Molecular pharmacology of xylazine at adrenoceptors: Relation to necrotic skin lesions associated with chronic intravenous administration

PROJECT SUMMARY/ABSTRACT The α2-adrenergic receptor (α2R) agonist xylazine has been used in veterinary medicine for decades owing to its anesthetic, muscle relaxant, and analgesic properties. Since the mid-2000’s however, illicit use of xylazine has grown dramatically and over the last few years alone, the illicit use of xylazine-

adulterated opioids, including fentanyl, has increased exponentially. Xylazine use in humans is linked to well-documented adverse effects including respiratory depression, hypotension, or bradycardia, which can all be directly coupled to its mechanism as an α2R agonist. Significantly, repeated intravenous injection of xylazine in humans has been associated with necrotizing skin ulceration that is

independent of bacterial etiology and is xylazine-specific. Despite the recent increase in illicit xylazine use, the molecular mechanisms related to xylazine’s action at α2Rs remain poorly understood, as is the mechanism of xylazine-mediated skin necrosis. This project is directly linked to the basic research

areas of interest in this Notice of Special Interest from NIDA. We hypothesize that xylazine exhibits unique agonist binding and functional profiles at peripheral α2 and α1Rs and that its ulcerative effects on the skin associated with intravenous injection are due to peripheral α2/α1R-mediated vasoconstriction, which causes hypoperfusion and hypoxia upon repeated exposure leading to skin

necrosis. We propose three specific aims to address this hypothesis. In Aim 1, we will assess the binding affinity of xylazine compared to the structurally distinct αR agonists clonidine, moxonidine, α- methylnorepinephrine and epinephrine at agonist-occupied conformations of each of the six α2/α1R

subtypes (α1A, α1B, α1D, α2A, 2B, 2C), expressed in translationally-relevant venous and arterial smooth muscle cells (VSMC). In Aim 2, we will utilize novel TRUPATH G protein-biosensors in both venous and arterial VSMC to assess the functional effects of xylazine compared to the other agonists at α1A, α1B, α1D, α2A, α2B, and α2CRs engaged with differing Gαβγ combinations. This aim will inform

on xylazine-induced α1R/α2R signaling and will gauge distinct G protein bias that the agent may possess, compared to the other α1R/α2R agonists. In Aim 3, we will directly test the hypothesis that IV xylazine-induced skin necrosis in vivo is caused by a peripheral α2R-mediated mechanism associated

with vasoconstriction and resulting hypoperfusion and hypoxia. Together, these aims will fill gaps in the literature related to xylazine binding and function in both venous and arterial VSMC as well as mechanisms of IV xylazine-induced skin necrosis.