RNA Scope assays for enzymes of the catecholamine synthesis pathway

Abstract

The endothelium is an organ capable of regulating vascular tone through the release of endothelium-derived relaxing factors (nitric oxide and prostacyclin - PGI2) and endothelium-derived contracting factors (endothelin - ET1 and prostanoid - PGH2). In addition, the current physiological concept poses that such regulation can also occur through the release of exclusively neural-derived catecholamines. Our group has demonstrated in tissues isolated from reptiles that the endothelium is also a source of catecholamines, since the contraction of the snake aorta induced by electrical field stimulation (EFS) is abolished by adrenergic antagonists and by removing the endothelium. The observation that the endothelium could be a source of catecholamines was supported by the identification of the enzyme tyrosine hydroxylase in the snake endothelium by immunohistochemistry. Accidentally, this observation was also extended for human endothelial cells. As described in the project, the physiological and pathophysiological role of endothelial catecholamines in humans is being evaluated through in vitro pharmacological approaches using human arteries and veins, and an in vivo pharmacological approach through clinical trials. The aim of this grant application is to embase the observations obtained in in vitro and in vivo pharmacological tests with a morphological substrate through the identification and possible quantification of the metabolic pathway of catecholamines by immunohistochemistry and by RNA in situ hybridization with RNA Scope® technology. It is important to emphasize that this morphological approach can be expanded for research also in experimental models of laboratory animals. It is expected that the information collected will demonstrate the physiological and pathophysiological relevance of endothelial catecholamines in humans and will determine its importance in relation to neural adrenergic control, underpinning a new research field. (AU)