The effects of the chaotrophic anion perchlorate (ClO4-) on glucose-induced electrical activity, exocytosis and ion channel activity in mouse pancreatic B-cells were investigated by patch-clamp recordings and capacitance measurements. ClO4- stimulated glucose-induced electrical activity and increased the action potential frequency by 70% whilst not affecting the membrane potential when applied in the presence of a subthreshold concentration of the sugar. ClO4- did not influence ATP-dependent K (KATP) channel activity and voltage-gated delayed K+ current. Similarly, ClO4- had no effect on Ca2+-dependent exocytosis. The stimulation of electrical activity and insulin secretion was instead attributable to an enhancement of the whole-cell Ca2+ current. This effect was particularly pronounced at voltages around the threshold for action potential initiation and a doubling of the current amplitude was observed at -30 mV. This was due to a 7-mV shift in the gating of the Ca2+ current towards negative voltages. The action of ClO4- was more pronounced when added in the presence of 0.1 mM BAY K8644, whereas no stimulation was observed when applied at a maximal concentration of the agonist (1 mM). Single-channel recordings revealed that the effect of ClO4- on whole-cell currents was principally due to a 60% increase in the mean duration of the long openings and the number of active channels. We propose that ClO4- stimulates insulin secretion and electrical activity by exerting a BAY K8644-like action on Ca2+ channel gating.

Non-inbred ob/ob mice were used to study the dynamics of cytoplasmic Ca2+ concentration ([Ca2+]i) in isolated pancreatic beta-cells using microfluorimetry with fura 2/AM as probe, and the dynamics of insulin secretion in isolated pancreatic islets. D-Glucose (20 mM) caused a transient peak increase in [CA2+]i which changed to either an oscillating or a flat, elevated phase. The lag-time before the first peak increase in [Ca2+]i was markedly shortened by 12 mM ClO4- and the glucose-stimulated level of [Ca2+]i after the first peak was clearly elevated by the anion. ClO4- did not change the basal [Ca2+]i at 3 mM glucose. Extracellular Ca2+ deficiency abolished the effect of high glucose and ClO4- on [Ca2+]i. This suggests that ClO4- acts as an amplifier of transmembrane Ca2+ inflow. The L-type Ca2+ channel agonist, Bay K 8644 (0.01-1.0 microM), strictly reproduced all the effects of perchlorate on the glucose-stimulated beta-cell [Ca2+]i. Both phases of insulin release (20 mM glucose) were markedly enhanced by ClO4- (12 mM) or Bay K 8644 (1.0 microM). The lag-time for glucose-stimulated insulin release was shortened by both agents. Taken together, these data strengthen the idea that perchlorate amplifies the glucose-stimulation of [Ca2+]i and insulin release by directly modifying the function of the L-type Ca2+ channel. This effect can induce both a more prompt onset of and an amplified level of beta-cell secretory activity.

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In order to elucidate the mechanisms responsible for the stimulatory effect of perchlorate (ClO4-) on insulin secretion, we have investigated the interaction between this chaotropic anion and the organic calcium antagonist nifedipine. This drug, known as a blocker of L-type calcium channels, was chosen as a tool to test the idea that ClO4- acts on insulin secretion by stimulating the gating of voltage-controlled Ca2+ channels. ClO4- amplified the stimulatory effect of D-glucose on insulin release from perfused pancreas (first and second phases) as well as from isolated islets incubated in static incubations for 60 min. This indicates that ClO4- amplifies physiologically regulated insulin secretion. Nifedipine reduced D-glucose-induced (20 mM) insulin release in a dose-dependent manner with half-maximum effect at about 0.8 microM and apparent maximum effect at 5 microM nifedipine. In the presence of 20 mM D-glucose, the inhibitory effects of 0.5, 1 or 5 microM nifedipine were only slightly, if at all, counteracted by perchlorate. When 12 mM ClO4- and 20 mM D-glucose were combined, calculation of the specific effect of ClO4- revealed that nifedipine produced almost maximum inhibition already at 0.05 microM. Thus, the perchlorate-induced amplification of D-glucose-stimulated insulin release shows higher sensitivity to nifedipine than the D-glucose-effect as such. This supports the hypothesis that perchlorate primarily affects the voltage-sensitive L-type calcium channel in the beta-cell.

A cell-specific response may be induced at several steps of beta-cell stimulus-secretion coupling. Mitochondrial metabolism generates a cell-specific response in normal beta cells but not in db/db and ob/ob mouse beta cells.