Nitric oxide (NO) is an important messenger molecule in the vascular system, immune system, and central nervous system. In brain, NO can act as a neurotransmitter/neuromodulator and increasing evidence suggests that NO, or closely related molecules, may also play an important pathophysiological role. The Broad, Long-Term objective of this project is to investigate the effects of ethanol on NO formation in brain. Glial cells and a subpopulation of neurons possess two distinctly different forms of NO synthase (NOS) that produces NO from the precursor amino acid L-arginine. Neuronal NOS is constitutively expressed and is activated by calcium. In contrast, glial NOS is not normally present in the cell but is synthesized de novo in response to stimulating agents such as cytokines and bacterial endotoxins. Once synthesized, the activity of this """"""""inducible"""""""" form of the enzyme is independent of calcium and is regulated at the level of gene transcription. Our preliminary studies in cell culture show that chronic ethanol exposure can increase NO formation in neurons but reduce NO formation in astroglia. The potentiating effect of chronic ethanol on NO formation in neurons is most likely due to an increase in NMDA receptor function while its inhibitory effect on NO formation in astroglia appears to be due to reduced expression of the NOS protein. Given the apparent importance of NO during both normal and pathological conditions, it is important to clearly understand how both acute and chronic ethanol exposure affects NO formation in brain.
The Specific Aims of this project are to: (1) identify NMDA receptor subunits that colocalize with and couple to activation of NOS in neurons; (2) determine the effects of chronic ethanol on expression of NMDA receptors that couple to NOS activation and the role of NO in the neuropathology of alcohol abuse; (3) determine the mechanism(s) of how chronic ethanol exposure can inhibit NOS expression in cultured astroglia; and (4) determine the effects of ethanol on NOS expression in reactive astrocytes during response to injury. The project involves studies to be performed in primary cultures of astroglia and neurons as well as studies with brain slice preparations. We hypothesize that ethanol induced alterations in brain function and pathology relate, in part, to altered NMDA receptor function and NO formation. These studies are likely to yield important and clinically significant information on the role of NO in the actions of ethanol in the brain, and may lead to more effective pharmacological treatments for acute alcohol intoxication, withdrawal and neurodegeneration associated with chronic alcohol abuse.
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