Muscarinic activation and calcium permeation of nonselective cation currents in airway myocytes

We examined the activation and Ca2+ permeation of nonselective cation channels in voltage-damped (nystatin), fura 2-loaded equine tracheal myocytes at 35 degrees C. Methacholine (50 mu M) induced a biphasic increase in intracellular Ca2+ concentration ([Ca2+](i)) and a biphasic inward current consisting of a large, rapidly inactivating Ca2+-activated Cl- current [I-Cl(Ca)] and a smaller, sustained nonselective cation current (I-cat). I-Cl(Ca) but not I-cat was activated by caffeine. Neither I-cat nor the sustained rise in [Ca2+](i) was blocked by nisoldipine, whereas both were rapidly blocked by Ni2+. I-cat was determined to be Ca2+ permeant, since 1) a sustained elevation of [Ca2+](i) occurred when I-cat was activated, and blockade of I-cat produced a rapid decline in [Ca2+](i); 2) increasing extracellular Ca2+ during I-cat increased [Ca2+](i); 3) 110 mM extracellular Ca2+ shifted the reversal potential of I-cat to 12 mV (Ca2+-to-Cs+ permeability ratio = 3.6); and 4) instantaneous voltage-clamp steps to negative potentials during I-cat increased the current and [Ca2+](i), whereas depolarizing steps decreased the current and [Ca2+](i). The fraction of I-cat carried by Ca2+ under physiological conditions was estimated to be 14% at -60 mV.