GABA transporter type 1 (GAT1) function in epilepsy

GABA 转运蛋白 1 型 (GAT1) 在癫痫中的功能

• 批准号: 8422868
• 负责人: Christopher Bruce Ransom
• 金额: --
• 依托单位: VA PUGET SOUND HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-01-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8422868
• 关键词: 3-aminobutyric acid; Acute; Affect; Afghanistan; Aging; American; Animals; Anions; Anticonvulsants; Antiepileptic Agents; Biological Assay; Brain; Caring; Cell physiology; Characteristics; Chemicals; Conflict (Psychology); Connexins; Data; Disease; Dose; Down-Regulation; Drug effect disorder; Electric Stimulation; Electroconvulsive Shock; Epilepsy; Extracellular Space; Failure; Frequencies; GABA Transporter 1; GABA transporter; Genetic; High Pressure Liquid Chromatography; Hippocampus (Brain); Iraq; Kainic Acid; Knock-out; Measurement; Mediating; Membrane Transport Proteins; Mentorship; Modeling; Molecular; Mus; Neurons; Neurotransmitters; Patients; Pentylenetetrazole; Pharmaceutical Preparations; Process; Publishing; Recurrence; Refractory; Research; Research Personnel; Residual state; Seizures; Severities; Signal Transduction; Slice; Status Epilepticus; Stroke; Surface; Synapses; Synaptic Vesicles; Testing; Therapeutic; Traumatic Brain Injury; Veratridine; Veterans; Vigabatrin; base; design; extracellular; gabapentin; gamma-Aminobutyric Acid; improved; in vivo; neuronal excitability; postsynaptic; receptor; relating to nervous system; research study; response; skills; synaptic inhibition; tiagabine; uptake

DESCRIPTION (provided by applicant): Epilepsy affects nearly 3 million Americans and approximately 30% of these patients continue to have seizures despite treatment with multiple antiepileptic drugs. Improved understanding of the cellular and molecular changes underlying seizures, and antiepileptic drug action, is essential to advance the care of veterans suffering from medically refractory epilepsy. Among the changes seen in epilepsy is a reduction of neuronal membrane transporters for the inhibitory neurotransmitter 3-aminobutyric acid (GABA) [GABA transporter type 1 (GAT1)] (During et al., 1995; Patrylo et al., 2001). Classically, GABA transporters were thought to function by removing GABA from the extracellular space, thereby terminating GABA actions. It is now clear that GABA transporters can also operate in the reverse direction to release GABA into the extracellular space, a process termed nonvesicular release. Nonvesicular GABA release is augmented by neuronal depolarization and alterations of ionic gradients, conditions that exist during epileptic seizures. Nonvesicular GABA release may be particularly important during prolonged seizures when vesicular GABA release at synapses is compromised by reductions in extracellular [Ca2+], GABA packaging into synaptic vesicles is limited by energy failure, and GABA synthesis is stimulated by low [ATP], causing an increase in cytoplasmic [GABA] that favors nonvesicular release (Erecinska et al., 1984; Hablitz and Heinemann, 1987; Allen et al., 2004). This proposal will test the hypothesis that nonvesicular GABA release has an important antiepileptic action and GAT1 downregulation in epilepsy is maladaptive. Specifically, it is hypothesized that the loss of nonvesicular GABA release mediated by GAT1 will exacerbate seizures despite elevated basal concentrations of ambient GABA. Consistent with this, studies on animals with genetic deletion of GABA transporter type 1 [GAT1 knockout (GAT1 KO)] have identified a heightened sensitivity to chemoconvulsants (Chiu et al., 2005). GAT1 function in epilepsy will be evaluated by three specific aims: Specific Aim 1 will test the hypothesis that nonvesicular GABA release in brain slices from GAT1 KO animals is reduced compared to wild type littermates. Specific Aim 2 will test the hypothesis that AEDs that increase GABA concentration or GAT1 surface expression will enhance nonvesicular release from wild type but not GAT1 KO neurons. Specific Aim 3 will test the hypothesis that nonvesicular release mediated by GAT1 affects seizure threshold in vivo and limits seizure frequency/severity in epileptic mice. Identification of changes in nonvesicular release in GAT1 knockout neurons and determination of AED effects on nonvesicular release in Specific Aims 1-2 will provide a basis for interpretation of in vivo data.

描述（由申请人提供）： 癫痫影响近300万美国人，尽管使用多种抗癫痫药物治疗，但这些患者中约30%仍继续癫痫发作。进一步了解癫痫发作的细胞和分子变化，以及抗癫痫药物的作用，对于促进患有难治性癫痫的退伍军人的护理至关重要。在癫痫中观察到的变化之一是抑制性神经递质3-氨基丁酸（GABA）[GABA转运蛋白1型（GAT 1）]的神经元膜转运蛋白的减少（During等人，1995; Patrylo等人，2001年）的报告。传统上，GABA转运蛋白被认为通过从细胞外空间去除GABA来起作用，从而终止GABA作用。现在清楚的是，GABA转运蛋白也可以在相反的方向将GABA释放到细胞外空间，这一过程称为非囊泡释放。非囊泡GABA释放增加神经元去极化和离子梯度的改变，癫痫发作期间存在的条件。当突触处的囊泡GABA释放受到细胞外[Ca 2 +]减少的损害时，非囊泡GABA释放在延长的癫痫发作期间可能特别重要，GABA包装到突触囊泡中受到能量衰竭的限制，并且GABA合成受到低[ATP]的刺激，导致有利于非囊泡释放的细胞质[GABA]增加（Erecinska et al.，1984; Hablitz和Heinemann，1987;艾伦等人，2004年）。这项建议将测试的假设，非囊泡GABA释放有重要的抗癫痫作用和GAT 1下调癫痫是适应不良。具体而言，假设由GAT 1介导的非囊泡GABA释放的丧失将加剧癫痫发作，尽管环境GABA的基础浓度升高。与此一致，对具有GABA转运蛋白1型[GAT 1敲除（GAT 1 KO）]基因缺失的动物的研究已经鉴定出对化学惊厥剂的敏感性提高（Chiu et al.，2005年）。将通过三个特定目的评价GAT 1在癫痫中的功能：特定目的1将检验GAT 1 KO动物脑切片中的非囊泡GABA释放与野生型同窝仔相比减少的假设。具体目标2将检验以下假设：增加GABA浓度或GAT 1表面表达的AED将增强野生型而非GAT 1 KO神经元的非囊泡释放。具体目标3将检验由GAT 1介导的非囊泡释放影响体内癫痫发作阈值并限制癫痫小鼠的癫痫发作频率/严重程度的假设。GAT 1基因敲除神经元中非囊泡释放的变化的鉴定和特定目的1-2中AED对非囊泡释放的影响的测定将为解释体内数据提供基础。