Transient receptor potential vanilloid type 1 channels contribute to reflex cutaneous vasodilation in humans

Mechanisms underlying the cutaneous vasodilation in response to an increase in core temperature remain unresolved. The purpose of this study was to determine a potential contribution of transient receptor potential vanilloid type 1 (TRPV-1) channels to reflex cutaneous vasodilation. Twelve subjects were equipped with four microdialysis fibers on the ventral forearm, and each site randomly received 1) 90% propylene glycol + 10% lactated Ringer (vehicle control); 2) 10 mM l-NAME; 3) 20 mM capsazepine to inhibit TRPV-1 channels; 4) combined 10 mM l-NAME + 20 mM capsazepine. Whole body heating was achieved via water-perfused suits sufficient to raise oral temperature at least 0.8°C above baseline. Maximal skin blood flow was achieved by local heating to 43°C and infusion of 28 mM nitroprusside. Systemic arterial pressure (SAP) was measured, and skin blood flow was monitored via laser-Doppler flowmetry (LDF). Cutaneous vascular conductance (CVC) was calculated as LDF/SAP and normalized to maximal vasodilation (%CVC(max)). Capsazepine sites were significantly reduced compared with control (50 ± 4%CVC(max) vs. 67 ± 5%CVC(max), respectively; P < 0.05). l-NAME (33 ± 3%CVC(max)) and l-NAME + capsazepine (30 ± 4%CVC(max)) sites were attenuated compared with control (P < 0.01) and capsazepine (P < 0.05); however, there was no difference between l-NAME and combined l-NAME + capsazepine. These data suggest TRPV-1 channels participate in reflex cutaneous vasodilation and TRPV-1 channels may account for a portion of the NO component. TRPV-1 channels may have a direct neural contribution or have an indirect effect via increased arterial blood temperature. Whether the TRPV-1 channels directly or indirectly contribute to reflex cutaneous vasodilation remains uncertain.     

Endothelial nitric oxide synthase mediates cutaneous vasodilation during local heating and is attenuated in middle-aged human skin

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 [N(G)-nitro-l-arginine methyl ester (l-NAME)], inducible NOS (iNOS) inhibited (1400W), and neuronal NOS (nNOS) inhibited (N(ω)-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 (%CVC(max): 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%CVC(max); 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%CVC(max); P = 0.013) and ACh perfusion (50 mmol/l: 83 ± 3 vs. 93 ± 2%CVC(max); 100 mmol/l: 83 ± 4 vs. 92 ± 3%CVC(max); 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.     

Decreased nitric oxide- and axon reflex-mediated cutaneous vasodilation with age during local heating.

Cutaneous vasodilation is reduced in healthy older vs. young subjects; however, the mechanisms that underlie these age-related changes are unclear. Our goal in the present study was to determine the role of nitric oxide (NO) and the axon reflexes in the skin blood flow (SkBF) response to local heating with advanced age. We placed two microdialysis fibers in the forearm skin of 10 young (Y; 22 +/- 2 yr) and 10 older (O; 77 +/- 5 yr) men and women. SkBF over each site was measured by laser-Doppler flowmetry (LDF; Moor DRT4). Both sites were heated to 42 degrees C for ~60 min while 10 mM N(G)-nitro-L-arginine methyl ester (L-NAME) was infused throughout the protocol to inhibit NO synthase (NOS) in one site and 10 mM L-NAME was infused after 40 min of local heating in the second site. Data were expressed as a percentage of maximal vasodilation (%CVC(max); 28 mM nitroprusside infusion). Local heating before L-NAME infusion resulted in a significantly reduced initial peak (Y: 61 +/- 2%CVC(max) vs. O: 46 +/- 4%CVC(max)) and plateau (Y: 93 +/- 2%CVC(max) vs. O: 82 +/- 5%CVC(max)) CVC values in older subjects (P < 0.05). When NOS was inhibited after 40 min of heating, CVC declined to the same value in the young and older groups. Thus the overall contribution of NO to the plateau phase of the SkBF response to local heating was less in the older subjects. The initial peak response was significantly lower in the older subjects in both microdialysis sites (Y: 52 +/- 4%CVC(max) vs. O: 38 +/- 5%CVCmax; P < 0.05). These data suggest that age-related changes in both axon reflex-mediated and NO-mediated vasodilation contribute to attenuated cutaneous vasodilator responses in the elderly.     

Oral atorvastatin therapy increases nitric oxide-dependent cutaneous vasodilation in humans by decreasing ascorbate-sensitive oxidants

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% CVC(max), P < 0.01) and NO-dependent vasodilation (HC: 40 ± 4 vs. NC: 54 ± 4% CVC(max), P < 0.01) was reduced in the HC group. Acute L-ascorbate alone (91 ± 5% CVC(max), P < 0.001) or combined with arginase inhibition (96 ± 3% CVC(max), P < 0.001) augmented the plateau in vasodilation in the HC group but not the NC group (ascorbate: 96 ± 2; combo: 93 ± 4% CVC(max), both P > 0.05). After the atorvastatin intervention NO-dependent vasodilation was augmented in the HC group (HC postatorvastatin: 64 ± 4% CVC(max), P < 0.01), and there was no further effect of ascorbate alone (58 ± 4% CVC(max,) P > 0.05) or combined with arginase inhibition (67 ± 4% CVC(max,) P > 0.05). Increased ascorbate-sensitive oxidants contribute to hypercholesteromic associated cutaneous microvascular dysfunction which is partially reversed with atorvastatin therapy.     

Local ascorbate administration augments NO- and non-NO-dependent reflex cutaneous vasodilation in hypertensive humans

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 N(G)-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 N(omega)-hydroxy-nor-L-arginine]. Oral temperature was increased by 0.8 degrees C via a water-perfused suit. N(G)-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 (%CVC(max); 28 mM sodium nitroprusside + local heating to 43 degrees C). During the plateau in skin blood flow (Delta T(or) = 0.8 degrees C), cutaneous VD was attenuated in HT skin (NT: 42 +/- 4, HT: 35 +/- 3 %CVC(max); P < 0.05). Asc and Asc+A-I augmented cutaneous VD in HT (Asc: 57 +/- 5, Asc+A-I: 53 +/- 6 %CVC(max); P < 0.05 vs. control) but not in NT. %CVC(max) 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 %CVC(max) 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.     

Acute ascorbate supplementation alone or combined with arginase inhibition augments reflex cutaneous vasodilation in aged human skin

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 (T(or)) by 0.8 degrees 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 (CVC(max)). VD during heating was attenuated in O (Y: 37 +/- 3 vs. O: 28 +/- 3% CVC(max); P < 0.05). NOS-I decreased VD in both groups compared with Co (Y: 20 +/- 4; O: 15 +/- 2% CVC(max); P < 0.05 vs. Co within group). Asc and Asc + A-I increased VD beyond Co in O (Asc: 35 +/- 4% CVC(max); Asc + A-I: 41 +/- 3% CVC(max); P < 0.001) but not in Y (Asc: 36 +/- 3% CVC(max); Asc + A-I: 40 +/- 5% CVC(max); 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.     

Ascorbic acid or L-arginine improves cutaneous microvascular function in chronic kidney disease

We sought to determine whether oxidative stress or a relative deficit of l-arginine plays a role in reducing cutaneous vasodilation in response to local heating in chronic kidney disease (CKD). Eight patients with stage 3-4 CKD and eight age- and sex-matched healthy control (HC) subjects were instrumented with four microdialysis (MD) fibers for the local delivery of 1) Ringers solution (R), 2) 20 mM ascorbic acid (AA), 3) 10 mM l-arginine (l-Arg), and 4) 10 mM N(G)-nitro-l-arginine methyl ester (l-NAME). Red blood cell (RBC) flux was measured via laser Doppler flowmetry. A standardized nonpainful local heating protocol (42°C) was used. Cutaneous vascular conductance (CVC) was calculated as RBC flux/MAP and all data were expressed as a percentage of the maximum CVC at each site (28 mM sodium nitroprusside, T(loc) = 43°C). The plateau %CVC(max) was attenuated in CKD (CKD: 76 ± 4 vs. HC: 91 ± 2%CVC(max); P < 0.05) and the NO contribution to the plateau was lower in CKD (CKD: 39 ± 7, HC: 54 ± 5; P < 0.05). The plateau %CVC(max) in the CKD group was significantly greater at the AA and l-Arg sites compared with R (AA: 89 ± 2; l-Arg: 90 ± 1; R: 76 ± 4; P < 0.05) and did not differ from HC. Initial peak %CVC(max) was also significantly attenuated at the R and l-Arg sites in CKD (P < 0.05) but did not differ at the AA site. These results suggest that cutaneous microvascular function is impaired in stage 3-4 CKD and that oxidative stress and a deficit of l-arginine play a role in this impairment.     

Cutaneous constitutive nitric oxide synthase activation in postural tachycardia syndrome with splanchnic hyperemia

Models of microgravity are linked to excessive constitutive nitric oxide (NO) synthase (NOS), splanchnic vasodilation, and orthostatic intolerance. Normal-flow postural tachycardia syndrome (POTS) is a form of chronic orthostatic intolerance associated with splanchnic hyperemia. To test the hypothesis that there is excessive constitutive NOS in POTS, we determined whether cutaneous microvascular neuronal NO and endothelial NO are increased. We performed two sets of experiments in POTS and control subjects aged 21.4 ± 2 yr. We used laser-Doppler flowmetry to measure the cutaneous response to local heating as an indicator of bioavailable neuronal NO. To test for bioavailable endothelial NO, we infused intradermal acetylcholine through intradermal microdialysis catheters and used the selective neuronal NOS inhibitor l-N(ω)-nitroarginine-2,4-L-diamino-butyric amide (N(ω), 10 mM), the selective inducible NOS inhibitor aminoguanidine (10 mM), the nonspecific NOS inhibitor nitro-l-arginine (NLA, 10 mM), or Ringer solution. The acetylcholine dose response and the NO-dependent plateau of the local heating response were increased in POTS compared with those in control subjects. The local heating plateau was significantly higher, 98 ± 1%maximum cutaneous vascular conductance (%CVC(max)) in POTS compared with 88 ± 2%CVC(max) in control subjects but decreased to the same level with N(ω) (46 ± 5%CVC(max) in POTS compared with 49 ± 4%CVC(max) in control) or with NLA (45 ± 3%CVC(max) in POTS compared with 47 ± 4%CVC(max) in control). Only NLA blunted the acetylcholine dose response, indicating that NO produced by endothelial NOS was released by acetylcholine. Aminoguanidine was without effect. This is consistent with increased endothelial and neuronal NOS activity in normal-flow POTS.     

Tetrahydrobiopterin does not affect end-organ responsiveness to norepinephrine-mediated vasoconstriction in aged skin

We have recently demonstrated that tetrahydrobiopterin (BH(4)) 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 BH(4) 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-cm(2) forearm skin of 10 young (Y) and 10 older (O) subjects for perfusion of 1) Ringer (control) and 2) 5 mM BH(4). 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 (%ΔCVC(base)). 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%ΔCVC(base); P < 0.01), and this response was not affected by pretreatment with BH(4) (P > 0.05). Localized BH(4) did not affect end-organ responsiveness to exogenous NE, suggesting that the effects of BH(4) on cutaneous VC are primarily isolated to the NE biosynthetic pathway.     

Angiotensin II type 1 receptor blockade corrects cutaneous nitric oxide deficit in postural tachycardia syndrome

Low-flow postural tachycardia syndrome (POTS) is associated with increased plasma angiotensin II (ANG II) and reduced neuronal nitric oxide (NO), which decreases NO-dependent vasodilation. We tested whether the ANG II type 1 receptor (AT(1)R) antagonist losartan would improve NO-dependent vasodilation in POTS patients. Furthermore, if the action of ANG II is dependent on NO, then the NO synthase inhibitor nitro-L-arginine (NLA) would reverse this improvement. We used local heating of the skin of the left calf to 42 degrees C and laser-Doppler flowmetry to assess NO-dependent conductance [percent maximum cutaneous vascular conductance (%CVC(max))] in 12 low-flow POTS patients aged 22.5 +/- 0.8 yr and in 15 control subjects aged 22.0 +/- 1.3 yr. After measuring the baseline local heating response at three separate sites, we perfused individual intradermal microdialysis catheters at those sites with 2 microg/l losartan, 10 mM NLA, or losartan + NLA. The predrug heat response was reduced in POTS, particularly the plateau phase reflecting NO-dependent vasodilation (50 +/- 5 vs. 91 +/- 7 %CVC(max); P < 0.001 vs. control). Losartan increased baseline flow in both POTS and control subjects (from 6 +/- 1 to 21 +/- 3 vs. from 10 +/- 1 to 21 +/- 2 %CVC(max); P < 0.05 compared with predrug). The baseline increase was blunted by NLA. Losartan increased the POTS heat response to equal the control subject response (79 +/- 7 vs. 88 +/- 6 %CVC(max); P = 0.48). NLA decreased both POTS and control subject heat responses to similar conductances (38 +/- 4 vs. 38 +/- 3 %CVC(max); P < 0.05 compared with predrug). The addition of NLA to losartan reduced POTS and control subject conductances compared with losartan alone (48 +/- 3 vs. 53 +/- 2 %CVC(max)). The data suggest that the reduction in cutaneous NO-dependent vasodilation in low-flow POTS is corrected by AT(1)R blockade.     

Nitric oxide and attenuated reflex cutaneous vasodilation in aged skin

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 degrees 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 (CVC(max)) to 53 +/- 3% CVC(max) in the young subjects and from 64 +/- 5% CVC(max) to 29 +/- 2% CVC(max) in the older subjects with a 1.0 degrees C rise in core temperature. Thus the relative NO-dependent portion of cutaneous active vasodilation (AVD) accounted for approximately 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.     

Cyclooxygenase and nitric oxide synthase dependence of cutaneous reactive hyperemia in humans

We tested the hypothesis that cyclooxygenases (COXs) or COX products inhibit nitric oxide (NO) synthesis and thereby mask potential effects of NO on reactive hyperemia in the cutaneous circulation. We performed laser-Doppler flowmetry (LDF) with intradermal microdialysis in 12 healthy volunteers aged 19-25 yr. LDF was expressed as the percent cutaneous vascular conduction (%CVC) or as the maximum %CVC (%CVC(max)) where CVC is LDF/mean arterial pressure. We tested the effects of the nonisoform-specific NO synthase inhibitor nitro-L-arginine (NLA, 10 mM), the nonspecific COX inhibitor ketorolac (Keto, 10 mM), combined NLA + Keto, and NLA + sodium nitroprusside (SNP, 28 mM) on baseline and reactive hyperemia flow parameters. We also examined the effects of isoproterenol, a beta-adrenergic agonist that causes prostaglandin-independent vasodilation to correct for the increase in baseline flow caused by Keto. When delivered directly into the intradermal space, Keto greatly augments all aspects of the laser-Doppler flow response to reactive hyperemia: peak reactive hyperemic flow increased from 41 +/- 5 to 77 +/- 7%CVC(max), time to peak flow increased from 17 +/- 3 to 56 +/- 24 s, the area under the reactive hyperemic curve increased from 1,417 +/- 326 to 3,376 +/- 876%CVC(max).s, and the time constant for the decay of peak flow increased from 100 +/- 23 to 821 +/- 311 s. NLA greatly attenuates the Keto response despite exerting no effects on baseline LDF or on reactive hyperemia when given alone. Low-dose NLA + SNP duplicates the Keto response. Isoproterenol increased baseline and peak reactive flow. These results suggest that COX inhibition unmasks NO dependence of reactive hyperemia in human cutaneous circulation.     

Antagonism of soluble guanylyl cyclase attenuates cutaneous vasodilation during whole body heat stress and local warming in humans

We hypothesized that nitric oxide activation of soluble guanylyl cyclase (sGC) participates in cutaneous vasodilation during whole body heat stress and local skin warming. We examined the effects of the sGC inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), on reflex skin blood flow responses to whole body heat stress and on nonreflex responses to increased local skin temperature. Blood flow was monitored by laser-Doppler flowmetry, and blood pressure by Finapres to calculate cutaneous vascular conductance (CVC). Intradermal microdialysis was used to treat one site with 1 mM ODQ in 2% DMSO and Ringer, a second site with 2% DMSO in Ringer, and a third site received Ringer. In protocol 1, after a period of normothermia, whole body heat stress was induced. In protocol 2, local heating units warmed local skin temperature from 34 to 41°C to cause local vasodilation. In protocol 1, in normothermia, CVC did not differ among sites [ODQ, 15 ± 3% maximum CVC (CVC(max)); DMSO, 14 ± 3% CVC(max); Ringer, 17 ± 6% CVC(max); P > 0.05]. During heat stress, ODQ attenuated CVC increases (ODQ, 54 ± 4% CVC(max); DMSO, 64 ± 4% CVC(max); Ringer, 63 ± 4% CVC(max); P < 0.05, ODQ vs. DMSO or Ringer). In protocol 2, at 34°C local temperature, CVC did not differ among sites (ODQ, 17 ± 2% CVC(max); DMSO, 18 ± 4% CVC(max); Ringer, 18 ± 3% CVC(max); P > 0.05). ODQ attenuated CVC increases at 41°C local temperature (ODQ, 54 ± 5% CVC(max); DMSO, 86 ± 4% CVC(max); Ringer, 90 ± 2% CVC(max); P < 0.05 ODQ vs. DMSO or Ringer). sGC participates in neurogenic active vasodilation during heat stress and in the local response to direct skin warming.     

Ascorbate improves circulation in postural tachycardia syndrome

Low flow postural tachycardia syndrome (LFP) is associated with vasoconstriction, reduced cardiac output, increased plasma angiotensin II, reduced bioavailable nitric oxide (NO), and oxidative stress. We tested whether ascorbate would improve cutaneous NO and reduce vasoconstriction when delivered systemically. We used local cutaneous heating to 42°C and laser Doppler flowmetry to assess NO-dependent conductance (%CVC(max)) to sodium ascorbate and the systemic hemodynamic response to ascorbic acid in 11 LFP patients and in 8 control subjects (aged 23 ± 2 yr). We perfused intradermal microdialysis catheters with sodium ascorbate (10 mM) or Ringer solution. Predrug heat response was reduced in LFP, particularly the NO-dependent plateau phase (56 ± 6 vs. 88 ± 7%CVC(max)). Ascorbate increased baseline skin flow in LFP and control subjects and increased the LFP plateau response (82 ± 6 vs. 92 ± 6 control). Systemic infusion experiments used Finometer and ModelFlow to estimate relative cardiac index (CI) and forearm and calf venous occlusion plethysmography to estimate blood flows, peripheral arterial and venous resistances, and capacitance before and after infusing ascorbic acid. CI increased 40% after ascorbate as did peripheral flows. Peripheral resistances were increased (nearly double control) and decreased by nearly 50% after ascorbate. Calf capacitance and venous resistance were decreased compared with control but normalized with ascorbate. These data provide experimental support for the concept that oxidative stress and reduced NO possibly contribute to vasoconstriction and venoconstriction of LFP.     

Aging and the skin blood flow response to the unloading of baroreceptors during heat and cold stress.

Control of skin blood flow (SkBF) is on the efferent arm of both thermoregulatory and nonthermoregulatory reflexes. To what extent aging may affect the SkBF response when these two reflex systems interact is unknown. To determine the response of aged skin to the unloading of baroreceptors in thermoneutral, cold stress, and heat stress conditions, sequential bouts of nonhypotensive lower body negative pressure (LBNP) were applied at -10, -20, and -30 mmHg in 14 young (18-25 yr) and 14 older (63-78 yr) men. SkBF was measured by laser-Doppler velocimetry (averaged over 2 forearm sites), and data are expressed as percentage of maximal cutaneous vascular conductance (%CVC(max)). Total forearm blood flow was measured by venous occlusion plethysmography, and forearm vascular conductance (FVC) was calculated as the ratio of forearm blood flow to mean arterial pressure. In young men, all three intensities of LBNP in thermoneutrality decreased FVC significantly (P < 0.05), but FVC at -10 mmHg did not change in the older men. There were no significant LBNP effects on %CVC(max). Application of LBNP during cold stress did not significantly change %CVC(max) or FVC in either age group. During heat stress, -10 to -30 mmHg of LBNP decreased FVC significantly (P < 0.05) in both age groups, but these decreases were attenuated in the older men (P < 0.05). %CVC(max) decreased at -30 mmHg in the younger men only. These results suggest that older men have an attenuated skin vasoconstrictor response to the unloading of baroreceptors in heat stress conditions. Furthermore, the forearm vasoconstriction elicited by LBNP in older men reflects that of underlying tissue (i.e., muscle) rather than that of skin, whereas -30 mmHg LBNP also decreases SkBF in young hyperthermic men.     

Aging and aerobic fitness affect the contribution of noradrenergic sympathetic nerves to the rapid cutaneous vasodilator response to local heating

Sedentary aging results in a diminished rapid cutaneous vasodilator response to local heating. We investigated whether this diminished response was due to altered contributions of noradrenergic sympathetic nerves by assessing 1) the age-related decline and 2) the effect of aerobic fitness. Using laser-Doppler flowmetry, we measured skin blood flow (SkBF) in young (24 ± 1 yr) and older (64 ± 1 yr) endurance-trained and sedentary men (n = 7 per group) at baseline and during 35 min of local skin heating to 42°C at 1) untreated forearm sites, 2) forearm sites treated with bretylium tosylate (BT), which prevents neurotransmitter release from noradrenergic sympathetic nerves, and 3) forearm sites treated with yohimbine + propranolol (YP), which antagonizes α- and β-adrenergic receptors. SkBF was converted to cutaneous vascular conductance (CVC = SkBF/mean arterial pressure) and normalized to maximal CVC (%CVC(max)) achieved by skin heating to 44°C. Pharmacological agents were administered using microdialysis. In the young trained group, the rapid vasodilator response was reduced at BT and YP sites (P < 0.05); by contrast, in the young sedentary and older trained groups, YP had no effect (P > 0.05), but BT did (P > 0.05). Neither BT nor YP affected the rapid vasodilator response in the older sedentary group (P > 0.05). These data suggest that the age-related reduction in the rapid vasodilator response is due to an impairment of sympathetic-dependent mechanisms, which can be partly attenuated with habitual aerobic exercise. Rapid vasodilation involves noradrenergic neurotransmitters in young trained men and nonadrenergic sympathetic cotransmitters (e.g., neuropeptide Y) in young sedentary and older trained men, possibly as a compensatory mechanism. Finally, in older sedentary men, the rapid vasodilation appears not to involve the sympathetic system.     

Cutaneous neuronal nitric oxide is specifically decreased in postural tachycardia syndrome

Low flow postural tachycardia syndrome (POTS), is associated with reduced nitric oxide (NO) activity assumed to be of endothelial origin. We tested the hypothesis that cutaneous microvascular neuronal NO (nNO) is impaired, rather than endothelial NO (eNO), in POTS. We performed three sets of experiments on subjects aged 22.5 +/- 2 yr. We used laser-Doppler flowmetry response to sequentially increase acetylcholine (ACh) doses and the local cutaneous heating response of the calf as bioassays for NO. During local heating we showed that when the selective neuronal nNO synthase (nNOS) inhibitor N(omega)-nitro-L-arginine-2,4-L-diaminobutyric amide (N(omega), 10 mM) was delivered by intradermal microdialysis, cutaneous vascular conductance (CVC) decreased by an amount equivalent to the largest reduction produced by the nonselective NO synthase (NOS) inhibitor nitro-L-arginine (NLA, 10 mM). We demonstrated that the response to ACh was minimally attenuated by nNOS blockade using N(omega) but markedly attenuated by NLA, indicating that eNO largely comprises the receptor-mediated NO release by ACh. We further demonstrated that the ACh dose response was minimally reduced, whereas local heat-mediated NO-dependent responses were markedly reduced in POTS compared with control subjects. This is consistent with intact endothelial function and reduced NO of neuronal origin in POTS. The local heating response was highly attenuated in POTS [60 +/- 6 percent maximum CVC(%CVC(max))] compared with control (90 +/- 4 %CVC(max)), but the plateau response decreased to the same level with nNOS inhibition (50 +/- 3 %CVC(max) in POTS compared with 47 +/- 2 %CVC(max)), indicating reduced nNO bioavailability in POTS patients. The data suggest that nNO activity but not NO of endothelial NOS origin is reduced in low-flow POTS.     

Nitric oxide and neurally mediated regulation of skin blood flow during local heating.

The mechanisms underlying the skin blood flow (SkBF) response to local heating are complex and poorly understood. Our goal was to examine the role of axon reflexes and nitric oxide (NO) in the SkBF response to a local heating protocol. We performed 40 experiments following a standardized heating protocol with different interventions, including blockade of the axon reflex (EMLA cream), antebrachial nerve blockade (0.5% bupivacaine injection), and NO synthase (NOS) inhibition (> or =10 mM N(G)-nitro-L-arginine methyl ester; microdialysis). Appropriate controls were performed to verify the efficacy of the various blocks. Values are expressed as a percentage of maximal SkBF (SkBF(max); 50 mM sodium nitroprusside). At the initiation of local heating, SkBF rose to an initial peak, followed by a brief nadir, and a secondary, progressive rise to a plateau. Axon reflex block decreased the initial peak from 75+3 to 32 +/- 2% SkBF(max) (P < 0.01 vs. control) but did not affect the plateau. NOS inhibition before and throughout local heating reduced the initial peak from 75 +/- 3 to 56 +/- 3% SkBF(max) (P < 0.01) and the plateau from 87 +/- 4 to 40 +/- 5%. NOS inhibition during axon reflex block did not further reduce the initial SkBF peak compared with axon reflex block alone. Antebrachial nerve block did not affect the local heating SkBF response. The primary finding of these studies is that there are at least two independent mechanisms contributing to the rise in SkBF during nonpainful local heating: a fast-responding vasodilator system mediated by the axon reflexes and a more slowly responding vasodilator system that relies on local production of NO.     

Intradermal angiotensin II administration attenuates the local cutaneous vasodilator heating response

The vasodilation response to local cutaneous heating is nitric oxide (NO) dependent and blunted in postural tachycardia but reversed by angiotensin II (ANG II) type 1 receptor (AT(1)R) blockade. We tested the hypothesis that a localized infusion of ANG II attenuates vasodilation to local heating in healthy volunteers. We heated the skin of a calf to 42 degrees C and measured local blood flow to assess the percentage of maximum cutaneous vascular conductance (%CVC(max)) in eight healthy volunteers aged 19.5-25.5 years. Initially, two experiments were performed; in one, Ringer solution was perfused in three catheters, the response to heating was measured, 2 microg/l losartan, 10 mM nitro-l-arginine (NLA), or NLA + losartan was added to perfusate, and the heat response was remeasured; in another, 10 microM ANG II was given, the heat response was measured, losartan, NLA, or NLA + losartan was added to ANG II, and the heat response was reassessed. The heat response decreased with ANG II, particularly the plateau phase (47 +/- 5 vs. 84 +/- 3 %CVC(max)). Losartan increased baseline conductance in both experiments (from 8 +/- 1 to 20 +/- 2 and 12 +/- 1 to 24 +/- 3). Losartan increased the ANG II response (83 +/- 4 vs. 91 +/- 6 in Ringer). NLA decreased both angiotensin and Ringer responses (31 +/- 4 vs. 43 +/- 3). NLA + losartan blunted the Ringer response (48 +/- 2), but the ANG II response (74 +/- 5) increased. In a second set of experiments, we used dose responses to ANG II (0.1 nM to 10 microM) with and without NLA + losartan to confirm graded responses. Sodium ascorbate (10 mM) restored the ANG II-blunted heating plateau. NO synthase and AT(1)R inhibition cause an NO-independent angiotensin-mediated vasodilation with local heating. ANG II mediates the AT(1)R blunting of local heating, which is not exclusively NO dependent, and is improved by antioxidant supplementation.     

Vasoactive intestinal peptide fragment VIP10-28 and active vasodilation in human skin.

A recent study reported the vasoactive intestinal peptide (VIP) fragment VIP(10-28) inhibited the rise in skin blood flow during heat stress. Our laboratory has reported that the nitric oxide (NO) pathway and histamine receptor-1 (H1)-receptor activation is common to both exogenous VIP-mediated dilation and active vasodilation (AVD). The present study aimed to further examine the specific role for VIP in AVD by using VIP(10-28) to antagonize VIP-mediated dilation in the presence of NO synthase (NOS) inhibition and an H1 antagonist. Study 1 (n = 12) examined whether VIP(10-28) antagonizes vasodilation to exogenous VIP via inhibition of NO-dependent mechanisms. Study 2 (n = 6) investigated AVD in skin sites receiving VIP(10-28) alone and in combination with NOS inhibition. Study 3 (n = 6) examined AVD in sites receiving VIP(10-28) alone and combined VIP(10-28) and H1 antagonism. Due to differences in our findings and those previously published, study 4 (n = 6) investigated whether an increase in baseline skin blood flow could result in a diminished rise in AVD. Red blood cell flux was measured using laser Doppler flowmetry, and cutaneous vascular conductance (flux/mean arterial pressure) was normalized to maximal vasodilation (28 mM sodium nitroprusside). VIP(10-28) augmented vasodilation to exogenous VIP (P < 0.05 vs. control) and hyperthermia (P < 0.05 vs. control). NOS inhibition had no effect on the augmented dilation during exogenous VIP or hyperthermia (P > 0.05). Similarly, H1-receptor antagonists had no effect on the augmented dilation during hyperthermia (P > 0.05 vs. VIP(10-28)). In study 4, percentage of maximal cutaneous vascular conductance was attenuated when baseline skin blood flow was elevated before whole body heating. Our results suggest that VIP(10-28) may be an unsuitable antagonist for examining a role for VIP-mediated dilation in human skin.