DETERMINATION OF DNA SEQUENCE OF NEUROPEPTIDE Y Y3-RECEPTOR

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In order to investigate a mechanism involved in the development of neurogenic pulmonary edema, we utilized three experiments: (1) cloning of neuropeptide Y Y3-receptor and determination of its DNA sequenc* (2) involvement of vascular endothelial cells; (3) action of central nervous system.We have reported that neuropeptide Y is a neurotransmitter, released from the sympathetic nerve terminal, which might increa* the pulmonary vascular permeability in the development of fibrin-induced pulmonary edema, a model of neurogenic pulmonary edema* So far, DNA sequence of NPY Y3-receptor has not been elucidated. Therefore it disturbs a generation of new drug. We extracted mR* and created cDNA. The cDNA was ligated to ZAP Express Vector, creating cDNA library. E.coli DNA was infected by phage which h* the cDNA. After incubation, E. coli successfully made plaques on agar plate. Blotted nitrocellulose membrane was treated with ant* NPY antibody and anti PYY antibody. Visualized with colorization, c … More loning is now under study.Secondly, we utilized rat aortic endothelial cells, making the cells alive in agar plates. Endothelial cells are now investigated f* detecting the vascular permeability with permeating molecules such as fluorescent molecule-binding albumin. At the same time, we * also studying the development of vascular endothelial cell growth factor (VEGF) which makes the vascular bed permeable to large * molecule, albumin. Venous anesthetics, ketamine, propofol, and thiopental, neurotransmitters, neuropeptide Y, norepinephrine and * acetylcholine, and other bioactive substances. Ketamine and thiopental have an activity to increase the release ofVEGF from the * vascular endothelial cells.We already reported that NO release in the medulla oblongata has enhanced the development of neurogenic pulmonary edema. * Since unilateral left-vagotomy increase the NO synthase 2 weeks later, we have examined the fibrin-induced pulmonary edema, obtain * the results demonstrating the inhibitory action of NO on the edema development. However, the increase in vascular permeability was * maximum at 2 weeks time point, but rather increased more at 4 weeks. Now we are studying the effects of 1-glutamate, an excitable * neurotransmitter, on the fibrin-induced pulmonary edema. Less