basis of a K+-controlled regulatory mechanism for cerebellar InsP3R.