Kisspeptin activation of TRPC4 channels in female GnRH neurons requires PIP2 depletion and cSrc kinase activation

Endocrinology. 2013 Aug;154(8):2772-83. doi: 10.1210/en.2013-1180. Epub 2013 Jun 6.


Kisspeptin signaling via its Gαq-coupled receptor GPR54 plays a crucial role in modulating GnRH neuronal excitability, which controls pituitary gonadotropins secretion and ultimately reproduction. Kisspeptin potently depolarizes GnRH neurons primarily through the activation of canonical transient receptor potential (TRPC) channels, but the intracellular signaling cascade has not been elucidated. Presently, we have established that kisspeptin activation of TRPC channels requires multiple membrane and intracellular signaling molecules. First, phosphatidylinositol-4,5-bisphosphate (PIP(2)) hydrolysis by phospholipase Cβ is required because whole-cell dialysis of Dioctanoylglycerol-PIP(2) (DiC8-PIP(2)) inhibited the kisspeptin activation of TRPC channels, and the phosphatidylinositol 4-kinase inhibitor wortmannin, which attenuates PIP(2) synthesis, prolonged TRPC channel activation. Using single cell RT-PCR, we identified that the mRNA for the PIP(2)-interacting TRPC channel subunit, TRPC4α, is expressed in GnRH neurons. Depletion of intracellular Ca(2+) stores by thapsigargin and inositol 1,4,5-trisphosphate had no effect, indicating that the TRPC channels are not store-operated. Neither removing extracellular Ca(2+) nor buffering intracellular Ca(2+) with EGTA or BAPTA had any effect on the kisspeptin activation of the TRPC channels. However, the Ca(2+) channel blocker Ni(2+) inhibited the kisspeptin-induced inward current. Moreover, inhibition of protein kinase C by bisindolylmaleimide-I or calphostin C had no effect, but activation of protein kinase C by phorbol 12,13-dibutyrate occluded the kisspeptin-activated current. Finally, inhibition of the cytoplasmic tyrosine kinase cSrc by genistein or the pyrazolo-pyrimidine PP2 blocked the activation of TRPC channels by kisspeptin. Therefore, TRPC channels in GnRH neurons are receptor-operated, and kisspeptin activates TRPC channels through PIP(2) depletion and cSrc tyrosine kinase activation, which is a novel signaling pathway for peptidergic excitation of GnRH neurons.

Publication types

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

MeSH terms

  • Androstadienes / pharmacology
  • Animals
  • CSK Tyrosine-Protein Kinase
  • Calcium / metabolism
  • Enzyme Activation / drug effects
  • Female
  • Genistein / pharmacology
  • Gonadotropin-Releasing Hormone / genetics
  • Gonadotropin-Releasing Hormone / metabolism*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Kisspeptins / pharmacology*
  • Membrane Potentials / drug effects
  • Mice
  • Mice, Transgenic
  • Neurons / cytology
  • Neurons / metabolism*
  • Neurons / physiology
  • Ovariectomy
  • Patch-Clamp Techniques
  • Phosphatidylinositol 3-Kinases / metabolism
  • Phosphatidylinositol 4,5-Diphosphate / metabolism*
  • Phosphoinositide-3 Kinase Inhibitors
  • Protein Kinase C / antagonists & inhibitors
  • Protein Kinase C / metabolism
  • Protein Kinase Inhibitors / pharmacology
  • Pyrimidines / pharmacology
  • TRPC Cation Channels / genetics
  • TRPC Cation Channels / metabolism*
  • Wortmannin
  • src-Family Kinases / antagonists & inhibitors
  • src-Family Kinases / metabolism*


  • AG 1879
  • Androstadienes
  • Kisspeptins
  • Phosphatidylinositol 4,5-Diphosphate
  • Phosphoinositide-3 Kinase Inhibitors
  • Protein Kinase Inhibitors
  • Pyrimidines
  • TRPC Cation Channels
  • TRPC4 ion channel
  • Green Fluorescent Proteins
  • Gonadotropin-Releasing Hormone
  • Genistein
  • CSK Tyrosine-Protein Kinase
  • src-Family Kinases
  • Protein Kinase C
  • Calcium
  • Wortmannin