This competitive U01 renewal application, under the INIA-West consortium, is based on behavioral findings that the central amygdala (CeA) is a key brain area underlying stress reactions and alcohol dependence, and that these behaviors involve several CeA transmitters (GABA, glutamate) and neuropeptides (CRF, opioids and galanin). Our published electrophysiological studies of these systems in CeA provide an entry point for proposed new studies: a stated need to pursue physiological evaluation of the function of gene products suggested by the molecular components (e.g., of Y. Blednov and others) of the INIA-West to be involved in excessive alcohol drinking. Therefore, we now propose to use electrophysiological and cytochemical methods to investigate the hypothesis of a role for neuroinflammatory factors (lipopolysaccharide {LPS}, toll-like receptor 4 {TLR4}, CD14, cytokines) in alcohol preference and excessive drinking. To test this hypothesis at the cellular level, we propose 4 Specific Aims: 1) To assess the role of TLR4 activation in effects of ethanol and CRF on GABAergic and glutamatergic transmission in CeA slices by LPS superfusion or i.p. injection in CeA of wild type (WT) and CD14 knockout (KO) mice;2) To assess effects of the TLR4-g en e rated inflammatory cytokines IL-ip, TNFa, and IL-6 on membrane and synaptic measures in CeA of WT mice;3) To determine if the LPS, CRF or chronic ethanol increase cytochemical signs of inflammation in CeA;4) To determine if the electrophysiological or cytochemical effects of LPS, cytokines, ethanol or CRF on CeA seen in Specific Aims 1-3 can be reversed by pre-treatment with certain anti-inflammatory drugs. These proposed studies thus represent new steps toward evaluating the cellular sites and mechanisms of action of the emerging gene targets suggested by other INIA West components to underlie alcohol preference or excessive drinking, and may further validate drug targets for reversal or prevention of alcohol effects and excessive drinking.
This project will examine the cellular and synaptic mechanisms likely to underlie the recently discovered inflammatory effects on the brain of alcohol drinking. Because such neuro-inflammatory effects are also suggested to lead to excessive drinking, the present studies also represent a new direction in attempts to validate drug targets for the prevention or treatment of excessive drinking and alcoholism.
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