Modulation of Neocortical Interneuronal Function

描述(申请人提供)：控制大脑皮质兴奋性和抑制性突触传递的突触前机制在正常的信息处理中起着关键作用，也可能有助于各种大脑疾病的病理生理学，如认知能力下降和癫痫。这些实验主要集中在含有GABA的快速峰抑制神经元(FS)介导的抑制性突触传递以及钠-钾ATPase(NKA，钠泵)活性的调节。FS中间神经元介导最常见和最强烈的抑制形式，已知的是容易受到伤害。初步数据显示，在部分分离(下切或UC)的新皮质中，FS细胞轴突终末α3KA的免疫反应减弱。我们将测试假设，FS到兴奋性细胞GABA能传递将受到α3KA活性降低的影响，无论它发生在部分分离的致痫皮质，通过强心类固醇治疗，还是在具有基因改变的NKAα3亚单位的癫痫小鼠中。具体目的是1)评估强心激素或皮质损伤对泵电流的影响和FS对兴奋性细胞抑制的动力学；2)研究增强α3KA表达的功能效应。 3)测试假设，即由于α3泵亚单位的突变，FS GABA能神经元的特性和FS细胞的传递将因遗传诱导癫痫的Myshkin小鼠而选择性地改变。这些技术将包括在大脑皮层切片中配对记录，以获得由FS细胞选择性产生的单一(U)抑制突触后电流(IPSCs)；在小白蛋白/ChR2小鼠中采用光遗传学方法选择性激活FS细胞；以及同时进行激光扫描光刺激和多个同时细胞外记录，以监测网络效应。部分皮质隔离模型将用于提供慢性损伤的致痫切片，并评估可能导致过度兴奋的泵功能变化。我们的目标是识别影响GABA能传递的泵活动，以及可能导致过度兴奋和癫痫发生的泵功能损伤或遗传改变；并测试增强NKA表达是否将挽救UC皮质中受损的FS细胞介导的抑制。

DESCRIPTION (provided by applicant): Presynaptic mechanisms controlling excitatory and inhibitory synaptic transmission in the cerebral cortex have critical roles in normal information processing and also may contribute to the pathophysiology of a variety of brain disorders such as cognitive decline and epilepsy. The proposed experiments focus primarily on inhibitory synaptic transmission mediated by gamma amino butyric acid (GABA)-containing fast-spiking (FS) inhibitory interneurons and its modulation by activities of sodium-potassium ATPase (NKA, the sodium pump). FS interneurons mediate the most common and strongest form of inhibition and are known to be vulnerable to injury. Preliminary data show that immunoreactivity for α3KA, the main pump isoform in interneurons, is decreased in axonal terminals of FS cells in partially isolated (undercut or UC) neocortex. We will test the hypothesis that FS to excitatory cell GABAergic transmission will be compromised by decreases in α3KA activity, whether it occurs in partially isolated epileptogenic cortex, via treatment with cardiotonic steroids, or in epileptic mice with genetically altered NKA alpha3 subunit. Specific aims are to 1) assess effects of cardiotonic steroids or cortical injury on pump current and the dynamics of FS to excitatory cell inhibition; 2) study functional effects of enhancing α3KA expression in injured or Myshkin mouse cortex with a small molecule partial TrkB agonist; and 3) test the hypothesis that properties of FS GABAergic neurons and transmission from FS cells will be selectively altered in Myshkin mice with a genetically induced epilepsy due to mutations in the α3 pump subunit. Techniques will include paired recordings in cortical slices to obtain unitary (u) inhibitry postsynaptic currents (IPSCs) generated selectively by FS cells; optogenetic approaches in parvalbumin/ChR2 mice to selectively activate FS cells; and both laser scanning photostimulation and multiple simultaneous extracellular recordings to monitor effects network effects. The partial cortical isolation model will be used to provide chronically injured, epileptogenic slices and assess changes in pump function that might contribute to hyperexcitability. The goals are to identify pump activities that influence GABAergic transmission, abnormalities induced by injury or genetic alterations in pump function that may contribute to hyperexcitability and epileptogenesis; and test whether enhancing NKA expression will rescue compromised FS cell-mediated inhibition in the UC cortex.