Comparison of the effects of perchlorate and Bay K 8644 on the dynamics of cytoplasmic Ca2+ concentration and insulin secretion in mouse β-cells

Abstract

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.

Insulin Trafficking in a Glucose Responsive Engineered Human Liver Cell Line is Regulated by the Interaction of ATP-Sensitive Potassium Channels and Voltage-Gated Calcium Channels

“…However, the amount of insulin secreted in the presence of BayK8644 was lower than that released in response to 20 mM glucose alone. Putatively, this concentration of glucose may have stimulated the influx of extracellular Ca Consistent with reports that BayK8644 is known to stimulate the opening of Ca V channels in pancreatic β-cells without altering the membrane potential [44], static stimulation of Huh7ins cells with 1 µM BayK8644 plus 20 mM glucose amplified glucose-stimulated insulin release. However, BayK8644 failed to amplify glucose-stimulated insulin secretion in MIN6 cells.…”

“…BayK8644 does not change the membrane potential of resting β-cells [43]. Rather, it acts on the Ca V channel in the open state, failing to affect basal insulin secretion at non-stimulatory glucose concentrations [43], but exaggerating glucose-stimulated insulin secretion [44,45]. The addition of BayK8644 increased insulin secretion by both the Huh7ins and MIN6 cells.…”

Disorganization of cytoplasmic Ca 2+ oscillations and pulsatile insulin secretion in islets from ob / ob mice

“…The experiments were carried out with 8 to 14 month old ob/ob mice because these Swedish obese mice are traditionally used as islet donors at this advanced age [26,29,30,31]. For comparison, we also studied islets from their lean littermates at a similar age.…”

“…The Swedish ob/ob mice are traditionally used as islet donors at an age between 8 and 13 months [26,29,30,31]. The experiments were thus carried out with 23 non-inbred female obese ob/ob mice (7.5-14 months, mean age 10.3 months) and 22 lean littermates (7.5-14 months, mean age 10.1 months) from the Umea colony.…”

Cell specificity of the cytoplasmic Ca2+ response to tolbutamide is impaired in β-cells from hyperglycemic mice

“…The cells were distributed on polylysine-coated cover glasses (Sigma Chemicals, Stockholm, Sweden) placed in Petri dishes and maintained in tissue culture for 24-48 h at 11$1 mM glucose in medium (RPMI 1640; Roche) supplemented with 10% heat-inactivated fetal calf serum (Gibco, Stockholm, Sweden), 60 mg/ml garamycin (Schering Co., Kenilworth, NJ, USA), 60 mg/ml benzylpenicillin (Schering Co.), and 2 mM L-glutamine (Labkemi AB, Stockholm, Sweden). Subsequent experimental handling was performed with a Krebs-Ringer medium (KRH, Gibco) having the following composition in mM: 130 NaCl, 4$7 KCl, 1$2 KH 2 PO 4 , 1$2 MgSO 4 , and 2$56 CaCl 2 and supplemented with 1 mg/ml BSA (Sigma Chemicals) and 3 mM D-glucose (Larsson-Nyrén & Sehlin 1996). The medium was buffered with 20 mM HEPES (Roche) and NaOH to pH of 7$4 and equilibrated with ambient air.…”