cGMP inhibits IP3-induced Ca2+ release in intact rat megakaryocytes via cGMP- and cAMP-dependent protein kinases
Author's present address S. Tertyshnikova: Bristol-Myers Squibb Company, Pharmaceutical Reseach Institute, Walingford, CT 06492-7660, USA.
Corresponding author A. Fein: Department of Physiology, University of Connecticut Health Center, 263 Farmington Avenue, Farmington, CT 06030-3505, USA. Email: firstname.lastname@example.org
- 1Inhibition of inositol 1,4,5-trisphosphate (IP3) receptor-mediated Ca2+ release by cGMP was examined in intact rat megakaryocytes, by using a combination of single cell fluorescence microscopy to monitor intracellular free calcium ([Ca2+]i) and flash photolysis of caged second messengers.
- 2Sodium nitroprusside (SNP), a nitric oxide (NO) donor, and the hydrolysis-resistant cGMP analogue 8-(4-chlorophenylthio)guanosine 3′,5′-cyclic monophosphate (pCPT-cGMP) inhibited Ca2+ release induced by photolysis of caged IP3. Neither of them affected the rate of Ca2+ removal from the cytoplasm following photolysis of caged Ca2+.
- 3Photolysis of the caged NO donor 3-morpholinosydnonimine (SIN-1) during agonist-induced [Ca2+]i oscillations inhibited Ca2+ release without affecting the rate of Ca2+ uptake and/or extrusion.
- 4We conclude that the inhibition of IP3-induced Ca2+ release is the principal mechanism of NO-cGMP-dependent inhibition of [Ca2+]i mobilization.
- 5IPG, a specific peptide inhibitor of cGMP-dependent protein kinase (cGMP-PK), blocked the inhibitory effect of pCPT-cGMP, indicating that the inhibition of IP3-induced Ca2+ release by pCPT-cGMP is mediated by cGMP-PK. However, the simultaneous application of both IPG and IP20, a specific peptide inhibitor of cAMP-dependent protein kinase (cAMP-PK), was required to block the inhibitory effect of SNP. These data strongly suggest that NO-cGMP-dependent inhibition of [Ca2+]i mobilization is mediated via the activation of both cGMP-PK and cAMP-PK.