Functional Analysis of SV2 by Targeted Gene Disruption

描述(申请人提供)：确定产生和调节神经递质释放的分子事件是分子神经生物学的中心目标之一。定义这些事件，以及它们是如何随着经验和疾病而改变的，将提供对学习等正常过程的理解，并作为开发治疗精神疾病、神经障碍和成瘾的新疗法的生物学基础。 这里提出的工作涉及突触囊泡蛋白2(SV2)的功能，它是所有神经递质(突触)囊泡的糖蛋白成分。在上一个资助阶段，我们证明了突触囊泡蛋白2(SV2)是神经传递的重要调节器。SV2A的缺失减少了海马神经元和皮质神经元的神经传递，并减少了嗜铬细胞中容易释放的囊泡池的大小。神经传递的减少不是由于形态的改变或形态上“停靠”的突触小泡的减少。综上所述，这些结果表明SV2是囊泡启动的正向调节器，它在囊泡附着在质膜上之后起作用。我们建议使用Sv2基因敲除小鼠来确定Sv2‘S在快速神经传递中的分子功能，并解决异构体功能上的差异。我们的具体目标是： 1.探讨SV2KO对培养的海马神经元神经传递功能的影响。 2.利用海马神经元外源性蛋白表达来验证SV2功能假说。 3.验证SV2B在带状突触维持和/或功能中起关键作用的假说。

DESCRIPTION (provided by applicant): Determining the molecular events that produce and regulate the release of neurotransmitters is one of the central goals of molecular neurobiology. Defining these events, and how they are altered with experience and in disease, will provide an understanding of normal processes such as learning and serve as the biological basis for developing new therapies for mental illness, neurological disorders and addiction. The work proposed here addresses the function of Synaptic Vesicle Protein 2 (SV2), a glycoprotein constituent of all neurotransmitter-containing (synaptic) vesicles. In the last funding period we demonstrated that Synaptic Vesicle Protein 2 (SV2) is an essential modulator of neurotransmission. Loss of SV2A reduces neurotransmission in hippocampal and cortical neurons, and reduces the size of the readily releasable pool of vesicles in chromaffin cells. Reduced neurotransmission is not due to morphological changes or to a decrease in morphologically "docked" synaptic vesicles. Together these results suggest that SV2 is a positive modulator of vesicle priming that acts after vesicle attachment at the plasma membrane. We propose to use SV2 knockout mice to determine SV2's molecular function in fast neurotransmission and to resolve differences in isoform functioning. Our specific aims are: 1. To determine how neurotransmission is altered in cultured hippocampal neurons from SV2 KOs. 2. To use exogeneous protein expression in hippocampal neurons to test hypotheses of SV2 function. 3. To test the hypothesis that SV2B plays a crucial role in ribbon synapse maintenance and/or functioning.