Relative Importance of Extracellular and Intracellular Ca2+ for Acetylcholine Stimulation of Insulin Release in Mouse Islets

Abstract

Mouse islets were used to study whether mobilization of intracellular Ca2+ is sufficient to account for acetylcholine (ACh) amplification of glucose-induced insulin release. In the presence of 15 mM glucose, the acceleration of 45Ca efflux and insulin release by 1-100 microM ACh increased with the concentration of extracellular Ca2+ (0.25-2.5 mM). Low concentrations of the Ca2+-channel blockers D 600 (1 microM) or nifedipine (0.1 microM) partially inhibited glucose-induced insulin release and its amplification by ACh. At higher concentrations, D 600 (25 microM) or nifedipine (2 microM) practically abolished the ionic and secretory effects of 1 microM ACh. However, 100 microM ACh still caused a fast, large, but transient acceleration of 45Ca efflux, accompanied by a small, short-lived release of insulin. Similar results were obtained in a Ca2+-free medium, indicating that this peak of 45Ca efflux reflects Ca2+ mobilization. Addition of nifedipine or omission of Ca2+ during ACh stimulation rapidly and strongly inhibited 45Ca efflux and insulin release. Both glucose and ACh-induced 45Ca uptake were inhibited by D 600. Only high concentrations of ACh (100 microM) mobilize enough cellular Ca2+ to trigger a small and transient insulin release when Ca2+ influx is prevented or impossible. A continuous influx of Ca2+ is necessary for low ACh concentrations to increase release and for high concentrations to have a sustained effect. The amplification of release by the neurotransmitter results from a slight enhancement of Ca2+ influx associated with a marked increase in the effectiveness of incoming Ca2+ on the releasing machinery.

Two distinct modes of Ca2+ signalling by ACh in rat pancreatic beta‐cells: concentration, glucose dependence and Ca2+ origin.

“…This action of ACh in fl-cells may be mediated by IP3 in view of the following reports: (1) muscarinic activation stimulates formation of IP3 in mouse islets (Nilsson et al 1987;Gao et al 1994) and RINm5F cells (Wollheim & Biden, 1986), (2) IP3 mobilizes Ca2P in permeabilized mouse islet cells (Nilsson et al 1987;Hellman et al 1992), RINm5F cells (Prentki & Matschinsky, 1987) and mouse islet cells in which caged IP3 is injected and liberated (Ammiilii et al 1993), and (3) an IP3 receptor subtype is expressed in rat islets and RINm5F cells (Blondel, Takeda, Janssen, Seino & Bell, 1993 (Table 1). This is in accord with the observation that ACh stimulation of insulin release requires that the f-cell membrane be sufficiently depolarized to reach the threshold potential at which Ca2+ channels are activated (Hermans et al 1987 (Hermans & Henquin, 1989), and that only a transient increase in…”

Ghrelin Attenuates cAMP-PKA Signaling to Evoke Insulinostatic Cascade in Islet β-Cells

“…In an apparent discrepancy, ghrelin is reported to suppress carbachol-induced insulin secretion at 5.5 mmol/L glucose, a concentration over the threshold for insulin release, in the perfused rat pancreas (36). ACh stimulates β-cells exclusively via muscarinic M 3 receptor (37,38) and induces [Ca 2+ ] i increases and insulin secretion in a glucose-dependent manner (39,40). Collectively, it is likely that ghrelin suppresses M 3 receptor-mediated glucose-dependent insulin release by interfering with glucose signaling but not with muscarinic receptor signaling.…”

Protein kinase C isoform specificity of cholinergic potentiation of glucose-induced pulsatile 5-HT/ insulin release from mouse pancreatic islets

“…The overall cholinergic response resembles that observed in terms of electrical activity recorded from whole mouse islets, where acetylcholine evoked a membrane depolarization with an initial phase of continuous spiking followed by bursts of electrical activity with a lower duration and higher frequency (Hermans et al, 1987;Santos and Rojas, 1989). The initial response is likely to reflect Ca 2+ release from internal stores via the PLC/ 1,4,5-IP 3 system (Gilon and Henquin, 2001;Hermans and Henquin, 1989). Significantly, this response remained unaffected by chronic TMX pretreatment (a procedure known to cause down-regulation of cPKC isoforms (Yanagita et al, 2000) and was not present when the islets were subjected to acute PKC stimulation with TMX.…”