The overall objective of this project is to reveal the cellular mechanisms underlying opiate effects and opiate dependence, and the role of central opioid peptides in both naive and opiate-dependent subjects. We propose to study the central amygdala nucleus (CeA), thought to be critically involved in stress and drug reward and dependence, and to contain opioids, nociceptin (orphanin FQ) and their relevant receptors. In pilot electrophysiological studies we found unusual actions of nociceptin on evoked and spontaneous IPSPs in a CeA slice, and in blocking the effects of ethanol on these IPSPs. We also found a mediator role for CRF receptors, and a regulator role for mu opiate receptors, in the ethanol effects. Also, withdrawal from chronic morphine treatment increased spontaneous activity and caused depolarization shifts in CeA neurons. Thus, we propose the following 3 Specific Aims: 1) To examine the membrane and synaptic effects of mu, delta, kappa and ORL-1 selective agonists on rat CeA neurons and determine if chronic morphine treatment alters these effects or baseline properties of CeA neurons; 2) To determine if chronic morphine causes a re-composition of NMDAR subunits in rat CeA as it appears to do in nucleus accumbens, by using a battery of multidisciplinary tests of NMDARs, including electrophysiological, pharmacological, molecular and histochemical methods for profiling NR2 subunits; 3) To assess the possibility of interactions between opioids and CRF on synaptic and membrane properties of CeA neurons, and determine if chronic morphine alters the interactions.
These aims will test several hypotheses, including those concerning neuroadaptative mechanisms following chronic opiate treatment, and that some actions of opiates in CeA involve CRF receptors. To accomplish these aims, we will record from CeA neurons of brain slices using intracellular or whole-cell patch methods and measure pharmacologically-isolated, evoked and spontaneous synaptic currents and responses to exogenous agonists. We also will use molecular (Western blots, quantitative RT-PCR) and histochemical (in situ hybridization) methods to determine relative levels of different NR2 subunits. These studies may help define the role of opioids and their receptors in behavior and provide rational targets for therapeusis of opiate """"""""craving. ? ?
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