Properties and calcium‐dependent inactivation of calcium currents in cultured mouse pancreatic B‐cells.

Abstract

SUMMARY1. Ca2+ currents were recorded using the whole-cell mode of the patch-clamp technique from mouse pancreatic B-cells kept in culture for 1-4 days. B-cells were identified in the cell-attached mode by their response to a change in the glucose concentration from 3 to 15 or 20 mm or by their inward currents.2. Only one component of Ca2+ current was observed in these cells, which activated at potentials > -50 mV and was blocked by nitrendipine (5/M), and increased in amplitude by CGP 28392 (5 /tM).3. During maintained depolarizations the Ca2+ current inactivated considerably but not completely. Inactivation was most marked at potentials where the Ca2+ currents were large, but in general was slower for currents at potentials > 0 mV than at more negative potentials.4. Two-pulse experiments showed that the inactivation curve for the Ca2+ current was U-shaped, returning to unity at potentials approaching the Ca2+ equilibrium potential. Measurements of Ca2+ entry showed that inactivation was dependent on the amount of Ca2+ entering during the pre-pulse, independent of the pre-pulse potential.5. Ca2+ currents were not appreciably slowed when BAPTA, a faster buffer of Ca2+, replaced EGTA in the pipette solution.6. Replacement of Ca2+ in the external solution by Ba2+ increased the amplitude of the inward current and largely abolished inactivation. Large inward currents through Ca2+ channels were observed in the absence of divalent cations in the external solution (+ EGTA), which were presumably carried by Na+. These currents did not inactivate during 150 ms depolarizations, but were increased in amplitude by CGP 28392 (5 /M) and blocked by D600 (30 FM).7. The observations suggest that normal mouse pancreatic B-cells have only one type of Ca2+ channel which is dihydropyridine sensitive and inactivates by a mechanism which is almost purely Ca2+ dependent. Inactivation of the Ca2+ current will probably be important in the control of Ca2+ entry during glucose-induced electrical activity.

Clonidine inhibits ATP‐sensitive K+channels in mouse pancreatic β‐cells

“…(Horn & Marty, 1988;Korn & Horn, 1989;Falke et al, 1989;Smith et , 1989). With this method calcium currents were much more stable than with the conventional whole cell technique (Plant, 1988), but did show a slow run-down in some cells. They were decreased by 100pM Cd2`and increased in amplitude by the dihydropyridine, CGP 28392, at a concentration of 5pMm (data not shown).…”

“…After isolation of the islets, islet cells were dispersed and cultured for 1-2 days as previously described (Plant, 1988 (Garrino et al, 1989;Plant & Henquin, 1990 (Horn & Marty, 1988 (Smith et al, 1989). The tip of the pipette was first filled by dipping the pipette briefly in this solution, then back-filled with the same solution supplemented with 100 or 150 pg ml-1 nystatin in 0.2% dimethylsulphoxide (DMSO).…”

Linoleic acid induces Ca2+-induced inactivation of voltage-dependent Ca2+ currents in rat pancreatic β-cells

“…Some reports suggest that activation of VDCC may be responsible for the FFA-induced increase in [Ca 2C ] i in b-cells (Remizov et al 2003, Fujiwara et al 2005. Increases in [Ca 2C ] i have an inhibitory effect on VDCC currents in many cell types including pancreatic b-cells, commonly known as Ca 2C -induced VDCC inactivation (Plant 1988, Findlay 2004, Yang & Berggren 2005. To further study the action of FFAs on insulin secretion, we investigated the effects of linoleic acid on VDCC in b-cells.…”

A single non-L-, non-N-type Ca2+ channel in rat insulin-secreting RINm5F cells

“…The channel inactivates little and very slowly irrespective of whether it is recorded in the cell-attached or outside-out configuration, suggesting a marked insensitivity to intracellular Ca 2+ buffers and a marginal role of Ca 2+ in the regulation of the channel inactivation gate [20,25]. In contrast, L-type channels of insulin-secreting cells exhibit marked Ca 2+ -dependent inactivation properties [20,22,25].…”