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Local skin heating is used to assess microvascular function in clinical populations because NO is required for full expression of the response; however, controversy exists as to the precise NO synthase (NOS) isoform producing NO. Human aging is associated with attenuated cutaneous vasodilation but little is known about the middle aged, an age cohort used for comparison with clinical populations. We hypothesized that endothelial NOS (eNOS) is the primary isoform mediating NO production during local heating, and eNOS-dependent vasodilation would be reduced in middle-aged skin. Vasodilation was induced by local heating (42°C) and during acetylcholine dose-response (ACh-DR: 0.01, 0.1, 1.0, 5.0, 10.0, 50.0, 100.0 mmol/l) protocols. Four microdialysis fibers were placed in the skin of 24 men and women; age cohorts were 12 middle-aged (53 ± 1 yr) and 12 young (23 ± 1 yr). Sites served as control, nonselective NOS inhibited [ NG-nitro-l-arginine methyl ester (l-NAME)], inducible NOS (iNOS) inhibited (1400W), and neuronal NOS (nNOS) inhibited ( Nω-propyl-l-arginine). After full expression of the local heating response, l-NAME was perfused at all sites. Cutaneous vascular conductance was measured and normalized to maximum (%CVCmax: Nitropress). l-NAME reduced %CVCmax at baseline, all phases of the local heating response, and at all ACh concentrations compared with all other sites. iNOS inhibition reduced the initial peak (53 ± 2 vs. 60 ± 2%CVCmax; P < 0.001); however, there were no other differences between control, nNOS-, and iNOS-inhibited sites during the phases of local heating or ACh-DR. When age cohorts were compared, NO-dependent vasodilation during local heating (52 ± 6 vs. 68 ± 4%CVCmax; P = 0.013) and ACh perfusion (50 mmol/l: 83 ± 3 vs. 93 ± 2%CVCmax; 100 mmol/l: 83 ± 4 vs. 92 ± 3%CVCmax; both P = 0.03) were reduced in middle-aged skin. There were no differences in NOS isoform expression obtained from skin biopsy samples between groups (all P > 0.05). These data suggest that eNOS mediates the production of NO during local heating and that cutaneous vasodilation is attenuated in middle-aged skin.

Functional constitutive nitric oxide synthase (NOS) is required for full expression of reflex cutaneous vasodilation that is attenuated in aged skin. Both the essential cofactor tetrahydrobiopterin (BH4) and adequate substrate concentrations are necessary for the functional synthesis of nitric oxide (NO) through NOS, both of which are reduced in aged vasculature through increased oxidant stress and upregulated arginase, respectively. We hypothesized that acute local BH4 administration or arginase inhibition would similarly augment reflex vasodilation in aged skin during passive whole body heat stress. Four intradermal microdialysis fibers were placed in the forearm skin of 11 young (22 ± 1 yr) and 11 older (73 ± 2 yr) men and women for local infusion of 1) lactated Ringer, 2) 10 mM BH4, 3) 5 mM ( S)-(2-boronoethyl)-l-cysteine + 5 mM Nω-hydroxy-nor-l-arginine to inhibit arginase, and 4) 20 mM NG-nitro-l-arginine methyl ester (l-NAME) to inhibit NOS. Red cell flux was measured at each site by laser-Doppler flowmetry (LDF) as reflex vasodilation was induced. After a 1.0°C rise in oral temperature (Tor), mean body temperature was clamped and 20 mM l-NAME was perfused at each site. Cutaneous vascular conductance was calculated (CVC = LDF/mean arterial pressure) and expressed as a percentage of maximum (%CVCmax; 28 mM sodium nitroprusside and local heat, 43°C). Vasodilation was attenuated at the control site of the older subjects compared with young beginning at a 0.3°C rise in Tor. BH4 and arginase inhibition both increased vasodilation in older (BH4: 55 ± 5%; arginase-inhibited: 47 ± 5% vs. control: 37 ± 3%, both P < 0.01) but not young subjects compared with control (BH4: 51 ± 4%CVCmax; arginase-inhibited: 55 ± 4%CVCmax vs. control: 56 ± 6%CVCmax, both P > 0.05) at a 1°C rise in Tor. With a 1°C rise in Tor, local BH4 increased NO-dependent vasodilation in the older (BH4: 31.8 ± 2.4%CVCmax vs. control: 11.7 ± 2.0%CVCmax, P < 0.001) but not the young (BH4: 23 ± 4%CVCmax vs. control: 21 ± 4%CVCmax, P = 0.718) subject group. Together these data suggest that reduced BH4 contributes to attenuated vasodilation in aged human skin and that BH4 NOS coupling mechanisms may be a potential therapeutic target for increasing skin blood flow during hyperthermia in older humans.

Essential hypertension is a proinflammatory, proconstrictor disease coinciding with endothelial dysfunction and inward vessel remodeling. Using the skin circulation, our aim was to determine whether inducible NO synthase (iNOS) upregulation attenuates NO-dependent cutaneous vasodilation in hypertensive humans. We hypothesized that, with hypertension, localized iNOS inhibition would restore vasodilation in response to NO-dependent stimuli, and iNOS expression would be increased and phosphorylated vasodilator-stimulated phosphoprotein would be decreased. For, in vivo protocols, 4 intradermal microdialysis fibers were placed in 9 hypertensive and 10 normotensive men and women (systolic blood pressure: 146±4 versus 113±2 mm Hg; P N ω - propyl - l -arginine), and nonselective NOS inhibited ( N G -nitro- l -arginine methyl ester). Cutaneous vascular conductance was calculated (percentage of sodium nitroprusside) during standardized local heating (42°C) and acetylcholine dose-response protocols (0.01, 0.10, 1.00, 5.00, 10.00, 50.00, 100.00 mmol/L). The NO-dependent local heating response was attenuated at control (95±2% versus 76±2% cutaneous vascular conductance; P P P P P P =0.006) and phosphorylated vasodilator-stimulated phosphoprotein was decreased in skin from hypertensive humans ( P =0.04). These data suggest that iNOS is upregulated in essential hypertensive humans and contributes to reduced NO-dependent cutaneous vasodilation.

Elevated oxidized low-density lipoproteins (LDL) are associated with vascular dysfunction in the cutaneous microvasculature, induced in part by upregulated arginase activity and increased globalized oxidant stress. Since tetrahydrobiopterin (BH(4)) is an essential cofactor for endothelial nitric oxide synthase (NOS3), decreased bioavailability of the substrate l-arginine and/or BH(4) may contribute to decreased NO production with hypercholesterolaemia. We hypothesized that (1) localized administration of BH(4) would augment NO-dependent vasodilatation in hypercholesterolaemic human skin, which would be further increased when combined with arginase inhibition and (2) the improvement induced by localized BH(4) would be attenuated after a 3 month oral atorvastatin intervention (10 mg). Four microdialysis fibres were placed in the skin of nine normocholesterolaemic (NC: LDL = 95 ± 4 mg dl(-1)) and nine hypercholesterolaemic (HC: LDL = 177 ± 6 mg dl(-1)) men and women before and after 3 months of systemic atorvastatin. Sites served as control, NOS inhibited, BH(4), and arginase inhibited + BH(4) (combo). Skin blood flow was measured while local skin heating (42°C) induced NO-dependent vasodilatation. After the established plateau l-NAME was perfused in all sites to quantify NO-dependent vasodilatation (NO). Data were normalized to maximum cutaneous vascular conductance (CVC). Vasodilatation at the plateau and NO-dependent vasodilatation were reduced in HC subjects (plateau HC: 70 ± 5% CVC(max) vs. NC: 95 ± 2% CVC(max); NO HC: 45 ± 5% CVC(max) vs. NC: 64 ± 5% CVC(max); both P 0.05 vs. control site). After the atorvastatin intervention (LDL = 98 ± mg * dl(-1)) there was an increase in the plateau in HC (96 ± 4% CVC(max), P < 0.001) and NO-dependent vasodilatation (68 ± 3% CVC(max), P < 0.001). Localized BH(4) alone or combo was less effective at increasing NO-dependent vasodilatation after the drug intervention (BH(4): 60 ± 5% CVC(max); combo 58 ± 2% CVC(max), both P < 0.001). These data suggest that decreased BH(4) bioavailability contributes in part to cutaneous microvascular dysfunction in hypercholesterolaemic humans and that atorvastatin is an effective systemic treatment for improving NOS coupling mechanisms in the microvasculature.

Elevated low-density lipoproteins (LDL) are associated with cutaneous microvascular dysfunction partially mediated by increased arginase activity, which is decreased following a systemic atorvastatin therapy. We hypothesized that increased ascorbate-sensitive oxidant stress, partially mediated through uncoupled nitric oxide synthase (NOS) induced by upregulated arginase, contributes to cutaneous microvascular dysfunction in hypercholesterolemic (HC) humans. Four microdialysis fibers were placed in the skin of nine HC (LDL = 177 ± 6 mg/dl) men and women before and after 3 mo of a systemic atorvastatin intervention and at baseline in nine normocholesterolemic (NC) (LDL = 95 ± 4 mg/dl) subjects. Sites served as control, NOS inhibited, L-ascorbate, and arginase-inhibited+L-ascorbate. Skin blood flow was measured while local skin heating (42°C) induced NO-dependent vasodilation. After the established plateau in all sites, 20 mM ≪ngname≫ was infused to quantify NO-dependent vasodilation. Data were normalized to maximum cutaneous vascular conductance (CVC) (sodium nitroprusside + 43°C). The plateau in vasodilation during local heating (HC: 78 ± 4 vs. NC: 96 ± 2% CVCmax, P < 0.01) and NO-dependent vasodilation (HC: 40 ± 4 vs. NC: 54 ± 4% CVCmax, P < 0.01) was reduced in the HC group. Acute L-ascorbate alone (91 ± 5% CVCmax, P < 0.001) or combined with arginase inhibition (96 ± 3% CVCmax, P < 0.001) augmented the plateau in vasodilation in the HC group but not the NC group (ascorbate: 96 ± 2; combo: 93 ± 4% CVCmax, both P > 0.05). After the atorvastatin intervention NO-dependent vasodilation was augmented in the HC group (HC postatorvastatin: 64 ± 4% CVCmax, P < 0.01), and there was no further effect of ascorbate alone (58 ± 4% CVCmax,P > 0.05) or combined with arginase inhibition (67 ± 4% CVCmax,P > 0.05). Increased ascorbate-sensitive oxidants contribute to hypercholesteromic associated cutaneous microvascular dysfunction which is partially reversed with atorvastatin therapy.

Heat is the most abundant byproduct of cellular metabolism. As such, dynamic exercise in which a significant percentage of muscle mass is engaged generates thermoregulatory demands that are met in part by increases in skin blood flow. Increased skin blood flow during exercise adds to the demands on cardiac output and confers additional circulatory strain beyond that associated with perfusion of active muscle alone. Endurance exercise training results in a number of physiological adaptations which ultimately reduce circulatory strain and shift thermoregulatory control of skin blood flow to higher levels of blood flow for a given core temperature. In addition, exercise training induces peripheral vascular adaptations within the cutaneous microvasculature indicative of enhanced endothelium-dependent vasomotor function. However, it is not currently clear how (or if) these local vascular adaptations contribute to the beneficial changes in thermoregulatory control of skin blood flow following exercise training. The purpose of this Hot Topic Review is to synthesize the literature pertaining to exercise training-mediated changes in cutaneous microvascular reactivity and thermoregulatory control of skin blood flow. In addition, we address mechanisms driving changes in cutaneous microvascular reactivity and thermoregulatory control of skin blood flow, and pose the question: what (if any) is the functional role of increased cutaneous microvascular reactivity following exercise training?

Elevated low-density lipoproteins (LDLs) are associated with vascular dysfunction evident in the cutaneous microvasculature. We hypothesized that uncoupled endothelial nitric oxide synthase (NOS3) through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic (HC) humans and that a statin intervention would decrease arginase activity. Five microdialysis fibres were placed in the skin of nine normocholesterolaemic (NC: LDL level 95±4 mg dl⁻¹) and nine hypercholesterolaemic (HC: LDL: 177±6 mg dl⁻¹) men and women before and after 3 months of systemic atrovastatin. Sites served as control, NOS inhibited, arginase inhibited, L-arginine supplemented and arginase inhibited plus L-arginine supplemented. Skin blood flow was measured while local skin heating (42°C) induced NO-dependent vasodilatation. L-NAME was infused after the established plateau in all sites to quantify NO-dependent vasodilatation. Data were normalized to maximum cutaneous vascular conductance (CVC(max)). Skin samples were obtained to measure total arginase activity and arginase I and arginase II protein. Vasodilatation was reduced in hyperocholesterolaemic subjects (HC: 76±2 vs. NC: 94±3%CVC(max), P 0.05). Arginase activity and protein were increased in HC skin (P < 0.05 vs. NC) and activity decreased with atrovastatin treatment (P < 0.05). Reduced NOS3 substrate availability through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic humans, which is corrected with atorvastatin therapy.

The attenuated reflex vasoconstriction in aged skin may be partly mediated by oxidant-induced reduction in functional substrate and cofactor availability for noradrenaline biosynthesis. We hypothesized that localized supplementation of tyrosine and tetrahydrobiopterin (BH4) in aged human skin could augment reflex- (whole-body cooling) and pharmacologically (tyramine, which displaces noradrenaline from axon terminals) induced vasoconstriction. Four microdialysis fibres were placed in the forearm skin of 10 young and 10 older subjects for infusion of (1) Ringer solution (control), (2) 0.5 mm l-tyrosine, (3) 5 mm BH4, and (4) BH4+l-tyrosine. Cutaneous vascular conductance (CVC) was calculated (laser Doppler flux/mean arterial pressure) and normalized to baseline (%ΔCVCbase). Vasoconstriction was attenuated at the control site in the older subjects during both whole-body cooling (young: −39 ± 3, older: −17 ± 3%ΔCVCbase; P < 0.01) and tyramine infusion (young: −41 ± 3, older: −21 ± 4%ΔCVCbase; P < 0.01). BH4 (cold, young: −37 ± 3, older: −36 ± 3; tyramine, young: −41 ± 2, older: −36 ± 3%ΔCVCbase) and tyrosine (cold, young: −37 ± 4, older: −34 ± 4; tyramine, young: −40 ± 4, older: −45 ± 4%ΔCVCbase) both resolved the age-related decrease in cutaneous vasoconstriction, but BH4+ tyrosine did not further augment vasoconstriction (cold, young: −38 ± 4, older: −31 ± 3; tyramine, young: −36 ± 3, older: −36 ± 5%ΔCVCbase). These data are consistent with the concept that reduced bioavailability of BH4 and/or tyrosine may impair noradrenaline synthesis and contribute to the attenuated vasoconstrictor response in aged skin.

Primary human aging may be associated with augmented Rho kinase (ROCK)-mediated contraction of vascular smooth muscle and ROCK-mediated inhibition of nitric oxide synthase (NOS). We hypothesized that the contribution of ROCK to reflex vasoconstriction (VC) is greater in aged skin. Cutaneous VC was elicited by 1) whole body cooling [mean skin temperature (Tsk) = 30.5°C] and 2) local norepinephrine (NE) infusion (1 × 10−6 M). Four microdialysis fibers were placed in the forearm skin of eight young (Y) and eight older (O) subjects for infusion of 1) Ringer solution (control), 2) 3 mM fasudil (ROCK inhibition), 3) 20 mM NG-nitro-l-arginine methyl ester (NOS inhibition), and 4) both ROCK + NOS inhibitors. Red cell flux was measured by laser-Doppler flowmetry over each site. Cutaneous vascular conductance (CVC) was calculated as flux/mean arterial pressure and normalized to baseline CVC (%ΔCVCbaseline). VC was reduced at the control site in O during cooling (Y, −34 ± 3; and O, −18 ± 3%ΔCVCbaseline; P < 0.001) and NE infusion (Y, −53 ± 4, and O, −41 ± 9%ΔCVCbaseline; P = 0.006). Fasudil attenuated VC in both age groups during mild cooling; however, this reduction remained only in O but not in Y skin during moderate cooling (Y, −30 ± 5; and O, −7 ± 1%ΔCVCbaseline; P = 0.016) and was not altered by NOS inhibition. Fasudil blunted NE-mediated VC in both age groups (Y, −23 ± 4; and O, −7 ± 3%ΔCVCbaseline; P < 0.01). Cumulatively, these data indicate that reflex VC is more reliant on ROCK in aged skin such that approximately half of the total VC response to whole body cooling is ROCK dependent.

In young healthy humans full expression of reflex cutaneous vasodilation is dependent on cyclooxygenase (COX)- and nitric oxide synthase (NOS)-dependent mechanisms. Chronic low-dose aspirin therapy attenuates reflex cutaneous vasodilation potentially through both platelet and vascular COX-dependent mechanisms. We hypothesized the contribution of COX-dependent vasodilators to reflex cutaneous vasodilation during localized acute COX inhibition would be attenuated in healthy middle-aged humans due to a shift toward COX-dependent vasoconstrictors. Four microdialysis fibers were placed in forearm skin of 13 middle-aged (53 ± 2 yr) normotensive healthy humans, serving as control (Ringer), COX-inhibited (10 mM ketorolac), NOS-inhibited (10 mM NG-nitro-l-arginine methyl ester), and combined NOS- and COX-inhibited sites. Red blood cell flux was measured over each site by laser-Doppler flowmetry as reflex vasodilation was induced by increasing oral temperature (Tor) 1.0°C using a water-perfused suit. Cutaneous vascular conductance was calculated (CVC = flux/mean arterial pressure) and normalized to maximal CVC (CVCmax; 28 mM sodium nitroprusside). CVCmax was not affected by localized microdialysis drug treatment ( P > 0.05). Localized COX inhibition increased baseline (18 ± 3%CVCmax; P < 0.001) compared with control (9 ± 1%CVCmax), NOS-inhibited (7 ± 1%CVCmax), and combined sites (10 ± 1%CVCmax). %CVCmax in the COX-inhibited site remained greater than the control site with ΔTor ≤ 0.3°C; however, there was no difference between these sites with ΔTor ≥ 0.4°C. NOS inhibition and combined COX and NOS inhibition attenuated reflex vasodilation compared with control ( P < 0.001), but there was no difference between these sites. Localized COX inhibition with ketorolac significantly augments baseline CVC but does not alter the subsequent skin blood flow response to hyperthermia, suggesting a limited role for COX-derived vasodilator prostanoids in reflex cutaneous vasodilation and a shift toward COX-derived vasoconstrictors in middle-aged human skin.

Reflex vasoconstriction (VC) is attenuated in aged skin resulting in greater skin blood flow and heat loss during cold exposure. We hypothesized that adrenergic function is compromised due to depletion of tetrahydrobiopterin (BH4), an essential cofactor required for catecholamine synthesis, and therefore local BH4 supplementation would functionally augment reflex and pharmacologically induced VC elicited by gradual whole-body cooling (skin temperature (Tsk) = 30.5°C) and tyramine infusion, respectively. Four microdialysis (MD) fibres were placed in the forearm skin of 11 young (Y) and 11 older (O) human subjects for infusion of (1) Ringer solution (control), (2) 5 mm BH4, (3) 5 mm BH4+ 10 mm ascorbate, and (4) 5 mm BH4+ adrenoreceptor blockade (5 mm yohimbine + 1 mm propranolol). Laser Doppler flux (LDF) was measured over each MD site and cutaneous vascular conductance was calculated as CVC = LDF/MAP and expressed as per cent change from baseline (%ΔCVCbase). The VC response was lower at the control site in O during cooling (Y: −34 ± 2%ΔCVCbase, O: −17 ± 2%ΔCVCbase; P < 0.001) and tyramine infusion (Y: −33 ± 4%ΔCVCbase, O: −15 ± 3%ΔCVCbase; P < 0.001). BH4 infusion normalized O to Y values during both cooling (Y: −34 ± 4%ΔCVCbase, O: −34 ± 2%ΔCVCbase; P < 0.001) and tyramine (Y: −38 ± 4%ΔCVCbase, O: −35 ± 3%ΔCVCbase; P < 0.001), however, adding adrenoreceptor blockade abolished VC in aged skin indicating that BH4 acts through adrenergic, not cotransmitter, mechanisms. Local BH4 supplementation augments reflex and tyramine-induced VC in aged skin, suggesting that reduced BH4 bioavailability may contribute to attenuated VC during whole-body cooling.

Human skin blood flow increases in response to increased body core and local skin temperature via distinct reflex and local mechanisms requiring functional nitric oxide (NO) for full expression. The mechanisms mediating cutaneous vasodilation are impaired with primary aging, resulting in attenuated vasodilation. This article highlights recent findings of how age-related vascular impairments in NO signaling contribute to attenuated cutaneous vasodilation.

Reflex cutaneous vasodilatation is dependent on nitric oxide (NO), which is diminished in hypertension (HTN). Arginase may be up-regulated with HTN, which preferentially metabolizes L-arginine (L-arg), competing with NO-synthase (NOS)-mediated pathways and limiting NO synthesis. We hypothesized that NO-dependent vasodilatation would be attenuated in HTN skin, and arginase inhibition (A-I) alone or with concurrent l-arginine supplementation, would augment vasodilatation. Five microdialysis fibres were placed in skin of eight unmedicated subjects with HTN (mean arterial pressure (MAP), 112 +/- 1 mmHg) and nine age-matched normotensive (AMN) (MAP: 87 +/- 1 mmHg) men and women to serve as: control (C, Ringer solution), NOS inhibited (NOS-I, 10 mM L-NAME), A-I (5 mM BEC + 5 mM nor-NOHA), L-arg supplemented (L-arg, 10 mM L-arg), and combined A-I + L-arg. Reflex vasodilatation was induced by using a water-perfused suit to increase oral temperature (T(or)) 1.0 degrees C. Red cell flux was measured by laser-Doppler flowmetry over each site. Cutaneous vascular conductance was calculated (CVC = flux/MAP) and normalized to maximal CVC (28 mM SNP + local heating to 43 degrees C). The Delta%CVC(max) between the control and NOS-I site was calculated as the difference between C and NOS-I sites. Maximal CVC was attenuated in the HTN subjects by approximately 25% compared with AMN subjects (P 0.05). NOS-I significantly decreased %CVC(max) in both groups but %CVC(max) was greater in the HTN group (HTN, 32 +/- 4%CVC(max) versus AMN, 23 +/- 3%CVC(max), P 0.5 degrees C in the HTN group (P 0.05). Combined A-I + L-arg augmented %CVC(max) in both subject groups compared with their respective control sites (HTN, 60 +/- 7%CVC(max) versus AMN, 61 +/- 3%CVC(max), both P

Full expression of reflex cutaneous vasodilation (VD) is dependent on nitric oxide (NO) and is attenuated in older humans. NO may be decreased by an age-related increase in reactive oxygen species or a decrease in l-arginine availability via upregulated arginase. The purpose of this study was to determine the effect of acute antioxidant supplementation alone and combined with arginase inhibition on reflex VD in aged skin. Eleven young (Y; 22 ± 1 yr) and 10 older (O; 68 ± 1 yr) human subjects were instrumented with four intradermal microdialysis (MD) fibers. MD sites were control (Co), NO synthase inhibited (NOS-I), l-ascorbate supplemented (Asc), and Asc + arginase-inhibited (Asc + A-I). After baseline measurements, subjects underwent whole body heating to increase oral temperature (Tor) by 0.8°C. Red blood cell flux was measured by using laser-Doppler flowmetry, and cutaneous vascular conductance (CVC) was calculated (CVC = flux/mean arterial pressure) and normalized to maximal (CVCmax). VD during heating was attenuated in O (Y: 37 ± 3 vs. O: 28 ± 3% CVCmax; P < 0.05). NOS-I decreased VD in both groups compared with Co (Y: 20 ± 4; O: 15 ± 2% CVCmax; P < 0.05 vs. Co within group). Asc and Asc + A-I increased VD beyond Co in O (Asc: 35 ± 4% CVCmax; Asc + A-I: 41 ± 3% CVCmax; P < 0.001) but not in Y (Asc: 36 ± 3% CVCmax; Asc + A-I: 40 ± 5% CVCmax; P > 0.05). Combined Asc + A-I resulted in a greater increase in VD than Asc alone in O ( P = 0.001). Acute Asc supplementation increased reflex VD in aged skin. Asc combined with arginase inhibition resulted in a further increase in VD above Asc alone, effectively restoring CVC to the level of young subjects.

Skin blood flow is neurally controlled by two distinct branches of the sympathetic nervous system: an adrenergic vasoconstrictor system and an active vasodilator system (Grant & Holling, 1938). Under normothermic conditions, the cutaneous vasculature is under tonic adrenergic control. However, with rising body core temperature, skin blood flow initially increases through a withdrawal of tonic vasoconstriction, and upon reaching a specific threshold skin blood flow further increases by the active vasodilator system (Roddie et al. 1957). Cutaneous active vasodilatation is mediated by sympathetic cotransmission of acetylcholine and an unknown neurotransmitter (Kellogg et al. 1995). Vasoactive intestinal peptide (VIP) and histamine 1 (H1) receptor activation contribute to reflex cutaneous vasodilatation (Bennett et al. 2003; Wilkins et al. 2004; Wong et al. 2004). Moreover, these vasodilator pathways mediate downstream nitric oxide (NO)-dependent vasodilatation, which is required for full expression of the reflex vasodilatory response (Kellogg et al. 1998a; Shastry et al. 1998). Aged humans exhibit attenuated cutaneous vasodilatory responses during hyperthermia (Kenney et al. 1997), resulting from a diminished neurogenic cotransmitter contribution and an increased reliance on impaired NO-dependent vasodilatation (Holowatz et al. 2003). Attenuated NO-dependent vasodilatation is associated with decreased NO bioavailability. In aged skin, decreased NO bioavailability is probably multifaceted, involving several potential signalling pathways, including dysregulated utilization of the NO substrate l-arginine (l-Arg). One potential mechanism that has been implicated in reduced l-Arg availability for NOS is augmented vascular arginase activity (Hecker et al. 1995; Berkowitz et al. 2003). Arginase is constitutively expressed in two isoforms (I and II), which catalyse the conversion of l-Arg to l-ornithine and urea during the final step of the urea cycle. Arginase I is most likely the predominant isoform in the vasculature and is capable of reciprocally regulating endothelial NOS by competing for the common substrate l-Arg (Berkowitz et al. 2003; White et al. 2006) (Fig. 1). Figure 1 Schematic representation of the l-arginine pathway in the cutaneous vasculature In aged and hypertensive animal models of microvasculature dysfunction, augmented arginase activity contributes to impaired NO bioavailability (Berkowitz et al. 2003; John & Schmieder, 2003; White et al. 2006), and both acute and chronic inhibition of arginase restore endothelial NO-dependent vasodilatation. Furthermore, augmented arginase activity may serve as a potential explanation for the ‘l-arginine paradox’, which states that despite the high intracellular concentrations of l-Arg that far exceed the Michaelis constant for endothelial NOS (2 × 10−6m) (Grody et al. 1987; Griffith & Stuehr, 1995), NO-dependent vasodilatation in vivo can be further increased by the administration of exogenous l-Arg. Taken together, these findings suggest that increased arginase activity may limit the available pool of l-Arg for NOS and therefore the bioavailability of NO. However, the role of arginase in regulating intracellular stores of l-Arg and subsequent NO-dependent vasodilatation in humans remains unclear. Therefore, the purpose of this study was to determine the role of arginase in reflex cutaneous vasodilatation in aged humans. We hypothesized that arginase inhibition alone and with concurrent l-Arg supplementation would augment cutaneous vasodilatation during passive whole-body heat stress by increasing l-Arg availability for NO synthesis.

Cutaneous vasoconstriction (VC) is the initial thermoregulatory response to skin cooling, effectively minimizing heat loss to the environment. Cutaneous VC can be stimulated by either whole-body skin cooling (reflex-mediated VC) or local skin cooling (locally mediated VC); these two mechanisms are not mutually exclusive and often operate in concert during cold exposure to maximize VC. However, human ageing substantially impairs cutaneous VC (Khan et al. 1992; Richardson et al. 1992; Kenney & Armstrong, 1996; Thompson & Kenney, 2004), rendering older people more susceptible to excessive heat loss and, potentially, hypothermia (Collins et al. 1977; Budd et al. 1991; Inoue et al. 1992). While several thermoregulatory studies have addressed age-related changes in reflex-mediated VC, it is unclear whether this blunted VC response in aged skin is likewise seen with local cooling, which involves different downstream mechanisms. Local cooling of cutaneous blood vessels evokes several vascular responses that alter responses to noradrenaline (NA) (Vanhoutte & Shepherd, 1970; Janssens & Vanhoutte, 1978; Flavahan et al. 1985). In vivo, local cooling leads to substantial VC mediated through α2-adrenergic receptors (ARs) (Flavahan et al. 1985; Ekenvall et al. 1988; Cankar et al. 2004). In vitro, local cooling increases intracellular Ca2+ mobilization as well as Ca2+ sensitivity. Additionally, local cooling increases the number of α2-ARs on the vascular smooth muscle cell surface by stimulating the translocation of α2C-ARs from the Golgi apparatus to the cell membrane (Chotani et al. 2000; Jeyaraj et al. 2001; Bailey et al. 2004). These intracellular responses to local cooling enhance the capacity for NA-mediated VC by augmenting the sensitivity of cutaneous vessels to the effects of NA. However, studies that have addressed the changes evoked by local cooling have only used young animal or human thermoregulatory models; very little is known regarding how local cooling affects VC in aged human skin. The purpose of the present study was to examine the effects of local cooling on aged skin. Specifically, we tested the hypotheses that (1) VC due to local cooling alone is blunted in aged skin, and (2) the heightened sensitivity to NA induced by local cooling observed in young skin is attenuated in aged skin.

Thermoregulatory cutaneous vasodilatation (VD) is attenuated in aged skin. While acetylcholine (ACh) plays a role in thermally mediated VD, the precise mechanisms through which ACh-mediated VD acts and whether those downstream mechanisms change with ageing are unclear. We tested the hypotheses that both nitric oxide (NO)- and prostanoid-mediated pathways contribute to exogenous ACh-mediated VD, and that both are attenuated with advanced age. Twelve young (Y: 23 ± 1 years) and 10 older (O: 69 ± 1 years) subjects underwent infusions of 137.5 μm ACh at four intradermal microdialysis sites: control (C, Ringer solution), NO synthase inhibited (NOS-I, 10 mml-NAME), cyclooxygenase inhibited (COX-I, 10 mm ketorolac) and NOS-I + COX-I. Red blood cell flux was monitored using laser-Doppler flowmetry, and cutaneous vascular conductance (CVC) was calculated (laser-Doppler flux/mean arterial pressure) and normalized to maximal CVC (%CVCmax) (28 mm sodium nitroprusside + local heating to 43°C). Baseline %CVCmax was increased in the O at COX-I sites (COX-I 16 ± 1, NOS-I + COX-I 16 ± 2 versus C 10 ± 1%CVCmax; P < 0.001) but not in the young, suggesting an age-related shift toward COX vasoconstrictors contributing to basal cutaneous vasomotor tone. There was no difference in peak %CVCmax during ACh infusion between age groups, and the response was unchanged by NOS-I (O: NOS-I 35 ± 5 versus C 38 ± 5%CVCmax; P= 0.84) (Y: NOS-I 41 ± 4 versus C 39 ± 4%CVCmax; P= 0.67). COX-I and NOS-I + COX-I attenuated the peak CVC response to ACh in both groups (COX-I O: 29 ± 3, Y: 22 ± 2%CVCmaxversus C; P < 0.001 both groups; NOS-I + COX-I O: 32 ± 3 versus Y: 29 ± 2%CVCmax; versus C; P < 0.001 both groups). ACh mediates cutaneous VD through prostanoid and non-NO-, non-prostanoid-dependent pathways. Further, older subjects have a diminished prostanoid contribution to ACh-mediated VD.

In normally active individuals, postexercise hypotension after a single bout of aerobic exercise occurs due to an unexplained peripheral vasodilation. Prostaglandin production has been suggested to contribute to the increases in blood flow during and after exercise; however, its potential contribution to postexercise hypotension has not been assessed. The purpose of this study was to determine the potential contribution of a prostaglandin-dependent vasodilation to changes in systemic vascular conductance underlying postexercise hypotension; this was done by inhibiting production of prostaglandins with the cyclooxygenase inhibitor ibuprofen. We studied 11 healthy normotensive men (aged 23.7 ± 4.2 yr) before and during the 90 min after a 60-min bout of cycling at 60% peak O2 uptake on a control and a cyclooxygenase inhibition day (randomized). Subjects received 10 mg/kg of oral ibuprofen on the cyclooxygenase inhibition day. On both study days, arterial blood pressure (automated auscultation) and cardiac output (acetylene uptake) were measured, and systemic vascular conductance was calculated. Inhibition of cyclooxygenase had no effect on baseline values of mean arterial pressure or systemic vascular conductance ( P > 0.2). After exercise on both days, mean arterial pressure was reduced (−2.2 ± 1.0 mmHg change with the control condition and −3.8 ± 1.5 mmHg change with the ibuprofen condition, both P < 0.05 vs. preexercise) and systemic vascular conductance was increased (5.2 ± 5.0% change with the control condition and 8.7 ± 4.1% change with the ibuprofen condition, both P < 0.05 vs. preexercise). There were no differences between study days ( P > 0.6). These data suggest that prostaglandin-dependent vasodilation does not contribute to the increased systemic vascular conductance underlying postexercise hypotension.

1. Human life is sustainable only below an internal temperature of roughly 42–44 °C. Yet our ability to survive at severe environmental extremes is testimony to the marvels of integrative human physiology. 2. One approach to understanding human thermoregulatory capacity is to examine the upper limits of thermal balance between man and the air environment, i.e. the maximal environmental conditions under which humans can maintain a steady-state core temperature. Heat acclimation expands the zone of thermal balance. 3. Human beings can and do, often willingly, tolerate extreme heat stresses well above these thermal balance limits. Survival in all such cases is limited to abbreviated exposure times, which in turn are limited by the robustness of the thermoregulatory response. 4. Figures are provided that relate tolerance time and the rate of change in core temperature to environmental characteristics based on data compiled from the literature.

Thermoregulatory cutaneous vasodilation is diminished in the elderly. The goal of this study was to test the hypothesis that a reduction in nitric oxide (NO)-dependent mechanisms contributes to the attenuated reflex cutaneous vasodilation in older subjects. Seven young (23 ± 2 yr) and seven older (71 ± 6 yr) men were instrumented with two microdialysis fibers in the forearm skin. One site served as control (Ringer infusion), and the second site was perfused with 10 mM N G-nitro-l-arginine methyl ester to inhibit NO synthase (NOS) throughout the protocol. Water-perfused suits were used to raise core temperature 1.0°C. Red blood cell (RBC) flux was measured with laser-Doppler flowmetry over each microdialysis fiber. Cutaneous vascular conductance (CVC) was calculated as RBC flux per mean arterial pressure, with values expressed as a percentage of maximal vasodilation (infusion of 28 mM sodium nitroprusside). NOS inhibition reduced CVC from 75 ± 6% maximal CVC (CVCmax) to 53 ± 3% CVCmax in the young subjects and from 64 ± 5% CVCmax to 29 ± 2% CVCmax in the older subjects with a 1.0°C rise in core temperature. Thus the relative NO-dependent portion of cutaneous active vasodilation (AVD) accounted for ∼23% of vasodilation in the young subjects and 60% of the vasodilation in the older subjects at this level of hyperthermia ( P < 0.001). In summary, NO-mediated pathways contributed more to the total vasodilatory response of the older subjects at high core temperatures. This suggests that attenuated cutaneous vasodilation with age may be due to a reduction in, or decreased vascular responsiveness to, the unknown neurotransmitter(s) mediating AVD.

cardiovascular diseases affect ∼80 million people and are collectively the leading cause of death in the United States ([3][1]). Impaired endothelial vascular signaling leading to endothelial dysfunction is one of the earliest vascular changes in the pathogenesis of cardiovascular disease.

In 10 men and nine women aged 20-23 years, we aimed to establish whether endogenous prostanoids synthesised by cyclooxygenase (COX) affect responses evoked in the finger by α(1)- or α(2)-adrenoceptor agonists. Cutaneous red cell flux (cRCF) was recorded in dorsal finger during iontophoresis of phenylephrine (PE) or clonidine (0.5 mm, seven 0.1 mA pulses followed by one 0.2 mA pulse: 30 s each at 60 s intervals) before and after the COX inhibitor aspirin (600 mg p.o.). In men, PE evoked a biphasic mean increase/decrease in cRCF before but a monophasic mean decrease in cRCF of 30-40% after aspirin (P0.05). In women in the low oestrogen (E(2)) phase of the menstrual cycle, PE evoked a decrease in cRCF (30-40%; P0.05) that was unchanged by aspirin, whereas in the high E(2) phase, PE evoked no change before but a graded decrease in cRCF (30-40%; P0.05) after aspirin. Clonidine evoked a decrease in cRCF (∼30%; P0.05) in men before, but not after, aspirin. Clonidine evoked both increases and decreases in cRCF before and after aspirin in women in the low and high E(2) phases (P0.05). We propose that finger vasoconstriction evoked by extraluminal α(1)-adrenoceptor stimulation is blunted by vasodilator COX products in young men and overcome by their action in women in the high, but not low E(2), phase of the menstrual cycle. By contrast, α(2)-adrenoceptor stimulation evokes finger vasoconstriction that is mediated by vasoconstrictor COX products in young men, but evokes no consistent response in women in the low or high E(2) phases of the menstrual cycle.

Heat is the most abundant byproduct of cellular metabolism. As such, dynamic exercise in which a significant percentage of muscle mass is engaged generates thermoregulatory demands that are met in part by increases in skin blood flow. Increased skin blood flow during exercise adds to the demands on cardiac output and confers additional circulatory strain beyond that associated with perfusion of active muscle alone. Endurance exercise training results in a number of physiological adaptations which ultimately reduce circulatory strain and shift thermoregulatory control of skin blood flow to higher levels of blood flow for a given core temperature. In addition, exercise training induces peripheral vascular adaptations within the cutaneous microvasculature indicative of enhanced endothelium-dependent vasomotor function. However, it is not currently clear how (or if) these local vascular adaptations contribute to the beneficial changes in thermoregulatory control of skin blood flow following exercise training. The purpose of this Hot Topic review is to synthesize the literature pertaining to exercise training-mediated changes in cutaneous microvascular reactivity and thermoregulatory control of skin blood flow. In addition, we address mechanisms driving changes in cutaneous microvascular reactivity and thermoregulatory control of skin blood flow, and pose the question: what (if any) is the functional role of increased cutaneous microvascular reactivity following exercise training?

Elevated low-density lipoproteins (LDLs) are associated with vascular dysfunction evident in the cutaneous microvasculature. We hypothesized that uncoupled endothelial nitric oxide synthase (NOS3) through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic (HC) humans and that a statin intervention would decrease arginase activity. Five microdialysis fibres were placed in the skin of nine normocholesterolaemic (NC: LDL level 95±4 mg dl⁻¹) and nine hypercholesterolaemic (HC: LDL: 177±6 mg dl⁻¹) men and women before and after 3 months of systemic atrovastatin. Sites served as control, NOS inhibited, arginase inhibited, L-arginine supplemented and arginase inhibited plus L-arginine supplemented. Skin blood flow was measured while local skin heating (42°C) induced NO-dependent vasodilatation. L-NAME was infused after the established plateau in all sites to quantify NO-dependent vasodilatation. Data were normalized to maximum cutaneous vascular conductance (CVC(max)). Skin samples were obtained to measure total arginase activity and arginase I and arginase II protein. Vasodilatation was reduced in hyperocholesterolaemic subjects (HC: 76±2 vs. NC: 94±3%CVC(max), P0.001) as was NO-dependent vasodilatation (HC: 43±5 vs. NC: 62±4%CVC(max), P0.001). The plateau and NO-dependent vasodilatation were augmented in HC with arginase inhibition (92±2, 67±2%CVC(max), P0.001), L-arginine (93±2, 71±5%CVC(max), P0.001) and combined treatments (94±4, 65±5%CVC(max), P0.001) but not in NC. After statin intervention (LDL: 98±5 mg dl⁻¹) there was no longer a difference between control sites (88±4, 61±5%CVC(max)) and localized microdialysis treatment sites (all P0.05). Arginase activity and protein were increased in HC skin (P0.05 vs. NC) and activity decreased with atrovastatin treatment (P0.05). Reduced NOS3 substrate availability through upregulated arginase contributes to cutaneous microvascular dysfunction in hyperocholesterolaemic humans, which is corrected with atorvastatin therapy.

We have recently demonstrated that tetrahydrobiopterin (BH4) augments reflex vasoconstriction (VC) in aged skin. Although this appears to occur through its role in norepinephrine (NE) biosynthesis, the extent with which vascular mechanisms are affected are unknown. We hypothesized that localized BH4 supplementation would not affect the VC response to exogenous NE when sympathetic nerves were blocked. Two microdialysis fibers were placed in bretylium tosylate pretreated (presynaptically blocks neurotransmitter release from sympathetic adrenergic nerve terminals; iontophoresis, 200 μA for 20 min) 3-cm2 forearm skin of 10 young (Y) and 10 older (O) subjects for perfusion of 1) Ringer (control) and 2) 5 mM BH4. While local skin temperature was clamped at 34°C, six concentrations of NE (10−12, 10−10, 10−8, 10−6, 10−4, 10−2 M) were infused at each drug-treated site. Cutaneous vascular conductance (CVC) was calculated (CVC = laser Doppler flux/mean arterial pressure) and normalized to baseline (%ΔCVCbase). Despite prejunctional adrenergic blockade, NE-mediated VC was blunted in aged skin at each NE dose (10−12: −12 ± 2 vs. −21 ± 2; 10−10: −15 ± 2 vs. −27 ± 1; 10−8: −22 ± 2 vs. −32 ± 2; 10−6: −27 ± 2 vs. −38 ± 1; 10−4: −52 ± 3 vs. −66 ± 5; 10−2: −62 ± 3 vs. −75 ± 4%ΔCVCbase; P < 0.01), and this response was not affected by pretreatment with BH4 ( P > 0.05). Localized BH4 did not affect end-organ responsiveness to exogenous NE, suggesting that the effects of BH4 on cutaneous VC are primarily isolated to the NE biosynthetic pathway.

Human skin blood flow is controlled via dual innervation from the sympathetic nervous system. Reflex cutaneous vasoconstriction and vasodilation are both impaired with primary aging, rendering the aged more vulnerable to hypothermia and cardiovascular complications from heat-related illness. Age-related alterations in the thermoregulatory control of skin blood flow occur at multiple points along the efferent arm of the reflex, including 1) diminished sympathetic outflow, 2) altered presynaptic neurotransmitter synthesis, 3) reduced vascular responsiveness, and 4) impairments in downstream (endothelial and vascular smooth muscle) second-messenger signaling. This mechanistic review highlights some of the recent findings in the area of aging and the thermoregulatory control of skin blood flow.

Chronic systemic platelet cyclooxygenase (COX) inhibition with low-dose aspirin [acetylsalicylic acid (ASA)] significantly attenuates reflex cutaneous vasodilation in middle-aged humans, whereas acute, localized, nonisoform-specific inhibition of vascular COX with intradermal administration of ketorolac does not alter skin blood flow during hyperthermia. Taken together, these data suggest that platelets may be involved in reflex cutaneous vasodilation, and this response is inhibited with systemic pharmacological platelet inhibition. We hypothesized that, similar to ASA, specific platelet ADP receptor inhibition with clopidogrel would attenuate reflex vasodilation in middle-aged skin. In a double-blind crossover design, 10 subjects (53 ± 2 yr) were instrumented with four microdialysis fibers for localized drug administration and heated to increase body core temperature [oral temperature (Tor)] 1°C during no systemic drug (ND), and after 7 days of systemic ASA (81 mg) and clopidogrel (75 mg) treatment. Skin blood flow (SkBF) was measured using laser-Doppler flowmetry over each site assigned as 1) control, 2) nitric oxide synthase inhibited (NOS-I; 10 mM NG-nitro-l-arginine methyl ester), 3) COX inhibited (COX-I; 10 mM ketorolac), and 4) NOS-I + COX-I. Data were normalized and presented as a percentage of maximal cutaneous vascular conductance (%CVCmax; 28 mM sodium nitroprusside + local heating to 43°C). During ND conditions, SkBF with change (Δ) in Tor = 1.0°C was 56 ± 3% CVCmax. Systemic low-dose ASA and clopidogrel both attenuated reflex vasodilation (ASA: 43 ± 3; clopidogrel: 32 ± 3% CVCmax; both P < 0.001). In all trials, localized COX-I did not alter SkBF during significant hyperthermia (ND: 56 ± 7; ASA: 43 ± 5; clopidogrel: 35 ± 5% CVCmax; all P > 0.05). NOS-I attenuated vasodilation in ND and ASA (ND: 28 ± 6; ASA: 25 ± 4% CVCmax; both P < 0.001), but not with clopidogrel (27 ± 4% CVCmax; P > 0.05). NOS-I + COX-I was not different compared with NOS-I alone in either systemic treatment condition. Both systemic ASA and clopidogrel reduced the time required to increase Tor 1°C (ND: 58 ± 3 vs. ASA: 45 ± 2; clopidogrel: 39 ± 2 min; both P < 0.001). ASA-induced COX and specific platelet ADP receptor inhibition attenuate reflex vasodilation, suggesting platelet involvement in reflex vasodilation through the release of vasodilating factors.

The attenuated reflex vasoconstriction in aged skin may be partly mediated by oxidant-induced reduction in functional substrate and cofactor availability for noradrenaline biosynthesis. We hypothesized that localized supplementation of tyrosine and tetrahydrobiopterin (BH(4)) in aged human skin could augment reflex- (whole-body cooling) and pharmacologically (tyramine, which displaces noradrenaline from axon terminals) induced vasoconstriction. Four microdialysis fibres were placed in the forearm skin of 10 young and 10 older subjects for infusion of (1) Ringer solution (control), (2) 0.5 mm L-tyrosine, (3) 5 mm BH(4), and (4) BH(4) + L-tyrosine. Cutaneous vascular conductance (CVC) was calculated (laser Doppler flux/mean arterial pressure) and normalized to baseline (% Delta CVC(base)). Vasoconstriction was attenuated at the control site in the older subjects during both whole-body cooling (young: 39 +/- 3, older: 17 +/- 3% Delta CVC(base); P0.01) and tyramine infusion (young: 41 +/- 3, older: 21 +/- 4% Delta CVC(base); P0.01). BH(4) (cold, young: 37 +/- 3, older: 36 +/- 3; tyramine, young: 41 +/- 2, older: 36 +/- 3% Delta CVC(base)) and tyrosine (cold, young: 37 +/- 4, older: 34 +/- 4; tyramine, young: 40 +/- 4, older: 45 +/- 4% Delta CVC(base)) both resolved the age-related decrease in cutaneous vasoconstriction, but BH(4) + tyrosine did not further augment vasoconstriction (cold, young: 38 +/- 4, older: 31 +/- 3; tyramine, young: 36 +/- 3, older: 36 +/- 5 Delta %CVC(base)). These data are consistent with the concept that reduced bioavailability of BH(4) and/or tyrosine may impair noradrenaline synthesis and contribute to the attenuated vasoconstrictor response in aged skin.

Full expression of reflex cutaneous vasodilation is dependent on cyclooxygenase- (COX) and nitric oxide synthase- (NOS) dependent mechanisms. Low-dose aspirin therapy is widely prescribed to inhibit COX-1 in platelets for atherothrombotic prevention. We hypothesized that chronic COX inhibition with daily low-dose aspirin therapy (81 mg) would attenuate reflex vasodilation in healthy human skin. Two microdialysis fibers were placed in forearm skin of seven middle-aged (57 ± 3 yr), normotensive, healthy humans with no preexisting cardiovascular disease, taking daily low-dose aspirin therapy (aspirin: 81 mg), and seven unmedicated, healthy, age-matched control (no aspirin, 55 ± 3 yr) subjects, with one site serving as a control (Ringer) and the other NOS inhibited (NOS inhibited: 10 mM NG-nitro-l-arginine methyl ester). Red cell flux was measured over each site by laser-Doppler flowmetry, as reflex vasodilation was induced by increasing core temperature (oral temperature) 1.0°C using a water-perfused suit. Cutaneous vascular conductance (CVC) was calculated (CVC = flux/mean arterial pressure) and normalized to maximal CVC (CVCmax; 28 mM sodium nitroprusside). CVCmax was not affected by either aspirin or NOS inhibition. The plateau in cutaneous vasodilation during heating (change in oral temperature = 1.0°C) was significantly attenuated in the aspirin group (aspirin: 25 ± 3% CVCmax vs. no aspirin: 50 ± 7% CVCmax, P < 0.001 between groups). NOS inhibition significantly attenuated %CVCmax in both groups (aspirin: 17 ± 2% CVCmax, no aspirin: 23 ± 3% CVCmax; P < 0.001 vs. control), but this attenuation was less in the no-aspirin treatment group ( P < 0.001). This is the first observation that chronic low-dose aspirin therapy attenuates reflex cutaneous vasodilation through both COX- and NOS-dependent mechanisms.

Ageing is associated with impaired endothelium-derived nitric oxide (NO) function in human microvessels. We investigated the impact of cardiorespiratory fitness and exercise training on physiological and pharmacological NO-mediated microvascular responses in older subjects. NO-mediated vasodilatation was examined in young, older sedentary and older fit subjects who had two microdialysis fibres embedded into the skin on the ventral aspect of the forearm and laser Doppler probes placed over these sites. Both sites were then heated to 42 degrees C, with Ringer solution infused in one probe and N-nitro-L-arginine methyl ester (L-NAME) through the second. In another study, three doses of ACh were infused in the presence or absence of L-NAME in similar subjects. The older sedentary subjects then undertook exercise training, with repeat studies at 12 and 24 weeks. The NO component of the heat-induced rise in cutaneous vascular conductance (CVC) was diminished in the older sedentary subjects after 30 min of prolonged heating at 42 degrees C (26.9 +/- 3.9%CVC(max)), compared to older fit (46.2 +/- 7.0%CVC(max), P0.05) and young subjects (41.2 +/- 5.2%CVC(max), P0.05), whereas exercise training in the older sedentary group enhanced NO-vasodilator function in response to incremental heating (P0.05). Similarly, the NO contribution to ACh responses was impaired in the older sedentary versus older fit subjects (low dose 3.2 +/- 1.3 versus 6.6 +/- 1.3%CVC(max); mid dose 11.4 +/- 2.4 versus 21.6 +/- 4.5%CVC(max); high dose 35.2 +/- 6.0 versus 52.6 +/- 7.9%CVC(max), P0.05) and training reversed this (12 weeks: 13.7 +/- 3.6, 28.9 +/- 5.3, 56.1 +/- 3.9%CVC(max), P0.05). These findings indicate that maintaining a high level of fitness, or undertaking exercise training, prevents age-related decline in indices of physiological and pharmacological microvascular NO-mediated vasodilator function. Since higher levels of NO confer anti-atherogenic benefit, this study has potential implications for the prevention of microvascular dysfunction in humans.

Human exposure to cold stimulates cutaneous vasoconstriction by activating both sympathetic reflex and locally mediated pathways. Older humans are vulnerable to hypothermia because primary aging impairs thermoregulatory cutaneous vasoconstriction. This article highlights recent findings discussing how age-related decrements in sympathetic neurotransmission contribute directly to thermoregulatory impairment, whereas changes in local cold-induced intracellular signaling suggest a more generalized age-associated vascular dysfunction.

Full expression of reflex cutaneous vasodilation (VD) is dependent on nitric oxide (NO) and is attenuated with essential hypertension. Decreased NO-dependent VD may be due to 1) increased oxidant stress and/or 2) decreased l-arginine availability through upregulated arginase activity, potentially leading to increased superoxide production through uncoupled NO synthase (NOS). The purpose of this study was to determine the effect of antioxidant supplementation (alone and combined with arginase inhibition) on attenuated NO-dependent reflex cutaneous VD in hypertensive subjects. Nine unmedicated hypertensive [HT; mean arterial pressure (MAP) = 112 ± 1 mmHg] and nine age-matched normotensive (NT; MAP = 81 ± 10 mmHg) men and women were instrumented with four intradermal microdialysis (MD) fibers: control (Ringer), NOS inhibited (NOS-I; 10 mM NG-nitro-l-arginine), l-ascorbate supplemented (Asc; 10 mM l-ascorbate), and Asc + arginase inhibited [Asc+A-I; 10 mM l-ascorbate + 5 mM ( S)-(2-boronoethyl)-l-cysteine-HCl + 5 mM Nω-hydroxy- nor-l-arginine]. Oral temperature was increased by 0.8°C via a water-perfused suit. NG-nitro-l-arginine was then ultimately perfused through all MD sites to quantify the change in VD due to NO. Red blood cell flux was measured by laser-Doppler flowmetry over each skin MD site, and cutaneous vascular conductance (CVC) was calculated (CVC = flux/MAP) and normalized to maximal CVC (%CVCmax; 28 mM sodium nitroprusside + local heating to 43°C). During the plateau in skin blood flow (ΔTor = 0.8°C), cutaneous VD was attenuated in HT skin (NT: 42 ± 4, HT: 35 ± 3 %CVCmax; P < 0.05). Asc and Asc+A-I augmented cutaneous VD in HT (Asc: 57 ± 5, Asc+A-I: 53 ± 6 %CVCmax; P < 0.05 vs. control) but not in NT. %CVCmax after NOS-I in the Asc- and Asc+A-I-treated sites was increased in HT (Asc: 41 ± 4, Asc+A-I: 40 ± 4, control: 29 ± 4; P < 0.05). Compared with the control site, the change in %CVCmax within each site after NOS-I was greater in HT (Asc: −19 ± 4, Asc+A-I: −17 ± 4, control: −9 ± 2; P < 0.05) than in NT. Antioxidant supplementation alone or combined with arginase inhibition augments attenuated reflex cutaneous VD in hypertensive skin through NO- and non-NO-dependent mechanisms.

Cutaneous vasoconstriction (VC) is the initial thermoregulatory response to cold exposure and can be elicited through either whole body or localized skin cooling. However, the mechanisms governing local cold-induced VC are not well understood. We tested the hypothesis that Rho kinase participates in local cold-induced cutaneous VC. In seven men and women (20–27 yr of age), up to four ventral forearm skin sites were instrumented with intradermal microdialysis fibers for localized drug delivery during cooling. Skin blood flow was monitored at each site with laser-Doppler flowmetry while local skin temperature was decreased and maintained at 24°C for 40 min. Cutaneous vascular conductance (CVC; laser-Doppler flowmetry/mean arterial pressure) was expressed as percent change from 34°C baseline. During the first 5 min of cooling, CVC decreased at control sites (lactated Ringer solution) to −45 ± 6% ( P < 0.001), increased at adrenoceptor-antagonized sites (yohimbine + propranolol) to 15 ± 14% ( P = 0.002), and remained unchanged at both Rho kinase-inhibited (fasudil) and adrenoceptor-antagonized + Rho kinase-inhibited sites (yohimbine + propranolol + fasudil) (−9 ± 1%, P = 0.4 and −6 ± 2%, P = 0.4, respectively). During the last 5 min of cooling, CVC further decreased at all sites when compared with baseline values (control, −77 ± 4%, P < 0.001; adrenoceptor antagonized, −61 ± 3%, P < 0.001; Rho kinase inhibited, −34 ± 7%, P < 0.001; and adrenoceptor antagonized + Rho kinase inhibited sites, −35 ± 3%, P < 0.001). Rho kinase-inhibited and combined treatment sites were significantly attenuated when compared with both adrenoceptor-antagonized ( P < 0.01) and control sites ( P < 0.0001). Rho kinase mediates both early- and late-phase cold-induced VC, supporting in vitro findings and providing a putative mechanism through which both adrenergic and nonadrenergic cold-induced VC occurs in an in vivo human thermoregulatory model.

Reflex-mediated cutaneous vasoconstriction (VC) is impaired in older humans; however, it is unclear whether this blunted VC also occurs during local cooling, which mediates VC through different mechanisms. We tested the hypothesis that the sensitization of cutaneous vessels to noradrenaline (NA) during direct skin cooling seen in young skin is blunted in aged skin. In 11 young (18-30 years) and 11 older (62-76 years) men and women, skin blood flow was monitored at two forearm sites with laser Doppler (LD) flowmetry while local skin temperature was cooled and clamped at 24 degrees C. Cutaneous vascular conductance (CVC; LD flux/mean arterial pressure) was expressed as percentage change from baseline (% DeltaCVC(base)). At one site, five doses of NA (10(-10)-10(-2) m) were sequentially infused via intradermal microdialysis during cooling while the other 24 degrees C site served as control (Ringer solution + cooling). At control sites, VC due to cooling alone was similar in young versus older (-54 +/- 5 versus -56 +/- 3% DeltaCVC(base), P = 0.46). In young, NA infusions induced additional dose-dependent VC (10(-8), 10(-6), 10(-4) and 10(-2) m: -70 +/- 2, -72 +/- 3, -78 +/- 3 and -79 +/- 4% DeltaCVC(base); P0.05 versus control). In older subjects, further VC did not occur until the highest infused dose of NA (10(-2) m: -70 +/- 5% DeltaCVC(base); P0.05 versus control). When cutaneous arterioles are sensitized to NA by direct cooling, young skin exhibits the capacity to further constrict to NA in a dose-dependent manner. However, older skin does not display enhanced VC capacity until treated with saturating doses of NA, possibly due to age-associated decrements in Ca2+ availability or alpha2C-adrenoceptor function.

Cutaneous vasoconstriction (VC) in response to cooling is impaired with human aging. On the basis of previous findings that older humans rely predominantly on norepinephrine (NE) for reflex VC of skin blood vessels, and that the VC effects of NE are blunted with age in many vascular beds, we tested the hypothesis that cutaneous VC responses to exogenous NE are attenuated in aged skin compared with young skin. In 11 young (18–30 yr) and 11 older (62–76 yr) men and women, skin blood flow was monitored at two forearm sites with laser Doppler (LD) flowmetry, while local skin temperature was clamped at 34°C. At one site, five doses of NE (10−10 to 10−2 M) were sequentially infused via intradermal microdialysis while the other site served as control (C; Ringer). Cutaneous vascular conductance (CVC; LD flux/mean arterial pressure) was expressed as percent change from baseline (%ΔCVCbase). At 10−10, 10−8, and 10−6 M NE, older VC responses were attenuated compared with young [10−10: −35 (95% confidence interval: −16, −52) vs. −49 (−40, −58) %ΔCVCbase, P = 0.02; 10−8: −38 (−20, −56) vs. −50 (−40, −61) %ΔCVCbase, P = 0.03; 10−6: −52 (−35, −70) vs. −67 (−60, −74) %ΔCVCbase, P = 0.01]. Older maximal VC responses were also blunted compared with young [−80 (confidence interval: −73,−87) vs. −88 (confidence interval: −87, −90) %ΔCVCbase, P = 0.03]. NE-mediated cutaneous VC is blunted at both physiological and superphysiological doses in older subjects compared with young subjects. Considering that NE is the only functional neurotransmitter mediating reflex VC in aged skin, attenuated NE-mediated VC may further predispose older humans to excess heat loss in the cold.

Moderate exercise elicits a relative postexercise hypotension that is caused by an increase in systemic vascular conductance. Previous studies have shown that skeletal muscle vascular conductance is increased postexercise. It is unclear whether these hemodynamic changes are limited to skeletal muscle vascular beds. The aim of this study was to determine whether the splanchnic and/or renal vascular beds also contribute to the rise in systemic vascular conductance during postexercise hypotension. A companion study aims to determine whether the cutaneous vascular bed is involved in postexercise hypotension (Wilkins BW, Minson CT, and Halliwill JR. J Appl Physiol 97: 2071–2076, 2004). Heart rate, arterial pressure, cardiac output, leg blood flow, splanchnic blood flow, and renal blood flow were measured in 13 men and 3 women before and through 120 min after a 60-min bout of exercise at 60% of peak oxygen uptake. Vascular conductances of leg, splanchnic, and renal vascular beds were calculated. One hour postexercise, mean arterial pressure was reduced (79.1 ± 1.7 vs. 83.4 ± 1.8 mmHg; P < 0.05), systemic vascular conductance was increased by ∼10%, leg vascular conductance was increased by ∼65%, whereas splanchnic (16.0 ± 1.8 vs. 18.5 ± 2.4 ml·min−1·mmHg−1; P = 0.13) and renal (20.4 ± 3.3 vs. 17.6 ± 2.6 ml·min−1·mmHg−1; P = 0.14) vascular conductances were unchanged compared with preexercise. This suggests there is neither vasoconstriction nor vasodilation in the splanchnic and renal vasculature during postexercise hypotension. Thus the splanchnic and renal vascular beds neither directly contribute to nor attenuate postexercise hypotension.

Reactive hyperemia is the sudden rise in blood flow after release of an arterial occlusion. Currently, the mechanisms mediating this response in the cutaneous circulation are poorly understood. The purpose of this study was to 1) characterize the reactive hyperemic response in the cutaneous circulation and 2) determine the contribution of nitric oxide (NO) to reactive hyperemia. Using laser-Doppler flowmetry, we characterized reactive hyperemia after 3-, 5-, 10-, and 15-min arterial occlusions in 10 subjects. The total hyperemic response was calculated by taking the area under the curve (AUC) of the hyperemic response minus baseline skin blood flow (SkBF) {i.e., total hyperemic response = AUC - [baseline SkBF as %maximal cutaneous vascular conductance (CVCmax) × duration of hyperemic response in s]}. For the characterization protocol, the total hyperemic response significantly increased as the period of ischemia increased from 5 to 15 min ( P < 0.05). However, the 3-min response was not significantly different from the 5-min response. In the NO contribution protocol, two microdialysis fibers were placed in the forearm skin of eight subjects. One site served as a control and was continuously perfused with Ringer solution. The second site was continuously perfused with 10 mM NG-nitro-l-arginine methyl ester (l-NAME) to inhibit NO synthase. CVC was calculated as flux/mean arterial pressure and normalized to maximal blood flow (28 mM sodium nitroprusside). The total hyperemic response in control sites was not significantly different from l-NAME sites after a 5-min occlusion (3,261 ± 890 vs. 2,907 ± 531% CVCmax · s). Similarly, total hyperemic responses in control sites were not different from l-NAME sites (9,155 ± 1,121 vs. 9,126 ± 1,088%CVCmax · s) after a 15-min arterial occlusion. These data suggest that NO does not directly mediate reactive hyperemia and that NO is not produced in response to an increase in shear stress in the cutaneous circulation.