Abstract
Perivascular adipose tissue (PVAT) importantly affects the contractile function of conduit and resistance arteries. Some findings suggest that this effect of PVAT is controlled in part by sympathetic neural input directly to various PVAT depots. However, the degree of innervation of PVAT by the sympathetic nervous system remains in question. Studies of murine mesenteric PVAT suggest limited innervation by tyrosine hydroxylase (TH)-positive nerves. The purpose of the present study was to extend these studies to rat mesenteric PVAT, particularly in Dahl-salt sensitive (Dahl-SS) rats, an important model of hypertension after high-fat diet consumption. Whole-mounts of mesenteric PVAT, mesenteric resistance arteries or small intestine were fixed, blocked with donkey serum and incubated with primary antibodies directed against rat TH (rabbit) and for CD-31 (PECAM, mouse monoclonal) as a control for antibody penetration. After washing and re-blocking with donkey serum, tissues were incubated with fluorescently labeled secondary antibodies. Z-stacks of the fluorescently labeled tissues were then acquired and fluorescence of labeled structure quantified in background-subtracted, maximum-intensity z-projections after thresholding. Transmitted light images were also captured to measure the width of PVAT surrounding mesenteric resistance arteries and PVAT adipocyte diameters. In PVAT from male Sprague-Dawley rats and in male and female Dahl-SS rats we found limited innervation by TH-positive nerves with most TH-positive nerves tracking along small blood vessels in the PVAT. In contrast, the expected strong labeling of TH-positive nerves on the surface of mesenteric resistance arteries was observed. The low level of TH-labeling in PVAT was not due to lack of antibody penetration or inability to image, because CD-31 labeled blood vessels were readily detected. Both TH and CD-31 labeling were also always detected in small intestine whole mounts. These data support the hypothesis that there is sparse innervation of mesenteric PVAT by TH-positive nerves and suggest limited direct control of individual adipocytes by sympathetic nerves.