Introduction: Pericytes are contractile cells wrapped around capillaries. They can evoke capillary constriction and dilation. We have previously demonstrated the crucial role that cardiac pericytes play in ischaemia-induced no-reflow following coronary artery block (O'Farrell et al., 2017). Here, we report improvement of no-reflow after ischaemia following remote ischaemic preconditioning (RPc). RPc relaxes cardiac pericytes via a GLP-1R-mediated mechanism.

Methods: Pentobarbital anaesthetized Sprague-Dawley rats were subjected to 45 min LAD occlusion followed by 15 min reperfusion. RPc was induced by 15 min occlusion of both femoral arteries. Systemic GLP-1R blockade was achieved by i.v. injection of the specific antagonist Exendin(9-39) (50µg/kg, Tocris) prior to RPc. At the end of the experiment, rats were perfused with FITC-albumin, and cardiac capillary perfusion was analysed using FIJI software (ImageJ 1.53c, NIH) following immunostaining. For dissection of molecular mechanisms, live cardiac tissue from NG2-DsRed mice (right free ventricular wall) was perfused intralumenally with modified Tyrode’s solution via a glass cannula introduced into the right coronary artery, and imaged using a Zeiss confocal LSM780 microscope. The GLP-1R agonist Exendin-4 (Ex4, 100nM, Tocris), and/or the ATP-sensitive K⁺ (K_ATP) channel inhibitor glibenclamide (20µM, Insight Biotechnology) were applied in oxygenated Tyrode's solution. Oxygen/glucose deprivation was achieved by replacing glucose with sucrose in the Tyrode’s solution and bubbling with 95% N₂/5% CO₂. Capillary diameters were measured using FIJI software.

Results: LAD occlusion-reperfusion (n=5 rats) blocked 69±4% of cardiac capillaries near pericyte somata vs 9±4% in sham controls (n=6; p<0.001, Tukey multiple comparisons test), which resulted in a 40% reduction of perfused blood volume within the affected region. RPc prevented pericyte constriction and capillary blockage (reduced to 28±8%, n=5; p<0.01). Ex(9-39) prevented PRc-mediated relaxation of cardiac pericytes (64±10%, n=5). Ex vivo, 25 min of OGD resulted in cardiac pericyte constriction and a decrease of capillary diameter to 84.6±1.3% (n=24 capillaries in 7 mice) compared to the baseline diameter. GLP-1R activation with Ex4, applied from 25 min of a 50 min period of OGD, relaxed pericytes and dilated capillaries back to the baseline diameter (98.4±2.0%, n=9 capillaries in 4 mice, p<0.0001). In the presence of glibenclamide, GLP-1R-mediated pericyte relaxation was abolished, and capillaries remained constricted at 87.3±2.2% of their original diameter (n=13 capillaries in 3 mice, p<0.01).

Conclusion: Cardioprotective effects of GLP-1 agonists are mediated, at least in part, by relaxation of cardiac pericytes. The downstream molecular mechanism of GLP-1R activation involves opening of K_ATP channels. Cardiac pericytes are therefore a novel therapeutic target in ischaemic heart disease.