Daniel H. Craighead

Heat waves, aging and human cardiovascular health

This brief review is based on a Presidents Lecture presented at the Annual Meeting of the American College of Sports Medicine in 2013. The purpose of this review was to assess the effects of climate change and consequent increases in environmental heat stress on the aging cardiovascular system. The earths average global temperature is slowly but consistently increasing, and along with mean temperature changes come increases in heat wave frequency and severity. Extreme passive thermal stress resulting from prolonged elevations in ambient temperature and prolonged physical activity in hot environments creates a high demand on the left ventricle to pump blood to the skin to dissipate heat. Even healthy aging is accompanied by altered cardiovascular function, which limits the extent to which older individuals can maintain stroke volume, increase cardiac output, and increase skin blood flow when exposed to environmental extremes. In the elderly, the increased cardiovascular demand during heat waves is often fatal because of increased strain on an already compromised left ventricle. Not surprisingly, excess deaths during heat waves 1) occur predominantly in older individuals and 2) are overwhelmingly cardiovascular in origin. Increasing frequency and severity of heat waves coupled with a rapidly growing at-risk population dramatically increase the extent of future untoward health outcomes.

Topical menthol increases cutaneous blood flow

Menthol, the active ingredient in several topically applied analgesics, activates transient receptor potential melastatin 8 (TRPM8) receptors on sensory nerves and on the vasculature inducing a cooling sensation on the skin. Ilex paraguariensis is also a common ingredient in topical analgesics that has potential vasoactive properties and may alter the mechanisms of action of menthol. We sought to characterize the microvascular effects of topical menthol and ilex application and to determine the mechanism(s) through which these compounds may independently and combined alter cutaneous blood flow. We hypothesized that menthol would induce vasoconstriction and that ilex would not alter skin blood flow (SkBF). Three separate protocols were conducted to examine menthol and ilex-mediated changes in SkBF. In protocol 1, placebo, 4% menthol, 0.7% ilex, and combination menthol+ilex gels were applied separately to the skin and red cell flux was continuously measured utilizing laser speckle contrast imaging (LSCI). In protocol 2, seven concentrations of menthol gel (0.04%, 0.4%, 1%, 2%, 4%, 7%, 8%) were applied to the skin to model the dose-response curve. In protocol 3, placebo, menthol, ilex, and menthol+ilex gels were applied to skin under local thermal control (34°C) both with and without sensory nerve blockage (topical lidocaine 4%). Post-occlusive reactive hyperemia (PORH) and local heating (42°C) protocols were conducted to determine the relative contribution of endothelium derived hyperpolarizing factors (EDHFs)/sensory nerves and nitric oxide (NO), respectively. Red cell flux was normalized to mean arterial pressure expressed as cutaneous vascular conductance (CVC: flux·mmHg(-1)) in all protocols. Topical menthol application increased SkBF compared to placebo (3.41±0.33 vs 1.1±0.19CVC: p<0.001). During the dose-response, SkBF increased with increasing doses of menthol (main effect, p<0.05) with an ED50 of 1.0%. Similarly, SkBF was increased after menthol application during PORH (3.62±0.29 vs. 2.50±0.21flux·mmHg(-1); p<0.001), but not local heating (2.98±0.24 vs 2.86±0.32flux·mmHg(-1); p=0.44). Concurrent sensory nerve inhibition attenuated menthol-mediated vasodilation at thermoneutral baseline (1.29±0.19flux·mmHg(-1); p<0.001) and during PORH (2.79±0.28flux·mmHg(-1); p<0.001), but not during local heating (3.45±0.21flux·mmHg(-1); p=0.1). Topically applied menthol, but not ilex, dose-dependently increases blood flow in the cutaneous microvasculature. This increase in blood flow is mediated, in-part by sensory nerves and EDHFs.

Mechanisms and time course of menthol-induced cutaneous vasodilation

Menthol is a vasoactive compound that is widely used in topical analgesic agents. Menthol induces cutaneous vasodilation, however the underlying mechanisms are unknown. Determining the rates of appearance and clearance of menthol in the skin is important for optimizing topical treatment formulation and dosing. The purpose of this study was to determine the mechanisms contributing to menthol-mediated cutaneous vasodilation and to establish a time course for menthol appearance/clearance in the skin. Ten young (23±1years, 5 males 5 females) subjects participated in two protocols. In study 1, four intradermal microdialysis fibers were perfused with increasing doses of menthol (0.1-500mM) and inhibitors for nitric oxide (NO), endothelium derived hyperpolarizing factors (EDHFs), and sensory nerves. Skin blood flow was measured with laser Doppler flowmetry and normalized to %CVCmax. In study 2, two intradermal microdialysis fibers were perfused with lactated Ringers solution. 0.017mL·cm-2 of a 4% menthol gel was placed over each fiber. 5μL samples of dialysate from the microdialysis fibers were collected every 30min and analyzed for the presence of menthol with high performance gas chromatography/mass spectrometry. Skin blood flow (laser speckle contrast imaging) and subjective ratings of menthol sensation were simultaneously obtained with dialysate samples. In study 1, menthol induced cutaneous vasodilation at all doses ≥100mM (all p<0.05). However, inhibition of either NO, EDHFs, or sensory nerves fully inhibited menthol-mediated vasodilation (all p>0.05). In study 2, significant menthol was detected in dialysate 30min post menthol application (0.89ng, p=0.0002). Relative to baseline, cutaneous vasodilation was elevated from minutes 15-45 and ratings of menthol sensation were elevated from minute 5-60 post menthol application (all p<0.05). Menthol induces cutaneous vasodilation in the skin through multiple vasodilator pathways, including NO, EDHF, and sensory nerves. Topical menthol is detectable in the skin within 30min and is cleared by 60min. Skin blood flow and perceptual measures follow a similar time course as menthol appearance/clearance.

Blood pressure normalization via pharmacotherapy improves cutaneous microvascular function through NO-dependent and NO-independent mechanisms

Hypertension is associated with endothelial dysfunction and vascular remodeling.

Ingestion of transient receptor potential channel agonists attenuates exercise-induced muscle cramps

Introduction: Exercise‐associated muscle cramping (EAMC) is a poorly understood problem that is neuromuscular in origin. Ingestion of transient receptor potential (TRP) channel agonists has been efficacious in attenuating electrically induced muscle cramps. This study examines the effect of TRP agonist ingestion on voluntarily induced EAMC and motor function. Methods: Study 1: Thirty‐nine participants completed 2 trials after ingesting TRP agonist‐containing active treatment (A), or vehicle (V) control. Cramping in the triceps surae muscle was induced via voluntary isometric contraction. Study 2: After ingesting A or V, 31 participants performed kinematic and psychomotor tests of manual dexterity. Results: A increased precramp contraction duration (A, 36.9 ± 4.1 s; V, 27.8 ± 3.1 s), decreased cramp EMG area under the curve (A, 37.3 ± 7.7 %EMGmax·s; V, 77.2 ± 17.7 %EMGmax·s), increased contraction force to produce the cramp (A, 13.8 ± 1.8 kg; V, 9.9 ± 1.6 kg), and decreased postcramp soreness (A, 4.1 ± 0.3 arbitrary units (a.u.); V, 4.7 ± 0.3 a.u.). Kinematic and psychomotor tests were not affected. Discussion: TRP agonist ingestion attenuated EAMC characteristics without affecting motor function. Muscle Nerve 56: 379–385, 2017

Effects of vehicle microdialysis solutions on cutaneous vascular responses to local heating

Microdialysis is a minimally invasive technique often paired with laser Doppler flowmetry to examine cutaneous microvascular function, yet presents with several challenges, including incompatibility with perfusion of highly lipophilic compounds. The present study addresses this methodological concern, with an emphasis on the independent effects of commonly used vehicle dialysis solutions to improve solubility of pharmacological agents with otherwise low aqueous solubility. Four microdialysis fibers were placed in the ventral forearm of eight subjects (4 men, 4 women; 25 ± 1 yr) with sites randomized to serve as 1) control (lactated Ringers), 2) Sodium carbonate-bicarbonate buffer administered at physiological pH [SCB-HCl; pH 7.4, achieved via addition of hydrochloric acid (HCl)], 3) 0.02% ethanol, and 4) 2% dimethyl sulfoxide (DMSO). After baseline (34°C), vehicle solutions were administered throughout a standardized local heating protocol to 42°C. Laser Doppler flowmetry provided an index of blood flow. Cutaneous vascular conductance was calculated and normalized to maximum (%CVCmax, sodium nitroprusside and 43°C local heat). The SCB-HCl solution increased baseline %CVCmax (control: 9.7 ± 0.8; SCB-HCl: 21.5 ± 3.5%CVCmax; P = 0.03), but no effects were observed during heating or maximal vasodilation. There were no differences with perfusion of ethanol or DMSO at any stage of the protocol ( P > 0.05). These data demonstrate the potential confounding effects of some vehicle dialysis solutions on cutaneous vascular function. Notably, this study provides evidence that 2% DMSO and 0.02% ethanol are acceptable vehicles with no confounding local vascular effects to a standardized local heating protocol at the concentrations presented. NEW & NOTEWORTHY This study examined the independent effects of common vehicle solutions on cutaneous vascular responses. A basic buffer (SCB-HCl) caused baseline vasodilation; 2% DMSO and 0.02% ethanol had no effects. This highlights the need for considering potential confounding effects of solubilizing solutions when combined with low aqueous soluble pharmacological agents. Importantly, DMSO and ethanol do not appear to influence cutaneous vascular function during baseline or local heating at the concentrations studied, allowing their use without confounding effects.

Acute lysyl oxidase inhibition alters microvascular function in normotensive but not hypertensive men and women

The lysyl oxidase (LOX) family of enzymes regulates collagen cross-linking. LOX is upregulated in hypertension, increasing vascular stiffness. In vivo human research is sparse, as long-term LOX inhibition in animals causes vascular instability. Our aim was to evaluate the effects of LOX inhibition on cutaneous microvascular function to determine whether LOX function was upregulated in hypertensive humans. Four intradermal microdialysis fibers were placed in the forearm of 10 young [age: 24 ± 1 yr, mean arterial pressure (MAP): 87 ± 2 mmHg], 10 normotensive (age: 50 ± 2 yr, MAP: 84 ± 1 mmHg), and 10 hypertensive (age: 53 ± 2 yr, MAP: 112 ± 2 mmHg) subjects. Two sites were perfused with 10 mM β-aminopropionitrile (BAPN) to inhibit LOX. The remaining two sites were perfused with lactated Ringer solution (control). A norepinephrine dose response (10-12-10-2 M) was performed to examine receptor-mediated vasoconstrictor function. A sodium nitroprusside dose response (10-8-10-1.3 M) was performed to examine vascular smooth muscle vasodilator function. Red blood cell flux was measured via laser-Doppler flowmetry and normalized to cutaneous vascular conductance (flux/MAP). LogEC50 values were calculated to determine changes in vasosensitivity. Skin tissue samples were analyzed for both extracellular matrix-bound and soluble LOX. LOX inhibition augmented vasoconstrictor sensitivity in young (control: -6.0 and BAPN: -7.1, P = 0.03) and normotensive (control: -4.8 and BAPN: -7.0, P = 0.01) but not hypertensive (control: -6.0 and BAPN: -6.1, P = 0.79) men and women. Relative to young subjects, extracellular matrix-bound LOX expression was higher in hypertensive subjects (young: 100 ± 8 and hypertensive: 162 ± 8, P = 0.002). These results suggest that upregulated LOX may contribute to the vascular stiffness and microvascular dysfunction characteristic in hypertension. NEW & NOTEWORTHY Matrix-bound lysyl oxidase (LOX) and LOX-like 2 expression are upregulated in the microvasculature of hypertensive men and women. Microvascular responsiveness to exogenous stimuli is altered with localized LOX inhibition in healthy men and women but not hypertensive adults. The LOX family differentially affects microvascular function in hypertensive and normotensive men and women.

Menthol-induced Cutaneous Vasodilation Is Preserved in Essential Hypertensive Men and Women

BACKGROUND Menthol is a selective transient receptor potential melastatin 8 (TRPM8) channel agonist that induces cutaneous vasodilation in young, normotensive men and women through nitric oxide synthase (NOS)-, endothelium-derived hyperpolarizing factor (EDHF)-, and sensory nerve-mediated mechanisms. Microvascular dysfunction is present in essential hypertension and whether menthol induces vasodilation is men and women with essential hypertension is equivocal. METHODS Four intradermal microdialysis fibers were placed in the forearm of 9 essential hypertensive and 10 age-matched normotensive control subjects. Sites were pretreated with lactated Ringers (control), l-NAME (NOS inhibited), TEA (EDHF inhibited), and lidocaine (sensory nerve inhibited). The microdialysis fibers were then perfused with 7 increasing doses of menthol (0.1-500 mM). Red cell flux in response to menthol was measured with laser Doppler flowmetry. Data were normalized to mean arterial pressure and presented as a percentage of site-specific maximum vasodilation (%CVCmax). RESULTS At the control site, menthol caused vasodilation in both the normotensive and hypertensive groups (menthol doses 100, 250, and 500 mM; all P < 0.05 compared to baseline). There were no differences between groups (P = 0.58, main effect). There was no effect of either NOS or sensory nerve inhibition on menthol-induced vasodilation in the normotensive group; however, menthol-induced vasodilation was attenuated with NOS and sensory nerve inhibition in the hypertensive group. EDHF inhibition attenuated menthol-induced vasodilation in both groups. CONCLUSIONS Menthol-induced vasodilation has NO, EDHF, and sensory nerve components. Menthol-induced cutaneous vasodilation is preserved in hypertensive subjects. However, the hypertensive subjects exhibited a loss of redundant vasodilator systems.

Kinesiology Tape Modestly Increases Skin Blood Flow Regardless of Tape Application Technique

**Background and Purpose:** Kinesiology tape (KNT) is commonly used for prevention of injury and as part of rehabilitation to treat muscle and joint pain. KNT is purported to increase local blood flow by reducing local pressure. Whether skin blood flow is increased by KNT is not presently known. **Methods:** We conducted 2 experiments to elucidate the effects of KNT on skin blood flow as follows: in protocol 1, KNT was applied to the skin at 0%, 25%, 50%, and 100% relative ten- sion, and in protocol 2, KNT was applied to the skin with and without convolutions. Red cell flux, an index of skin blood flow, was measured by laser Doppler flowmetry at each site and at a no-tape control site. **Results:** For protocol 1: there was an overall effect of tape on cutaneous vascular conduct- ance (CVC: laser Doppler flowmetry/MAP; KNT: 0.12, 95% confidence interval: 0.10, 0.14; control: 0.08, 95% confidence interval: 0.07, 0.10 flux·mmHg 1; P .05). KNT was retained for 3 days after which the patients returned for follow-up testing. CVC remained unchanged after 3 days of KNT appli- cation (P = 0.07). For protocol 2: there was an overall effect of tape on CVC (KNT: 0.30, 95% confidence interval: 0.21, 0.39; control: 0.15 (0.09, 0.21 flux·mmHg 1; P = .03), but there was no difference between KNT applied with or without convolutions (all P > .05). **Conclusion:** These data suggest that KNT modestly increases microvascular blood flow regardless of tension or presence of convolutions.