Supported by Awards AA11085 (S.M.S.) and AA12057 (D.R.A.) from the NIH.
Ethanol Triggers Neural Crest Apoptosis through the Selective Activation of a Pertussis Toxin–Sensitive G Protein and a Phospholipase Cβ–Dependent Ca2+ Transient
Version of Record online: 3 MAY 2006
Alcoholism: Clinical and Experimental Research
Volume 29, Issue 7, pages 1237–1246, July 2005
How to Cite
Garic-Stankovic, A., Hernandez, M. R., Chiang, P. J., Debelak-Kragtorp, K. A., Flentke, G. R., Armant, D. R. and Smith, S. M. (2005), Ethanol Triggers Neural Crest Apoptosis through the Selective Activation of a Pertussis Toxin–Sensitive G Protein and a Phospholipase Cβ–Dependent Ca2+ Transient. Alcoholism: Clinical and Experimental Research, 29: 1237–1246. doi: 10.1097/01.ALC.0000172460.05756.D9
- Issue online: 3 MAY 2006
- Version of Record online: 3 MAY 2006
- Received for January 14, 2005; accepted April 22, 2005.
Alcohol is a potent neurotoxin that triggers the selective apoptosis of neuronal populations in the developing fetus. For neural crest cells, clinically relevant ethanol levels (0.3%) rapidly elicit a phospholipase C (PLC)–dependent intracellular Ca2+ transient that is sufficient to activate apoptosis. We investigated the biochemical origins of this Ca2+ transient.
Three somite chick embryos (stage 8-) were pretreated with agonists and antagonists of PLC signaling pathways before ethanol challenge. The resulting intracellular Ca2+ release was quantified using Fluo-3; apoptosis was assessed using vital dyes.
Pretreatment of embryos with PLC antagonists U73122 or ET-18-OCH3 confirmed that a phosphoinositide-specific PLC was required for both the ethanol-dependent Ca2+ transient and subsequent cell death. Ethanol rapidly elevated intracellular inositol-1,4,5-trisphosphate Ins(1,4,5)P3 levels in the rostral portion of the embryo that contains neural crest progenitors. The Ins(1,4,5)P3 receptor antagonist xestospongin C blocked the appearance of the ethanol-dependent Ca2+ transient. Pretreatment with the pan-Gα protein antagonist GDPβS, but not with the tyrosine kinase antagonist genistein, suppressed ethanol's ability to elicit the Ca2+ transient, suggesting that a rise in PLC activity and Ins(1,4,5)P3 concentration originates from stimulation of heterotrimeric G proteins. To probe the identity of this G protein, embryos were treated with G protein antagonists. Pertussis toxin and NF023 suppressed the ethanol-induced Ca2+ transient and subsequent neural crest apoptosis, whereas suramin was weakly inhibitory. C3 exoenzyme was embryolethal over a wide concentration range, consistent with suggestions that Rho family GTPases participate in neural crest development. Gαi2 was identified by immunostaining in the neural crest cells.
We propose a role for Gαi/o protein activation and subsequent interaction of Gβγ with PLCβ in mediating the proapoptotic effects of ethanol upon the developing neural crest.