Nitrergic interneurons and cue-induced cocaine seeking

项目总结 易复发仍然是治疗成瘾的一个重大临床障碍。来自操作员的数据 成瘾和复发的啮齿动物模型表明，暴露于可卡因条件下的线索唤起了明显的 伏隔核核心(NAcore)释放谷氨酸。提示谷氨酸释放与特定细胞类型有关 NAcore中棘神经元(MSN)的瞬时突触增强，这在提示过程中不会发生 寻找蔗糖。这种瞬时突触增强包括增加突触和结构的可塑性。 MSNS。重要的是，这种可塑性的大小与复发行为呈正相关。我们假设两者都是 结构成分(树突棘头部扩张)和突触成分(增加的插入 这种可塑性的谷氨酸受体)由一氧化氮(NO)参与；一氧化氮(NO)是一种由 表达神经元型一氧化氮合酶(NNOS)的中间神经元。与这些神经元在脑内的作用相一致 提示复发，我们最近证明，NAcore中NO释放的增加也与 线索可卡因寻找过程中谷氨酸的释放。拟议研究的目标是揭示以下方面的投入 复发时释放NO所需的NAcore，以确定NAcore nNOS神经元上的哪些受体 调节NO的释放和提示的可卡因寻找，并揭示nNOS如何参与诱导 MSN的暂时性结构和突触可塑性，驱动线索可卡因寻找。在目标1中，我们将使用 在提示可卡因期间记录谷氨酸和NO释放时对NAcore输入的化学发生抑制 寻找。我们预测，对NAcore的输入携带谨慎的线索和上下文显著程度将 辨证调控NO释放，提示复发。在目标2中，我们将阐明谷氨酸和多巴胺是如何 NAcore nNOS中间神经元上特异表达的受体协同作用调节NO的产生和IF 它们是寻找可卡因线索所必需的。为此，我们将使用在nNOS中表达Cre的转基因小鼠 神经元、依赖Cre的shRNA病毒载体以及可卡因自我给药和线索可卡因寻找 审判。我们预测nNOS神经元中的谷氨酸和多巴胺受体系统协同调节NO 释放并提示寻找可卡因。在目标3中，我们将确定NAcore中nNOS的丢失是否会阻止CUE- MSN中介导的突触和结构可塑性。为此，我们将使用shRNA载体来敲除nNOS NAcore中表达MSN的D1或D2受体的病毒标记。这将使用以下工具完成 分别由D1和D2启动子驱动的CRE大鼠队列。在目标3中，我们将测量形态和 电生理读数显示线索诱导的突触可塑性。我们预计nNOS的丢失将阻止 与复发有关的可塑性形式，主要见于D1MSN。总而言之，这些调查的结果 将揭示nNOS神经元如何将可卡因线索转化为复发行为的机制。 因此，通过阻止nNOS神经元的提示激活来防止这种复发的信号转导 代表了一种潜在的治疗策略，以减少对药物的渴望和防止复发。

PROJECT SUMMARY Vulnerability to relapse remains a significant clinical hurdle in the treatment of addiction. Data from operant rodent models of addiction and relapse indicate that cocaine-conditioned cue exposure evokes a pronounced glutamate release in the nucleus accumbens core (NAcore). Cued glutamate release engages a cell type-specific transient synaptic potentiation in NAcore medium spiny neurons (MSNs), which does not occur during cued sucrose seeking. This transient synaptic potentiation consists of increased synaptic and structural plasticity in MSNs. Importantly, the magnitude of this plasticity positively correlates with relapse behavior. We posit that both the structural component (dendritic spine head expansion) and the synaptic component (increased insertion of glutamate receptors) of this plasticity are engaged by nitric oxide (NO); a gaseous transmitter produced by interneurons that express neuronal nitric oxide synthase (nNOS). Consistent with a role for these neurons in cued relapse, we have recently demonstrated that elevated NO release in the NAcore also occurs in parallel with glutamate release during cued cocaine seeking. The objectives of the proposed study are to reveal the inputs to the NAcore required for NO release during relapse, to determine which receptors on NAcore nNOS neurons regulate NO release and cued cocaine seeking, and to reveal how nNOS participates in the induction of the transient structural and synaptic plasticity in MSNs that drives cued cocaine seeking. In Aim 1, we will use chemogenetic inhibition of inputs to the NAcore while recording glutamate and NO release during cued cocaine seeking. We predict that inputs to the NAcore carrying discreet aspects of cue and contextual salience will differentially regulate NO release and cued relapse. In Aim 2, we will elucidate how glutamate and dopamine receptors, specifically expressed on NAcore nNOS interneurons, act in concert to regulate NO production and if they are required for cued cocaine seeking. To do this we will use transgenic mice that express Cre in nNOS neurons, Cre-dependent shRNA viral vectors, as well as cocaine self-administration and cued cocaine seeking trials. We predict that glutamate and dopamine receptor systems in nNOS neurons cooperatively regulate NO release and cued cocaine seeking. In Aim 3, we will determine if loss of nNOS in the NAcore will prevent cue- mediated synaptic and structural plasticity in MSNs. To do this we will use an shRNA vector to knockdown nNOS and concomitant viral labeling of D1 or D2 receptor expressing MSNs in the NAcore. This will be done using separate cohorts of D1-and D2-promoter driven Cre rats. In Aim 3, we will measure morphological and electrophysiological readouts synaptic plasticity induced by cues. We expect that loss of nNOS will prevent both forms of plasticity linked to relapse, predominantly in D1 MSNs. In conclusion, findings from these investigations will reveal the mechanistic aspects of how nNOS neurons translate cocaine cue exposure to relapse behavior. Accordingly, preventing this relapse signal transduction by preventing cued activation of nNOS neurons represents a potential therapeutic strategy to reduce drug craving and prevent relapse.