Multiple conductance states of single Ca2+-activated Cl− channels in rabbit pulmonary artery smooth muscle cells

摘要

Ca²⁺-activated Cl⁻ channels contribute to agonist-evoked contraction and spontaneous activity in some smooth muscle preparations. Patch pipette techniques were used to study the properties of single Ca²⁺-activated Cl⁻ channels in freshly dispersed rabbit pulmonary artery myocytes. In the cell-attached recording mode, two conductance states of 3.5 and 1.8 pS were recorded either spontaneously or in response to increasing [Ca²⁺]i. With inside-out patches, the 3.5 pS channel current predominated at 50 nm [Ca²⁺]i, but at 500 nm [Ca²⁺]i most channels opened to the 1.8 pS level and an additional 1.2 pS channel conductance was resolved. At 1 µm [Ca²⁺]i all of the Cl⁻ channels opened either to the 1.8 pS or 1.2 pS level. In 0 [Ca²⁺]i, no channel activity was observed at −100 mV to +100 mV, but with 10–250 nm [Ca²⁺]i the total single channel open probability (NPo) increased with depolarisation. This voltage dependence was not seen at higher values of [Ca²⁺]i. The plot of NPovs.[Ca²⁺]i yielded Ca²⁺ affinity constants of 8 and 250 nm and Hill slopes of 1.3 and 2.3 at +100 and −100 mV, respectively. The distribution of open times was fitted by two exponentials of about 5 and 30 ms, which were neither voltage nor Ca²⁺ dependent. Replacement of external Cl⁻ by I⁻ shifted the reversal potential by about −30 mV and lengthened the longer of the two mean open times without significant effects on other kinetic parameters. Based on these data, a model for the activation of Ca²⁺-activated Cl⁻ channels is proposed.