The experiments in this proposal are aimed at uncovering adaptive changes in neuronal activity that result from chronic opioid treatment. Behaviorally it can be observed that a high degree of tolerance to and dependence on morphine develops with chronic use. It is curious that, at the cellular and biochemical level, the degree of tolerance and dependence is orders of magnitude smaller. The disparity in the observations may suggest that there are multiple mechanisms arranged in series that could account for the high level of behavioral tolerance. Another possibility is that regulation of ion conductances other than those directly affected by opioids are altered with chronic treatment and they may account for the increased level of tolerance and dependence. Opioids activate potassium channels and/or the inactive calcium channels to perturb cell excitability and function. In the continued presence of opioids, tolerance develops to these direct actions of opioids. There are, in addition, other adaptive changes in cell function and excitability not directly related to the specific ion channels gated by opioids. The regulation of those channels 'indirectly' gated by opioids is the subject of this proposal. Intracellular and whole cell recordings from neurons in the locus coeruleus and substantia gelatinosa of the spinal trigeminal nucleus will be made in brain slice preparations. With these two preparations, two opioid mediated responses will be investigated, the increase in potassium conductance and the presynaptic inhibition of glutamate release. One consistent observation made with chronic opioid treatment is an increase in the basal and stimulated level of cAMP. In this proposal, the effects of cAMP-dependent processes on transmitter release and ion conductances gated by voltage, transmitters and G- proteins will be identified and characterized. These cAMP-dependent processes will be studied after acute application (min or hours) of opioid agonists and in animals treated chronically (days) with morphine. By investigating the alteration in the regulation of ion conductances not directly affected by opioids, it may be possible to clarify the mechanisms by which the high levels of tolerance found at the behavioral level are generated.
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