Rotundifolone is the major constituent of the essential oil of Mentha x villosa Hudson. In preliminary studies, rotundifolone induced significant hypotensive, bradycardic and vasorelaxant effects in rats. Thus, to gain more insight into the pharmacology of rotundifolone, the aim of this study was to characterize the molecular mechanism of action involved in relaxation produced by rotundifolone. The relaxant effect was investigated in rat superior mesenteric arteries by using isometric tension measurements and whole-cell patch-clamp techniques. Rotundifolone relaxed phenylephrine-induced contractions in a concentration-dependent manner. Pre-treatment with KCl (20 mM), charybdotoxin (10 )7 M) or tetraethylammonium (TEA 10 )3 or 3 · 10 )3 M) significantly attenuated the relaxation effect induced by rotundifolone. Additionally, whole-cell patch-clamp recordings were made in mesenteric smooth muscle cells and showed that rotundifolone significantly increased K + currents, and this effect was abolished by TEA (10 )3 M), suggesting the participation of BK Ca channels. Furthermore, rotundifolone inhibited the vasoconstriction induced by CaCl 2 in depolarizing nominally Ca 2+-free medium and antagonized the contractions elicited by an L-type Ca 2+ channel agonist, S(-)-Bay K 8644 (2 · 10 )7 M), indicating that the vasodilatation involved inhibition of Ca 2+ influx through L-type voltage-dependent calcium channels (Ca v type-L). Additionally, rotundifolone inhibited L-type Ca 2+ currents (I Ca L), affecting the voltage-dependent activation of I Ca L and steady-state inactivation. Our findings suggest that rotundifolone induces vasodilatation through two distinct but complementary mechanisms that clearly depend on the concentration range used. Rotundifolone elicits an increase in the current density of BK Ca channels and causes a shift in the steady-state inactivation relationship for Ca v type-L towards more hyperpolarized membrane potentials.Terpenoids constitute the largest class of plant secondary metabolites and have been used in essential oils for centuries as therapeutically relevant compounds. However, little is known about their mechanism of action [1,2].The monoterpenoids, such as camphor, borneol, citronellol, alpha-terpineol or menthol, compose a group of naturally occurring organic compounds derived from two isoprene units. They are the major components of some essential oils presenting anaesthetic and analgesic activities [3,4] Rotundifolone (C 10 H 14 O 2 ; molecular weight 166) is a naturally occurring monoterpenic ketone of plant origin and an important chemical constituent of the essential oils of many Mentha species (Mentha rotundifolia, M. suaveolens, M. spicata L., M. longifolia and M. x villosa) [19,20]. A previous report from our group showed that rotundifolone induced marked hypotension and bradycardia in non-anaesthetized normotensive rats using an in vivo approach. Those effects were probably due to a decrease in peripheral vascular resistance [21], a hypothesis that was subsequently str...

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.

BackgroundIn northeastern Brazil, grape pomace has become a potential alternative byproduct because of the recover phenolic compounds from the vinification process. Comparative analyses were performed between lyophilized extract of grape skins from pomace, described as fermented (FGS), and fresh, unfermented (UGS) grape skins to show the relevant brand’s composition upon the first maceration in winemaking.MethodsThe use of in vitro testing such as Folin-Ciocalteu’s, DPPH free radical scavenger and HPLC methods were performed to evidence antioxidant effect and phenolic compounds. Additionally, vascular reactivity studies were performed in third-order branches of rat superior mesenteric arteries, which were obtained and placed in organ baths containing Krebs-Henseleit solution, maintained at 37 °C, gassed with a mixture of 95% O2 and 5% CO2, and maintained at pH 7.4. The in situ formation of reactive oxygen species (ROS) was evaluated in small mesenteric rings using oxidative fluorescent dihydroethidium dye.ResultsWe found higher phenolic content and antioxidant activity in FGS when compared to UGS. HPLC analyses identified a significant number of phenolic compounds with antioxidant potential in both samples. The vasorelaxant effect induced by FGS was more potent than that induced by UGS, and the activity was attenuated after removal of vascular endothelium or by blockade of endothelium-derived relaxing factors, such as NO and EDHF.ConclusionsThe FGS extract may be a great source of natural polyphenol products with potent antioxidant effects and endothelium-dependent vasodilatory actions involving NO and EDHF pathways.

Ethnopharmacological Relevance: The traditional herbal medicinal product Funchicórea® has been widely used in clinical practice for the treatment of intestinal colic and constipation in newborns. However, no scientific data on the herbal product to prove its efficacy is available. Aim of the Study: This study aimed to evaluate the laxative and spasmolytic actions of Funchicórea®. Materials and Methods: Wistar rats (Rattus norvegicus) and Swiss mice (Mus musculus) of both sex, were used. In vivo pharmacological assays were performed to evaluate the stimulating effect on the gastrointestinal tract, and in vitro studies to verify its spasmodic activity. Results: Funchicórea® increased the motility of the small intestine in male mice at doses of 100 mg/kg (161.66±14.86 %, n=6) and 200 mg/kg (151.04±17.17 %, n=6) compared to control (100.00±10.49 %, n=6). However, this drug did not induce any change in intestinal transit in female mice. The intestinal transit of male mice treated with loperamide (3 mg/kg/day, during three days) was reduced 66.25±7.49 % (n=8) compared to the control group (100.00±5.16%, n=8) and we observed the normalization of the intestinal transit in constipated animals treated with Funchicórea® 100 mg/Kg (98.42±6.33 %, and 200 mg/kg (99.32±8.47%, n=7). Similar results were observed in the quantification for 24 hours of male and female rats faeces constipated by loperamide (3 mg/kg/day three days), however, in both animals groups treated with Funchicórea® 100 mg/kg (1.24±2.90 g, male; 3.60±0.80 g, female, n=6) and 200 mg/Kg (8.70±2.01 g, male, 10.03±1.30 g, female, n=6) the levels of faeces returned to basal values compared to constipated group (4.01±1.43 g, male; 1.70±0.10 g, female, n=6). In addition, Funchicórea® (0.01-1000 μg/mL) elicited relaxation in rat ileum pre-contracted by KCl 40 mM (Emax=97.5±7.0 %, n=7) and carbachol (1 μM, Emax=100.0±7.0 %, n=7). Conclusion: The results obtained demonstrated that the herbal medicine Funchicórea® acts by stimulating the intestine of rats and mice and has spasmolytic activity in isolated rat ileum.

Benzoyltryptamine analogues act as neuroprotective and spasmolytic agents on smooth muscles. In this study, we investigated the ability of N-salicyloyltryptamine (STP) to produce vasorelaxation and determined its underlying mechanisms of action. Isolated rat mesenteric arteries with and without functional endothelium were studied in an isometric contraction system in the presence or absence of pharmacological inhibitors. Amperometric experiments were used to measure the nitric oxide (NO) levels in CD31+ cells using flow cytometry. GH3 cells were used to measure Ca2+ currents using the whole cell patch clamp technique. STP caused endothelium-dependent and -independent relaxation in mesenteric rings. The endothelial-dependent relaxations in response to STP were markedly reduced by L-NAME (endothelial NO synthase—eNOS—inhibitor), jHydroxocobalamin (NO scavenger, 30 µM) and ODQ (soluble Guanylyl Cyclase—sGC—inhibitor, 10 µM), but were not affected by the inhibition of the formation of vasoactive prostanoids. These results were reinforced by the increased NO levels observed in the amperometric experiments with freshly dispersed CD31+ cells. The endothelium-independent effect appeared to involve the inhibition of voltage-gated Ca2+ channels, due to the inhibition of the concentration-response Ca2+ curves in depolarizing solution, the increased relaxation in rings that were pre-incubated with high extracellular KCl (80 mM), and the inhibition of macroscopic Ca2+ currents. The present findings show that the activation of the NO/sGC/cGMP pathway and the inhibition of gated-voltage Ca2+ channels are the mechanisms underlying the effect of STP on mesenteric arteries.