Glutaminase Knockouts and drug dependence

描述（来自申请人摘要）： 反复暴露于精神兴奋剂会导致精神兴奋剂致敏， 增加对重复精神兴奋剂给药的运动反应， 被认为是药物依赖的动物模型。精神兴奋剂-以及 大多数药物滥用-似乎有其主要作用， 中脑多巴胺系统敏化涉及突触前和突触后 多巴胺和谷氨酸突触传递的适应， mesoadenbens系统因此，令人惊讶的是， 中胚层系统似乎共同释放谷氨酸。这就提出了一个 假设多巴胺神经元的谷氨酸突触传递是至关重要的 精神兴奋剂致敏，并提供了一种新的治疗靶点， 成瘾的药物治疗为了解决这一假设，转基因 技术将被用来产生三个系列的突变小鼠缺乏 转氨酶-主要负责合成 神经递质谷氨酸。第一行将是组成型敲除; 这应该有助于证实转氨酶是主要来源， 神经递质谷氨酸。第二批小鼠将涉及拯救 组成型基因敲除小鼠前脑神经元中转氨酶的表达 这将产生单胺能神经元选择性的精氨酸酶敲除，并且 为评价单胺能神经元谷氨酸的作用提供了初步的方法 致敏的共传递。在第三组小鼠中， 转运蛋白启动子将用于驱动选择性消除 多巴胺神经元中的转氨酶表达。在这三组小鼠中， 经典组织学、蛋白质印迹法、原位杂交，以及 将进行免疫细胞化学以评估敲除对细胞凋亡的影响。 大脑的细胞组织，转氨酶的细胞选择性 敲除，以及对细胞谷氨酸水平的影响。多巴胺神经元 从突变小鼠制备的培养物将用于显示消除 谷氨酰胺酶表达阻断谷氨酸能突触传递。行为 然后将对突变小鼠进行测试，以评估其运动功能的变化， 活动，食欲行为，表现出精神兴奋剂的倾向 敏感性和自我管理的脆弱性。如果精神兴奋剂 通过消除，敏感性和自我管理确实受到阻碍 多巴胺神经元谷氨酸的共同传递，那么这些突触将成为一个 药物依赖性治疗中治疗性干预的新靶点。

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.