Deficits in KCC2 activity and the pathophysiology of Autism spectrum disorders

 描述（由申请人提供）：自闭症谱系障碍（ASD）具有共同的核心症状，包括焦虑增加和重复行为，社交能力降低。人们普遍认为，ASD是由兴奋性和抑制性GABA能神经传递之间的“平衡”的细微差异引起的。电中性K+/Cl-共转运体2（KCC 2，或SLC 12 A5）在出生后选择性地在CNS中表达，并允许神经元维持低细胞内Cl-浓度，这是抑制性神经传递的出生后发育的基本先决条件。与其促进神经元抑制的作用一致，KCC 2表达的缺陷在ASD患者和多种自闭症动物模型中是明显的。KCC 2活性分别通过丝氨酸940和苏氨酸残基906和1007的磷酸化受到正和负调节。在这里，我们将直接测试这些调节过程是否影响GABA能抑制的出生后发育，以及ASD的病理生理学。为此，我们已经创建了小鼠，其中通过磷酸化对KCC 2活性的正向调节已经通过S940突变为丙氨酸（S940 A）而消除。T906/1007被无赖氨酸激酶（WNK）磷酸化，我们还获得了WNK活性缺陷的品系小鼠。使用这些新试剂的初步研究使我们能够制定一个新的假设，将在这里进行测试：出生后的KCC 2激活是由S940的相互磷酸化和T906/1007的去磷酸化促进的。损害这些过程会降低GABA能的功效，并直接导致ASD的病理生理学。我们将进行的实验来测试我们的假设在以下目标中详细说明。具体目标1。验证KCC 2中S940的磷酸化是GABA能抑制的出生后发展的关键决定因素的假设。具体目标2。验证WNK依赖性KCC 2磷酸化减缓超极化GABA能抑制的出生后发展的假设。具体目标3。为了验证这一假设，即减缓出生后GABA能抑制的发展再现了ASD的核心行为缺陷。总的来说，这些实验将为KCC 2磷酸依赖性调节的缺陷如何促进ASD的病理生理学提供关键的机制见解。这些信息可能会导致开发更有效的针对KCC 2活性的治疗方法，以最终改善ASD患者的预后。这些策略也可能与其他神经精神疾病有关，如癫痫、神经性疼痛和精神分裂症，其中KCC 2活性的缺陷被认为是重要的。

 DESCRIPTION (provided by applicant): Autism spectrum disorders (ASD) have common core symptoms which include increases in anxiety, and repetitive behaviors, with reduced sociability. It is widely believed that ASDs arise from subtle differences in the "equilibrium" between excitatory and inhibitory GABAergic neurotransmission. The electroneutral K+/Cl- co- transporter 2 (KCC2, or SLC12A5) is selectively expressed in the CNS after birth and allows neurons to maintain low intracellular Cl- concentrations an essential pre-requisite for the postnatal development of inhibitory neurotransmission. Consistent with its role in facilitating neuronal inhibition, deficits in KCC2 expression are evident in patients with ASDs and multiple autism animal models. KCC2 activity is subject to both positive and negative modulation via phosphorylation of serine's 940 and threonine residues 906 and 1007 respectively. Here, we will directly test if these regulatory processes influence the postnatal development of GABAergic inhibition, and the pathophysiology of ASDs. To do so we have created mice in which positive modulation of KCC2 activity by phosphorylation has been ablated via mutation of S940 to an alanine (S940A). T906/1007 are phosphorylated by with with-no-lysine kinases (WNKs), and we have also obtained line mouse lines with deficits in WNK activity. Preliminary studies using these new reagents have allowed us to formulate a novel hypothesis that will be tested here: The postnatal activation of KCC2 is facilitated by the reciprocal phosphorylation of S940 and dephosphorylation of T906/1007. Compromising these processes decreases the efficacy of GABAergic and directly contributes to the pathophysiology of ASDs. The experiments we will perform to test our hypothesis are detailed in the following aims. Specific Aim 1. To test the hypothesis that phosphorylation of S940 in KCC2 is a critical determinant for the postnatal development of GABAergic inhibition. Specific Aim 2. To test the hypothesis that WNK dependent phosphorylation of KCC2 slows the postnatal development of hyperpolarizing GABAergic inhibition. Specific Aim 3. To test the hypothesis that slowing the postnatal development of GABAergic inhibition reproduces the core behavioral deficits of ASDs. Collectively, these experiments will provide key mechanistic insights into how deficits in KCC2 phospho-dependent modulation contribute to the pathophysiology of ASD. This information may lead to the development of more effective therapies targeting KCC2 activity to ultimately improve patient outcomes for ASDs. Such strategies may also be relevant for other neuropsychiatric disorders, such as epilepsy, neuropathic pain, and schizophrenia in which deficits of KCC2 activity are believed to be of significance.