Evidence for a role for vasoactive intestinal peptide in active vasodilatation in the cutaneous vasculature of humans

Abstract

Active vasodilatation (AVD) in human, non-glabrous skin depends on functional cholinergic fibres but not on acetylcholine (ACh). We tested whether AVD is a redundant system in which ACh and vasoactive intestinal polypeptide (VIP) are co-released from cholinergic nerves. (1) We administered VIP by intradermal microdialysis to four discrete areas of skin in the presence of different levels of the VIP receptor antagonist, VIP(10-28), also delivered by microdialysis. Skin blood flow (SkBF) was continuously monitored by laser Doppler flowmetry (LDF). Mean arterial pressure (MAP) was measured non-invasively and cutaneous vascular conductance (CVC) calculated as LDF/MAP. Subjects were supine and wore water-perfused suits to control whole-body skin temperature (Tsk) at 34 degrees C. Concentrations of 54 microM, 107 microM, or 214 microM VIP(10-28) were perfused via intradermal microdialysis at 2 microl min-1 for approximately 1 h. Then 7.5 microM VIP was added to the perfusate containing VIP(10-28) at the three concentrations or Ringer solution and perfusion was continued for 45-60 min. At the control site, this level of VIP caused approximately the vasodilatation typical of heat stress. All VIP(10-28)-treated sites displayed an attenuated dilatation in response to the VIP. The greatest attenuation was observed at the site that received 214 microM VIP(10-28) (P < 0.01). (2) We used 214 microM VIP(10-28) alone and with the iontophoretically administered muscarinic receptor antagonist atropine (400 microA cm-2, 45 s, 10 mM) in heated subjects to test the roles of VIP and ACh in AVD. Ringer solution and 214 microM VIP(10-28) were each perfused at two sites, one of which in each case was pretreated with atropine. After 1 h of VIP(10-28) treatment, individuals underwent 45-60 min of whole-body heating (Tsk to 38.5 degrees C). VIP(10-28), alone or in combination with atropine, attenuated the increase in CVC during heat stress, suggesting an important role for VIP in AVD.

Figure 1. Protocol 1: blockade of 7.5 μm VIP-induced vasodilation

Sterile Ringer solution was perfused (2 μl min⁻¹) via intradermal microdialysis at all four sites for a baseline period of 5-15 min. Sites then received either VIP(10−28) at the concentrations indicated in the figure, or Ringer solution for approximately 1 h. Then, a combination of VIP(10−28) at the same concentration and 7.5 μm VIP was administered at the three sites previously infused with antagonist alone while the control site received VIP alone. This portion of the experiment lasted about 50 min. Finally, 25 mM sodium nitroprusside (SNP) was given at all sites to elicit a maximal dilatation of the vessels. T_sk was maintained at 34 °C throughout the protocol.

Figure 2. Protocol 2: instrumentation of subjects with four microdialysis probes

Sites 2 and 4 were pretreated with atropine via iontophoresis. Initially, T_sk was maintained at 34 °C. After a brief baseline period the VIP antagonist, VIP(10−28), was perfused through two of the microdialysis probes (sites 3 and 4), throughout the periods of normothermia and hyperthermia. During VIP(10−28) treatment, individuals were subjected to whole-body heating for 45-60 min, T_sk = 38.5 °C. T_sk was then returned to normothermic levels and sodium nitroprusside (SNP) was perfused through all four microdialysis membranes to elicit maximal vasodilation. Temperature at the sites of blood flow measurement was maintained at 34 °C throughout.

Figure 3. Results from eight subjects for protocol 1

Data are from the final 38 min of combination VIP + VIP(10−28) at three sites and VIP alone at the control sites. Treatment with VIP(10−28) attenuated the 7.5 μm VIP-induced vasodilation (P < 0.007). Dunnett's post hoc test showed responses at the control sites differed significantly from those at sites treated with 107 μm and 214 μm VIP(10−28) (P < 0.05 and P < 0.01, respectively). The greatest attenuation was detected at the sites treated with 214 μm VIP(10−28). Alternating pairs (i.e. control and 214 μm VIP(10−28) vs. 54 μm and 107 μm VIP(10−28)) of s.e.m. error bars are depicted for visual clarity.

Figure 4. Typical responses to body heating in protocol 2

Data are from a single subject. Shown are blood pressure (MAP), heart rate (HR), skin temperature (T_sk), oral temperature (T_or) and cutaneous vascular conductance (CVC) from a control site (continuous line) and from a site treated with atropine and the VIP receptor antagonist VIP(10−28) (dashed line). Atropine pretreatment took place 1 h earlier. Intradermal infusion of VIP(10−28) by microdialysis began after 10 min of baseline data collection. After 1 h, body heating commenced by raising the skin temperature to 38.5 °C. This was not accompanied by local warming of the areas at which blood flow was measured.

Figure 5. Results from protocol 2

Sites were treated with atropine, VIP(10−28), both antagonists, or Ringer solution only. These are the vasodilator responses, expressed as changes relative to maximal (Δ% Max), from the onset of AVD at the control site to the end of heating. A, repeated measures ANOVA on the magnitude of the change in CVC during the last 2 min of heat stress did not reveal a statistically significant difference among treatment sites (‡P = 0.08). However, one subject displayed a dramatic, unexplained vasodilatation during the last 7 min of heat stress at the VIP(10−28)-treatment site only. B, the average response with these data excluded. VIP(10−28) reduced the vasodilator response significantly (†P < 0.0001). As with the sensitivity, atropine did not significantly affect the magnitude of the change in CVC.

Figure 6. Results from linear regression analyses of the CVC: time relationships for protocol 2

Individual slopes are shown for all subjects for pairwise comparisons of the responses between treatment sites. Antagonist-treated sites showed a reduced slope compared to control sites (A-C). VIP(10−28) (B) and combination atropine and VIP(10−28) (C) caused a statistically significant reduction in slope compared to control (P < 0.036). Although statistical analysis of the group data did not show an effect of atropine treatment, the slopes of the responses for each individual were reduced relative to control (A). B, atropine vs. combination atropine + VIP(10−28)-treated sites, shows a consistent reduction in slope with VIP(10−28) treatment. E shows similar responses at the VIP(10−28) and the combination atropine + VIP(10−28)-treated sites, suggesting no significant effect of atropine in the presence of VIP antagonism. Note that in one subject the atropine-treated site was not useful for technical reasons and data from that site are not included here or in Fig. 5.