Vasopressin stimulates action potential firing by protein kinase C-dependent inhibition of KCNQ5 in A7r5 rat aortic smooth muscle cells.
[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.
Brueggemann
Lioubov I
LI
Loyola Univ Medical Center, Dept of Pharmacology, Maywood, IL 60153, USA.
Moran
Christopher J
CJ
Barakat
John A
JA
Yeh
Jay Z
JZ
Cribbs
Leanne L
LL
Byron
Kenneth L
KL
D000200:Action Potentials / Q000187:drug effects / Q000502:physiology*; D000818:Animals; D001011:Aorta; D002118:Calcium / Q000378:metabolism; D002138:Calibration; D002478:Cells, Cultured; D051656:KCNQ Potassium Channels / Q000037:antagonists & inhibitors*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques; D011493:Protein Kinase C / Q000378:metabolism*; D051381:Rats; D014667:Vasopressins / Q000494:pharmacology*
D051656:KCNQ Potassium Channels; C501028:Kcnq5 protein, rat; D014667:Vasopressins; D011493:Protein Kinase C; D002118:Calcium