Guanine Nucleotides Induce Ca2+-independent Insulin Secretion

The role of guanine nucleotides in insulin secretion was investigated in electrically permeabilized RINm5F cells. Ca2+ stimulated insulin release (EGO 2 p~ Ca2+). The GTP stable analog, GTPrS, elicited insulin secretion at vanishingly low Ca2+ concentrations (<lo-” M), slightly potentiated the response to intermediate Ca2+ levels, but exerted less than additive effects at maximal Ca2+ concentrations. The GDP analog, GDPfiS, inhibited both GTPrSand Ca2+-stimulated secretion. The action of GTPrS was not mediated by CAMP, as the latter only enhanced Caz+-induced secretion. In contrast, 12-O-tetradecanoylphorbol-13acetate, an activator of protein kinase C, promoted insulin release at nonstimulatory Ca2+ levels as well as potentiating the Caz+ response. GTP analogs stimulated hydrolysis of phosphatidylinositol 4,5-bisphosphate (PtdInsPz), as assessed by inositol phosphate generation. However, this could not fully explain guanine nucleotide-induced secretion because: 1) GTPTS-stimulated PtdInsPz breakdown was totally dependent on Ca2+ and abolished at Caz+ below lo-” M; 2) at these Ca2+ levels, activators of protein kinase C were weak or ineffective secretagogues; 3) the GTP analog Gpp(NH)p was much less effective than GTPrS in activating PtdInsPz hydrolysis, while fully mimicking the effect on Ca2+-independent secretion. Both GTPrSinduced PtdInsP2 hydrolysis and insulin release were insensitive to pertussis toxin and cholera toxin. The findings point to a guanine nucleotide-regulated site in the activation of insulin secretion different from the known transmembrane signalling systems.