NITRIC-OXIDE MODULATION OF AGONIST-EVOKED INTRACELLULAR CA2-12 CELLS - INHIBITION OF PHOSPHOLIPASE-C ACTIVITY VIA CYCLIC GMP-DEPENDENT PROTEIN-KINASE( RELEASE IN NEUROSECRETORY PC)

Nitric oxide is a signaling molecule involved in events crucial to neu ronal cell function, such as neurotransmitter release, gene transcript ion, and neurotoxicity, i.e., a number of processes in which a key rol e appears to be played by increases in intracellular Ca2+ concentratio n. In the neurosecretory/neuronal cell line PC-12, we have investigate d the role of nitric oxide in the modulation of Ca2+ release from intr acellular stares elicited by activation of three different receptors c oupled to phosphatidylinositol-4,5-bisphosphate hydrolysis, i.e., the purinergic P-2U, muscarinic M3, and bradykinin B-2 receptors. The resu lts obtained show that nitric oxide donors have an inhibitory effect o n agonist-evoked Ca2+ release. This effect is not due to nitric oxide- induced modifications of Ca2+ storage, because the total releasable Ca 2+ pool, measured as the radioactivity released by thapsigargin and io nomycin in cells loaded at equilibrium with Ca-45(2+), was unchanged. In contrast, nitric oxide donors decreased agonist-evoked inositol-1,4 ,5-trisphosphate generation and total inositol phosphate accumulation. Similarly, nitric oxide inhibited total inositol phosphate accumulati on stimulated by either aluminium fluoride or Ca2+. All of these effec ts were mimicked by the cGMP analogue 8-bromo-cGMP. When cells were in cubated with nitric oxide synthase inhibitors, the results observed we re opposite those produced by nitric oxide donors. All of the effects of nitric oxide were abolished when cells were treated with the cGMP-d ependent protein kinase I inhibitor KT5823. Furthermore, KT5823 mimick ed the effects of nitric oxide synthase inhibitors. We conclude that n itric oxide and Ca2+ signaling pathways are interconnected in PC-12 ce lls. Modulation of inositol phosphate generation and Ca2+ release by n itric oxide appears to be exerted primarily at the level of phospholip ase C functioning and to be mediated by the activation of cGMP-depende nt protein kinase I.