HYDROLYSIS-RESISTANT GTP ANALOGS STIMULATE CATECHOLAMINE RELEASE FROM DIGITONIN-PERMEABILIZED PC12 CELLS

Abstract: The effect of the hydrolysis‐resistant GTP analogs, guanosine 5′‐O‐(3‐thiotriphosphate) (GTPγS) and guanylyl imidodiphosphate (GMPPNP), on norepinephrine (NE) secretion from digitonin‐permeabilized rat pheochromocytoma cells, PC12, was examined. Although secretion in the presence of saturating Ca2+ (10 μM) was not affected by GTP7S or GMPPNP, secretion in the absence of Ca2+ was stimulated by these GTP analogs. Secretion induced by saturating concentrations of GTPγS or GMPPNP was approximately 80% of that induced by 10 μM Ca2+. Half‐maximum stimulation was induced by 30 μM GTPγS or GMPPNP. Both Ca2+‐stimulated and GTPγS‐stimulated secretion were ATP dependent and inhibited by N‐ethylmaleimide. The GTPγS‐stimulated secretion of NE from permeabilized PC12 cells does not appear to result from either the release of Ca2+ or the activation of protein kinase C. Activation of protein kinase C by pretreatment of intact cells with 12‐O‐tetradecanoyl‐phorbol 13‐acetate caused a 50% increase in both Ca2+‐stimulated and GTP7S‐stimulated secretion. Cholera and pertussis toxins did not affect Ca2+‐stimulated or GTPγS‐stim‐ulated NE secretion. Guanosine 5′‐O‐(2‐thiodiphosphate) (GDPβS) and GTP inhibited GTPγS‐stimulated secretion but not Ca2+‐stimulated secretion. The inability of GDPβS to inhibit Ca2+‐stimulated secretion indicates that the process affected by GTPγS is not an essential step in the Ca2+‐stimulated pathway. Copyright © 1990, Wiley Blackwell. All rights reserved