Hemichannels, astrocytic release, and neuropathic pain

描述(由申请者提供)：疼痛状况是美国的一个主要健康问题，导致数百万美国人的医疗发病率和生活质量下降。慢性神经性疼痛尤其难以治疗。一个在很大程度上尚未解决的挑战是，从急性疼痛到慢性神经病理性疼痛的转变是如何发生的，以及如何预防和逆转患者的这种转变。脊髓突触的可塑性和长时程增强(LTP)与慢性神经病理性疼痛的发生密切相关。越来越多的证据也表明，神经胶质细胞在神经病理性疼痛的发病机制中起着重要作用。星形胶质细胞是中枢神经系统中含量最丰富的细胞类型，维持着中枢神经系统的动态平衡。已有的研究表明，连接蛋白-43(Cx43)等星形细胞半通道是胶质递质释放的重要途径。虽然Cx43通常被认为调节星形胶质细胞之间的缝隙连接通讯，但在损伤条件下，这一功能可以通过ATP和谷氨酸释放切换到旁分泌信号。我们的中心假说是，神经损伤后半脑核介导的胶质递质释放通过调节脊髓突触的可塑性和LTP，有助于从急性疼痛向慢性神经病理性疼痛的转变。我们将通过解决以下四个具体目标来验证我们的中心假说：目标1将检验脊神经损伤增加脊髓星形胶质细胞释放谷氨酸和ATP的假说；目标2将检验Cx43介导的星形细胞ATP释放在脊髓神经损伤后小胶质细胞激活和小胶质细胞增生中起主要作用的假说；目标3将确定Cx43介导的星形细胞胶质递质释放在神经损伤后脊髓突触可塑性和长时程增强中的作用；目标4将确定星形细胞Cx43在神经损伤后神经病理性疼痛的发生和维持中的作用。这项建议将包括在纪万昌博士和Nedergaard博士之间建立创新的合作伙伴关系。纪万昌博士是一位疼痛科学家，擅长研究神经病理性疼痛中的神经元-神经胶质相互作用和神经可塑性，尼德加德博士擅长研究脊髓损伤和中风后星形细胞ATP和谷氨酸的释放。这一应用将采用多学科方法，包括使用带有转基因星形胶质细胞的可诱导转基因小鼠，对脊髓中ATP释放(生物发光)和小胶质细胞运动及钙变化(双光子)的体内成像，对神经损伤后诱发和持续的神经病理性疼痛的行为测试，以及脊髓的体外和体内电生理学。拟议的研究将提供由神经损伤启动的神经元-神经胶质细胞相互作用的逐步分析，并可能包括预防和治疗慢性疼痛的有效手段。 公共卫生相关性：神经损伤后脊髓星形胶质细胞功能障碍可能通过释放神经递质，如三磷酸腺苷和谷氨酸，促进神经病理性疼痛的发展。我们将采用多学科的方法，如转基因星形胶质细胞小鼠，双光子成像，诱发和持续疼痛的行为分析，以及体外和体内电生理学，以确定脊髓星形胶质细胞表达的连接蛋白-43半通道在调节突触可塑性和神经病理性疼痛中的作用。鉴于目前对神经病理性疼痛发生机制的认识尚不完全，对神经病理性疼痛的治疗还不够充分，本研究将为神经病理性疼痛从急性疼痛向慢性神经病理性疼痛的转变提供新的信息，为神经病理性疼痛的治疗提供新的策略。

DESCRIPTION (provided by applicant): Pain conditions are a major health problem in the US and lead to medical morbidity and a reduced quality of life for millions of Americans. Chronic neuropathic pain conditions are especially difficult to treat. A largely unaddressed challenge is how the transition from acute pain to chronic neuropathic pain occurs and how to prevent and reverse this transition in patients. Spinal cord synaptic plasticity and long-term potentiation (LTP) have been strongly implicated in chronic neuropathic pain development. Accumulating evidence also points to an important role of glial cells in the pathogenesis of neuropathic pain. Astrocytes are the most abundant cell type in the CNS and maintain the homeostasis of the CNS. It is well established that astrocytic hemichannels such as connexin-43 (Cx43) constitute an important pathway for gliotransmitter release. Although Cx43 was typically thought to regulate gap junction communication between astrocytes, this function could be switched to paracrine signaling via ATP and glutamate release under injury conditions. Our central hypothesis is hemichannels-mediated gliotransmitter release after nerve injury contributes to transition from acute pain to chronic neuropathic pain by modulating spinal cord synaptic plasticity and LTP. We will test our central hypothesis by addressing the following 4 specific aims: Aim 1 will test the hypothesis that spinal nerve injury increases glutamate and ATP release from spinal cord astrocytes; Aim 2 will test the hypothesis that Cx43-mediated astrocytic ATP release plays a chief role in microglia activation and microgliosis in the spinal cord after spinal nerve injury; Aim 3 will determine the role of Cx43-medicated astrocytic gliotransmitter release in spinal cord synaptic plasticity and LTP after nerve injury; Aim 4 will define the role o astrocytic Cx43 in neuropathic pain development and maintenance after nerve injury. This proposal will involve formation of an innovative partnership between Dr. Ji, a pain scientist with expertise in studying neuronal-glial interactions and neural plasticity in neuropathic pain, and Dr Nedergaard with expertise in studying astrocytic ATP and glutamate release after spinal cord injury and stroke. This application will employ a multidisciplinary approach including the use of inducible transgenic mice with genetically modified astrocytes, in vivo imaging of ATP release (bioluminescence) and microglia motility and Ca2+ changes (2-photon) in the spinal cord, behavioral testing of evoked and ongoing neuropathic pain after nerve injury, and ex vivo and in vivo electrophysiology in the spinal cord. The proposed studies will provide a step-by-step analysis of neuron- glia interactions initiated by nerve injury and may comprise an efficient means to prevent and treat chronic pain. PUBLIC HEALTH RELEVANCE: Dysfunction of spinal cord astrocytes after nerve injury may contribute to the development of neuropathic pain via releasing gliotransmitters such as ATP and glutamate. We will employ multidisciplinary approaches such as transgenic mice with genetically modified astrocytes, two-photon imaging, behavioral analyses of evoked and ongoing pain, and ex vivo and in vivo electrophysiology to define the role of connexin-43 hemichannels expressed by spinal astrocytes in regulating synaptic plasticity and neuropathic pain. Given the incomplete understanding of neuropathic pain mechanisms and insufficient treatment of neuropathic pain, this application will provide new information about transition of acute pain to chronic neuropathic pain and offer new strategy for neuropathic pain management.