Cholinergic-induced [³H] noradrenaline release in rat brain cortical slices is mediated via a pertussis toxin sensitive GTP binding protein and involves activation of protein kinase C

The involvement of a GTP-binding protein (G-protein) in the process of neurotransmitter release was examined using pertussis toxin and cholerra toxin. Cholinergic agonists are shown to mediate [³H]noradrenaline release in rat brain slices via a pertussis toxin (1.2 μg/ml) sensitive, and cholera toxin (0.5 μg/ml) insenstive G. protein. An indication for the involvement of a G-protein and phospholipase C activation in the release process was implied from the inhibitory effect of neomycin on K⁺-, veratridine, and carbachol-induced-norepinephrine release. Depolarizing agents mediate a neomycin-senstive release, which is not affected either by pertussis toxin or cholera toxin, suggesting a different mode of phospholipase C activation, unlike carbachol-induced release, which is both neomycin and pertussis toxin senstive. Similarly, a hormone-sensitive carrier activated by phenylephrine not via α₁-adrenergic receptors, mediates a non-exocytosis efflux which is not affected by neomycin and is shown to be pertussis toxin-insensitive. The inhibitory action of protein kinase C inhibitors polymyxin B, K252_a and H-7 [(1-(1-(5-isoquinolinesulphonyl)-2-methyl-piperazine] on release, strongly suggests its participation in the process. Polymyxin B, a relatively selective protein kinase C inhibitor, inhibted carbachol-induced release (ic₅₀ = 0.53 μM) as well as the K⁺ and the veratridine induced [³H] noradrenaline release. K252_a, an inhibitor of various protein kinases at the ATP site, and H-7, another protein kinase C inhibitor, inhibited carbachol-induced noradrenaline released with ic = 35 nM and 3 μM respectively. Consistent with its inability to activate phospholipase C, phenylephrine-induced noradrenaline efflux was unaffected by polymyxin B (> 70 μM). The results offer more supportive evidence for a major role played by the dual messengers inositol triphosphate and diacylglycerol (IP₃/DG) in the mechanisms of neuronal release.