Vagus nerve stimulation modulates synaptic plasticity in the rat prefrontal cortex during the extinction of drug-seeking

药物使用导致形成强烈的暗示/奖励联系，这种联系在停止吸毒后长期存在， 会增加复发的长期风险。打破这些联系是物质治疗的一个重要目标 使用障碍。灭绝是一种学习形式，它抑制了对学习刺激的行为反应。调制 灭绝过程和巩固新形成的记忆具有重塑适应不良行为的临床潜力 防止复发我们最近发现，迷走神经刺激（VNS）促进灭绝学习， 减少可卡因自我给药大鼠中线索诱导的药物寻求恢复。这些变化与 改变了以边缘下（IL）皮层为中心的网络的活动。我们的初步数据还显示，吸毒 和恢复减少AMPA受体电流在第5层IL锥体神经元，和VNS逆转这些变化。 在消退过程中全身阻断脑源性神经营养因子（BDNF）的TrkB受体可消除VNS。 对提示诱导的恢复和IL中的神经递质传递的影响。我们假设灭绝训练 逆转药物诱导的IL中突触AMPAR的变化。配对灭绝与VNS导致（额外的）BDNF释放 这巩固了这些变化并减少了恢复。在这个应用程序中，我们将使用行为， 电生理学和细胞类型特异性形态学分析，以1）进一步研究时间和电路特异性 IL中导致药物寻求和复发的功能失调的神经适应，以及2）确定机制 通过VNS巩固消退记忆以减少复发。目标1中的实验将使用膜片钳 从基底外侧杏仁核（BLA）到IL的传入的电生理学和光遗传学刺激，以确定 这些输入是如何被药物寻求、灭绝和VNS改变的。我们将从2种类型进行电压钳记录 IL投射神经元（BLA和核壳）的变化，并确定VNS诱导的突触后神经元的变化。 谷氨酸受体功能和突触前释放在灭绝和恢复。这些实验将 在相同类型的IL投射神经元中进行形态学分析以确定VNS诱导的变化 特别是在通过恢复激活的细胞中（用活性标记物pCREB标记），以检验我们的假设， VNS优先调节与VNS配对的行为相关的网络。在目标2中，我们将确定 VNS诱导的灭绝巩固依赖于IL或其输入的内源性BDNF水平。为此，我们将 使用CRISPR/Cas9敲除IL内细胞或海马体细胞产生的BDNF， 分别投射到IL，以确定VNS诱发的BDNF如何调节消退和恢复。 目的3中的实验将确定VNS诱导的BDNF释放对突触可塑性变化的重要性， 调节复发的细胞形态。我们还将记录BLA和NAC壳中的体内局部场电位， 确定BDNF敲低如何影响VNS调节IL输出中突触可塑性的能力。采取 总之，我们的研究将提供关于可卡因和吗啡诱导的突触变化的重要新信息， IL，他们将确定VNS可以减少复发的神经递质和BDNF依赖性机制。

Drug use causes the formation of strong cue/reward associations which persist long after cessation of drug-taking and contribute to the long-term risk of relapse. Breaking these associations is an important goal in the treatment of substance use disorders. Extinction is a form of learning that inhibits behavioral responses to a learned stimulus. Modulating extinction processes and consolidating the newly-formed memories has clinical potential to reshape maladaptive behavior and to prevent relapse. We have recently shown that vagus nerve stimulation (VNS) facilitates extinction learning and reduces cue-induced reinstatement of drug-seeking in cocaine self-administering rats. These changes correlate with altered activity in a network that centers on the infralimbic (IL) cortex. Our preliminary data also show that drug-taking and reinstatement reduce AMPA-receptor currents at layer 5 IL pyramidal neurons, and that VNS reverses these changes. Systemic blockade of TrkB receptors for the brain-derived neurotrophic factor (BDNF) during extinction abolishes VNS’ effects on cue-induced reinstatement and on glutamatergic transmission in the IL. We hypothesize that extinction training reverses drug-induced changes in synaptic AMPARs in the IL. Pairing extinction with VNS leads to (additional) BDNF release which consolidates these changes and reduces reinstatement. In this application we will use a combination of behavioral, electrophysiological, and celltype-specific morphological analyses to 1) further investigate the time- and circuit-specific dysfunctional neuroadaptations in the IL that contribute to drug-seeking and relapse, and 2) to determine the mechanisms through which VNS consolidates extinction memory to reduce relapse. Experiments in Aim 1 will use patch-clamp electrophysiology and optogenetic stimulation of afferents from the basolateral amygdala (BLA) to the IL to determine how these inputs are altered by drug-seeking, extinction, and VNS. We will perform voltage-clamp recordings from 2 types of IL projection neurons (to the BLA and Nucleus accumbens shell) and determine VNS-induced changes in postsynaptic glutamate receptor function and presynaptic release during extinction and reinstatement. These experiments will be supported with morphological analyses in the same type of IL projection neurons to determine VNS-induced changes specifically in cells that were activated by reinstatement (labeled by the activity marker pCREB) to test our hypothesis that VNS preferentially modulates networks relevant in behaviors paired with VNS. In Aim 2 we will determine whether the VNS-induced consolidation of extinction depends on endogenous BDNF levels in the IL or its inputs. To this end we will use CRISPR/Cas9 to knock down BDNF produced either from cells within the IL or from cells in the hippocampus that project to the IL, respectively, in order to determine how VNS-evoked BDNF modulates extinction and reinstatement. Experiments in Aim 3 will determine the importance of VNS-induced BDNF release to changes in synaptic plasticity and morphology in cells that regulate relapse. We will also record in-vivo local field potentials in the BLA and the NACshell to determine how BDNF knockdown affects VNS’ ability to modulate synaptic plasticity in the outputs of the IL. Taken together, our studies will provide important novel information about cocaine- and extinction-induced synaptic changes in the IL, and they will identify glutamatergic and BDNF-dependent mechanisms through which VNS can reduce relapse.