Mast cells are an important source of histamine (H) and are involved in several common pathological conditions including various urticarias, atopic eczema, and inflammation. Neurotensin (NT) has been shown to bind to specific receptor sites on the isolated mast cell surface, to elicit H release from isolated mast cells, to evaluate H levels in the blood when given intravenously, and to increase cutaneous vascular permeability. Preliminary experiments in our laboratories have demonstrated that very low concentrations of NT (Less than 10 to the -10M) can significantly increase H release from perfused slices of skin, that there is a means to form NT-related peptides in extracts of skin, and that these NT-related peptides can release H from isolated mast cells. It is the objective of this project to study in detail this NT-mast cell system in order to ascertain its possible physiological and/or pathological significance. Using perfused slices of skin and isolated peritoneal mast cells, the effect of NT on the release of H will be characterized in terms of ionic, metabolic, and pharmacologic requirements; the possible involvement of other effectors of H release in the response will be tested, whether additional mediators of inflammation accompany the H released by NT will be determined, and the cellular source of the released H will be confirmed. The chemical, immunochemical, and biological properties of the NT-related material that can be extracted from skin will be characterized. Finally, the effects of physiological and/or pathological stimuli to the skin on the level of these NT-related peptides and the level of H in the skin will be determined. Skin slices and mast cells will be isolated from adult rats. Histamine will be assayed fluorometrically and radioenzymatically. NT will be assayed by radioimmunoassay and by receptor binding-assay. NT-related material will be characterized by HPLC and biological activity assayed by ability to release H from skin slices and mast cells. Changes in blood flow and vascular permeability in vivo will be followed using radioactive microspheres and the escape of I125 albumin, respectively. The experiments proposed in this application will provide new and significant information concerning the biological role of NT and the relationship between this peptide and the mast cell in health as well as in disease states.
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