Effects of TrkB Activation on Abnormalities in Neocortical FS Interneurons

描述(申请人提供)：创伤性脑损伤(TBI)导致大脑皮层异常、感觉和运动功能障碍、认知异常和癫痫。在最近的冲突中从严重脑震荡损伤中幸存下来的大量个体可能罹患癫痫，这强调了理解潜在的病理生理过程和制定预防策略的必要性(Garga和Lowenstein，2006年)。该项目的一个目标是获得有关可改善或预防这类创伤后异常情况的方法的基本信息。损伤释放化学递质GABA的神经细胞是创伤性脑损伤的常见结果。改善这些被称为“中间神经元”的抑制性细胞的结构和功能，可能会预防包括癫痫在内的一些损伤后果。初步结果表明，皮层中最常见的中间神经元类型--快峰抑制中间神经元，在部分切断与周围大脑的连接所致的皮质损伤区域，存在神经突起异常和GABA释放缺陷。由于这一缺陷和其他缺陷，大脑皮质变得过度兴奋，经常产生类似于人类局灶性癫痫的脑电活动的癫痫样电活动。神经营养蛋白BDNF及其受体TrkB对中间神经元的发育、生长和维持起重要作用，并在损伤区域减少，导致假说TrkB激活可能纠正FS或其他中间神经元的异常，改善受损皮质的功能。为了验证这一假说，将在麻醉的啮齿动物和用新设计的“小”分子LM22A-4治疗的动物身上进行切开，这种小分子进入大脑并激活TrkB受体。治疗2周后，再次麻醉大鼠，采用标准的体外切片和膜片钳技术，从药物处理动物和生理盐水对照组的损伤区域获得单个中间神经元和兴奋细胞的记录。单个神经细胞的活动 大量的神经元(“场电位”)将用适当的软件进行分析。激光激活的兴奋性化学递质谷氨酸的释放将被用来绘制单个切片内神经网络中兴奋性和抑制性连接的变化，以及慢性LM22A-4治疗的影响。单细胞的结构将在填充一种名为生物细胞素的标记，用抗体染色切片，并使用计算机控制的显微成像技术，包括光学和共聚焦显微镜后进行测量。这些实验的目的是确定激活TrkB受体是否会改善FS中间神经元的解剖和功能异常，恢复GABA的正常释放，并有利地影响受损皮质的神经回路。

DESCRIPTION (provided by applicant): Traumatic brain injury (TBI) results in abnormalities in cerebral cortex and disorders of sensory and motor function, cognitive abnormalities and epilepsy. The potential development of epilepsy by the large number of individuals, who have survived severe concussive injury during recent conflicts, emphasizes the need for understanding the underlying pathophysiological processes and the development of prophylactic strategies (Garga and Lowenstein, 2006). A goal of this project is to obtain basic information on approaches that may improve or prevent such posttraumatic abnormalities. Injury to nerve cells that release the chemical transmitter GABA is a common result of TBI. Improvement in the structure and function of these inhibitory cells, "interneurons", may prevent some of the consequences of injury, including epilepsy. Preliminary results show that fast-spiking (FS) inhibitory interneurons, the most common type of interneuron in cortex, have abnormal nerve processes and defects in releasing GABA in areas of cortical injury produced by partially cutting connections with surrounding brain ("undercuts"). Undercut cortex becomes hyperexcitable due to this and other defects and often generates epileptiform electrical activity that resembles EEG activity in human focal epilepsy. A neurotrophic protein BDNF, and its receptor TrkB, are important for development, growth and maintenance of interneurons, and are reduced in the injured area, leading to the hypothesis that TrkB activation may correct abnormalities in FS or other interneurons and improve function in the injured cortex. To test this hypothesis, undercuts will be made in anesthetized rodents, and animals treated with a newly- engineered "small" molecule, LM22A-4, that enters the brain and activates the TrkB receptor. After treatment for ~2 weeks, rodents are re-anesthetized, and standard in vitro slice and patch clamp techniques used to obtain recordings from single interneurons and excitatory cells in areas of injury from drug-treated animals and saline controls. Activities of individual nerve cells and large groups of neurons ("field potentials") will be analyzed with appropriate software. Laser-activated release of the excitatory chemical transmitter, glutamate, will be used to map changes in excitatory and inhibitory connections in neural networks within individual slices, and effects of chronic LM22A-4 treatment. The structure of single cells will be measured after filling them with a marker called biocytin, staining slices with antibodies, and using computer-controlled microscopic imaging techniques, including light and confocal microscopy. The aims of these experiments are to determine whether activation of the TrkB receptor will improve the anatomical and functional abnormalities in FS interneurons, restore normal release of GABA and favorably affect nerve circuits in the injured cortex.