Role Of TRPV1 Channels Research Articles

UPLC-QTOF-MS/MS-based metabolomic approach and gastroprotective effect of two chemotypes of Egletes viscosa (L.) less. against ethanol-induced gastric ulcer in mice.

Egletes viscosa (L.) (macela) is a native wild herb that can be found in different states of northeastern Brazil. The infusions of its flower buds are traditionally used for the treatment of gastrointestinal disorders. E. viscosa possesses two chemotypes (named A and B), distinguishable by the composition of the essential oil from the flower buds. Although there are previous studies of the gastroprotective effect of the isolated constituents of E. viscosa, its infusions have not been investigated yet. The present study aimed to evaluate and compare the chemical composition and the gastroprotective effect of flower bud infusions of E. viscosa from chemotype A (EVCA) and chemotype B (EVCB). Sixteen infusions were brewed with flower buds according to the traditional preparation mode and were analyzed through a UPLC-QTOF-MS/MS based metabolomic approach for determination of their metabolic fingerprints and quantification of bioactive compounds. Afterward, these data were analyzed by chemometric methods (OPLS-DA) for discrimination of the two chemotypes. Additionally, infusions of EVCA and EVCB (50, 100 and 200mg/kg, p.o.) were evaluated on gastric ulcers induced by absolute ethanol (96%, 0.2mL, p.o.) in mice. To elucidate the gastroprotective mechanisms, the effect of EVCA and EVCB on gastric acid secretion and gastric wall mucus was determined and the role of TRPV1 channels, prostaglandins, nitric oxide and KATP channels were assessed. Moreover, the oxidative stress-related parameters and the histological aspects of the stomach tissue were analyzed. The chemotypes can be discriminated from each other using UPLC-QTOF-MS/MS chemical fingerprints. Both chemotypes presented similar chemical compositions, consisting basically of caffeic acid derivatives, flavonoids and diterpenes. The quantification of bioactive compounds demonstrated that chemotype A possesses more ternatin, tanabalin and centipedic than chemotype B. EVCA and EVCB (50, 100 and 200mg/kg, p.o.) significantly decreased the severity of ethanol-induced gastric lesions, as shown by a reduction in histological alterations and leucocyte infiltration in gastric tissue. The gastroprotective mechanism of both infusions involves an antioxidant effect, maintenance of gastric mucus and reduction gastric secretion. Stimulation of endogenous prostaglandins and nitric oxide release, activation of TRPV1 channels, and KATP channels are also involved in the gastroprotection of the infusions. The gastroprotective effect of EVCA and EVCB was equivalent and mediated through antioxidant and antisecretory actions, including the activation of TRPV1 receptors, stimulation of endogenous prostaglandins and nitric oxide, and opening of KATP channels. The presence of caffeic acid derivatives, flavonoids and diterpenes in both infusions is involved in mediating this protective effect. Our findings support the traditional use of infusions of E. viscosa for gastric disorders regardless of the chemotype.

TRPV4 Inhibition Exerts Protective Effects Against Resistive Breathing Induced Lung Injury.

IntroductionTRPV4 channels are calcium channels, activated by mechanical stress, that have been implicated in the pathogenesis of pulmonary inflammation. During resistive breathing (RB), increased mechanical stress is imposed on the lung, inducing lung injury. The role of TRPV4 channels in RB-induced lung injury is unknown.Materials and MethodsSpontaneously breathing adult male C57BL/6 mice were subjected to RB by tracheal banding. Following anaesthesia, mice were placed under a surgical microscope, the surface area of the trachea was measured and a nylon band was sutured around the trachea to reduce area to half. The specific TRPV4 inhibitor, HC-067047 (10 mg/kg ip), was administered either prior to RB and at 12 hrs following initiation of RB (preventive) or only at 12 hrs after the initiation of RB (therapeutic protocol). Lung injury was assessed at 24 hrs of RB, by measuring lung mechanics, total protein, BAL total and differential cell count, KC and IL-6 levels in BAL fluid, surfactant Protein (Sp)D in plasma and a lung injury score by histology.ResultsRB decreased static compliance (Cst), increased total protein in BAL (p < 0.001), total cell count due to increased number of both macrophages and neutrophils, increased KC and IL-6 in BAL (p < 0.001 and p = 0.01, respectively) and plasma SpD (p < 0.0001). Increased lung injury score was detected. Both preventive and therapeutic HC-067047 administration restored Cst and inhibited the increase in total protein, KC and IL-6 levels in BAL fluid, compared to RB. Preventive TRPV4 inhibition ameliorated the increase in BAL cellularity, while therapeutic TRPV4 inhibition exerted a partial effect. TRPV4 inhibition blunted the increase in plasma SpD (p < 0.001) after RB and the increase in lung injury score was also inhibited.ConclusionTRPV4 inhibition exerts protective effects against RB-induced lung injury.

Role of TRPV1 channels on glycogen synthase kinase-3β and oxidative stress in ouabain-induced bipolar disease

Bipolar disorder (BD) is a multifactorial chronic and refractory disease characterized by manic, depressive, and mixed mood episodes. Although epidemiological, and pathophysiological studies demonstrated a strong correlation between bipolar disorder and oxidative stress, precise etiology is still missing. Recent studies suggested the possible role of transient receptor potential channels (TRP) in the BD but, current knowledge is limited. Therefore, the current study investigates the possible role of TRPV1 in the ouabain-induced model of BD. The model was created with intracerebroventricular single dose ouabain (10−3 M) administration. Animals were treated with capsaicin, capsazepine, and lithium for seven days. Mania and depressive-like states were investigated with open-field, sucrose preference, and elevated plus maze tests. Oxidative stress was assessed by measuring total antioxidant and oxidant states, spectrophotometrically. The phosphorylation Glycogen synthase kinase-3β (GSK-3β) evaluated by western blotting. Our results demonstrated that capsaicin dose-dependently inhibited the ouabain-induced hyperlocomotion and depression. Although capsazepine exacerbated behavioral impairment, it did not show a significant effect on the antioxidant and oxidant states, and the effects of capsazepine on behaviors were abolished by combination with capsaicin. Additionally, capsaicin potently prevented the ouabain-induced decrease in GSK-3β phosphorylation. In contrast, capsazepine potentiated ouabain-induced decrease in GSK-3β phosphorylation and combination with capsaicin, suppressed the effect of capsazepine on GSK-3β phosphorylation. The effects of TRPV1 activation on oxidative stress and mania-like behaviors in the ouabain-induced BD model might be regulated by GSK-3β phosphorylation.

ІОННІ КАНАЛИ В КОНТЕКСТІ ПОШУКУ МОЛЕКУЛЯРНИХ МІШЕНЕЙ РЕГУЛЯЦІЇ СКОРОТЛИВОСТІ МІОМЕТРІЮ

Many women now have complications in childbirth due to poor labor, which often threatens both mother and fetus. Also,the problem of prevention and treatment of premature uterine contractions is unresolved. Therefore, in this work we investigated the influence of ion channels as the end stage effectors of the regulatory cascades in the contractility of myometrium. To better understand the participation of TRPC4, TRPV4 and BKCa ion channels in myometrial contractility, we conducted experiments, keeping in mind the fact that changes in ionic conductivity of the plasma membrane regulate spontaneous and agonist-induced contractions. On the myometrial preparations of pregnant rats usingisolated tissue tensiometry, the amplitude of contractile force was recorded under the activation of these ion channels by their selective agonists. Obtained results allow us to consider (-) – englerin A as a way to stimulate uterine contractions in case of insufficient response to oxytocin, because at a concentration of 1 nM a significant increase in contraction force was developed and did not differ statistically from the response to oxytocin or carbacholin. The use of an agonist at concentrations of 30-100 nM causes some suppression of contractility. Based on the results describing the role of TRPV4 channels, namely the reduction of uterine smooth muscle contractions in response to their selective agonist GSK1016790A administration, we suggest that the main effect of activation of these channels depends on the expression and activity of adjacent calcium-dependent potassium channels. Our experiments found that the use of the liposomal form of quercetin to activate BKCa channels inhibits the excitability of myometrial cells more effectively than that dissolved in DMSO, which is promising for the correction of premature or excessive uterine activity.

TRPV4 activation in rat carotid artery in DOCA hypertension involves eNOS and endothelium-derived contractile factor (EDCF)

ABSTRACTAim: Role of TRPV4 channel in regulation of endothelial function in the carotid artery in deoxycorticosterone acetate (DOCA) model of hypertension in rat was studied.Methods: 8–10 weeks old albino Wistar rats divided into three groups namely Control, UNX and hypertensive animals. Vascular smooth muscle response was studied in isolated carotid artery of rat with acetylcholine, sodium nitroprusside, GSK1016790A (GSK) in presence and absence of L-NAME and indomethacin.Results: At the end of the 6th week, the mean systolic blood pressure was increased in DOCA-treated hypertensive rats (166 ± 8 mm Hg) compared to Control and UNX (125 ± 5 mm Hg). ACh (10–9 to 10–5 M) produced almost 100% relaxation in Control (Emax = 97.48 ± 1.06 %) and UNX animals (Emax = 93.16 ± 2.33 %) which was attenuated in DOCA-treated hypertensive animals (Emax = 70.85 ± 1.65 %). No significant changes seen in SNP (10–12 to 10–5 M) induced relaxation. GSK1016790A (10–12 to 10–7 M)-mediated relaxation was significantly attenuated in DOCA-treated hypertensive animals (Emax = 25.58 ± 13.60%) compared to the control (Emax = 80.59 ± 6.86%) and UNX (Emax = 87.32 ± 2.01%) animals. L-NAME (10–4 M) potently blocked GSK-induced relaxation, and a contractile response to GSK was observed in presence of L-NAME in all the three groups of animals which was sensitive to indomethacin (10–5 M).Conclusion: TRPV4 may regulate the vascular tone of rat carotid artery through an attenuated NO pathway and stimulation of the release of contractile prostanoids in the DOCA hypertensive rats.

Fluid-induced lung injury-role of TRPV4 channels.

Administration of bolus intravenous fluid is associated with respiratory dysfunction and increased mortality, findings with no clear mechanistic explanation. The objective of this study was to examine whether bolus intravenous (i.v.) fluid administration results in acute lung injury in a rat model and further, to examine whether this injury is associated with transient receptor potential vallinoid (TRPV)4 channel function and endothelial inflammatory response. Healthy male Sprague-Dawley rats were administered 60ml/kg 0.9% saline i.v. over 30min. Manifestation of acute lung injury was assessed by lung physiology, morphology, and markers of inflammation. The role of TRPV4 channels in fluid-induced lung injury was subsequently examined by the administration of ruthenium red (RR) in this established rat model and again in TRPV4 KO mice. In endothelial cell culture, permeability and P-selectin expression were measured following TRPV4 agonist with and without antagonist; 0.9% saline resulted in an increase in lung water, lavage protein and phospholipase A2, and plasma angiopoietin-2, with worsening in arterial blood oxygen (PaO2), lung elastance, surfactant activity, and lung histological injury score. These effects were ameliorated following i.v. fluid in rats receiving RR. TRPV4 KO mice did not develop lung edema. Expression of P-selectin increased in endothelial cells following administration of a TRPV4 agonist, which was ameliorated by simultaneous addition of RR. Bolus i.v. 0.9% saline resulted in permeability pulmonary edema. Data from ruthenium red, TRPV4 KO mice, and endothelial cell culture suggest activation of TRPV4 and release of angiopoietin 2 and P-selectin as the central mechanism.

Investigating the involvement of TRPV1 ion channels in remote hind limb preconditioning-induced cardioprotection in rats.

Remote ischemic preconditioning (RIPC) treatment strategy is a breakthrough in the field of cardiovascular pharmacology as it has the potential to attenuate myocardial ischemia-reperfusion injury. However, the underlying intracellular pathways have not been widely explored. The present study intends to explore the possible role of TRPV1 channels in mediating remote hind limb preconditioning-induced cardioprotection. Remote hind limb preconditioning stimulus (4cycles in succession) was delivered by tying the blood pressure cuff at the inguinal level of the rat. The Langendorff system was used to perfuse the isolated heart and afterward was subjected to 30min of global ischemia and 120min of reperfusion. Sustained ischemia and, thereafter, reperfusion led to cardiac injury that was assessed in terms of infarct size, lactate dehydrogenase (LDH) release, creatine kinase (CK) release, left ventricular end diastolic pressure (LVEDP), left ventricular developed pressure (LVDP), +dp/dtmax, -dp/dtmin, heart rate, rate pressure product, and coronary flow rate. The pharmacological modulators employed included capsaicin as TRPV1 agonist and capsazepine as TRPV1 antagonist. Remote hind limb preconditioning stimulus and capsaicin preconditioning (5 and 10mg/kg) led to significant reduction in infarct size, LVEDP, LDH release, CK release, and significant improvement in LVDP, +dp/dtmax, -dp/dtmin, heart rate, rate pressure product, and coronary flow rate. However, remote hind limb preconditioning-induced cardioprotective effects were considerably abolished in the presence of capsazepine (2.5 and 5mg/kg). This indicates that remote hind limb preconditioning stimulus possibly activates TRPV1 channels to produce cardioprotective effects.

Role of TRPV4 channels in regulation of eNOS expression in brain microvascular endothelial cells under the condition of mechanical stretch

To identify the subtype of transient receptor potential (TRPs) channel involved in stretch-induced injury of human brain microvascular endothelial cells (HBMEC) and to explore the mechanism responsible for eNOS expression. TRPs expression was examined by Western blot and immunocytofluoresence in the cultured HBMEC. Mechanical stretch was performed by mini-type multi-functional bio-impact machine. The levels of free calcium ion in cells were examined by the flow cytometry. The eNOS expression was detected by Western blot. The mRNA and protein expression of TRPV4 was detected in HBMEC by qRT-PCR, Western blot and immunocytofluoresence. The levels of free calcium ion in the stretch-treated HBMEC was significantly decreased in the presence of TRPV4 specific inhibitor (P<0.001), but there was no difference in calcium levels between the stretch and the control or unspecific inhibitor group (P=0.072 or 0.308). The levels of eNOS protein in the stretch-treated HBMEC were reduced in the presence of TRPV4 specific inhibitor or NOS inhibitor (P<0.05), but it was not changed compared with that in the control group (P>0.05). The eNOS expression is up-regulated under the condition of mechanic stretch, which is related to the activation of TRPV4, resulting in the influx of calcium.

Role of TRPV1 Channels in Glioma Cell Viability and Survival

High grade primary brain tumors are among the deadliest malignancies known to medicine. Anaplastic gliomas (WHO grade 3) and Glioblastoma Multiforme, or GBM, (WHO grade 4) are prevalent primary brain tumors in adults. Exploration of new avenues in treatment of GBM is absolutely crucial when taking into consideration the short (12-17 months) survival time after the diagnosis. Annually there are around 20,000 new cases, including over 4,000 pediatric (age 0-19) cases. The heat and capsaicin receptor TRPV1 is highly expressed in malignant gliomas, where the channels are thought to play a role in inducing apoptotic cell death. We evaluated TRPV1 expression in different glioma cell lines and determined the correlation of the protein expression with age and sex in the tissue samples obtained from patients with GMB. We found that age dependent correlation of TRPV1 expression is more characteristic for male patients with GBM than for females. However, in both groups TRPV1 expression was higher in patients of age 35-55 years old.We also investigated the sensitivity of different glioma cell lines to the TRPV1 agonists, including endovanilloid NADA and capsaicin. We found that 50 μM of capsaicin inhibited the cell growth and proliferation of U87 but not U251 glioma cells. The presence of TRPV1 on the plasma membrane was higher in U87 cells in comparison to U251, as confirmed by biotinylation, and may account for the effect of capsaicin on the cell viability. These results indicate that TRPV1 could play an important role in regulating Ca2+ homeostasis of primary brain tumors.

Correction: Role of TRPV1 Channels in Ischemia/Reperfusion-Induced Acute Kidney Injury

Correction: Role of TRPV1 Channels in Ischemia/Reperfusion-Induced Acute Kidney Injury

The role of TRPV1 channels in gastric vagal afferent satiety signalling in lean and obese mice

The role of TRPV1 channels in gastric vagal afferent satiety signalling in lean and obese mice

TRPV1 inhibition attenuates IL-13 mediated asthma features in mice by reducing airway epithelial injury

Even though neurogenic axis is well known in asthma pathogenesis much attention had not been given on this aspect. Recent studies have reported the importance of TRP channels, calcium-permeable ion channels and key molecules in neurogenic axis, in asthma therapeutics. The role of TRPV1 channels has been underestimated in chronic respiratory diseases as TRPV1 knockout mice of C57BL/6 strains did not attenuate the features of these diseases. However, this could be due to strain differences in the distribution of airway capsaicin receptors. Here, we show that TRPV1 inhibition attenuates IL-13 induced asthma features by reducing airway epithelial injury in BALB/c mice. We found that IL-13 increased not only the lung TRPV1 levels but also TRPV1 expression in bronchial epithelia in BALB/c rather than in C57BL/6 mice. TRPV1 knockdown attenuated airway hyperresponsiveness, airway inflammation, goblet cell metaplasia and subepithelial fibrosis induced by IL-13 in BALB/c mice. Further, TRPV1 siRNA treatment reduced not only the cytosolic calpain and mitochondrial calpain 10 activities in the lung but also bronchial epithelial apoptosis indicating that TRPV1 siRNA might have corrected the intracellular and intramitochondrial calcium overload and its consequent apoptosis. Knockdown of IL-13 in allergen induced asthmatic mice reduced TRPV1, cytochrome c, and activities of calpain and caspase 3 in lung cytosol. Thus, these findings suggest that induction of TRPV1 with IL-13 in bronchial epithelia could lead to epithelial injury in in vivo condition. Since TRPV1 expression is correlated with human asthma severity, TRPV1 inhibition could be beneficial in attenuating airway epithelial injury and asthma features.

Coupling of Coronary Blood Flow to Cardiac Metabolism: Role of TRPV1 Channels and Local Changes in pH

TRPV1 channels, known to be activated by noxious stimuli such as pH changes, are thought to play a role in the regulation of the vascular tone. Accordingly, we hypothesized that TRPV1 activation by local pH changes during increased metabolic demand and/or ischemia would lead to modulation of myocardial blood flow (MBF). Further we will examine if TRPV1 channel dysfunction could contribute to the microvascular impairment in diabetic cardiomyopathy. To test our hypothesis, control (C57/B6), db/db and TRPV1 null mice (TRPV1−/−) were subjected to contrast stress echocardiography for quantification of MBF at baseline and during the infusion of the TRPV1 agonist capsaicin (CAP; 1–100 μg/kg). Arterial pressures (AP) and heart rate (HR) were simultaneously recordedfor double product (DP) measurement as a surrogate of myocardial oxygen consumption. In controls, CAP produced a dose‐dependent increase in MBF, which was significantly blunted in db/db mice, despite db/db having a higher DP. Low pH changes induced vasodilation in isolated mesenteric microvessels from controls, which was blunted in microvessels from db/db, TRPV1−/− and control after TRPV1 inhibition (SB366791), suggesting a portion of low pH induced vasodilation may be due to TRPV1 activation. We conclude that TRPV1 channels may be involved in the metabolic regulation of myocardial blood flow and this mechanism appears to be corrupted in diabetic mice.

Role of TRPV4 channels in agonist‐induced endothelial Ca2+ entry and vasodilation: Evidence from TRPV4‐deficient mice

Transient receptor potential vallinoid 4 (TRPV‐4), is a Ca2+‐permeant cation channel implicated in endothelial‐dependent vasorelaxation to mechanical stimuli but limited information exists for agonist‐induced responses. We hypothesized that TRPV‐4 is required for agonist‐induced vasodilation by regulating Ca2+ entry. Isometric tension was measured in isolated mesenteric arteries (MA) from wild type (WT) and TRPV‐4 knockout mice using a wire myograph. Dilation to acetylcholine (Ach, 10−9–10−5 M) was diminished in TRPV‐4−/− vs. WT MA (% max relaxation, 44±12 and 86±5, respectively n=4, P&lt;0.05). L‐NAME, blocked Ach relaxation in TRPV‐4−/− and WT MA (11±4 and 35±11, respectively n=4, P&lt;0.05). Responses to papaverine were similar in WT and TRPV‐4−/− MA. Intracellular calcium ([Ca2+]i) was assessed after intraluminal fura‐2 (fluorescence) in MA. Ach (10−5 M) increased endothelial [Ca2+]i in WT and TRPV‐4−/− MA (Δ[Ca2+]i, 49±4.6nM and 26±2.1nM, respectively n=4). The addition of 4α‐PDD, a TRPV4 opener, increased [Ca2+]i in WT but not in TRPV‐4−/− (Δ[Ca2+]i, 164±30nM and 15±1.3nM, respectively n=4). In endothelium of WT MA the increase in [Ca2+] was abolished by the TRPV‐4 blocker ruthenium red (Δ[Ca2+]i, RR, 19±1.2nM n=4). These data suggest an important role for the TRPV‐4 channels in vasodilation to Ach, and regulation of endothelial intracellular Ca2+in mice MA.