Nitric oxide underlies the differentiation of PC12 cells induced by depolarization with high KCl.
Nitric oxide (NO) acts as a cytostatic agent to induce neuronal differentiation of PC12 cells after nerve growth factor (NGF) treatment. We newly subcloned PC12K cells that extended neurites after depolarization with high KCl. Here we present evidence that the neuronal differentiation of PC12K cells caused by depolarization with high KCl is mediated by endogenous NO. The outgrowth of neurites was significantly inhibited by 2 mM N-nitro-L-arginine methyl ester (L-NMAE), and 10 mM L-NAME was necessary for complete inhibition. The inhibition of NGF-dependent neurite outgrowth by L-NAME was abolished by depolarization of cells with KCl. The expression of neuronal- and endothelial-NO-synthase in PC12K cells was confirmed by immuno-cytochemical and immuno-blotting analyses with the respective monoclonal antibodies. However, the expression of inducible-NO synthase was not observed in PC12K cells cultured with high KCl under the depolarization conditions with 45 mM KCl. We observed the increase of NO in the differentiated PC12K cells using diaminofluorescein, a novel fluorescent indicator for NO.
Nakagawa
H
H
Department of Biochemical Science, Faculty of Computer and Systems, Engineering, Kyushu Institute of Technology, Iizuka, Fukuoka 820-8502, Japan.
Yoshida
M
M
Miyamoto
S
S
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division; D016764:Cell Polarity / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009414:Nerve Growth Factors / Q000494:pharmacology; D016501:Neurites / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D009474:Neurons / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D009569:Nitric Oxide / Q000096:biosynthesis / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D016716:PC12 Cells; D011189:Potassium Chloride / Q000494:pharmacology*; D051381:Rats
D004791:Enzyme Inhibitors; D009414:Nerve Growth Factors; D009569:Nitric Oxide; D011189:Potassium Chloride; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; C496322:Nos1 protein, rat; C496318:Nos2 protein, rat; C496316:Nos3 protein, rat; D019331:NG-Nitroarginine Methyl Ester