Blockade by NS-7, a neuroprotective compound, of both L-type and P/Q-type Ca2+ channels involving depolarization-stimulated nitric oxide synthase activity in primary neuronal culture.
The effect of 4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy)pyrimidine hydrochloride (NS-7), a neuroprotective compound, on Ca2+ channels involving the activation of nitric oxide synthase (NOS) was investigated in primary neuronal culture. The NOS activity was estimated from the cyclic GMP formation. The KCl (25 mM)-stimulated cyclic GMP formation was totally abolished by a combined treatment with nifedipine and omega-agatoxin IVA (omega-Aga), whereas spontaneous cyclic GMP formation was partially but significantly reduced by nifedipine. In contrast to nifedipine, NS-7 blocked KCl-stimulated cyclic GMP formation without affecting spontaneous cyclic GMP formation. Subsequently, the effects of nifedipine and NS-7 on L-type Ca2+ channels were compared. Nifedipine blocked equally the cyclic GMP formation stimulated by various concentrations of (+/-)-Bay K 8644, whereas NS-7 inhibited the maximal response without affecting the responses induced by low concentrations of (+/-)-Bay K 8644. The effects of NS-7 on L-type and P/Q-type Ca2+ channels involving KCl-stimulated cyclic GMP formation were subsequently examined. NS-7 suppressed the KCl-stimulated cyclic GMP formation measured in the presence of omega-Aga to almost the same extent as that determined in the presence of nifedipine. In contrast, NS-7 had no influence on ionomycin-induced enhancement of cyclic GMP formation. Finally, NS-7 reversed KCl-induced elevation of the intracellular free Ca2+ concentration. These findings suggest that NS-7 inhibits NOS activation in primary neuronal culture by reducing Ca2+ entry through L-type and P/Q-type Ca2+ channels, in which the inhibition is largely dependent on Ca2+ channel activity.
Research Laboratories, Nippon Shinyaku Co., Ltd., Kyoto, Japan.
M
M
Oka
Y
Y
Itoh
Y
Y
Ukai
K
K
Kimura
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester / Q000494:pharmacology; D060848:Agatoxins; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002120:Calcium Channel Agonists / Q000494:pharmacology; D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000187:drug effects*; D020746:Calcium Channels, L-Type; D020864:Calcium Channels, N-Type; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D004594:Electrophysiology; D051379:Mice; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009543:Nifedipine / Q000494:pharmacology; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D011189:Potassium Chloride / Q000037:antagonists & inhibitors; D011743:Pyrimidines / Q000494:pharmacology*; D013111:Spider Venoms / Q000494:pharmacology
D001498:3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethyl-5-nitro-4-(2-(trifluoromethyl)phenyl)-, Methyl ester; D060848:Agatoxins; D002118:Calcium; D002120:Calcium Channel Agonists; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D020864:Calcium Channels, N-Type; D006152:Cyclic GMP; D009419:Nerve Tissue Proteins; D018696:Neuroprotective Agents; D009543:Nifedipine; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D011189:Potassium Chloride; D011743:Pyrimidines; D013111:Spider Venoms; C106144:4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy)pyrimidine hydrochloride; C060465:omega-agatoxin I; C496321:Nos1 protein, mouse