Genetic and Optogenetic Models to Dissect the Role of the Serotonin 1B Receptor

描述(由申请人提供)：攻击性和冲动是行为的两个未被充分研究的方面，是许多精神障碍的重要症状。5-羟色胺1B受体(5-HT1B R)已被药理学、结构性基因敲除小鼠模型和人类基因多态关联研究发现与攻击性和冲动行为有关。鉴于5-羟色胺能调节大脑发育的证据，5-HT1B R很可能在建立攻击和冲动行为的基础回路方面发挥发育作用。5-HT1B受体是一种抑制5-羟色胺能和非5-羟色胺能神经元轴突终末神经递质释放的GPCR，广泛分布于基底节、海马、丘脑、小脑和中缝。因此，需要一种诱导性和组织特异性的策略来描述5-HT1B受体调节攻击性和冲动性的敏感期和神经回路。因此，我们建立了一个允许5-HT1B R的时间和空间调节的小鼠系，在5-HT1B R(TeO1B)的启动子和编码区之间插入了一个四环素操作符(Teto)。受体结合和原位杂交表明，与129只WT小鼠相比，这些TetO1B小鼠在大脑中具有正常的基础5-HT1B R表达。将MitO1B小鼠与表达四环素依赖转录沉默蛋白(TTS)的转基因小鼠杂交，在肌动蛋白(无处不在的)、CaMKII(前脑特异的)或Pet(中缝特异的)启动子的控制下，产生组织特异性的和可诱导的5-HT1B R下调的小鼠系。使用TetO1B：：？-Actin-TTS+小鼠的初步行为分析显示，与结构性基因敲除小鼠相似的行为表型--即显著增加冲动攻击性。因此，我们有工具在空间和时间上操纵5-HT1B R的表达，并检查由此产生的对攻击和冲动行为的影响。使用光遗传学的后续研究将通过确定成年后攻击性和冲动回路中的哪些大脑区域可能在关键时期被干预改变来解决电路水平的机制。假说是，在发育的关键时期缺乏5HT1b受体会导致过度活跃的“攻击回路”，因此通过抑制这些过度活跃的大脑区域，可以挽救正常的表型。这是有可能使用光敏的抑制性氯泵，卤视紫质，通过病毒载体注射到目标脑区，随后通过光纤植入黄光激活。总体而言，了解5-HT1B受体如何调节攻击性和冲动的神经回路将对治疗有意义 几种神经精神障碍，包括成瘾、注意力缺陷多动障碍和双相情感障碍，以及自杀抑郁。

DESCRIPTION (provided by applicant): Aggression and impulsivity are two understudied aspects of behavior that are significant symptoms in a number of psychiatric disorders. The serotonin 1B receptor (5-HT1B R) has been implicated in aggressive and impulsive behavior using pharmacology, constitutive knock-out mouse models, and human gene polymorphism association studies. Given evidence of serotonergic modulation of brain development, it is likely that 5-HT1B R plays a developmental role in establishing circuits underlying aggressive and impulsive behavior. 5-HT1B R is a GPCR that inhibits neurotransmitter release in axon terminals of serotonergic and non-serotonergic neurons localized broadly in the basal ganglia, hippocampus, thalamus, cerebellum and raphe. Thus, an inducible and tissue specific strategy is necessary to delineate the sensitive period and neural circuits through which 5-HT1B Rs regulate aggression and impulsivity. We have therefore generated a mouse line that permits temporal and spatial regulation of 5-HT1B R. A tetracycline operator (tetO) has been inserted between the promoter and coding region of the 5-HT1B R (tetO1B). Receptor binding and in situ hybridization reveals that these tetO1B mice have normal basal 5-HT1B R expression in the brain compared to 129 WT mice. Crossing tetO1B mice to transgenic mice expressing the tetracycline-dependent transcriptional silencer (tTS) under the control of ¿-Actin (ubiquitous), CaMKII (forebrain-specific) or Pet (raphe-specific) promoters yields mouse lines with tissue specific and inducible knock-down of 5-HT1B R. Initial characterization with autoradiography and in situ hybridization reveals promoter-specific 5-HT1B R expression patterns that can be modulated with doxycycline administration. Preliminary behavioral assays using tetO1B:: ¿-Actin-tTS+ mice reveal a similar behavioral phenotype to the constitutive knock-out mouse-i.e., significantly increased impulsive aggression. Therefore, we have the tools to manipulate the expression of 5-HT1B R spatially and temporally and examine the resulting impact on aggressive and impulsive behaviors. Follow-up studies using optogenetics will address the circuit level mechanisms by determining which brain regions in the aggression and impulsivity circuitry in adulthood may have been altered by the intervention during the critical period. The hypothesis is that a lack of 5HT1B Rs during a critical period of development results in an overactive 'aggression circuit', and so by inhibiting these overactive brain regions, a normal phenotype can be rescued. This is possible using the light-sensitive inhibitory Cl- pump, halorhodopsin, introduced through viral vector injection into target brain regions, with subsequent activation by yellow light administered via fiber optic implant. Overall, understanding how 5-HT1B R mediates the neural circuits underlying aggression and impulsivity will have implications for the treatments of several neuropsychiatric disorders, including addiction, attention-deficit hyperactivity disorde and bipolar disorder, and suicidal depression.