Acetylcholine-induced vasodilation is mediated by nitric oxide and prostaglandins in human skin.

Acetylcholine (ACh) can effect vasodilation by several mechanisms, including activation of endothelial nitric oxide (NO) synthase and prostaglandin (PG) production. In human skin, exogenous ACh increases both skin blood flow (SkBF) and bioavailable NO levels, but the relative increase is much greater in SkBF than NO. This led us to speculate ACh may dilate cutaneous blood vessels through PGs, as well as NO. To test this hypothesis, we performed a study in 11 healthy people. We measured SkBF by laser-Doppler flowmetry (LDF) at four skin sites instrumented for intradermal microdialysis. One site was treated with ketorolac (Keto), a nonselective cyclooxygenase antagonist. A second site was treated with NG-nitro-L-arginine methyl ester (L-NAME) to inhibit NO synthase. A third site was treated with a combination of Keto and L-NAME. The fourth site was an untreated control site. After the three treated sites received the different inhibiting agents, ACh was administered to all four sites by intradermal microdialysis. Finally, sodium nitroprusside (SNP) was administered to all four sites. Mean arterial pressure (MAP) was monitored by Finapres, and cutaneous vascular conductance (CVC) was calculated (CVC = LDF/MAP). For data analysis, CVC values for each site were normalized to their respective maxima as effected by SNP. The results showed that both Keto and L-NAME each attenuated the vasodilation induced by exogenous ACh (ACh control = 79 +/- 4% maximal CVC, Keto = 55 +/- 7% maximal CVC, L-NAME = 46 +/- 6% maximal CVC; P < 0.05, ACh vs. Keto or L-NAME). The combination of the two agents produced an even greater attenuation of ACh-induced vasodilation (31 +/- 5% maximal CVC; P < 0.05 vs. all other sites). We conclude that a portion of the vasodilation effected by exogenous ACh in skin is due to NO; however, a significant portion is also mediated by PGs.     

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.     

Bradykinin does not mediate cutaneous active vasodilation during heat stress in humans.

To test the hypothesis that bradykinin effects cutaneous active vasodilation during hyperthermia, we examined whether the increase in skin blood flow (SkBF) during heat stress was affected by blockade of bradykinin B(2) receptors with the receptor antagonist HOE-140. Two adjacent sites on the forearm were instrumented with intradermal microdialysis probes for local delivery of drugs in eight healthy subjects. HOE-140 was dissolved in Ringer solution (40 microM) and perfused at one site, whereas the second site was perfused with Ringer alone. SkBF was monitored by laser-Doppler flowmetry (LDF) at both sites. Mean arterial pressure (MAP) was monitored from a finger, and cutaneous vascular conductance (CVC) was calculated (CVC = LDF/MAP). Water-perfused suits were used to control body temperature and evoke hyperthermia. After hyperthermia, both microdialysis sites were perfused with 28 mM nitroprusside to effect maximal vasodilation. During hyperthermia, CVC increased at HOE-140 (69 +/- 2% maximal CVC, P < 0.01) and untreated sites (65 +/- 2% maximal CVC, P < 0.01). These responses did not differ between sites (P > 0.05). Because the bradykinin B(2)-receptor antagonist HOE-140 did not alter SkBF responses to heat stress, we conclude that bradykinin does not mediate cutaneous active vasodilation.     

Effect of nonsteroidal anti-inflammatory drugs with varying extent of COX-2-COX-1 selectivity on urinary sodium and potassium excretion in the rat

Nonsteroidal anti-inflammatory drugs (NSAIDs) have different selectivity to inhibit cyclooxygenase-1 (COX-1) and COX-2. Treatment with NSAIDs has been associated with kidney side effects. We compared the effect of a selected group of NSAIDs with different COX-2--COX-1 selectivities on urinary sodium and potassium excretion in rats. Each treatment with rofecoxib, celecoxib, meloxicam, diclofenac, and flurbiprofen (30, 120, 9, 30, and 125 mg/kg, respectively) and placebo was administered orally once daily for 4 days. Urine was collected 0-8 h after each dose. Urinary sodium and potassium excretion and urine flow rate were compared with placebo. As compared with placebo, rofecoxib, celecoxib, diclofenac, and flurbiprofen significantly reduced excretion rate of sodium (rofecoxib, 0.28 +/- 0.02 vs. 0.41 +/- 0.03; celecoxib, 0.23 +/- 0.03 vs. 0.48 +/- 0.04; diclofenac, 0.09 +/- 0.02 vs. 0.46 +/- 0.03; and flurbiprofen, 0.11 +/- 0.02 vs. 0.47 +/- 0.02 micromol/(min x 100 g)) and potassium (rofecoxib, 0.55 +/- 0.04 vs. 0.68 +/- 0.04; celecoxib, 0.50 +/- 0.06 vs. 0.72 +/- 0.06; diclofenac, 0.26 +/- 0.05 vs. 0.67 +/- 0.04; and flurbiprofen, 0.35 +/- 0.05 vs. 0.62 +/- 0.03 micromol/ (min x 100 g)). Rofecoxib and flurbiprofen significantly reduced urine flow rate. Meloxicam had no significant effect on either sodium and potassium excretion or on the urine flow rate. At the examined dosage level, no relationship was found between reported COX-2--COX-1 selectivity and urinary electrolytes excretion.     

Nitric oxide synthase inhibition does not alter the reactive hyperemic response in the cutaneous circulation.

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 (CVC(max) x 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 (3261 +/- 890 vs. 2907 +/- 531% CVC(max. s). Similarly, total hyperemic responses in control sites were not different from l-NAME sites (9155 +/- 1121 vs. 9126 +/- 1088% CVC(max. 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.     

Cyclooxygenase-2 inhibitors constrict the fetal lamb ductus arteriosus both in vitro and in vivo

Nonselective cyclooxygenase (COX) inhibitors are potent tocolytic agents; however, they also have adverse fetal effects such as constriction of the fetal ductus arteriosus. Recently, selective COX-2 inhibitors have been used in the management of preterm labor in the hope of avoiding fetal complications. However, both COX-1 and -2 are expressed by cells of the ductus arteriosus. We used fetal lambs (0.88 gestation) to assess the ability of selective COX-2 inhibitors celecoxib and NS398 to affect the ductus arteriosus. Both selective COX-2 inhibitors decreased PGE(2) and 6ketoPGF(1alpha) production in vitro; both inhibitors constricted the isolated ductus in vitro. The nonselective COX-1/COX-2 inhibitor indomethacin produced a further reduction in PG release and an additional increase in ductus tension in vitro. We used a prodrug of celecoxib to achieve 1.4 +/- 0.6 microg/ml, mean +/- standard deviation, of the active drug in vivo. This concentration of celecoxib produced both an increase in pressure gradient and resistance across the ductus; celecoxib also decreased fetal plasma concentrations of PGE(2) and 6ketoPGF(1alpha). Indomethacin (0.7 +/- 0.2 microg/ml) produced a significantly greater fall in ductus blood flow than celecoxib and tended to have a greater effect on ductus resistence in vivo. We conclude that caution should be used when recommending COX-2 inhibitors for use in pregnant women, because COX-2 appears to play a significant role in maintaining patency of the fetal ductus arteriosus.     

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.     

Age-specific modification of local cutaneous vasodilation by capsaicin-sensitive primary afferents

Age-related changes in neurogenic vasodilation mediated by sensory nerves may alter local skin blood flow (SkBF) responses in older individuals. The purpose of this study was to determine the age-specific modification of cutaneous vasodilation by capsaicin-sensitive primary afferent (CSPA) nerves during local heating. Nine young (18-30 yr), eight middle-aged (40-55 yr), and eight older (65-80 yr) healthy men participated in the experiments. Two local-heating protocols (rapid and slow) were performed before and after 1 wk of capsaicin pretreatment (CP), used to desensitize CSPAs. All temperatures were below those that elicit pain. SkBF was measured with a laser-Doppler imager and indexed to percentage of maximal cutaneous vascular conductance (%CVCmax). CP caused a significant reduction in %CVCmax in the middle-aged and older groups during slow heating (P < 0.05), without affecting %CVCmax in the young group. During rapid heating, CP significantly reduced (53.9 +/- 4.4 vs. 74.4 +/- 7.4% CVCmax, P < 0.05), but did not abolish, the initial sensory nerve-mediated rise in SkBF in the young group. No significant effects of CP on SkBF were observed during rapid heating in the middle-aged or older groups. These results indicate that, with advanced age, CSPA activity is more important to the maximal SkBF response during prolonged local heating, whereas it has a reduced role in the initial SkBF peak elicited by rapid local heating. In summary, CSPA activity contributes modestly to the overall SkBF response to local heating in an age-specific manner.     

Prolonged aspirin inhibition of anodal vasodilation is not due to the trafficking delay of neural mediators

We previously reported that forearm vasodilation to a delivered all-at-once over 5 min or a 1-min repeated monopolar anodal 0.10-mA current application is aspirin sensitive and that a single high-dose aspirin exerts a long-lived effect in the former case. We hypothesized that 1) in the latter case, the effect of aspirin would also be long lived and 2) the time required to resupply nerve endings with unblocked cyclooxygenase through axonal transport could explain this phenomenon. We studied the time course for the recovery of vasodilation to repeated current application after placebo or 1-g aspirin treatment. We then searched for a difference at a proximal vs. distal site in the recovery of the response. Aspirin abolished current-induced vasodilation at 2 h, 10 h, and 3 days, with a progressive recovery thereafter, but no difference between distal and proximal site was observed for the recovery of the response. This suggests that, although neural cyclooxygenase could participate in the response, the time course of aspirin inhibition of current-induced cutaneous vasodilation is not due to the time required through neural transport to resupply nerve endings with unblocked proteins.     

Cathodal current-induced vasodilation to single application and the amplified response to repeated application in humans rely on aspirin-sensitive mechanisms.

Assumed to rely on an axon reflex, the current-induced vasodilation (CIV) interferes with the microvascular response to iontophoretic drug delivery. Mechanisms resulting in CIV are likely different at the anode and at the cathode. While studies have been conducted to understand anodal CIV, little information is available on cathodal CIV. The present study investigates CIV observed following 0.1-mA cathodal applications on forearms of healthy volunteers and the possible mechanisms involved. Results are expressed in percentage of the cutaneous heat-induced maximal vascular conductance [%MVC (means +/- SE)]. 1) The amplitude of CIV was proportional to the duration of cathodal currents for periods of <1 min: r = 0.99. 2) Two current applications of 10 s, with 10-min interstimulation interval, induced a higher peak value of CIV (79.1 +/- 8.6% MVC) than the one obtained with all-at-once 20-s current application (39.5 +/- 4.3% MVC, P < 0.05). This amplified vascular response due to segmental application was observed for all tested interstimulation intervals (up to 40 min). 3) Two hours and 3 days following pretreatment with 1-g oral aspirin, the CIV observed following cathodal application, as well as the difference of cathodal CIV amplitude between all-at-once and segmented applications, were reduced. These findings suggest a role of prostaglandins, not only released from endothelial or smooth muscle cells, as direct vasodilator and/or as a sensitizer. Thus aspirin pretreatment could be used to decrease CIV resulting from all-at-once and repeated cathodal application and facilitate the study of the specific vascular effect induced by the drug delivered.     

Relative roles of local and reflex components in cutaneous vasoconstriction during skin cooling in humans.

The reduction in skin blood flow (SkBF) with cold exposure is partly due to the reflex vasoconstrictor response from whole body cooling (WBC) and partly to the direct effects of local cooling (LC). Although these have been examined independently, little is known regarding their roles when acting together, as occurs in environmental cooling. We tested the hypothesis that the vasoconstrictor response to combined LC and WBC would be additive, i.e., would equal the sum of their independent effects. We further hypothesized that LC would attenuate the reflex vasoconstrictor response to WBC. We studied 16 (7 women, 9 men) young (30.5+/-2 yr) healthy volunteers. LC and WBC were accomplished with metal Peltier cooler-heater probe holders and water-perfused suits, respectively. Forearm SkBF was monitored by laser-Doppler flowmetry (LDF). Cutaneous vascular conductance (CVC) was calculated as LDF/blood pressure. Subjects underwent 15 min of LC alone or 15 min of WBC with and without simultaneous LC, either at equal levels (34-31 degrees C) or as equipotent stimuli (34-28 degrees C LC; 34-31 degrees C WBC). The fall in CVC with combined WBC and LC was greater (P<0.05) than for either alone (57.0+/-5% combined vs. 39.2+/-6% WBC; 34.4+/-4% LC) with equipotent cooling, but it was only significantly greater than for LC alone with equal levels of cooling (51.3+/-8% combined vs. 29.5+/-4% LC). The sum of the independent effects of WBC and LC was greater than their combined effects (74.9+/-4 vs. 51.3+/-8% equal and 73.6+/-7 vs. 57.0+/-5% equipotent; P<0.05). The fall in CVC with WBC at LC sites was reduced compared with control sites (17.6+/-2 vs. 42.4+/-8%; P<0.05). Hence, LC contributes importantly to the reduction in SkBF with body cooling, but also suppresses the reflex response, resulting in a nonadditive effect of these two components.     

Cholinergic mechanisms of cutaneous active vasodilation during heat stress in cystic fibrosis.

To test the hypothesis that cutaneous active vasodilation in heat stress is mediated by a redundant cholinergic cotransmitter system, we examined the effects of atropine on skin blood flow (SkBF) increases during heat stress in persons with (CF) and without cystic fibrosis (non-CF). Vasoactive intestinal peptide (VIP) has been implicated as a mediator of cutaneous vasodilation in heat stress. VIP-containing cutaneous neurons are sparse in CF, yet SkBF increases during heat stress are normal. In CF, augmented ACh release or muscarinic receptor sensitivity could compensate for decreased VIP; if so, active vasodilation would be attenuated by atropine in CF relative to non-CF. Atropine was administered into skin by iontophoresis in seven CF and seven matched non-CF subjects. SkBF was monitored by laser-Doppler flowmetry (LDF) at atropine treated and untreated sites. Blood pressure [mean arterial pressure (MAP)] was monitored (Finapres), and cutaneous vascular conductance was calculated (CVC = LDF/MAP). The protocol began with a normothermic period followed by a 3-min cold stress and 30-45 min of heat stress. Finally, LDF sites were warmed to 42 degrees C to effect maximal vasodilation. CVC was normalized to its site-specific maximum. During heat stress, CVC increased in both CF and non-CF (P < 0.01). CVC increases were attenuated by atropine in both groups (P < 0.01); however, the responses did not differ between groups (P = 0.99). We conclude that in CF there is not greater dependence on redundant cholinergic mechanisms for cutaneous active vasodilation than in non-CF.     

Cutaneous vasoconstrictor response to whole body skin cooling is altered by time of day.

To test for a diurnal difference in the vasoconstrictor control of the cutaneous circulation, we performed whole body skin cooling (water-perfused suits) at 0600 (AM) and 1600 (PM). After whole body skin temperature (T(sk)) was controlled at 35 degrees C for 10 min, it was progressively lowered to 32 degrees C over 18-20 min. Skin blood flow (SkBF) was monitored by laser-Doppler flowmetry at three control sites and at a site that had been pretreated with bretylium by iontophoresis to block noradrenergic vasoconstriction. After whole body skin cooling, maximal cutaneous vascular conductance (CVC) was measured by locally warming the sites of SkBF measurement to 42 degrees C for 30 min. Before whole body skin cooling, sublingual temperature (T(or)) in the PM was significantly higher than that in the AM (P < 0.05), but CVC, expressed as a percentage of maximal CVC (%CVC(max)), was not statistically different between AM and PM. During whole body skin cooling, %CVC(max) levels at bretylium-treated sites in AM or PM were not significantly reduced from baseline. In the PM, %CVC(max) at control sites fell significantly at T(sk) of 34.3 +/- 0.01 degrees C and lower (P < 0.05). In contrast, in the AM %CVC(max) at control sites was not significantly reduced from baseline until T(sk) reached 32.3 +/- 0.01 degrees C and lower (P < 0.05). Furthermore, the decrease in %CVC(max) in the PM was significantly greater than that in AM at T(sk) of 33.3 +/- 0.01 degrees C and lower (P < 0.05). Integrative analysis of the CVC response with respect to both T(or) and T(sk) showed that the cutaneous vasoconstrictor response was shifted to higher internal temperatures in the PM. These findings suggest that during whole body skin cooling the reflex control of the cutaneous vasoconstrictor system is shifted to a higher internal temperature in the PM. Furthermore, the slope of the relationship between CVC and T(sk) is steeper in the PM compared with that in the AM.     

Oral vitamin C enhances the adrenergic vasoconstrictor response to local cooling in human skin

Local administration of ascorbic acid (Asc) at a supraphysiological concentration inhibits the cutaneous vasoconstrictor response to local cooling (LC). However, whether orally ingesting Asc inhibits the LC-induced vasoconstrictor response remains unknown. The purpose of the present study was to examine the acute influence of oral Asc on the adrenergic vasoconstrictor response to LC in human skin. In experiment 1, skin blood flow (SkBF) was measured by laser-Doppler flowmetry at three sites (forearm, calf, palm). The three skin sites were locally cooled from 34 to 24°C at -1°C/min and maintained at 24°C for 20 min before (Pre) and 1.5 h after (Post) oral Asc (2-g single dose) or placebo supplementation. Cutaneous vascular conductance (CVC) was calculated as the ratio of SkBF to blood pressure and expressed relative to the baseline value before LC. Oral Asc enhanced (P < 0.05) the reductions in CVC in the forearm (Pre, -50.3 ± 3.3%; Post, -57.8 ± 2.2%), calf (Pre, -52.6 ± 3.7%; Post, -66.1 ± 4.3%), and palm (Pre, -46.2 ± 6.2%; Post, -60.4 ± 5.6%) during LC. The placebo did not change the responses at any site. In experiment 2, to examine whether the increased vasoconstrictor response caused by oral Asc is due to the adrenergic system, the release of neurotransmitters from adrenergic nerves in forearm skin was blocked locally by iontophoresis of bretylium tosylate (BT). Oral Asc enhanced (P < 0.05) the reductions in CVC at untreated control sites but did not change the responses at BT-treated sites during LC. In experiment 3, to further examine whether adrenergically mediated vasoconstriction is enhanced by oral Asc, 0.1 mM tyramine was administered using intradermal microdialysis in the forearm skin at 34°C in the Pre and Post periods. Oral Asc increased (P < 0.05) the tyramine-induced reduction in CVC. These findings suggest that oral Asc acutely enhances the cutaneous vasoconstrictor responses to LC through the modification of adrenergic sympathetic mechanisms.     

Graded cutaneous vascular responses to dynamic leg exercise

The cutaneous vascular conductance-esophageal temperature (CVC-Tes) relationship was examined at five work loads (75-200 W) in each of four men to find whether there is a role for exercise intensity in the control of skin blood flow (SkBF). Several factors contributed to our evaluation of the CVC-Tes relationship during work. Laser-Doppler velocimetry (LDF) provided a continuous measure of SkBF that is not influenced by underlying muscle blood flow. Local warming to 39 degrees C at the site of measurement of SkBF provided a consistent skin temperature and facilitated observation of changes in LDF. Mean arterial pressure was measured noninvasively once per minute to calculate CVC. Supine exercise minimized baroreceptor-induced cutaneous vasoconstriction. Our major finding was that the internal temperature at which CVC began to rise during exercise (CVC threshold) was graded with work load beyond 125 W (P less than 0.05). In that range the CVC threshold increased by 0.16 degrees C for every increment of 25 W. The CVC threshold was never reached at the highest work load in three of the four subjects. There was no consistent effect of work load on the slope of the CVC-Tes relationship or on the internal temperature at which sweating began during exercise (sweat rate threshold). We conclude that the level of work beyond 125 W affects the CVC-Tes relationship in a graded fashion, principally through shifts in threshold.     

Reflex control of cutaneous vasoconstrictor system is reset by exogenous female reproductive hormones.

To determine whether cardiovascular influences of exogenous female steroid hormones include effects on reflex thermoregulatory control of the adrenergic cutaneous vasoconstrictor system, we conducted ramp decreases in skin temperature (T(sk)) in eight women in both high- and low (placebo)-progesterone/estrogen phases of oral contraceptive use. With the use of water-perfused suits, T(sk) was held at 36 degrees C for 10 min (to minimize initial vasoconstrictor activity) and was then decreased in a ramp, approximately 0.2 degrees C/min for 12-15 min. Subjects rested supine for 30-40 min before each experiment, and the protocol was terminated before the onset of shivering. Skin blood flow was monitored by laser-Doppler flowmetry and arterial pressure by finger photoplethysmography. In all experiments, cutaneous vasoconstriction began immediately with the onset of cooling, and cutaneous vascular conductance (CVC) decreased progressively with decreasing T(sk). Regression analysis of the relationship of CVC to T(sk) showed no difference in slope between phases (low-hormone phase: 17.67 +/- 5.57; high-hormone phase: 17.40 +/- 8.00 %baseline/ degrees C; P > 0.05). Additional studies involving local blockade confirmed this response as being solely due to the adrenergic vasoconstrictor system. Waking oral temperature (T(or)) was significantly higher on high-hormone vs. low-hormone days (36.60 +/- 0.11 vs. 36.37 +/- 0.09 degrees C, respectively; P < 0.02). Integrative analysis of CVC in terms of simultaneous values for T(sk) and T(or) showed that the cutaneous vasoconstrictor response was shifted in the high-hormone phase such that a higher T(or) was maintained throughout cooling (P < 0.05). Thus reflex thermoregulatory control of the cutaneous vasoconstrictor system is shifted to higher internal temperatures by exogenous female reproductive hormones.     

The effects of selective and nonselective cyclooxygenase inhibitors on endothelin-1-induced fever in rats

It was previously shown that sustained fever can be induced in rats by central injection of endothelin-1 (ET-1). This peptide appears to participate in the mechanism(s) of LPS-induced fever, which is reduced by pretreatments with ET(B) receptor antagonists. In this study, we compared the effects of a nonselective cyclooxygenase (COX) inhibitor, indomethacin, with those of two selective COX-2 inhibitors, celecoxib and lumiracoxib, on ET-1-induced fever in rats. Fever induced in conscious animals by ET-1 (1 pmol icv) or LPS (5 mug/kg iv) was prevented by pretreatments with celecoxib (5 and 10 mg/kg) or lumiracoxib (5 mg/kg) given by oral gavage 1 h before stimuli. Lower doses of celecoxib had partial (2.5 mg/kg) or no effect (1 mg/kg). Indomethacin (2 mg/kg ip) partially inhibited fever induced by LPS but had no effect on ET-1-induced fever. The levels of PGE(2) and PGF(2alpha) in the cerebrospinal fluid (CSF) of pentobarbital sodium-anesthetized rats were significantly increased 3 h after the injection of LPS or ET-1. The latter increase was abolished by celecoxib at all tested doses and by indomethacin. In conclusion, selective COX-2 inhibitors were able to prevent ET-1-induced fever, indicating a role for COX-2 in this phenomenon. However, the fact that reduced CSF PG levels obtained with indomethacin and a low dose of celecoxib are not accompanied by changes in fever induced by ET-1, along with the lack of inhibitory effects of indomethacin on ET-1 fever, suggests that the latter might also involve COX-2-independent mechanisms.     

Effects of selective COX-1 and -2 inhibition on formalin-evoked nociceptive behaviour and prostaglandin E(2) release in the spinal cord.

Nociception evoked prostaglandin (PG) release in the spinal cord considerably contributes to the induction of hyperalgesia and allodynia. To evaluate the relative contribution of cyclooxygenase-1 (COX-1) and COX-2 in this process we assessed the effects of the selective COX-1 inhibitor SC560 and the selective COX-2 inhibitor celecoxib on formalin-evoked nociceptive behaviour and spinal PGE(2) release. SC560 (10 and 20 mg/kg) significantly reduced the nociceptive response and completely abolished the formalin-evoked PGE(2) raise. In contrast, celecoxib (10 and 20 mg/kg) was ineffective in both regards, i.e. the flinching behaviour was largely unaltered and the formalin-induced PGE(2) raise as assessed using microdialysis was only slightly, not significantly reduced. This suggests that the formalin-evoked rapid PG release was primarily caused by COX-1 and was independent of COX-2. Mean free spinal cord concentrations of celecoxib during the formalin assay were 32.0 +/- 4.5 nM, thus considerably higher than the reported IC50 for COX-2 (3-7 nM). Therefore, the lack of efficacy of celecoxib is most likely not to be a result of poor tissue distribution. COX-2 mRNA and protein expression in the spinal cord were not affected by microdialysis alone but the mRNA rapidly increased following formalin injection and reached a maximum at 2 h. COX-2 protein was unaltered up to 4 h after formalin injection. The time course of COX-2 up-regulation suggests that the formalin-induced nociceptive response precedes COX-2 protein de novo synthesis and may therefore be unresponsive to COX-2 inhibition. Considering the results obtained with the formalin model it may be hypothesized that the efficacy of celecoxib in early injury evoked pain may be less than that of unselective NSAIDs.     

Platelet inhibition by low-dose aspirin but not by clopidogrel reduces the axon-reflex current-induced vasodilation in humans

We previously showed a prolonged inhibition of current-induced vasodilation (CIV) after a single oral high dose of aspirin. In this study, we tested the hypothesis of platelet involvement in CIV. Nine healthy volunteers took 75 mg aspirin/day, 98 mg of clopidogrel bisulfate/day, or placebo for 4 days. CIV was induced by two consecutive 1-min anodal current applications (0.08 mA/cm(2)) through deionized water with a 10-min interval. CIV was measured with laser Doppler flowmetry and expressed as a percentage of baseline cutaneous vascular conductance: %C(b). In a second experiment in 10 volunteers, aspirin and placebo were given as in experiment 1, but a 26-h delay from the last aspirin intake elapsed before ACh iontophoresis and postocclusive hyperemia were studied in parallel to CIV. In experiment 1, the means +/- SE amplitude of CIV was 822 +/- 314, 313 +/- 144, and 746 +/- 397%C(b) with placebo, aspirin (P < 0.05 from placebo and clopidogrel), and clopidogrel (NS from placebo), respectively. In experiment 2, CIV impairment with aspirin was confirmed: CIV amplitudes were 300 +/- 99, and 916 +/- 528%C(b) under aspirin and placebo, respectively (P < 0.05), whereas vasodilation to ACh iontophoresis (322 +/- 74 and 365 +/- 104%C(b)) and peak postocclusive hyperemia (491 +/- 137 and 661 +/- 248%C(b)) were not different between aspirin and placebo, respectively. Low-dose aspirin, even 26 h after oral administration, impairs CIV, while ACh-mediated vasodilation and postocclusive hyperemia are preserved. If platelets are involved in the neurovascular mechanism triggered by galvanic current application in humans, it is likely to occur through the cyclooxygenase but not the ADP pathway.