Regulatory Role of the GTP‐Binding Protein, Go, in the Mechanism of Exocytosis in Adrenal Chromaffin Cells

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Abstract
To elucidate the possible involvement of GTP‐binding proteins (G proteins) in the mechanism of exocytosis, we studied effects of pertussis toxin (PTX), guano‐sine 5′‐O‐(3‐thiotriphosphate) (GTP‐γ‐S), and antibodies against the G proteins (Gi and Go) on the secretory function of bovine adrenal chromaffin cells. Pretreatment of chromaffin cells with PTX resulted in an increase in acetylcholine‐evoked catecholamine release. High K+‐, histamine‐, or γ‐aminobutyric acid‐evoked catecholamine release was also potentiated by PTX pretreatment. The concentration of extracellular Ca2+ required for maximal release by 10−4M acetylcholine was decreased significantly in PTX‐treated cells. In digitonin‐permeabilized cells, PTX pretreatment resulted in a decrease of the half‐maximal concentration (Km) of Ca2+ required for exocytosis with no significant change in the maximal stimulation (Vmax). Exposure of permeabilized cells to GTP‐γ‐S (a nonhydrolyzable GTP analogue) inhibited Ca2+‐dependent exocytosis by reducing the affinity for Ca2+. The effects of PTX pretreatment were mimicked by treatment of permeabilized cells with polyclonal antibodies selective for the α subunit of the PTX‐sensitive G protein, Go. Treatment with similar antibodies against the α subunit of Gi had no effect. These findings suggest that Go directly controls the Ca2+‐triggered process in the machinery of exocytosis by lowering the affinity of the unknown target for Ca2+.