Calcium‐dependent chloride currents in isolated cells from rat lacrimal glands.

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Abstract
1. Isolated cells from rat lacrimal glands were studied with the tight-seal whole-cell recording technique. Cells were dialysed with K-free solutions containing a high concentration of Ca2+ buffer in order to record Ca-dependent Cl- currents at a Ca2+ level fixed between 0.1 and 10 .mu.M. 2. HEDTA was preferable to EGTA as a Ca2+ buffer, even under conditions of equivalent equilibrium buffering power. 3. After replacement of all internal K+ with Na+, the cells displayed a small conductance component which could be abolished by removal of external K+ or by external application of 2 mM-tetraethylammonium. It is suggested that this conductance is due to Ca-dependent K+ channels. 4. The main part of the cell current was Cl selective. The Cl- conductance was negligible at 0.5 .mu.M-Ca2+, and fully activated at 2 .mu.M-Ca2+. The dose-response curve relating Cl- currents to the internal Ca2+ concentration, [Ca]i, was steeper than predicted by a simple binding isotherm reaction. 5. Relaxations observed in response to voltage jumps could, in most cases, be fitted with single exponentials. At [Ca]i 0.5 .mu.M, the curve relating the relaxation time constant, .tau., to the membrane potential, displayed a maximum near +20 mV and 250 ms. At hyperpolarized potentials, .tau. varied by an e-fold factor in 130 mV. At [Ca]i 1 .mu.M, .tau. decreased from 100 ms at -120 mV to 60 ms at +60 mV. 6. Relaxation analysis gave an estimate of the variation of the channel open state probability, po, with potential. At [Ca]i 0.5 .mu.M, po varied by an e-fold factor in 50-70 mV at hyperpolarized potentials, and saturated above +60 mV. At [Ca]i 1 .mu.M, po varied e-fold in 100 to 110 mV at hyperpolarized potentials, and saturated near +20 mV. 7. External Cl- was substituted with various anions. From reversal potential measurements, the following permeablility sequence was obtained; I- > NO3- > Br- > Cl- > F- > isethionate, methanesulphonate > glutamate. The corresponding normalized permeability coefficients were 2.7, 2.4, 1.6, 1, 0.2, 0.1, 0.1, 0.05. 8. Replacement of external Cl- with Br-, isethionate, methanesulphonate or glutamate did not alter current kinetics as obtained during or after a depolarizing voltage jump. However, replacement of Cl- with NO3- or I- led to complex alterations of the current kinetics. In addition, external application of I- led to long-term changes of the currents which were only partially reversible. Both NO3- and I- effects were enhanced under conditions that drove these ions into the cells, suggesting that they alter channel properties by binding to a site located on the intracellular side of the membrane. The poor reversibility of the I- effects may be related to the formation of lipid soluble polymeric forms of the halide. 9. Comparison of current kinetics at low and high internal Cl- suggests that internal Cl- also induces channel activation, but less efficiently than NO3- or I-.