Opposite regulation of KCNQ5 and TRPC6 channels contributes to vasopressin-stimulated calcium spiking responses in A7r5 vascular smooth muscle cells

Cell Calcium. 2009 Apr;45(4):400-11. doi: 10.1016/j.ceca.2009.01.004. Epub 2009 Feb 25.

Abstract

Physiologically relevant concentrations of [Arg(8)]-vasopressin (AVP) induce repetitive action potential firing and Ca(2+) spiking responses in the A7r5 rat aortic smooth muscle cell line. These responses may be triggered by suppression of KCNQ potassium currents and/or activation of non-selective cation currents. Here we examine the relative contributions of KCNQ5 channels and TRPC6 non-selective cation channels to AVP-stimulated Ca(2+) spiking using patch clamp electrophysiology and fura-2 fluorescence measurements in A7r5 cells. KCNQ5 or TRPC6 channel expression levels were suppressed by short hairpin RNA constructs. KCNQ5 knockdown resulted in more positive resting membrane potentials and induced spontaneous action potential firing and Ca(2+) spiking. However physiological concentrations of AVP induced additional depolarization and increased Ca(2+) spike frequency in KCNQ5 knockdown cells. AVP activated a non-selective cation current that was reduced by TRPC shRNA treatment or removal of external Na(+). Neither resting membrane potential nor the AVP-induced depolarization was altered by knockdown of TRPC6 channel expression. However, both TRPC6 shRNA and removal of external Na(+) delayed the onset of Ca(2+) spiking induced by 25pM AVP. These results suggest that suppression of KCNQ5 currents alone is sufficient to excite A7r5 cells, but AVP-induced activation of TRPC6 contributes to the stimulation of Ca(2+) spiking.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Calcium Signaling / drug effects*
  • Cell Line
  • Culture Media
  • Diglycerides / metabolism
  • Diglycerides / pharmacology
  • Ion Channel Gating / drug effects
  • KCNQ Potassium Channels / metabolism*
  • Membrane Potentials / drug effects
  • Muscle, Smooth, Vascular / cytology*
  • Myocytes, Smooth Muscle / drug effects*
  • Myocytes, Smooth Muscle / metabolism*
  • Protein Subunits / metabolism
  • RNA, Small Interfering / metabolism
  • Rats
  • Sodium
  • TRPC Cation Channels / metabolism*
  • Vasopressins / pharmacology*

Substances

  • Culture Media
  • Diglycerides
  • KCNQ Potassium Channels
  • Kcnq5 protein, rat
  • Protein Subunits
  • RNA, Small Interfering
  • TRPC Cation Channels
  • Trpc6 protein, rat
  • Vasopressins
  • 1-oleoyl-2-acetylglycerol
  • Sodium