Double injection vs skin microdialysis technique in minimally invasive in vivo pharmacology

Wenzel and colleagues recently concluded that an endothelin-A (ET-A) receptor antagonist attenuates the skin vasoconstriction produced by angiotensin II (Ang II) and noradrenaline (NA) [1]. The postulate that the selective ET-A antagonist, BQ123, blocks Ang II and NA mediated vasoconstriction challenges well-established physiological and pharmacological concepts, in particular, the notion of specific receptors for these peptidergic and nonpeptidergic substances. We believe that the double injection technique (DIT) used to assess blood flow of the skin microcirculation in this study has some drawbacks that might have affected the data generated from it. In this study [1], skin blood flow was assessed immediately after injection of the study substance, which can give rise to confounding results as injection trauma can produce significant vasodilation [2]. Studies using microdialysis probes to visualize the changes in skin blood flow by laser-Doppler perfusion imaging demonstrate that a blood flow equilibrium period of about 60 min is required for the effects of trauma to dissipate [2]. The vasodilatory effects of local histamine released from injection trauma can last up to 40 min [2]. Hence, the effects of injection trauma cannot be distinguished from the effects of the study drug. It can be assumed that, in this study, the dilator effects of the control drug (i.e. saline or the antagonist) were subtracted from the vasodilation or vasoconstriction induced by the study drug. Regardless, subtracting the value does not ensure that the effects of histamine or other vasodilatory substances released by injection trauma will not be present during the study period of 30 min For example, BQ123 could have induced a nonspecific vasodilatory effect unrelated to blockade of ET-A receptors. Similarly, trauma could have affected the vasoconstrictor effects of Ang II and NA by release of local vasodilatory substances. The combination of microdialysis and laser-Doppler flowmetry (Figure 1), with an equilibrium period of 2 h after insertion of the microdialysis probes, allows similar studies without the confounding effects of trauma. This technique allows for the study of vasodilatory and vasoconstrictor substances on skin blood flow even at the already low baseline flows found during normothermia and after resolution of trauma [3]. Using this technique, we have shown that the ET-B receptor antagonist (BQ788) at a dose of 1.5 × 10−7 mol l−1 produced cutaneous vasodilation in healthy men and vasoconstriction in healthy females. Therefore, we concluded that the contribution of ET-B receptors to resting vascular tone differs between males and females in the human cutaneous vasculature [4]. In contrast, the authors’ in the present study [1] demonstrated a mild vasoconstriction with BQ788 in men at doses ranging from 10-8-10-10 mol l−1. This could result from an effect of BQ788 on the vasodilatory response produced by histamine or other local vasodilators, like nitric oxide, in response to trauma. Figure 1 Diagram of the setup for measurement of skin blood flow in combination with microdialysis. The laser-Doppler probe is taped on the skin directly above the microdialysis fiber. The endsof the microdialysis fiber are exteriorized and connected to a microinfusion ... We agree with the comments of the senior author, in a recent review [5], regarding the need for a minimally invasive tool to assess human in vivo pharmacology especially, because other techniques are either more invasive (forearm blood flow studies using venous occlusion plethysmography and intra-arterial injections) or are unpredictable due to variable drug penetration (iontophoresis). Interpretation of data generated using DIT can be difficult due to the confounding effects of vasoactive and inflammatory substances released locally in response to trauma. Since the use of skin microcirculation as a surrogate for other areas of circulation is relatively new, it is imperative that investigators interested in this area agree on using techniques that minimize such confounding effects.