Noradrenaline inhibits exocytosis via the G protein βγ subunit and refilling of the readily releasable granule pool via the αi1/2 subunit

The molecular mechanisms responsible for the 'distal' effect by which noradrenaline (NA) blocks exocytosis in the beta-cell were examined by whole-cell and cell-attached patch clamp capacitance measurements in INS 832/13 beta-cells. NA inhibited Ca2+-evoked exocytosis by reducing the number of exocytotic events, without modifying vesicle size. Fusion pore properties also were unaffected. NA-induced inhibition of exocytosis was abolished by a high level of Ca2+ influx, by intracellular application of antibodies against the G protein subunit G beta and was mimicked by the myristoylated beta gamma-binding/activating peptide mSIRK. NA-induced inhibition was also abolished by treatment with BoNT/A, which cleaves the C-terminal nine amino acids of SNAP-25, and also by a SNAP-25 C-terminal-blocking peptide containing the BoNT/A cleavage site. These data indicate that inhibition of exocytosis by NA is downstream of increased [Ca2+](i) and is mediated by an interaction between G beta gamma and the C-terminus of SNAP-25, as is the case for inhibition of neurotransmitter release. Remarkably, in the course of this work, a novel effect of NA was discovered. NA induced a marked retardation of the rate of refilling of the readily releasable pool (RRP) of secretory granules. This retardation was specifically abolished by a G alpha(i1/2) blocking peptide demonstrating that the effect is mediated via activation of G alpha(i1) and/or G alpha(i2).