Spinal GABA Receptors in Neuropathic Pain

描述（由申请人提供）：脊髓中gaba介导的抑制丧失被认为有助于神经性疼痛的发展和维持。然而，支持数据出人意料地疲弱。此外，这些研究没有考虑GABA受体的复杂分子生物学，以及亚基组成和药理学如何在疾病状态和神经损伤后发生的重复性活动条件下发生变化。我们假设神经损伤引起脊髓中GABAA或GABAB受体的表达、分布和亚基组成的即时和长期变化。我们建议在传入疼痛通路中定义突触前和突触后元素的背景下描述这些变化，并建立一个具有良好特征的脊髓神经结扎模型，该模型显示神经性疼痛行为的时间依赖性变化。特别重要的是，神经损伤后，这些受体在未损伤、损伤的初级传入神经元和背角神经元上的表达和分子组成随时间的变化。Western blot检测结扎大鼠和假手术大鼠脊髓背角、同侧和对侧L4、L5 DRG中GABAA和GABAB受体亚基的表达和亚基组成的变化。采用双色间接免疫荧光法和体视学方法检测结扎和假手术大鼠L4和L5 DRG不同免疫组织化学鉴定的初级传入神经元群中GABAA和GABAB受体亚基的分布。免疫组织化学方法也将与结扎大鼠和假手术大鼠的脊髓丘脑和脊髓旁臂神经元逆行标记相结合，以确定神经损伤后脊髓背角突触后GABAA和GABAB受体的分布和组成是否也发生了改变。伤情分析的时间点将在伤后7 - 140天。这些研究将为GABAA和GABAB受体在幼龄动物的初级传入神经元和背角神经元上的分布提供新的信息。他们将描述神经损伤后这些受体的分布和分子组成如何随着时间的变化而变化，并为脊髓中抑制突触传递的分子和神经解剖学基础提供新的见解。

DESCRIPTION (provided by applicant): Loss of GABA-mediated inhibition in the spinal cord is proposed to contribute to the development and maintenance of neuropathic pain. However, the supporting data are surprisingly weak. Further, these studies have not considered the complex molecular biology of GABA receptors, and how subunit composition and pharmacology change in disease states and under conditions of repetitive activity as occurs after nerve injury. We hypothesize that nerve injury causes immediate, as well as long-term changes in the expression, distribution and subunit composition of GABAA or GABAB receptors in the spinal cord. We propose to characterize these changes in the context of defined presynaptic and postsynaptic elements of the afferent pain pathways and a well-characterized model of spinal nerve ligation that exhibits time-dependent changes in neuropathic pain behaviors. Of particular importance are the changes induced in the expression and molecular composition of these receptors on uninjured, and on injured primary afferent neurons and on dorsal horn neurons as a function of time after nerve injury. Changes in the expression and subunit composition of GABAA and GABAB receptor subunits in the spinal cord dorsal horn, and in ipsilateral and contralateral L4 and L5 DRG of ligated and sham rats will be determined by Western blot. Two-color indirect immunofluorescence methods will be used with stereological measurements to examine the distribution of GABAA and GABAB receptor subunits on different populations of immunohistochemically-identified primary afferent neurons in the L4 and L5 DRG of ligated and sham-operated rats. Immunohistochemical methods will also be used in conjunction with retrograde labeling of spinothalamic and spinoparabrachial neurons in ligated and sham rats to determine whether the distribution and composition of postsynaptic GABAA and GABAB receptors in the spinal cord dorsal horn is also altered after nerve injury. Time points for analysis will range 7 to 140 days after injury. These studies will provide new information about the distribution of GABAA and GABAB receptors on identified populations of primary afferent neurons and dorsal horn neurons in the naive animal. They will describe how the distribution and molecular composition of these receptors changes as a function of time after nerve injury and provide new insights into the molecular and neuroanatomical bases of inhibitorv svnaptic transmission in the spinal cord.