Neurotrophin protection against toxicity induced by low potassium and nitroprusside in cultured cerebellar granule neurons.
Long-term survival of cultured rat cerebellar granule neurons requires depolarizing concentrations of potassium (high potassium; 25 mM KCl). A high-potassium culturing condition has been reported to increase the intracellular calcium concentration ([Ca2+]i) and the expression of brain-derived neurotrophic factor (BDNF), which in turn induces the expression of neurotrophin-3 (NT-3) in these neurons. We therefore examined the neurotrophic effect of these two neurotrophins in low-potassium (5 mM) cultures and their neuroprotective capabilities against sodium nitroprusside-induced neurotoxicity in both low- and high-potassium cultures. Neuronal survival and neurotrophic effects were monitored by [3H]ouabain binding and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. In low-potassium cultures, the neurotrophic effect of BDNF approached that found in high-potassium cultures but was much more robust than that of NT-3. In contrast, undifferentiated neurons cultured in high-potassium medium were much less responsive to BDNF and not responsive at all to NT-3. Induction of nitroprusside neurotoxicity occurred more readily in low- than in high-potassium cultures. BDNF, NT-3, and a high potassium concentration, alone or in combination, were unable to protect neurons treated with nitroprusside at 50 or 100 microM. However, the neurotoxicity of a lower dose of nitroprusside (10 microM) was reversed by the combined actions of these two neurotrophins in low-potassium cultures and by BDNF alone in high-potassium cultures. Because nitroprusside neurotoxicity is less robust in high-potassium cultures, high-potassium-induced BDNF expression and subsequent NT-3 expression may participate in its neuroprotection and neurotrophism in these cultures. Also, we found that toxic doses of nitroprusside antagonized KCl- and NMDA-induced rises in [Ca2+]i, suggesting that this effect is related to nitroprusside-induced neurotoxicity.
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Section on Molecular Neurobiology, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland 20892-1272, USA.
Chuang
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D000818:Animals; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology*; D016923:Cell Death; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D003470:Culture Media; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D009414:Nerve Growth Factors / Q000494:pharmacology*; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D020933:Neurotrophin 3; D009599:Nitroprusside / Q000008:administration & dosage* / Q000494:pharmacology; D009994:Osmolar Concentration; D011188:Potassium / Q000008:administration & dosage* / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley
D019208:Brain-Derived Neurotrophic Factor; D003470:Culture Media; D009414:Nerve Growth Factors; D018696:Neuroprotective Agents; D020933:Neurotrophin 3; D016222:Fibroblast Growth Factor 2; D009599:Nitroprusside; D011188:Potassium