PKMzeta involvement in cocaine sensitization

描述(由申请人提供)：可卡因敏化是对随后的可卡因挑战所引起的运动刺激效应的渐进和持久的增强。这项工作的目的是阐明PKM的持续活动是否？在VTA和NAC中维持可卡因敏化的发展和表达。具体目标有三个：1)评价PKM的效果；抑制可卡因行为敏化的发展和表达。我们将使用行为敏化范式，在VTA内和NAC内微量注射ZIP(PKM？选择性抑制剂)在敏化的不同阶段 确定PKM的过程？在行为敏感化的发展和表达中的作用。假设在VTA和NAC中给予ZIP将阻断可卡因敏化的发展和表达2)以确定PKM？可卡因敏化对VTA、DA和NAC介质刺神经元AMPA电流和AMPA/NMDA比值的抑制作用。利用VTA DA和NAC培养的棘神经元的全细胞记录，我们将测量反复暴露于可卡因的大鼠AMPA介导的EPSCs和AMPA/NMDA的比率。ZIP是否会被用来评估PKM？是维持可卡因诱导的增强所必需的。我们假设ZIP将导致AMPA电流的减少，因此，将降低可卡因致敏动物的AMPA/NMDA比率至基础水平。3)评估PKM的细胞底物。抑制物对成瘾相关的可塑性起作用。利用蛋白质印迹分析，我们将检测可卡因致敏发展和表达过程中VTA和NAC中PKM和AMPA受体亚单位的水平。其次，我们将使用合成肽TAT-GluR23y的显微注射和全细胞斑片记录中的整流指数曲线来检测依赖和缺失GluR2的AMPA受体在维持PKM？对可卡因致敏的影响。假设可卡因致敏与PKM增加有关。通过抑制GluR2依赖的AMPA受体从突触后部位移除而起作用的水平。这项研究将为更好地了解药物诱导的中枢神经系统可塑性提供更好的基础，并可能为治疗性药物干预提供可能的途径。

DESCRIPTION (provided by applicant): Cocaine sensitization is a progressive and long-lasting enhancement of the motor stimulant effect induced by a subsequent cocaine challenge. The goal of this work is to elucidate if the persistent activity of PKM? in the VTA and NAc sustains the development and expression of cocaine sensitization. There are three specific aims: 1) to evaluate the effect of PKM? inhibition in the development and expression of cocaine behavioral sensitization. We will use a behavioral sensitization paradigm with intra VTA and intra NAc microinfusions of ZIP (PKM? selective inhibitor) at different stages of the sensitization process to determine PKM? 's role in the development and expression of behavioral sensitization. The hypothesize is that ZIP administration in the VTA and NAc will block the development and expression of cocaine sensitization 2) To determine the role of PKM? inhibition on AMPA currents and AMPA/NMDA ratios in VTA DA and NAc medium spiny neurons after cocaine sensitization. Using whole cell recordings from VTA DA and NAc medium spiny neurons, we will measure AMPA mediated EPSCs and AMPA/NMDA ratios from rats repeatedly exposed to cocaine. ZIP will be used to assess whether PKM? is necessary for the maintenance of the cocaine-induced potentiation. We hypothesize that ZIP will induce a decrease in AMPA currents and therefore, will diminish AMPA/NMDA ratios to basal levels in cocaine sensitized animals. 3) To assess the cellular substrate by which the PKM? inhibitor exerts its effects on addiction related plasticity. Using western blot analysis we will measure PKM¿ and AMPA receptor subunit levels in the VTA and NAc during development and expression of cocaine sensitization. Second, we will use microinjections of the synthetic peptide Tat-GluR23y and rectification index curves in whole cell patch recordings to examine the contribution of GluR2-dependent and GluR2-lacking AMPA receptors in maintaining PKM? effects in cocaine sensitization. The hypothesis is that cocaine sensitization involves an increase in PKM? levels which is acting by inhibiting GluR2-dependent AMPA receptor removal from postsynaptic sites. This research will provide a better fundamental understanding of drug-induced plasticities in the CNS and might present possible avenues for therapeutic pharmacological interventions.