Hypercholesterolemia impairs reactive hyperemic vasodilation of 2A but not 3A arterioles in mouse cremaster muscle

Abstract

Hypercholesterolemia and atherosclerosis have been associated with changes in the microvasculature, in particular with endothelial dysfunction. In the present study, the impact of atherogenic conditions on arteriolar vasomotor control was determined. Arteriolar [second-order (2A) and third-order (3A) arterioles; diameter range: 9-37 microm] responses during reactive hyperemia (RH) were determined in cremaster muscle of anesthetized mice. C57Bl/6 mice on normal rodent chow were used as controls and high-fat/high-cholesterol (HFC)-fed C57Bl/6 and ApoE3-Leiden mice as hypercholesterolemic mice. The HFC diet resulted in time-dependent increases in plasma cholesterol and triglyceride concentrations (P < 0.001), which were more pronounced in ApoE3-Leiden mice (P < 0.001). In control mice, inhibition of nitric oxide (NO) synthesis with Nomega-nitro-L-arginine (L-NNA) reduced baseline diameter from 17.9 +/- 1.2 to 15.9 +/- 1.3 microm (P < 0.05) and decreased the duration of RH [time to 50% (t50) of recovery: 23.3 +/- 3.6 vs. 12.5 +/- 1.3 s (P = 0.003)]. t50 was longer in 2A versus 3A arterioles (33 +/- 3 vs. 18 +/- 2 s, P < 0.001) and increased with wall shear rate at the beginning of RH in 2A arterioles only. Compared with control mice, RH duration was reduced in 2A arterioles of HFC mice (t50: 11 +/- 2 s, P < 0.001 vs. control) but not affected in 3A vessels. L-NNA did not affect baseline diameter in HFC mice and reduced t50 only in "slow" responders (t50 > or = 10 s). It is concluded that hypercholesterolemia results in an impairment of NO-mediated vasomotor control in 2A but not 3A arterioles during dynamic changes of perfusion like RH. 2A arterioles likely therefore represent the functional locus of endothelial dysfunction during atherogenic conditions.

Heparin Impairs Glycocalyx Barrier Properties and Attenuates Shear Dependent Vasodilation in Mice

“…18,27 Shear dependent vasodilation was assessed in 2A arterioles of the cremaster muscle from the diameter and red blood cell velocity response during a reactive hyperemia (RH). 28 We found in a previous study that this maneuver elicits robust NO-mediated vasodilation in these vessels, with the duration of dilation being related to the wall shear rate (WSR) increase on release of the occlusion. 28 To test for a change in NO bioavailability after heparin administration, NO production was blocked with an L-arginine analogue, and involvement of an enhanced NO degradation by superoxide anions was checked by adding free radical scavengers to the muscle.…”

“…28 We found in a previous study that this maneuver elicits robust NO-mediated vasodilation in these vessels, with the duration of dilation being related to the wall shear rate (WSR) increase on release of the occlusion. 28 To test for a change in NO bioavailability after heparin administration, NO production was blocked with an L-arginine analogue, and involvement of an enhanced NO degradation by superoxide anions was checked by adding free radical scavengers to the muscle. For reference, additional experiments were performed in which hyaluronidase was infused.…”

“…The mouse was placed in a supine position on a custom-built animal platform, and the right cremaster muscle was prepared 28 (for details, see the online data supplement available at http://hyper.ahajournals. org).…”

“…In each experiment, 30-second occlusions of a first-order (1A) or second-order (2A) arteriole in the proximal and central region of the muscle were performed and responses during reactive hyperemia measured in the 2A arteriole distal from the occlusion. 28 One observation site was examined per experiment. Occlusions were performed every 10 to 15 minutes; this period of time between repetitive occlusions was sufficient for resting tone to recover while obtaining reproducible responses.…”

“…Hereafter, animals were given a bolus of saline, heparin, or hyaluronidase (as described earlier) after which measurements were resumed for another 90 minutes (saline and heparin) and 120 minutes (hyaluronidase). At 90 minutes postheparin, NO synthesis was blocked in 1 group of animals by adding N -nitro-L-arginine (L-NNA; 10 Ϫ4 mol/L; Sigma-Aldrich) to the physiological salt solution (PSS) superfusing the muscle, 28 whereas in a second group, SOD (50 U/mL; Sigma-Aldrich) and catalase (CAT; 50 U/mL; Sigma-Aldrich) were added to the physiological salt solution. 31 Arteriolar responses were re-evaluated Ն30 minutes after initial addition of the blocker or free radical scavenger.…”