Rapid inactivation of depletion-activated calcium current (ICRAC) due to local calcium feedback.

Abstract

Rapid inactivation of Ca 2+ release-activated Ca 2+ (CRAG) channels was studied in Jurkat leukemic T lymphocytes using whole-ceU patch clamp recording and [Ca2+]i measurement techniques. In the presence of 22 mM extracellular Ca 2 § the Ca z+ current declined with a biexponential time course (time constants of 8-30 ms and 50-150 ms) during hyperpolarizing pulses to potentials more negative than -40 inV. Several lines of evidence suggest that the fast inactivation process is Ca 2+ but not voltage dependent. First, the speed and extent of inactivation are enhanced by conditions that increase the rate of Ca 2+ entry through open channels. Second, inactivation is substantially reduced when Ba 2+ is present as the charge carrier. Third, inactivation is slowed by intracellular dialysis with BAPTA (12 raM), a rapid Ca 2+ buffer, but not by raising the cytoplasmic concentration of EGTA, a slower chelator, from 1.2 to 12 raM. Recovery from fast inactivation is complete within 200 ms after repolarization to -12 mV. Rapid inactivation is unaffected by changes in the number of open CRAC channels or global [Ca2+]i . These results demonstrate that rapid inactivation of lc~c results from the action of Ca z+ in close proximity to the intracellular mouths of individual channels, and that Ca 2+ entry through one CRAC channel does not affect neighboring channels. A simple model for Ca z+ diffusion in the presence of a mobile buffer predicts multiple Ca z+ inactivation sites situated 3-4 nm from the intracellular mouth of the pore, consistent with a location on the CRAC channel itself.

Orai1 pore residues control CRAC channel inactivation independently of calmodulin

“…Using this protocol, WT Orai1 consistently displayed CDI kinetics and extent similar to that of native CRAC channels (Fig. 3 A; Zweifach and Lewis, 1995). At the same time, we observed phenotypes for several CDI mutants that differed somewhat from those described previously, presumably due to higher STIM1/ Orai1 ratio (see Discussion).…”

“…We assume that N is constant immediately after a switch from 2 mM Ca 2+ to DVF (though it will decline subsequently due to depotentiation); hence N Ca = N Na . P o is reduced in 2 mM Ca 2+ relative to DVF because of CDI, which should reverse within milliseconds (Zweifach and Lewis, 1995) after Ca 2+ flux is terminated upon the switch to DVF. Thus, P o,Ca /P o , Na is given by the extent of CDI (from 0 to 1) that occurs in 2 mM Ca 2+ at 100 mV.…”

“…Unitary properties of Orai1 channels with charged substitutions at position Y80 CRAC channel CDI is triggered by local Ca 2+ flux through individual channels (Zweifach and Lewis, 1995). In principle, mutations that inhibit CDI could act by reducing Ca 2+ accumulation near the pore or by limiting the channel's response to a given local [Ca 2+ ].…”

Voltage Gating at the Selectivity Filter of the Ca2+ Release-activated Ca2+ Channel Induced by Mutation of the Orai1 Protein

“…4, A and B). This partial initial inactivation of the current at the time of break-in, later recovered by BAPTA perfusion, is likely because of saturation of the endogenous Ca 2ϩ buffers and CRAC inactivation by elevated cytosolic Ca 2ϩ (37). To measure the fully activated CRAC current, we perfused the cell cytosol after break-in for at least 2 min with 10 mM BAPTA to allow diffusion of BAPTA into the cell before taking any statistical measurements.…”

New factors contributing to dynamic calcium regulation in the skeletal muscle triad—a crowded place

“…Before, SOCE had been identified in T-lymphocytes (Zweifach and Lewis 1995) and endothelial tissue (Holder and Blatter 1997). In their original study, Kurebayashi and Ogawa used intact thin mouse EDL fiber bundles that they subjected to short (30 s) high K + pulses in Ca 2+ -free solution and recorded force transients and fura-2 Mn 2+ influx.…”