NYP INTERACTION WITH MICROVASCULR ENDOTHELIUM

神经肽Y是1982年发现的一种由36个氨基酸组成的多肽 脑组织的提取物 自其被发现以来， 一直在积累，以支持其在监管中的作用， 心血管系统 大部分活动都被认为是 直接的血管收缩和突触后 增强其他血管收缩剂的作用。 其他报告的监管 神经肽Y调节脑干区域的功能， 心脏和血管成分的自主流出及其能力 调节去甲肾上腺素和其他递质的释放 血管周围神经和交感神经和内在神经在 心脏水平。 虽然有限，但有证据支持NPY 在内皮水平的调节。 在大血管内皮中 模型 一方面，已经观察到NPY增强白细胞 粘附以时间和浓度依赖性的方式，并在 另一方面改变前列环素的产生。 使用体外 牛毛细血管内皮细胞培养模型 我们已经报告了肾上腺髓质存在高亲和力 NPY及其类似物的特异性结合位点，并观察到， 所述神经肽能够诱导信号传导事件 与其他模型系统中的NPY受体激活有关。 而且我们 有初步的数据来支持一个可比的信号响应， 牛视网膜微血管组织（不含内皮） 来源于牛主动脉。 我们目前的目标是： 一个建议是有助于定义NPY相互作用的性质 与微血管内皮细胞相连。 我们的假设是 NPY受体优先分布于毛细血管内皮 并且它们的激活导致了 内皮源性血管扩张剂。 为了证明这个假设 我们提出的具体目标如下：（1） 评估NPY受体是否优先在其他细胞中表达。 微血管 这将通过执行放射性配体 与培养的内皮细胞的结合研究， 血管和几种组织的微血管系统。 (2)测试 假设NPY诱导NO和PGI 2的释放 微血管内皮通过检测基底和 在存在和不存在这些血管扩张剂的情况下， 的神经肽。 (3)评估NPY诱导的相互作用 钙反应与其他信号级联暗示在 调节内皮功能，特别是（a）参与 蛋白酪氨酸激酶途径的研究 蛋白激酶抑制剂的作用及其免疫检测 在NPY受体活化后磷酸化蛋白质。(b)的作用 Ca ~（2+）激活的K ~+通道的膜片钳表现 检测NPY对电生理的影响 与这些通道的表达相关联的参数。 （四） 测试NPY负偶联腺苷酸环化酶是否调节 NO和PGI_2的刺激释放，通过测量 这些药物在改变cAMP水平的操作后。 完全 这些研究应该提供深入了解神经肽Y的功能作用， 微血管内皮的水平扩展了我们对 对心血管系统的调节 神经肽

NPY is a 36 amino acid peptide originally discovered in 1982 romextracts of brain tissue. Since its identification, evidence has been accumulating to favor its role in the regulationof the cardiovascular system. Most of this activity has been ascribed to both direct vasoconstriction and its ability to post-synaptically potentiate other vasoconstrictors action. Other reported regulatory functions concern NPY modulationof brainstem areas controlling the autonomic outflow to cardiac and vacular components and its ability to moduate the release of norepinephrine and other transmitters from perivascular nerves and the sympathetic and intrinsic nerves at the heart level. Although limited, there is evidence to support a NPY modulation at the endothelial level. In large vessel endothelial models. NPY has been observed in one hand to potentiate leukocyte adhesion in a time and concentration dependent manner and in the other hand to modify prostacyclin production. Using an in vitro model of cultured capillary endothelial cells derived from bovine adrenal medulla we have reported the presence of high affinity specific binding sites for NPY and its analogs and have observed that the neuropeptide is capable of inducing signaling events associated with NPY receptor activation in other model systems. Moreover, we have preliminary data to support a comparable signaling responses in microvascular tissue from bovine retina but not with endothelium derived from bovine aorta. Our present goal with the following proposal is to contribute to define the nature of NPY interaction with the microvascular endothelium. Our general hypothesis is that NPY receptors are preferentially distributed in capillary endothelium and that their activation results inthe formation of endothelialderived vasodilators. In order to prove this hypothesis we are propsing to carry out the following specific aims: (1) Evaluate whether NPY receptors are preferentially expressed in other microvasculatures. This will be achieved by performing radioligand binding studies with cultured endothelial cells dervied from large vessels and the microvasculature of several tissues. (2) Test the hypothesis that NPY induces the release of NO and PGI2 from microvascular endothelium through the detection of the basal and stimulated release of these vasodilators in the presence and absence of the neuropeptide. (3) Evaluate the interaction of NPY induced calcium response with other signaling cascades implied in the modulation of endothelial function, specifically (a) the involvement of protein tyrosine kinase pathways through the assessment of the effect of proteinkinases inhibitors and the immunodetectionof phosphorylated proteins upon NPY receptor activation. (b) therole of Ca2+-activated K+ channels through the performance of patch-clamp experiments to detect the effect of NPY on the electrophysiologic parameters associated with the expression of these channels. (4) Test whether the NPY negative coupling to adenylate cyclase modulates the stimulated release of NO and PGI2 by measuring the relese of these agents after manipulations to alter cAMP levels. Altogether the studies should provide insight into the functional role of NPY at the level of the microvscular endothelium expanding our understanding of the regulation of the cardiovascular system provided by this neuropeptide.