Mechanisms underlying opposing neuronal responses to brief vs. prolonged dopamine

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Drug abuse is a chronic and devastating disease, costing society over 200 billion per year. The problem is widespread; in 2004, over 34 million Americans reported lifetime use of cocaine. One of NIDA's top research priorities is finding drugs to block cocaine?s effects, which will require an understanding of the molecular mechanisms of addiction. Persistent use of cocaine leads to maladaptations in reward-related learning such that the drug is prized above all other rewards, and is compulsively sought after and used, despite severe negative consequences (addiction). Cocaine prevents dopamine (DA) reuptake. A single dose of an addictive drug can elevate synaptic DA for hours. It is not clear how repeated, prolonged elevations in [DA] disrupt the normal mechanisms of associative learning and memory. Such processes depend upon the flow of ions through channels in the neuronal membrane (ion currents); therefore, the densites and characteristics of ion channels in the neuronal membrane help to determine the neuron?s capacity to engage in mechanisms of learning and memory. Both cocaine and DA are known to alter ion current densities. Perhaps cocaine-induced aberrations in learning and memory are due to changes in ion current densities resulting from prolonged elevations in DA. Elucidating the processes by which prolonged elevations in synaptic DA lead to changes in ion current densities may lead to a deeper appreciation for how addiction usurps the normal mechanisms of reward related learning and memory. The transient potassium current (IA) is important for learning and memory. Kv4 channels mediate IA. Using a model circuit, the crustacean pyloric network, we found that when DA binds to its receptors, D1 and D2, they produce global biochemical signals that have different effects on IA density over the short- and long-term. For example, in response to a brief application of DA, D2 receptors mediate an increase in IA density. On the other hand, a prolonged 4hr. application of DA produces a D2 mediated, persistent decrease in IA density 10-12 hrs. after DA has been removed. This proposal focuses on the mechanism(s) by which brief versus prolonged applications of DA produce opposing effects on IA density. We specifically test the hypothesis that DA induces global changes in [cAMP] that then alter the phosphorylation state of both Kv4 channels and a transcription factor named CREB. Whereas changes in Kv4 channels are relatively short-lived, modifications in CREB activity are long-lived and result in alterations in Kv4 transcript number. Here we propose to use molecular biology and electrophysiology techniques to measure and correlate changes in global [cAMP], IA density and shal transcript number. Furthermore, pharmacological tools will be used to antagonize or mimic global changes in [cAMP] to determine if they underlie the changes in IA density and shal transcript number. Additionally, expression of a dominant-negative CREB protein and visualization of protein kinase A translocation using confocal microscopy will help to determine if CREB is involved in mediating the long-term response.