Specific binding of inositol hexakisphosphate (phytic acid) to adrenal chromaffin cell membranes and effects on calcium-dependent catecholamine release

Abstract

In a membrane preparation of cultured bovine adrenal chromaffin cells, [ 3H]inositol hexakisphosphate ([ 3H]InsP 6) was shown to bind specifically with a k d of 90 nM and a B max of 700 fmol/mg protein. The Hill coefficient was not significantly different from unity. The association of [ 3H]InsP 6 was slow, with equilibrium binding being reached within 10 min. The dissociation of [ 3H]InsP 6 showed monophasic kinetics. In [ 3H]InsP 6 competition binding experiments, we found that reduction in the number of phosphorylated sites in inositol resulted in a gradual loss of binding potency. In intact bovine adrenal chromaffin cells, InsP 6 elicited a concentration-dependent facilitation of 45C 2+ influx along with the release of the catecholamines, epinephrine and norepinephrine. The latter responses were slower and longer-lasting than responses to depolarizing stimuli, such as nicotine and high K +. The catecholamine release required the presence of extracellular Ca 2+. In good agreement with the binding studies, lower inositol phosphates displayed reduced secretagogue potency. In conclusion, in bovine adrenal chromaffin cells, InsP 6 appeared to bind to specific sites and elicited Ca 2+ influx and catecholamine release.