Ethanol and Plasticity of Tripartite Synapses

描述(申请人提供)：现在广泛认识到，星形胶质细胞在控制神经元的兴奋性方面发挥着核心作用，并对与学习和形成新记忆相关的细胞机制做出了贡献。学习和记忆形成的可塑性涉及神经元上谷氨酸能突触的树突棘的变化，而星形胶质细胞在大多数成熟的棘突上形成三部分突触结构。星形胶质细胞调节细胞外谷氨酸水平，并分泌影响三部分突触结构功能和结构的神经递质、神经营养因子和蛋白水解酶。正是这种星形胶质细胞与谷氨酸能突触的紧密物理定位，使星形胶质细胞突起处于控制神经元兴奋性和突触可塑性的位置。此外，星形胶质细胞在谷氨酸能突触处的存在调节了脊髓的结构，并控制了海马CA1区锥体神经元的NMDA受体依赖性的可塑性。众所周知，长期酒精暴露于海马神经元会增加谷氨酸能突触上NMDA受体的数量，也有研究表明，这种增加可能促进了脊椎大小依赖肌动蛋白的增加。这些变化被认为是导致酒精依赖和耐受性的原因。然而，目前尚不清楚星形胶质细胞在调节与慢性酒精暴露和戒断相关的谷氨酸能突触的结构和功能可塑性方面起到了什么作用。在这个探索性的R21应用中，我们将研究慢性酒精诱导的三部分突触的结构重构。最重要的假设是，酒精暴露会诱导神经元和星形胶质细胞释放某些控制脊椎-星形胶质细胞结构和功能可塑性的蛋白酶。我们提出了三个具体的目标来测试我们提出的酒精诱导的三体突触结构可塑性模型：目标1将测试急性和慢性体外酒精暴露改变活细胞制剂中三部分突触的结构动力学的假说；目标2将测试长期体内酒精暴露改变三方突触的形态和密度的假说；以及目标3将测试酒精诱导的三体突触结构可塑性与作用于突触细胞外基质的蛋白酶的释放有关的假说。这些研究将利用新的双脂酸染料负载程序和荧光蛋白的慢病毒感染来可视化三方突触结构，并结合共聚焦成像和复杂的3D图像分析酒精诱导的脊柱-星形胶质细胞结构的变化。这项提议的广泛、长期的目标是确定星形胶质细胞是否有助于酒精诱导的谷氨酸能突触的可塑性，以及星形胶质细胞是否可以被确定为治疗酒精滥用和依赖的新的治疗靶点。 公共卫生相关性：酒精中毒的特征是对酒精的渴求和强迫性饮酒行为。因此，确定酒精暴露导致大脑异常和不适当的突触连接的过程可能会导致有效治疗酒精中毒和酒精相关障碍的新方法。

DESCRIPTION (provided by applicant): It is now widely recognized that astrocytes (astroglia) play a central role in controlling excitability of neurons and contribute to the cellular mechanisms associated with learning and the formation of new memories. The plasticity of learning and memory formation involve changes in dendritic spines at glutamatergic synapses on neurons, and astrocytes form tripartite synaptic structures at a majority of mature spines. Astrocytes regulate extracellular glutamate levels and secrete gliotransmitters, neurotrophic factors, and proteases that affect the function and structure of the tripartite synaptic structure. It is this close physical localization of astroglia with glutamatergic synapses that places the astroglial process in a position to control neuronal excitability and synaptic plasticity. Moreover, the presence of astrocytes at glutamatergic synapses modulated the structure of the spine and controls NMDA receptor-dependent metaplasticity of hippocampal CA1 pyramidal neurons. It is well- established that chronic exposure of alcohol to hippocampal neurons increases the number of NMDA receptors at glutamatergic synapses, and it has also been suggested that this increase may promote actin-dependent increases in the size of the spine. These changes are suggested to contribute to alcohol dependence and tolerance. However, it is unknown what role astrocytes play in regulating the structural and functional plasticity of glutamatergic synapses associated with chronic alcohol exposure and withdrawal. In this exploratory R21 application, we will examine chronic alcohol-induced structural remodeling of the tripartite synapse. The over-arching hypothesis is that alcohol exposure induces the release of certain proteases from both neurons and astroglia that control spine-astroglial structural and functional plasticity. Three specific aims are proposed that will test our proposed model of alcohol- induced plasticity of tripartite synaptic structure: Aim 1 will test the hypothesis that acute and chronic in-vitro alcohol exposure alters the structural dynamics of the tripartite synapse in a live-cell preparation; Aim 2 will test the hypothesis that chronic in-vivo ethanol exposure alters the morphology and density of tripartite synapses; and Aim 3 will test the hypothesis that ethanol-induced structural plasticity of the tripartite synapse is associated with the release of proteases that act upon the synaptic extracellular matrix. These studies will utilize novel diolistic dye loading procedures and lentiviral infection of fluorescent proteins to visualize tripartite synaptic structures coupled with confocal imaging and sophisticated 3D-image analysis of alcohol-induced alterations in spine-astrocyte structures. The broad, long term objectives of this proposal are to determine if astrocytes contribute to alcohol-induced plasticity of glutamatergic synapses and whether astrocytes can be identified as a novel therapeutic target for the treatment of alcohol abuse and dependence. PUBLIC HEALTH RELEVANCE: Alcoholism is characterized by craving for alcohol and compulsive alcohol-seeking behavior. Thus, determining the processes by which alcohol exposure leads to aberrant and inappropriate synaptic connections of the brain may lead to novel approaches to effective treatments of alcoholism and alcohol related disorders.