BRAIN GALANIN RECEPTORS AND EPILEPSY

描述（由申请人提供）：本项目的长期目标是了解神经肽甘丙肽抑制癫痫发作活动的机制。我们的实验室是第一个表明癫痫发作会改变海马体中的甘丙肽，并且甘丙肽在癫痫持续状态中具有深刻的神经保护作用。该项目有三个目标。首先，我们将开发一种新的方法来下调甘丙肽受体在幼稚动物，以研究甘丙肽受体在癫痫发作中的作用。这将通过开发和优化针对甘丙肽受体亚型1和2的反义肽核酸（PNA）来实现。PNA对甘丙肽受体的影响将通过研究甘丙肽受体结合、免疫细胞化学和Western印迹法在甘丙肽受体转染的细胞系和原代神经元培养物中进行检查。其次，我们将测试的假设，下调甘丙肽受体亚型在大脑中选择性地增加动物的癫痫发作的倾向，并增加癫痫相关的神经元损伤。我们将研究幼稚和甘丙肽受体PNA治疗的大鼠之间的差异，在自我持续癫痫持续状态期间的癫痫发作活动的启动和维持，并将比较控制和PNA治疗的大鼠之间的神经元损伤的程度。第三，我们将研究潜在的甘丙肽受体刺激的神经保护作用的机制。具体而言，我们将研究甘丙肽受体PNA治疗如何影响第二信使系统（环磷酸腺苷，磷酸肌醇营业额，促分裂原活化蛋白激酶活性，兴奋性氨基酸谷氨酸释放），并将这些影响与PNA对癫痫持续状态的影响相关联。拟议的项目将产生重要的数据，了解参与内源性抑制癫痫发作的大脑机制，并将证明使用甘丙肽受体的活性配体作为治疗癫痫的新方法。

DESCRIPTION (provided by applicant): The long-term objective of this project is to understand the mechanisms involved in inhibition of seizure activity by the neuropeptide galanin. Our lab was the first to show that seizures alter galanin in the hippocampus and that galanin has a profound seizure-protective effect in status epilepticus. The project has three goals. First, we will develop a novel approach to down-regulate galanin receptors in naive animals, in order to study the role of galanin receptors in seizures. This will be achieved by developing and optimizing peptide nucleic acid (PNA) antisense targeted against galanin receptor subtypes 1 and 2. The effects of PNA on galanin receptors will be examined by studying galanin receptor binding, immunocytochemistry, and Western blot in galanin receptor-transfected cell lines and primary neuronal cultures. Second we will test the hypothesis that down-regulation of galanin receptor subtypes in the brain selectively increases animals' predisposition to seizures, and increases seizure-related neuronal injury. We will examine the differences between naive and galanin receptor PNA-treated rats in the initiation and maintenance of seizure activity during self-sustaining status epilepticus, and will compare the degree of neuronal injury between control and PNA-treated rats. Third, we will study putative mechanisms that underlie seizure-protective effects of galanin receptor stimulation. Specifically, we will study how galanin receptor PNA treatment affects second messenger systems (cyclic adenosine monophosphate, phosphoinisitide turnover, mitogen-activated protein kinase activity, excitatory amino acid glutamate release) and will correlate these effects with the effects of PNA on status epilepicus. The proposed project will yield important data for understanding brain mechanisms that participate in endogenous suppression of seizures, and will justify the use of active ligands of galanin receptors as a new approach for the treatment of epilepsy.