The pre-synaptic Munc13-1 binds alcohol and modulates alcohol self-administration in Drosophila
Article first published online: 10 JUN 2013
© 2013 International Society for Neurochemistry
Journal of Neurochemistry
Volume 126, Issue 6, pages 715–726, September 2013
How to Cite
J. Neurochem. (2013) 126, 715–726.
- Issue published online: 6 SEP 2013
- Article first published online: 10 JUN 2013
- Accepted manuscript online: 20 MAY 2013 11:37AM EST
- Manuscript Accepted: 17 MAY 2013
- Manuscript Revised: 9 MAY 2013
- Manuscript Received: 25 MAR 2013
- University of Houston. Grant Number: NIH 7R21AA016140
- Drosophila ;
- mass spectrometry;
- protein kinase C
Munc13-1 is a pre-synaptic active-zone protein essential for neurotransmitter release and involved in pre-synaptic plasticity in brain. Ethanol, butanol, and octanol quenched the intrinsic fluorescence of the C1 domain of Munc13-1 with EC50s of 52 mM, 26 mM, and 0.7 mM, respectively. Photoactive azialcohols photolabeled Munc13-1 C1 exclusively at Glu-582, which was identified by mass spectrometry. Mutation of Glu-582 to alanine, leucine, and histidine reduced the alcohol binding two- to five-fold. Circular dichroism studies suggested that binding of alcohol increased the stability of the wild-type Munc13-1 compared with the mutants. If Munc13-1 plays some role in the neural effects of alcohol in vivo, changes in the activity of this protein should produce differences in the behavioral responses to ethanol. We tested this prediction with a loss-of-function mutation in the conserved Dunc-13 in Drosophila melanogaster. The Dunc-13P84200/+ heterozygotes have 50% wild-type levels of Dunc-13 mRNA and display a very robust increase in ethanol self-administration. This phenotype is reversed by the expression of the rat Munc13-1 protein within the Drosophila nervous system. The present studies indicate that Munc13-1 C1 has binding site(s) for alcohols and Munc13-1 activity is sufficient to restore normal self-administration to Drosophila mutants deficient in Dunc-13 activity.
The pre-synaptic Mun13-1 protein is a critical regulator of synaptic vesicle fusion and may be involved in processes that lead to ethanol abuse and addiction. We studied its interaction with alcohol and identified Glu-582 as a critical residue for ethanol binding. Munc13-1 can functionally complement the Dunc13 haploinsufficient ethanol self-administration phenotype in Drosophila melanogaster, indicating that this protein participates in alcohol-induced behavioral plasticity.