TRPM8 Channel Activation Induced by Monoterpenoid Rotundifolone Underlies Mesenteric Artery Relaxation

Abstract

In this study, our aims were to investigate transient receptor potential melastatin-8 channels (TRPM8) involvement in rotundifolone induced relaxation in the mesenteric artery and to increase the understanding of the role of these thermosensitive TRP channels in vascular tissue. Thus, message and protein levels of TRPM8 were measured by semi-quantitative PCR and western blotting in superior mesenteric arteries from 12 week-old Spague-Dawley (SD) rats. Isometric tension recordings evaluated the relaxant response in mesenteric rings were also performed. Additionally, the intracellular Ca2+ changes in mesenteric artery myocytes were measured using confocal microscopy. Using PCR and western blotting, both TRPM8 channel mRNA and protein expression was measured in SD rat mesenteric artery. Rotundifolone and menthol induced relaxation in the isolated superior mesenteric artery from SD rats and improved the relaxant response induced by cool temperatures. Also, this monoterpene induced an increase in transient intracellular Ca2+. These responses were significantly attenuated by pretreatment with capsazepine or BCTC, both TRPM8 channels blockers. The response induced by rotundifolone was not significantly attenuated by ruthenium red, a non-selective TRP channels blocker, or following capsaicin-mediated desensitization of TRPV1. Our findings suggest that rotundifolone induces relaxation by activating TRPM8 channels in rat superior mesenteric artery, more selectively than menthol, the classic TRPM8 agonist, and TRPM8 channels participates in vasodilatory pathways in isolated rat mesenteric arteries.

Current Knowledge on the Vascular Effects of Menthol

“…The rising cytosolic calcium concentration induces calmodulin-mediated myosin light chain kinase (MLCK) activation, which results in myosin light chain (MLC) phosphorylation and consequently in VSM contraction and vasoconstriction (Davis and Hill, 1999). This response has been demonstrated in endothelial-denuded vessels (Macpherson et al, 2006;Johnson et al, 2009;Silva et al, 2015), and observed in other smooth muscles, including the bladder (Jun et al, 2012), vas deferens (Boldyrev et al, 2009;Vladymyrova et al, 2011) and gastric fundus (Mustafa and Oriowo, 2005). It has also been proposed that TRPM8 channels might exist in cutaneous sympathetic vasodilator fibers, whereby released acetylcholine could lead to NO-dependent VSM relaxation (Johnson et al, 2009), although no studies have effectively investigated this hypothesis.…”

“…The vascular expression of TRPM8 channels seems to differ according to animal species and age, as well as to the type of vascular bed. For example, TRPM8 were identified in rat pudendal (Silva et al, 2015), aortae, tail and mesenteric arteries (Johnson et al, 2009), while another study failed to detect their presence in the mesenteric arteries (Inoue et al, 2006). TRPM8 channels have been detected in different cellular locations, namely the plasma membrane (Gai-ying et al, 2014) and in the membrane of intracellular calcium stores, namely Golgi apparatus (Mahieu et al, 2007) and sarcoplasmic reticulum (SR) (Eccles et al, 1988;Lawrence et al, 2011;Schaefer, 2015), although there are conflicting data regarding the latter (Mahieu et al, 2007).…”

Smooth Muscle Ion Channels and Regulation of Vascular Tone in Resistance Arteries and Arterioles

“…In aorta, mesenteric artery, and femoral artery, TRPM8 activation results in a modest relaxation that is endothelium independent (712). Another study concluded that TRPM8 played a part in mesenteric artery vasodilation in rats, but some of the pharmacological tools used bring the specificity of this conclusion into question (1305). Thus, the role of TRPM8 channels is still relatively unknown.…”

Activation of host transient receptor potential (TRP) channels by praziquantel stereoisomers

“…Phasic vasorelaxation of mouse mesenteric arteries has previously been associated with the action of agonists of the transient receptor potential melastatin 8 channel (TRPM8, [ 24 , 25 ]) under a similar contractile paradigm. This observation has especial relevance given recent data showing that ±PZQ activates TRPM8 in both heterologous expression experiments, as well as in assays for endogenous TRP activity in dorsal root ganglion neurons [ 15 ].…”

“…Given the efficacy of ( S )-PZQ at causing vasorelaxation ( Fig 2 ), the stereoselectivity of ( S )-PZQ at hTRPM8 ( Fig 3 ) and the proposed role for TRPM8 in mesenteric vascular beds [ 24 , 25 ], secondary assays were performed using single cell confocal Ca 2+ imaging to validate ( S )-PZQ action at human TRPM8. Untransfected, and human TRPM8 transfected, HEK293 cells were challenged with ±PZQ, ( R )-PZQ and ( S )-PZQ and menthol (a TRPM8 agonist).…”