NEURAL MECHANISMS OF CENTRAL CARDIOVASCULAR CONTROL

在定义和理解方面取得了令人印象深刻的进展 向脊交感神经节前投射的棘上通路 神经元(SPN)。相比之下，在以下方面进展甚微 另一组基本中枢神经系统通路的特征 参与交感神经流出的直接调节，以及 因此在控制心血管功能方面有一定作用。几乎没有什么是 已知可能的神经递质由释放，或位置 产生脊髓内环路的起源细胞 与交感节前神经元的单突触联系。这个 本申请集中在该方向上的努力，并且具体地 与识别可能存在的抑制通路有关 对心血管功能的正常调节和 在至少一个慢性的、感同身受的 多动症。使用(1)包埋前后的免疫过氧化物酶， 免疫荧光和免疫金免疫细胞化学程序；和(2) 跨突触和跨神经元标记方法如下 电子显微镜项目已经被提出。具体目标1 会检验甘氨酸是一种抑制性神经递质的假设 包含在接触交感神经节前神经元的终末。这个 项目将决定：(1)细胞内、躯体树突状细胞 甘氨酸受体样免疫反应性的定位(GlyR-LIR，93kd) 在四个自主神经核团内逆行标记的SPN内) 胸段脊髓(ILP、ILF、IC、CA)；(2)终末节段是否 相对的突触后GlyR-LIR呈现均匀的形态； 如果含有γ-氨基丁酸样免疫反应性的牛顿 (GABA-LIR)在SPN内与突触后GlyR-LIR相对。特定目标 2将检验这一假设，即脊髓V层和第V层的中间神经元 胸段脊髓长出主轴突或副轴突 到SPN的投影。这些研究将确定：(1)节段性和 脊髓中间神经元的节间(固有脊髓)分布 投影到已被逆行标记为以下两种之一的SPN 跨神经元转运的示踪物质：小麦胚芽凝集素 (WGA)或破伤风毒素的无毒结合片段，片段C(TTC)； 以及(2)WGA的层流和节段性分布 TTC标记的脊髓中间神经元在种群中移动或保持不变 对具有不同突触后靶点的SPN进行逆行标记。 特定目标3将检验跨神经元WGA-OR的假设 TTC标记的脊髓-SPN通路起源于丘脑的第V层和第VII层 胸段脊髓合成和释放抑制 神经递质甘氨酸或GABA和/或兴奋性 神经递质/神经调质P物质(SP)：DO WGA或TTC标记 中间神经元含有GABA、甘氨酸或SP-LIR？具体目标4将 检验破伤风患者心血管功能亢进的假说 是接触SPN的抑制性突触中毒的结果。 这些实验将确定跨突触标记的TTC ILF、ILP、IC和CA中SPN上的末端外显子：(1)含有GABA-LIR； 和/或(2)是相对的突触后GlyR-LIR。所有的实验都将是 在大鼠身上进行。

Impressive progress has been made in defining and understanding supraspinal pathways with projections to spinal sympathetic preganglionic neurons (SPNs). In contrast, there has been little progress in characterizing another fundamental set of central nervous system pathways involved in the direct regulation of the sympathetic outflow, and therefore in the control of cardiovascular function. Almost nothing is known about putative neurotransmitters released by, or the locations of the cells of origin which give rise to, intraspinal circuits with monosynaptic connections to sympathetic preganglionic neurons. The present application focuses efforts in this direction and is particularly concerned with the identification of inhibitory pathways that are likely to be critical for the normal regulation of cardiovascular function and which may be compromised in at least one example of chronic, sympathetic hyperactivity. Using (1) pre- and postembedding immunoperoxidase, immunofluorescence, and immunogold immunocytochemical procedures; and (2) trans-synaptic and transneuronal labeling methods, the following light and electron microscopic projects have been proposed. Specific Aim 1 would test the hypothesis that glycine is an inhibitory neurotransmitter contained in terminals contacting sympathetic preganglionic neurons. The projects would determine: (1) the intracellular, somadendritic localization of glycine receptor-like immunoreactivity (GlyR-LIR, 93 kd subunit) within retrogradely labeled SPNs in four autonomic nuclei in thoracic spinal cord (Ilp, Ilf, IC, and CA); (2) whether terminal boutons opposite postsynaptic GlyR-LIR exhibit a homogeneous morphology; and (3) if boutons containing gamma-aminobutyric acid-like immunoreactivity (GABA-LIR) are opposite postsynaptic GlyR-LIR within SPNs. Specific Aim 2 would test the hypothesis that spinal' interneurons in laminae V and VII of thoracic spinal cord give rise to principal or collateral axon projections to SPNs. The studies would determine: (1) the segmental and intersegmental (propriospinal) distributions of spinal interneurons projecting to SPNs that have been retrogradely labeled with one of two transneuronally transported tracer substances: wheat germ agglutinin (WGA) or the atoxic binding fragment of tetanus toxin, Fragment C (TTC); and (2) whether the laminar and segmental distributions of WGA- or TTC-labeled spinal interneurons shift or remain constant when populations of SPNs with different postsynaptic targets are retrogradely labeled. Specific Aim 3 would test the hypothesis that transneuronally WGA- or TTC-labeled spinal-SPN pathways originating in laminae V and VII of thoracic spinal cord synthesize and release the inhibitory neurotransmitters glycine or GABA, and/or the excitatory neurotransmitter/ neuromodulator substance P(SP): do WGA- or TTC-labeled interneurons contain GABA-, glycine-, or SP-LIR? Specific Aim 4 would test the hypothesis that cardiovascular hyperactivity manifest in tetanus is a consequence of intoxication of inhibitory synapses contacting SPNs. The experiments would establish whether trans-synaptically TTC-labeled terminal boutons on SPNs in Ilf, Ilp, IC and CA: (1) contain GABA-LIR; and/or (2) are opposite postsynaptic GlyR-LIR. All experiments would be performed in rats.