Glutaminase Knockouts and drug dependence

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DESCRIPTION(From applicant's abstract): Repeated psychostimulant exposure causes psychostimulant sensitization, an increasing motor response to repeated psychostimulant administration that is thought to be an animal model of drug dependence. Psychostimulants - as well as most drugs of abuse - appear to have their principal action in the mesoaccumbens dopamine system. Sensitization involves pre- and postsynaptic adaptations in dopamine as well as glutamate synaptic transmission in the mesoaccumbens systems. So, it is striking that the dopamine neurons that form the mesoaccumbens system appear to corelease glutamate. This raises the hypothesis that glutamate synaptic transmission by dopamine neurons is crucial to psychostimulant sensitization and provides a novel therapeutic target for the pharmacotherapy of addiction. To address this hypothesis, transgenic techniques will be used to generate three lines of mutant mice lacking glutaminase - the enzyme principally responsible for the synthesis of neurotransmitter glutamate. The first line will be a constitutive knockout; this should serve to confirm that glutaminase is the principal source of neurotransmitter glutamate. The second line of mice will involve the rescue of glutaminase expression in forebrain neurons of the constitutive knockout mice; this will yield a monoaminergic neuron-selective glutaminase knockout, and provide an initial way to evaluate the role of monoaminergic neuron glutamate cotransmission in sensitization. In the third line of mice, the dopamine transporter promotor will be used to drive the selective elimination of glutaminase expression in dopamine neurons. In the three lines of mice, classical histology, western blotting, in situ hybridization, and immunocytochemistry will be done to evaluate the impact of the knockouts on the cellular organization of the brain, the cellular selectivity of the glutaminase knockout, and the effects on cellular glutamate levels. Dopamine neuron cultures prepared from the mutant mice will be used to show that eliminating glutaminase expression blocks glutamatergic synaptic transmission. Behavioral testing of the mutant mice will then be done to evaluate changes in their motor activity, appetitive behavior, propensity to manifest psychostimulant sensitization, and vulnerability to self-administration. If psychostimulant sensitization and self-administration are indeed blocked by eliminating dopamine neuron glutamate cotransmission, then these synapses would become a novel target for therapeutic intervention in the treatment of drug dependence.