Calcium channel activation facilitated by nitric oxide in retinal ganglion cells.
We investigated the modulation of voltage-gated Ca channels by nitric oxide (NO) in isolated salamander retinal ganglion cells with the goals of determining the type of Ca channel affected and the signaling pathway by which modulation might occur. The NO donors, S-nitroso-N-acetyl-penicillamine (SNAP, 1 mM) and S-nitroso-cysteine (1 mM) induced modest increases in the amplitude of Ca channel currents recorded with ruptured- and permeabilized-patch techniques by causing a subpopulation of the Ca channels to activate at more negative potentials. The Ca channel antagonists omega-conotoxin GVIA and nisoldipine each reduced the Ca channel current partially, but only omega-conotoxin GVIA blocked the enhancement by SNAP. The SNAP-induced increase was blocked by oxadiazolo-quinoxaline (50 microM), suggesting that the NO generated by SNAP acts via a soluble guanylyl cyclase to raise levels of cGMP. The membrane-permeant cGMP analog 8-(4-chlorophenylthio) guanosine cyclic monophosphate also enhanced Ca channel currents and 8-bromo guanosine cyclic monophosphate (1 mM) occluded enhancement by SNAP. Consistent with these results, isobutyl-methyl-xanthine (IBMX, 10 microM), which can raise cGMP levels by inhibiting phosphodiesterase activity, increased Ca channel current by the same amount as SNAP and occluded subsequent enhancement by SNAP. Neither IBMX, the cGMP analogs, nor SNAP itself, led to activation of cGMP-gated channels. N-[2-(methylamino)ethyl]-5-isoquinoline-sulfonamide (2 microM), a broad spectrum inhibitor of protein kinase activity, KT5823 (1 microM), a specific protein kinase G (PKG) inhibitor, and a peptide inhibitor of PKG (200 microM) blocked SNAP enhancement, as did 5'-adenylylimidophosphate (1.5 mM), a nonhydrolyzable ATP analog that prevents protein phosphorylation. A peptide inhibitor of protein kinase A (10 nM) did not block the facilitory effects of SNAP. Okadaic acid (1 microM), a phosphatase inhibitor, had no effect by itself but increased the enhancement of Ca channel current by SNAP. These results suggest that NO modulates retinal ganglion cell N-type Ca channels by facilitating their voltage-dependent activation via a mechanism involving guanylyl cyclase/PKG-dependent phosphorylation. This effect could fine-tune neural integration in ganglion cells or play a role in ganglion cell disease by modulating intracellular calcium signaling.
Hirooka
K
K
Departments of Physiology, Dalhousie University, Halifax, Nova Scotia B3H 4H7, Canada.
Kourennyi
D E
DE
Barnes
S
S
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D000557:Ambystoma; D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D003545:Cysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D066298:In Vitro Techniques; D007814:Larva; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D015737:Nisoldipine / Q000494:pharmacology; D009569:Nitric Oxide / Q000502:physiology; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D009603:Nitroso Compounds / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D012165:Retinal Ganglion Cells / Q000187:drug effects / Q000502:physiology*; D026423:S-Nitroso-N-Acetylpenicillamine; D026403:S-Nitrosothiols; D013779:Tetrodotoxin / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology; D020866:omega-Conotoxin GVIA / Q000494:pharmacology
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D026403:S-Nitrosothiols; D013873:Thionucleotides; D009569:Nitric Oxide; D013779:Tetrodotoxin; D015737:Nisoldipine; C055656:8-((4-chlorophenyl)thio)cyclic-3',5'-GMP; D026423:S-Nitroso-N-Acetylpenicillamine; D020866:omega-Conotoxin GVIA; C021315:S-nitrosocysteine; D010396:Penicillamine; D006152:Cyclic GMP; D003545:Cysteine; D015056:1-Methyl-3-isobutylxanthine