Control of Fear Behavior by Brain Derived Neurotrophic F

控制恐惧行为的大脑回路已经有了很好的描述，然而，在各种类型的恐惧形成和表达过程中，这些回路中发生的分子事件还没有被很好地理解。对这些事件的了解将有助于开发控制和调节精神疾病造成的恐惧的药理学工具。 随着一代前脑限制性敲除脑源性神经营养因子(BDNF)的小鼠的出生，我们注意到这些动物表达了不充分的高水平恐惧。这种恐惧被记录为几个特征，包括对各种刺激的惊吓和冰冻反应。这些行为特征使BDNF基因敲除小鼠成为包括创伤后应激障碍在内的几种精神疾病的潜在模型，创伤后应激障碍的特征是惊吓反应增加。 这个项目的总体目标是了解大脑恐惧回路中的分子变化如何控制先天和后天的恐惧反应，并确定潜在的治疗目标来纠正异常的恐惧。具体目标是确定脑源性神经营养因子水平的降低如何导致恐惧反应的增加。为实现这一目标确立了以下目标。 1)确定大脑中缺乏BDNF会导致恐惧反应升高的区域。为了描绘这些回路，我们将尝试通过使用表达BDNF或Cre重组酶的慢病毒载体在大脑的不同区域重新引入或消除BDNF来挽救行为缺陷。主要感兴趣的区域包括杏仁核和纹状床核终末。 2)缺乏脑源性神经营养因子引起的基因表达变化的特征。为了找出差异表达的基因，我们比较了携带和不携带BDNF基因的动物的基因表达谱。 3)描述大脑恐惧控制区神经元的电特性。脑源性神经营养因子基因敲除小鼠杏仁基底外侧核和中央杏仁核兴奋性和抑制性神经元的单细胞记录。 4)在确定缺乏BDNF引起的分子和生理变化后，将进行抢救性实验以确定这些分子变化，这些变化直接导致恐惧回路中的生理变化。 上个财政年度完成了以下工作： 1)杏仁核被确定为BDNF基因敲除动物中调节恐惧反应变化的主要部位。杏仁核损伤消除了野生型和BDNF基因敲除动物之间的惊吓差异。 2)构建并检测了表达BDNF和Cre重组酶的慢病毒载体。这些病毒将被用来识别大脑中BDNF影响恐惧表达的特定位置。慢病毒Cre载体将用于BDNF基因的局部缺失，而BDNF载体将用于将BDNF导入到基因敲除动物的脑中。 3)确定了扁桃体立体定向注射慢病毒的适宜条件。 4)建立了单细胞电生理系统。对BDNFKO小鼠的初步分析显示，杏仁核中央核中的非适应性神经元的抑制性突触后电流增加。 5)生长抑素被认为是在前脑BDNF消除后降低其表达的基因。

Brain circuits controlling fear behaviors are well characterized, however molecular events taking place in those circuits during formation and expression of various types of fear are not well understood. Understanding of these events will be useful for development of pharmacological tools to control and modulate fears caused by psychiatric illnesses. Following generation of mice with forebrain restricted knockout of brain derived neurotrophic factor (BDNF), we have noticed that these animals express inadequately high levels of fear. This fear is registered as several features including elevated startle and freezing responses to various stimuli. These behavioral features make BDNF knockout mice a potential model of several psychiatric illnesses including post traumatic stress disorder, which is characterized by increased startle response. The general goal of this project is to understand how molecular changes in the fear circuits of the brain control innate and learned fear responses and to define potential therapeutic targets for correcting the abnormal fear. The specific goal is to determine how a reduction in the level of BDNF causes increased fear responses. The following aims are established to achieve this goal. 1) Identification the areas in the brain where absence of BDNF leads to elevated fear responses. To map those circuits we will try to rescue behavioral deficits by reintroducing or eliminating BDNF in various regions of the brain using lentiviral vectors expressing BDNF or Cre recombinase. The primary areas of interest include amygdala and bed nucleus stria terminal. 2) Characterization of changes in gene expression resulting from the absence of BDNF. To identify differentially expressed genes we are comparing gene expression profiles between the animals with and without BDNF gene. 3) Characterization of the electrical properties of neurons in the fear controlling areas of the brain. Single cell recording from both excitatory and inhibitory neurons in basolateral and central amygdala of mice with BDNF knockout. 4) Following identification of molecular and physiological changes caused by the lack of BDNF, rescue experiments will be performed to identify those molecular changes, which are directly responsible for the physiological alterations within the fear circuits. The following has been accomplished during the past fiscal year: 1) Amygdala was identified as a primary site mediating changes in fear responses in BDNF knockout animals. Amygdala lesions resulted in the elimination of the startle differences between the wild type and BDNF knockout animals. 2) We have created and tested lentiviral vectors expressing BDNF and Cre recombinase. These viruses will be used for identification of specific sites in the brain where BDNF is influencing expression of fear. The lentivirus Cre vector will be used for local deletion of BDNF gene, whereas the BDNF vector will be used to introduce BDNF into the brain of the knockout animals. 3) Proper conditions have been worked out for stereatactic injections of lentivirus into the amygdala. 4) Single cell electrophysiology system has been set up. Initial analysis of BDNF KO mice revealed increased inhibitory postsynaptic currents in non-accommodating neurons in the central nucleus of the amygdala. 5) Somatostatin has been identified as a gene reducing its expression following elimination of BDNF in the forebrain.