Calcium-G protein interactions in the regulation of macrophage secretion

The interplay between activated G proteins and intracellular calcium ([Ca2+](i)) in the regulation of secretion was studied in the macrophage, coupling membrane capacitance with calcium-sensitive microfluorimetry. Intracellular elevation of either the nonhydrolyzable analogue of GTP, guanosine-5'-O-(3-thiotriphosphate) (GTP gammaS), or [Ca2+](i) enhanced the amplitude and shortened the time course of stimulus-induced secretion in a dose-dependent manner. Both the ionophore- and the stimulus-induced secretory response were abolished in the presence of guanosine-5'-O-(2-thiodiphosphate) (GDP betaS). The K-d of Ca2+-driven secretion was independent of GTP gammaS concentration, whereas the K-d of the GTP gammaS-driven response decreased from 63 to 31 muM in the presence of saturating concentrations of [Ca2+](i). The time course of stimulus-induced secretion was dependent upon the concentration of [Ca2+](i). The time course of GTP gammaS-driven secretion was concentration-independent at high levels of [Ca2+](i), suggesting that a calcium-dependent translocation/binding step was rate-limiting. Our data strongly support a model in which [Ca2+](i) and activated G proteins act independently of one another in the sequential regulation of macrophage secretion. [Ca2+](i) appears to play a role in the recruitment and priming of vesicles from reserve intracellular pools at a step that is upstream of G protein activation. While activated, G proteins appear to play a key role in fusion of docked vesicles. Thus, secretion can result either from activating more G proteins or from elevating [Ca2+](i) at basal levels of G protein activation.