In vivo measurement of flow-mediated vasodilation in living rats using high-resolution ultrasound

Abstract

In humans, endothelial vasodilator function serves as a surrogate marker for cardiovascular health and is measured as changes in conduit artery diameter after temporary ischemia [flow-mediated dilation (FMD)]. Here we present an FMD-related approach to study femoral artery (FA) vasodilation in anesthetized rats. Diameter and Doppler flow were monitored in the FA. Using high-resolution ultrasound (35 MHz) and automated analysis software, we detected dose-dependent vasodilation using established endothelium-independent [intravenous nitroglycerin EC(50) = 3.3 x 10(-6) mol/l, peak 21Delta% (SD 4)] and endothelium-dependent [intra-arterial acetylcholine EC(50) = 1.3 x 10(-6) mol/l, peak 27Delta% (SD 4)] pharmacological vasodilators. Wall shear stress induced by intra-aortic injection of adenosine and infusion of saline at increasing rates (1.5-4.5 ml/min) led to vasodilation at 1 to 2 min. Transient hindlimb ischemia by common iliac occlusion (5 min) led to reactive hyperemia with flow velocity and wall shear stress increase and was followed by FA dilation [16Delta% (SD 2)], the latter of which was completely abolished by nitric oxide synthase (NOS) inhibition with N(G)-monomethyl-L-arginine [1Delta% (SD 2)]. FMD was significantly reduced in adult 20-24-wk-old animals compared with 9- to 10-wk-old animals, consistent with age-dependent endothelial dysfunction [16Delta% (SD 3) vs. 10Delta% (SD 3), P < 0.05]. Whereas FMD was completely NOS dependent in 9- to 10-wk-old animals, NOS-dependent mechanisms accounted for only half of the FMD in 20-24-wk-old animals, with the remainder being blocked by charybdotoxin and apamin, suggesting a contribution of endothelium-derived hyperpolarizing factor. To our knowledge, this is the first integrative physiological model to reproducibly study FMD of conduit arteries in living rats.

Paclitaxel-Induced Endothelial Dysfunction in Living Rats Is Prevented by Nicorandil via Reduction of Oxidative Stress

“…Measurement of FMD in rats is considered a useful experimental tool for estimating endothelial dysfunction in serious vascular diseases. We previously reported the impairment of FMD in streptozotocin-induced diabetic rats (16) according to the FMD method described in the report by Heiss et al (35). Endothelial function could be successfully assessed by in vivo FMD measurement in our current experiment because 5-min ischemia/reperfusion or intra-arterial acetylcholine induced femoral vasodilation without affecting the reactive hyperemia or NTG-induced endothelium-independent vasodilation in anesthetized rats.…”

Brief Exposure to Secondhand Smoke Reversibly Impairs Endothelial Vasodilatory Function

“…FMD was measured in living rats as we have described (Heiss et al, 2008b) by a blinded investigator. See Supplementary Material online for details.…”

“…To study how SHS affects endothelial function in a physiologically consistent study population, we used a method that we previously developed to measure FMD by micro-ultrasound in the hindlimbs of living rats (Heiss et al, 2008b). FMD measured in rats is similar to human FMD with respect to factors that impair or improve the response and underlying mechanisms, and has provided us with unprecedented insight into the roles played by mediators of NO bioavailability (Chen et al, 2013;Heiss et al, 2008b).…”

Central Role of eNOS in the Maintenance of Endothelial Homeostasis

“…The most important clinical and experimental methods to measure endothelial function, in particular flow-mediated vasodilation (FMD), will be outlined. FMD has emerged as the most important and most widely used non-invasive methodology to measure endothelial function and is now even available in rodents (41). We will discuss how genetic and environmental factors, in essence cardiovascular risk factors, induce endothelial dysfunction, promoting the initiation and progression of cardiovascular diseases (CVDs), and we will also critically review markers of endothelial injury and regenerative capacity.…”