Endothelin-1 and Vasopressin Activate Ca2+-permeable Non-selective Cation Channels in Aortic Smooth Muscle Cells: Mechanism of Receptor-mediated Ca2+Influx

The effects of vasopressin and endothelin-1 on cultured aortic smooth muscle cell lines (A7r5) were investigated by measurements of intracellular calcium [Ca2+]iand the patch-clamp techniques. Vasopressin and endothelin-1 (100 n ) evoked an initial peak followed by a smaller sustained rise of [Ca2+]iin the presence of extracellular calcium [Ca2+]o. In the absence of [Ca2+]o, only the initial peak of [Ca2+]iwas observed. Therefore, the initial peak of [Ca2+]iwas mainly due to calcium release from the storage sites, whereas the later sustained rise of [Ca2+]iwas due to the calcium entry from outside. The sustained rise of [Ca2+]iwas unaffected by nifedipine (10μ ) significantly, but was completely abolished by La3+(1 m ). Under current clamp conditions with K+-internal solution, vasopressin and endothelin-1 (100 n ) produced hyperpolarization, then followed by depolarization. Under voltage clamp conditions at a holding potential of −40 mV, both vasopressin and endothelin-1 first activated the outward current, then followed by a long-lasting inward current with a high noise level. The first outward current was abolished by charybdotoxin (100 n ), Cs+in the patch pipette and high EGTA (10 m ) in the pipette, suggesting that it was a Ca2+-sensitive K+current (IK.Ca). The inward current was still elicited with the patch pipette containing Cs+-internal solution, and reversed at about 0 mV. The reversal potential was not significantly altered by the replacement of [Cl−]ior [Cl−]o, proposing that the inward current is a cation selective channel (IN.S.). The inward current was also observed even when extracellular cations are Ca2+. La3+(1 m ), Cd2+(1 m ) completely abolished the vasopressin-induced IN.S., however, nifedipine (10μ ) failed to inhibit it significantly. Single channel activities were recorded in the cell-attached configurations when vasopressin or endothelin-1 was applied to the bathing solution. The unitary conductance of the channels was approximately 20 pS with 140 m Na+, Cs+, or K+in the pipette, but was 15 pS with 110 m Ca2+in the pipette. Permeabilities sequence calculated from the reversal potentials was Na+ Cs+ K+>Ca+. These results provide evidence that calcium entry and membrane depolarization elicited by vasopressin or endothelin-1 are mediated by a receptor-mediated Ca2+-permeable non-selective cation channel in aortic smooth muscle cells.