Optogenetic Targeting of mGluR5 Receptor Signaling

描述（由申请人提供）：5型代谢型谷氨酸受体（mGluR 5）在药物成瘾的各个方面发挥关键作用。mGluR 5受体的药理学或遗传失活减少了药物奖赏、强化和复发样行为。另一方面，经由正变构调节剂（PAM）的mGluR 5功能的药理学增强产生促认知作用，例如促进消退学习和逆转药物诱导的认知障碍。然而，mGluR 5受体的药理学活化的各种挑战和限制显著限制了其作为详细研究药物成瘾中mGluR 5功能的研究工具的效用。这些限制包括时空控制差和缺乏受体活化的细胞类型特异性、正构mGluR 5激动剂的非选择性和mGluR 5 PAM的次优理化性质。这些技术限制可以通过光遗传学工具的成功开发来克服，所述光遗传学工具允许mGluR 5受体信号传导的精确时空和细胞类型特异性激活。我们最近开发并进行了表达光激活mGluR 5受体（OptoXR-mGluR 5）的慢病毒载体的初步表征。OptoXR-mGluR 5的表达受Ca 2 +/钙调蛋白依赖性激酶IIα（CaMKIIα）启动子的控制，允许在皮层神经元中选择性表达，该载体还编码增强型黄色荧光蛋白（eYFP）报告蛋白，用于表达的可视化。我们对这种慢病毒在大鼠大脑皮层体内的初步表征证明了它的细胞类型特异性和能力， 急性激活mGluR 5信号传导。然而，尚未评估OptoXR-mGluR 5激活在成瘾相关行为和认知功能障碍动物模型中的作用。此外，尚未探索重复刺激OptoXR-mGluR 5以持续激活mGluR 5信号传导的能力，这对于在数天或数周内进行的行为研究是必要的。因此，本申请中提出的研究的总体目标是检查药物诱导的认知功能障碍的啮齿动物模型中急性OptoXR-mGluR 5激活的行为效应，并开发适用于涉及重复刺激的行为范例的mGluR 5信号传导的光遗传学激活的新工具。这些目标将在两个独立的具体目标下实现。在具体目标1中，我们将评估OptoXR-mGluR 5急性激活逆转药物诱导的认知功能障碍的能力。在具体目标2中，我们将开发用于重复激活mGluR 5信号传导的新型光遗传学工具。成功开发用于mGluR 5激活的光遗传学工具将允许对这些受体在药物成瘾的各个方面的作用进行前所未有的研究。用于mGluR 5激活的光遗传学工具还可以为涉及mGluR 5受体的其他神经精神疾病（包括阿尔茨海默病、脆性X综合征和精神分裂症）提供对病理生理学机制和新的治疗途径的洞察。

DESCRIPTION (provided by applicant): The type 5 metabotropic glutamate receptor (mGluR5) plays a critical role in various aspects of drug addiction. Pharmacological or genetic inactivation of mGluR5 receptors reduces drug reward, reinforcement, and relapse- like behaviors. On the other hand, pharmacological potentiation of mGluR5 function via positive allosteric modulators (PAMs) produces pro-cognitive effects such as facilitated extinction learning and reversal of drug- induced cognitive impairments. However, various challenges and limitations to pharmacological activation of mGluR5 receptors significantly limit its utility as a research tool for detailed investigation of mGluR5 function in drug addiction. Such limitations include poor spatiotemporal control and lack of cell-type specificity of receptor activation, non-selectivity of orthosteric mGluR5 agonists, and suboptimal physiochemical properties of mGluR5 PAMs. These technical limitations can be surmounted by the successful development of optogenetic tools that allow for precise spatiotemporal and cell-type specific activation of mGluR5 receptor signaling. We have recently developed and performed a preliminary characterization of a lentiviral vector that expresses light- activated mGluR5 receptor (OptoXR-mGluR5). The expression of OptoXR-mGluR5 is under the control of Ca2+/calmodulin-dependent kinase IIα (CaMKIIα) promoter, allowing for selective expression in cortical glutamatergic neurons, and the vector also encodes enhanced yellow fluorescent protein (eYFP) reporter protein for visualization of expression. Our preliminary characterization of this lentivirus in the rat cerebral cortex in vivo demonstrates its cell-type specificity and ability to acutely activate mGluR5 signaling. However, the effects of OptoXR-mGluR5 activation in animal models of addiction-related behaviors and cognitive dysfunction have not yet been assessed. In addition, the ability of repeated stimulation of OptoXR-mGluR5 to consistently activate mGluR5 signaling, which would be necessary for behavioral studies conducted over the course of days or weeks, has not yet been explored. Therefore, the overarching goals of the studies proposed in this application are to examine the behavioral effects of acute OptoXR-mGluR5 activation in a rodent model of drug-induced cognitive dysfunction, and to develop novel tools for optogenetic activation of mGluR5 signaling that are amenable to behavioral paradigms involving repeated stimulation. These goals will be achieved under two independent Specific Aims. In Specific Aim 1, we will assess the ability of acute activation of OptoXR-mGluR5 to reverse drug-induced cognitive dysfunction. In Specific Aim 2, we will develop novel optogenetic tools for repeated activation of mGluR5 signaling. Successful development of optogenetic tools for mGluR5 activation will allow for an unprecedented level of investigation into the role of these receptors n various aspects of drug addiction. Optogenetic tools for mGluR5 activation may also provide insight into pathophysiological mechanisms and novel therapeutic avenues for other neuropsychiatric disorders in which mGluR5 receptors are implicated, including Alzheimer's disease, Fragile X syndrome, and schizophrenia.