| 17071736 |
Brueggemann LI, Moran CJ, Barakat JA, Yeh JZ, Cribbs LL, Byron KL: Vasopressin stimulates action potential firing by protein kinase C-dependent inhibition of KCNQ5 in A7r5 rat aortic smooth muscle cells. Am J Physiol Heart Circ Physiol. 2007 Mar;292(3):H1352-63. Epub 2006 Oct 27.
[Arg (8)]-vasopressin (AVP), at low concentrations (10-500 pM), stimulates oscillations in intracellular Ca (2+) concentration (Ca (2+) spikes) in A7r5 rat aortic smooth muscle cells. Our previous studies provided biochemical evidence that protein kinase C (PKC) activation and phosphorylation of voltage-sensitive K (+) (K (v)) channels are crucial steps in this process. In the present study, K (v) currents (I (Kv)) and membrane potential were measured using patch clamp techniques. Treatment of A7r5 cells with 100 pM AVP resulted in significant inhibition of I (Kv). This effect was associated with gradual membrane depolarization, increased membrane resistance, and action potential (AP) generation in the same cells. The AVP-sensitive I (Kv) was resistant to 4-aminopyridine, iberiotoxin, and glibenclamide but was fully inhibited by the selective KCNQ channel blockers linopirdine (10 microM) and XE-991 (10 microM) and enhanced by the KCNQ channel activator flupirtine (10 microM). BaCl (2) (100 microM) or linopirdine (5 microM) mimicked the effects of AVP on K (+) currents, AP generation, and Ca (2+) spiking. Expression of KCNQ5 was detected by RT-PCR in A7r5 cells and freshly isolated rat aortic smooth muscle. RNA interference directed toward KCNQ5 reduced KCNQ5 protein expression and resulted in a significant decrease in I (Kv) in A7r5 cells. I (Kv) was also inhibited in response to the PKC activator 4beta-phorbol 12-myristate 13-acetate (10 nM), and the inhibition of I (Kv) by AVP was prevented by the PKC inhibitor calphostin C (250 nM). These results suggest that the stimulation of Ca (2+) spiking by physiological concentrations of AVP involves PKC-dependent inhibition of KCNQ5 channels and increased AP firing in A7r5 cells. |
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