The protective effect of neuronal nitric oxide synthase (nNOS) against alcohol toxicity depends upon the NO-cGMP-PKG pathway and NF-kappaB.
Fetal alcohol syndrome (FAS) stems from maternal alcohol abuse during pregnancy and is an important cause of mental retardation and hyperactivity in children. In the developing brain, alcohol can kill neurons, leading to microencephaly. However, due to their genetic makeup, some individuals are less vulnerable than others to alcohol's neurotoxic effects. Animal studies have demonstrated that one particular gene, neuronal nitric oxide synthase (nNOS), protects developing neurons in vivo against alcohol-induced death. We utilized pharmacologic techniques to demonstrate that nNOS protects neurons against alcohol toxicity by activating the NO-cGMP-PKG signaling pathway. Cerebellar granule cell cultures derived from mice carrying a null mutation for nNOS (nNOS-/- mice) were substantially more vulnerable than cultures from wild-type mice to alcohol-induced cell death. However, activation of the pathway at sites downstream of nNOS protected the cultures against alcohol toxicity. Conversely, blockade of the pathway rendered wild-type cultures vulnerable to alcohol-induced death. We further identified NF-kappaB as the downstream effector through which nNOS and the NO-cGMP-PKG pathway signal their neuroprotective effects. Tumor necrosis factor-alpha (TNF-alpha), which activates NF-kappaB, ameliorated alcohol-induced cell death in nNOS-/- and wild-type cultures, while an NF-kappaB inhibitor (NFi) blocked the protective effects of TNF-alpha and worsened alcohol-induced cell death. Furthermore, NFi blocked the protective effects of NO-cGMP-PKG pathway activators, demonstrating that NF-kappaB is downstream of the NO-cGMP-PKG pathway. As wild-type neurons matured in culture, they became resistant to alcohol toxicity. However, this maturation-dependent alcohol resistance did not occur in nNOS-/- mice and could be reversed in wild-type mice with NFi, demonstrating that nitric oxide and NF-kappaB are crucial for the development of alcohol resistance with age. Thus, nNOS protects developing neurons against alcohol toxicity by activating the NO-cGMP-PKG-NF-kappaB pathway and is crucial for the acquisition of maturation-dependent alcohol resistance.
Bonthius
Daniel J
DJ
Department of Pediatrics, College of Medicine, University of Iowa, Iowa City, IA, USA. daniel-bonthius@uiowa.edu
Bonthius
Nancy E
NE
Li
Shenglan
S
Karacay
Bahri
B
D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D002492:Central Nervous System Depressants / Q000633:toxicity*; D002531:Cerebellum / Q000166:cytology; D006152:Cyclic GMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000431:Ethanol / Q000633:toxicity*; D051379:Mice; D018345:Mice, Knockout; D009474:Neurons / Q000187:drug effects*; D009569:Nitric Oxide / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I / Q000172:deficiency / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D000097782:NF-kappaB-Inducing Kinase
D002492:Central Nervous System Depressants; D006152:Cyclic GMP; D004791:Enzyme Inhibitors; D000431:Ethanol; D009569:Nitric Oxide; D052248:Nitric Oxide Synthase Type I; D017346:Protein Serine-Threonine Kinases; D014409:Tumor Necrosis Factor-alpha; D000097782:NF-kappaB-Inducing Kinase