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 脊髓上通路与脊髓交感节前神经的投射 神经元（SPNs）。 相比之下， 表征了另一组基本的中枢神经系统通路 参与交感神经流出的直接调节， 从而控制心血管功能。 几乎毫无所 已知推定的神经递质释放，或位置 产生脊髓内回路的起源细胞， 交感节前神经元的单突触连接。 的 本申请将努力集中在这个方向上， 与识别抑制途径有关， 对心血管功能的正常调节至关重要， 这可能在至少一个慢性、交感神经性 多动症。 使用（1）植入前和植入后免疫过氧化物酶， 免疫荧光和免疫金免疫细胞化学方法;和（2） 跨突触和跨神经元标记方法，以下光 和电子显微镜项目已经被提出。 具体目标1 将检验甘氨酸是一种抑制性神经递质的假设 包含在接触交感节前神经元的终末中。 的 项目将确定：（1）细胞内，体细胞 甘氨酸受体样免疫反应性（GlyR-LIR，93 kd）的定位 亚单位）内逆行标记的SPNs在四个自主神经核， 胸脊髓（Ilp、Ilf、IC和CA）;（2）终末终扣是否 相反的突触后GlyR-LIR表现出均匀的形态;和（3） 如果含有γ-氨基丁酸样免疫反应性的终扣 （GABA-LIR）与SPN内的突触后GlyR-LIR相反。 具体目标 2将检验假设，脊髓'中间神经元在板V和 胸段脊髓的第VII神经元产生主轴突或侧支轴突 对SPN的预测。 这些研究将确定：（1）分段和 脊髓中间神经元的节段间（脊髓本体）分布 投射到已经被逆行标记为两个之一的SPN 经神经元转运的示踪物质：麦胚凝集素 (WGA)或破伤风毒素的无毒结合片段，片段C（TTC）; 以及（2）WGA-或WGA-的层流和节段分布是否 TTC标记的脊髓中间神经元移位或保持不变， 对具有不同突触后靶点的SPNs进行了逆行标记。 具体目标3将检验跨神经元WGA-或 TTC标记的脊髓-SPN通路起源于脊髓的V层和VII层， 胸脊髓合成和释放抑制性 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）：WGA或TTC标记 interneurons含有GABA-，甘氨酸-，或SP-LIR？具体目标4 验证心血管活动过度表现为破伤风的假设 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TTC标记的 Ilf、Ilp、IC和CA的SPN终末结：（1）含有GABA-LIR; 和/或（2）与突触后GlyR-LIR相对。 所有的实验都是 在老鼠身上进行。

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.