Exercise and intravascular ATP elicit vasodilatation that is dependent on activation of inwardly rectifying potassium (K ) channels, with a modest reliance on nitric oxide (NO) and prostaglandin (PG) synthesis. Both exercise and intravascular ATP attenuate sympathetic α-adrenergic vasoconstriction (sympatholysis). However, K channels, NO, PGs and Na /K -ATPase activity are not obligatory to observe sympatholysis during exercise. To further determine similarities between exercise and intravascular ATP, we tested the hypothesis that inhibition of K channels, NO and PG synthesis, and Na /K -ATPase would not alter the ability of ATP to blunt α -adrenergic vasoconstriction. In healthy subjects, we measured forearm blood flow (Doppler ultrasound) and calculated changes in vascular conductance (FVC) to intra-arterial infusion of phenylephrine (PE; α -agonist) during ATP or control vasodilatator infusion, before and after K channel inhibition alone (barium chloride; n = 7; Protocol 1); NO (l-NMMA) and PG (ketorolac) inhibition alone, or combined NO, PGs, Na /K -ATPase (ouabain) and K channel inhibition (n = 6; Protocol 2). ATP attenuated PE-mediated vasoconstriction relative to adenosine (ADO) and sodium nitroprusside (SNP) (PE-mediated ΔFVC: ATP: -16 ± 2; ADO: -38 ± 6; SNP: -59 ± 6%; P < 0.05 vs. ADO and SNP). Blockade of K channels alone or combined with NO, PGs and Na /K -ATPase, attenuated ATP-mediated vasodilatation (∼35 and ∼60% respectively; P < 0.05 vs. control). However, ATP maintained the ability to blunt PE-mediated vasoconstriction (PE-mediated ΔFVC: K blockade alone: -6 ± 5%; combined blockade:-4 ± 14%; P > 0.05 vs. control). These findings demonstrate that intravascular ATP modulates α -adrenergic vasoconstriction via pathways independent of K channels, NO, PGs and Na /K -ATPase in humans, consistent with a role for endothelium-derived hyperpolarization in functional sympatholysis.

Key points The ability of contracting skeletal muscle to attenuate sympathetic vasoconstriction (functional sympatholysis) is critical for maintaining blood flow during exercise‐mediated sympathoexcitation. Functional sympatholysis and endothelial function are impaired with ageing, resulting in compromised blood flow and oxygen delivery to contracting skeletal muscle during exercise. In the present study, intra‐arterial infusion of ACh or ATP to augment endothelium‐dependent signalling during exercise attenuated α1‐adrenergic vasoconstriction in the contracting muscle of older adults. The vascular signalling mechanisms capable of functional sympatholysis are preserved in healthy ageing, and thus the age‐related impairment in functional sympatholysis probably results from the loss of a functional signal (e.g. plasma [ATP]) as opposed to an intrinsic endothelial dysfunction. Abstract The ability of contracting skeletal muscle to attenuate sympathetic α‐adrenergic vasoconstriction (‘functional sympatholysis’) is impaired with age. In young adults, increasing endothelium‐dependent vasodilatory signalling during mild exercise augments sympatholysis. In the present study, we tested the hypothesis that increasing endothelium‐dependent signalling during exercise in older adults can improve sympatholysis. In 16 older individuals (Protocol 1, n = 8; Protocol 2, n = 8), we measured forearm blood flow (Doppler ultrasound) and calculated changes in vascular conductance (FVC) to local intra‐arterial infusion of phenylephrine (PE; α1‐agonist) during (i) infusion of an endothelium‐dependent vasodilator alone (Protocol 1: ACh or Protocol 2: low dose ATP); (ii) mild handgrip exercise (5% maximum voluntary contraction; MVC); (iii) moderate handgrip exercise (15% MVC); and (iv) mild or moderate handgrip exercise + infusion of ACh or ATP to augment endothelium‐dependent signalling. PE caused robust vasoconstriction in resting skeletal muscle during control vasodilator infusions (ΔFVC: ACh: −31 ± 3 and ATP: −30 ± 4%). PE‐mediated vasoconstriction was not attenuated by mild or moderate intensity exercise (ΔFVC: 5% MVC: −30 ± 9; 15% MVC: −33 ± 8%; P > 0.05 vs. control ACh and ATP), indicative of impaired sympatholysis, and ACh or ATP infusion during mild exercise did not impact this response. However, augmentation of endothelium‐dependent signalling via infusion of ACh or ATP during moderate intensity exercise attenuated PE‐mediated vasoconstriction (ΔFVC: −13 ± 1 and −19 ± 5%, respectively; P < 0.05 vs. all conditions). Our findings demonstrate that, given a sufficient stimulus, endothelium‐dependent sympatholysis remains intact in older adults. Strategies aimed at activating such pathways represent a viable approach for improving sympatholysis and thus tissue blood flow and oxygen delivery in older adults.

Key pointsr In humans, the vasodilatory response to skeletal muscle contraction is mediated in part by activation of inwardly rectifying potassium (K IR ) channels. Evidence from animal models suggest that K IR channels serve as electrical amplifiers of endothelium-dependent hyperpolarization (EDH).r We found that skeletal muscle contraction amplifies vasodilatation to the endotheliumdependent agonist ACh, whereas there was no change in the vasodilatory response to sodium nitroprusside, an endothelium-independent nitric oxide donor.r Blockade of K IR channels reduced the exercise-induced amplification of ACh-mediated vasodilatation. Conversely, pharmacological activation of K IR channels in quiescent muscle via intra-arterial infusion of KCl independently amplified the vasodilatory response to ACh.r This study is the first in humans to demonstrate that specific endothelium-dependent vasodilatory signalling is amplified in the vasculature of contracting skeletal muscle and that K IR channels may serve as amplifiers of EDH-like vasodilatory signalling in humans.Abstract The local vasodilatory response to muscle contraction is due in part to the activation of inwardly rectifying potassium (K IR ) channels. Evidence from animal models suggest that K IR channels function as 'amplifiers' of endothelium-dependent vasodilators. We tested the hypothesis that contracting muscle selectively amplifies endothelium-dependent vasodilatation via activation of K IR channels. We measured forearm blood flow (Doppler ultrasound) and calculated changes in vascular conductance (FVC) to local intra-arterial infusion of ACh (endothelium-dependent dilator) during resting conditions, handgrip exercise (5% maximum voluntary contraction) or sodium nitroprusside (SNP; endothelium-independent dilator) which served as a high-flow control condition (n = 7, young healthy men and women). Trials were performed before and after blockade of K IR channels via infusion of barium chloride. Exercise augmented peak ACh-mediated vasodilatation ( FVC saline: 117 ± 14; exercise: 236 ± 21 ml min −1 (100 mmHg) −1 ; P < 0.05), C. M. Hearon and others J Physiol 597.5 whereas SNP did not impact ACh-mediated vasodilatation. Blockade of K IR channels attenuated the exercise-induced augmentation of ACh. In eight additional subjects, SNP was administered as the experimental dilator. In contrast to ACh, exercise did not alter SNP-mediated vasodilatation ( FVC saline: 158 ± 35; exercise: 121 ± 22 ml min −1 (100 mmHg) −1 ; n.s.). Finally, in a subset of six subjects, direct pharmacological activation of K IR channels in quiescent muscle via infusion of KCl amplified peak ACh-mediated vasodilatation ( FVC saline: 97 ± 15, KCl: 142 ± 16 ml min −1 (100 mmHg) −1 ; respectively; P < 0.05). These findings indicate that skeletal muscle contractions selectively amplify endothelium-dependent vasodilatory signalling via activation of K IR channels, and this may be an important mechanism contributing to the normal vasodilatory response to exercise in humans.

Aging is the primary risk factor for cardiovascular disease (CVD), and Rho‐kinase has emerged as an important contributor to the pathogenesis of CVD in preclinical and clinical studies. Therefore, we tested the hypothesis that acute, systemic Rho‐kinase inhibition would improve measures of CVD risk in older adults. We studied healthy young (YA; 25 ± 1 yrs; 6M/6F) and older adults (OA; 65 ± 1 yrs; 6M/7F) under control conditions (CON, intravenous saline) and during inhibition of Rho‐kinase via intravenous fasudil administration (60 mg) in a double‐blind, randomized, crossover design. In separate visits to the laboratory, saline and fasudil were infused over the course of 1 h, after which we measured blood pressure, carotid‐femoral pulse wave velocity (cfPWV; index of arterial stiffness), and central augmentation index (AIx; SphygmoCor XCEL). In a subset of participants (YA: 5M/5F; OA: 6M/3F), we assessed reactive hyperemia (RH) and flow‐mediated dilation (FMD) to evaluate microvascular function and conduit artery endothelial function, respectively. We measured brachial artery diameter and mean blood velocity (MBV; Doppler ultrasound) to calculate forearm vascular conductance (FVC; forearm blood flow/mean arterial pressure · 100). FMD was quantified as the percent change in brachial artery diameter following 5 min of forearm ischemia. Shear rate (8 · MBV/diameter) was expressed as area under the curve (SRAUC) from cuff release to peak diameter to quantify the stimulus for FMD. In OA, fasudil reduced systolic (SBP, 127 ± 4 vs. CON: 134 ± 4 mmHg, P < 0.05), diastolic (DBP, 76 ± 2 vs. CON: 79 ± 1 mmHg, P < 0.05), and mean arterial pressure (MAP, 93 ± 2 vs. CON: 97 ± 2 mmHg, P < 0.05), and BP remained lower 1 and 2 h postinfusion. Although fasudil initially reduced SBP (114 ± 2 vs. CON: 119 ± 2 mmHg, P < 0.05), DBP (68 ± 2 vs. CON: 72 ± 2 mmHg, P < 0.05), and MAP (84 ± 2 vs. CON: 88 ± 2 mmHg, P < 0.05) in YA, BP returned to control levels and was not different from CON at 1 or 2 h post‐infusion. Fasudil lowered cfPWV in OA (7.6 ± 0.4 vs. CON: 7.8 ± 0.4 m/s, P < 0.05) and tended to lower cfPWV in YA (5.2 ± 0.2 vs. CON: 5.4 ± 0.2 m/s, P = 0.07). However, AIx was unchanged by the fasudil treatment in both OA (27 ± 2 vs. CON: 31 ± 2%, P = NS) and YA (−2 ± 2 vs. CON: −5 ± 5%, P = NS). Baseline brachial diameter increased with fasudil in OA (3.29 ± 0.23 vs. CON: 3.16 ± 0.20 mm, P < 0.05), whereas fasudil had no effect on baseline diameter in YA (3.19 ± 0.19 vs. CON: 3.16 ± 0.17 mm, P = NS). Similarly, resting FVC was elevated under fasudil conditions in OA (36 ± 4 vs. CON: 25 ± 3 ml/min/100 mmHg, P < 0.05), while FVC was unchanged with fasudil in YA (36 ± 9 vs. CON: 32 ± 5 ml/min/100 mmHg, P = NS). Following forearm ischemia, there was no effect of fasudil on peak blood flow (OA, 288 ± 30 vs. CON: 286 ± 31 ml/min, P = NS; YA, 346 ± 65 vs. CON: 346 ± 45 ml/min, P = NS) or on FMD (OA, 6.2 ± 1.1 vs. CON: 7.0 ± 1.1%, P = NS; YA, 6.5 ± 1.1 vs. CON: 7.9 ± 1.4%, P = NS). Fasudil did not affect SRAUC (OA, 29 ± 5 vs. CON: 29 ± 4 s−1 ...