Transient Receptor Potential Vanilloid 1 Agonist Research Articles

Blockage of transient receptor potential vanilloid 4 inhibits brain edema in middle cerebral artery occlusion mice.

Brain edema is an important pathological process during stroke. Activation of transient receptor potential vanilloid 4 (TRPV4) causes an up-regulation of matrix metalloproteinases (MMPs) in lung tissue. MMP can digest the endothelial basal lamina to destroy blood brain barrier, leading to vasogenic brain edema. Herein, we tested whether TRPV4-blockage could inhibit brain edema through inhibiting MMPs in middle cerebral artery occlusion (MCAO) mice. We found that the brain water content and Evans blue extravasation at 48 h post-MCAO were reduced by a TRPV4 antagonist HC-067047. The increased MMP-2/9 protein expression in hippocampi of MCAO mice was attenuated by HC-067046, but only the increased MMP-9 activity was blocked by HC-067047. The loss of zonula occludens-1 (ZO-1) and occludin protein in MCAO mice was also attenuated by HC-067047. Moreover, MMP-2/9 protein expression increased in mice treated with a TRPV4 agonist GSK1016790A, but only MMP-9 activity was increased by GSK1016790A. Finally, ZO-1 and occludin protein expression was decreased by GSK1016790A, which was reversed by an MMP-9 inhibitor. We conclude that blockage of TRPV4 may inhibit brain edema in cerebral ischemia through inhibiting MMP-9 activation and the loss of tight junction protein.

Tramadol and its metabolite m1 selectively suppress transient receptor potential ankyrin 1 activity, but not transient receptor potential vanilloid 1 activity.

The transient receptor potential vanilloid 1 (TRPV1) and the transient receptor potential ankyrin 1 (TRPA1), which are expressed in sensory neurons, are polymodal nonselective cation channels that sense noxious stimuli. Recent reports showed that these channels play important roles in inflammatory, neuropathic, or cancer pain, suggesting that they may serve as attractive analgesic pharmacological targets. Tramadol is an effective analgesic that is widely used in clinical practice. Reportedly, tramadol and its metabolite (M1) bind to μ-opioid receptors and/or inhibit reuptake of monoamines in the central nervous system, resulting in the activation of the descending inhibitory system. However, the fundamental mechanisms of tramadol in pain control remain unclear. TRPV1 and TRPA1 may be targets of tramadol; however, they have not been studied extensively. We examined whether and how tramadol and M1 act on human embryonic kidney 293 (HEK293) cells expressing human TRPV1 (hTRPV1) or hTRPA1 by using a Ca imaging assay and whole-cell patch-clamp recording. Tramadol and M1 (0.01-10 μM) alone did not increase in intracellular Ca concentration ([Ca]i) in HEK293 cells expressing hTRPV1 or hTRPA1 compared with capsaicin (a TRPV1 agonist) or the allyl isothiocyanate (AITC, a TRPA1 agonist), respectively. Furthermore, in HEK293 cells expressing hTRPV1, pretreatment with tramadol or M1 for 5 minutes did not change the increase in [Ca]i induced by capsaicin. Conversely, pretreatment with tramadol (0.1-10 μM) and M1 (1-10 μM) significantly suppressed the AITC-induced [Ca]i increases in HEK293 cells expressing hTRPA1. In addition, the patch-clamp study showed that pretreatment with tramadol and M1 (10 μM) decreased the inward currents induced by AITC. These data indicate that tramadol and M1 selectively inhibit the function of hTRPA1, but not that of hTRPV1, and that hTRPA1 may play a role in the analgesic effects of these compounds.

The Role of TRPV1‐Expressing Neurons in the Control of Heart Rate During Dynamic Exercise in Rats

Heart rate (HR) increases at the beginning of dynamic exercise, then remains constant or gradually increases in response to the exercise intensity. Both central command and the exercise pressor reflex (EPR) are thought to play an important role for this tachycardic response. Recently, the contribution of transient receptor potential vanilloid 1 (TRPV1) channel to the EPR has been reported (Smith SA et al, J Physiol 2010). However, the role of the TRPV1 channel on the exercise tachycardia during dynamic exercise remains largely unknown. We therefore examined the effect of systemic TRPV1 channel blockade or activation on the HR response to treadmill exercise in rats. Male Wistar rats were previously implanted with telemetry transmitters (TRM54P, Millar) to measure HR and arterial blood pressure (AP), and trained to run on a treadmill. Both HR and mean AP (MAP) responses to treadmill running (20 or 40 m/min, 60 s) were compared before and after administration of TRPV1 channel agonists or antagonists. Neither the TRPV1‐selective (SB366791; 3 mg/kg) nor the nonselective (ruthenium red; 5 mg/kg) antagonists affected the HR response to exercise. However, another TRPV1 selective antagonist 5′‐iodoresiniferatoxin (IRTX; 0.3 mg/kg) or a TRPV1 agonist capsaicin (20 mg/kg, sc under anesthesia) markedly attenuated the HR response to exercise. MAP responses to exercise were not affected by these drugs. Given the partial agonist‐like effect of IRTX, systemic desensitization of the TRPV1 channel‐expressing neurons attenuated the EPR‐mediated HR response during dynamic exercise in rats. In conclusion, TRPV1‐expressing neurons play an important role for the HR response during dynamic exercise.

Mo1692 Vascular Endothelial Transient Receptor Potential Vanilloid 4 Links Colonic Inflammation in Dextran Sulfate Sodium-Induced Colitis Model Mice

BACKGROUND&AIMS: Transient receptor potential vanilloid 4 (TRPV4) is a nonselective cation channel, and involved in physical sensing in various types of tissues. TRPV4 is expressed in sensory neuron and vascular endothelial cells in various organs. In gastrointestinal tracts, TRPV4 activation in epithelial cells increases in intracellular calcium concentrations and induces colitis. However, the localization and the role of the TRPV4 channel expressed in vascular endothelial cells in colitis are still poorly defined. In the present study, we investigated the alteration of TRPV4 expression in the colonic vascular endothelial cells during dextran sulfate sodium (DSS)-induced colitis. METHODS: Eight week-old wild type (WT) and TRPV4 knock-out (TRPV4 KO) mice were used. Colitis was induced by 2% DSS solution given as drinking water for 7 days. The disease activity index (DAI) was evaluated during the treatment, while colon length, histological damage, and myeloperoxidase (MPO) activity were examined post-DSS treatment. Immunohistochemical analysis was performed in distal colon. TRPV4-immunoreactivity was detected by using immunohistochemical staining with fluorescein-conjugated tyramide amplification. CD31, CD105, CD11b, CD68, Ly6B.2, and VEGF receptor-2 were detected by indirect staining with their specific antibodies. Vascular leakage was investigated using Evans blue dye extrusion assay. RESULTS: TRPV4 KO mice protected against DSS-induced weight loss, neutrophil (MPO), colon shortening, diarrhea, occult-fecal blood, and histological damage. TRPV4 immunoreactivities were detected mostly in epithelial cells of normal distal colon. DSS treatment drastically increased TRPV4 expression in vascular endothelial cells of mucosa and submucosal layer which colocalized with CD31, CD105, and VEGF receptor-2, but not epithelial cells. Repeated intrarectal and intravenous administration of TRPV4 agonist GSK1016790A exacerbated the severity of DSS colitis. Increased vascular permeability was observed following single intravenous administration of GSK1016790A in DSS colitis and this effect was abolished with TRPV4 antagonist RN1734. CD11b, CD68, Ly6B.2 immunopositive cells were detected around TRPV4 positive vascular endothelial cells in mucosa and submucosal layer in DSS colitis. CONCLUSION: These findings indicate that the alteration of TRPV4 in vascular endothelial cells may contribute to progression of colonic inflammation via increasing vascular permeability.

Mo1693 Activation of Bone Marrow-Derived TRPV4-Expressing M2 Macrophages Promotes Intestinal Contraction

BACKGROUND&AIMS: Transient receptor potential vanilloid 4 (TRPV4) is a nonselective cation channel, and involved in physical sensing in various types of tissues. TRPV4 is expressed in sensory neuron and vascular endothelial cells in various organs. In gastrointestinal tracts, TRPV4 activation in epithelial cells increases in intracellular calcium concentrations and induces colitis. However, the localization and the role of the TRPV4 channel expressed in vascular endothelial cells in colitis are still poorly defined. In the present study, we investigated the alteration of TRPV4 expression in the colonic vascular endothelial cells during dextran sulfate sodium (DSS)-induced colitis. METHODS: Eight week-old wild type (WT) and TRPV4 knock-out (TRPV4 KO) mice were used. Colitis was induced by 2% DSS solution given as drinking water for 7 days. The disease activity index (DAI) was evaluated during the treatment, while colon length, histological damage, and myeloperoxidase (MPO) activity were examined post-DSS treatment. Immunohistochemical analysis was performed in distal colon. TRPV4-immunoreactivity was detected by using immunohistochemical staining with fluorescein-conjugated tyramide amplification. CD31, CD105, CD11b, CD68, Ly6B.2, and VEGF receptor-2 were detected by indirect staining with their specific antibodies. Vascular leakage was investigated using Evans blue dye extrusion assay. RESULTS: TRPV4 KO mice protected against DSS-induced weight loss, neutrophil (MPO), colon shortening, diarrhea, occult-fecal blood, and histological damage. TRPV4 immunoreactivities were detected mostly in epithelial cells of normal distal colon. DSS treatment drastically increased TRPV4 expression in vascular endothelial cells of mucosa and submucosal layer which colocalized with CD31, CD105, and VEGF receptor-2, but not epithelial cells. Repeated intrarectal and intravenous administration of TRPV4 agonist GSK1016790A exacerbated the severity of DSS colitis. Increased vascular permeability was observed following single intravenous administration of GSK1016790A in DSS colitis and this effect was abolished with TRPV4 antagonist RN1734. CD11b, CD68, Ly6B.2 immunopositive cells were detected around TRPV4 positive vascular endothelial cells in mucosa and submucosal layer in DSS colitis. CONCLUSION: These findings indicate that the alteration of TRPV4 in vascular endothelial cells may contribute to progression of colonic inflammation via increasing vascular permeability.

PD20-05 HERPES SIMPLEX VIRUS (HSV) VECTOR-MEDIATED GENE DELIVERY OF PROTEIN PHOSPHATASE 1α REDUCES BLADDER OVERACTIVITY AND

INTRODUCTION AND OBJECTIVES: Increased afferent excitability has been proposed as an important pathophysiological basis of overactive bladder (OAB) and hypersensitive bladder disorders such as interstitial cystitis/bladder pain syndrome (IC/BPS). Previous studies reported that transient receptor potential vanilloid-1 (TRPV1) receptors greatly contribute to afferent sensitization. We recently found Protein Phosphatase 1a (PP1a) as a modulator of TRPV1 receptor activity using an HSV vector cDNA library screen method to identify genes that antagonize TRPV1 activation. HSV vector-mediated PP1a expression significantly reduced capsaicin-induced thermal hyperalgesia. Therefore, we investigated the effect of HSV vectors-mediated gene delivery of PP1a on bladder overactivity and pain-related behavior in rats. METHODS: Replication-deficient HSV vectors encoding PP1a or green fluorescent protein (GFP) as control were injected into the bladder wall of adult female Sprague-Dawley rats. Cystometry (CMG) under urethane anesthesia was performed 1 week after viral injection to evaluate bladder overactivity induced by resiniferatoxin (RTX, a TRPV1 agonist). RTX-induced nociceptive behavior such as licking (lower abdominal licking) and freezing (motionless headturning) was observed 2 weeks after viral innoculation. GFP expression in the L6/S1 DRG and the bladder as well as c-Fos positive cells in the L6 spinal cord dorsal horn were evaluated. The expression of p-ERK, p-Inositol trisphosphate receptor (p-IP3R), and p-TRPV1 in the bladder was estimated by Western blot. RESULTS: GFP expression was seen in L6/S1 DRG as well as in the bladder. In CMG, the PP1a group showed a significantly (p1⁄40.03) smaller reduction (46 1%) in intercontraction intervals after RTX infusion than the GFP group (64 2%). The number of RTX-induced freezing behavior, which is correlated with bladder pain sensation, was significantly (p<0.001) decreased in the PP1a vs. GFP groups. The number of c-Fos positive cells in the L6 spinal dorsal horn was significantly (p<0.01) smaller in the PP1a vs. the GFP group (33 4 vs. 59 6 cells per section). Western blot revealed that p-ERK, p-IP3R, and p-TRPV1 was less expressed in the PP1a vs. the GFP group (p1⁄40.07, 0.01, and 0.04, respectively) CONCLUSIONS: These results indicate that HSV vectorsmediated gene delivery of PP1a can suppress TRPV1-mediated bladder overactivityandpainbehavior, possibly due todecreasedphosphorylation of TRPV1 channels. Thus, PP1a gene thetapy could be a novel treatment of OAB and/or hypersensitive bladder disorders such as IC/BPS.

Ablation and Regeneration of Peripheral and Central TRPV1 Expressing Nerve Terminals and the Consequence of Nociception

Transient Receptor Potential Vanilloid 1 (TRPV1) expressed in peripheral terminals is responsible for transducing thermal and chemical nociception. Role of TRPV1 expressed in the central terminals is not clear, however, its activation modulates synaptic transmission and contributes to central sensitization. In this study, we have determined the role of TRPV1 expressed in the peripheral and central terminals using resiniferatoxin (RTX), a potent TRPV1 agonist. A single intraplantar injection of RTX, within two days induced loss of capsaicin-induced nocifensive behavior and enhanced response latency to hot plate, which recovered over a period of two months. RTX treatment resulted in the ablation of peripheral TRPV1 expressing fibers in paw skin, which regenerated over the same time period. On the other hand, a single dose of intrathecal administration of RTX, within two days caused thermal hypoalgesia. RTX treatment ablated TRPV1 expressing central sensory nerve terminals. Intriguingly, in contrast to peripheral nerve terminal regeneration that occurred within two months, the central TRPV1 expressing nerve terminals did not regenerate even after five months. The present study demonstrates that TRPV1 in the peripheral and central terminals play a role in nociception and the peripheral terminals have the ability to regenerate, whereas the central terminals do not regenerate even after five months.

Interference of TRPV1 function altered the susceptibility of PTZ-induced seizures.

Transient receptor potential vanilloid 1 (TRPV1) is widely distributed in the central nervous system (CNS) including hippocampus, and regulates the balance of excitation and inhibition in CNS, which imply its important role in epilepsy. We used both pharmacological manipulations and transgenic mice to disturb the function of TRPV1 and then studied the effects of these alterations on the susceptibility of pentylenetetrazol (PTZ)-induced seizures. Our results showed that systemic administration of TRPV1 agonist capsaicin (CAP, 40 mg/kg) directly induced tonic-clonic seizures (TCS) without PTZ induction. The severity of seizure was increased in lower doses of CAP groups (5 and 10 mg/kg), although the latency to TCS was delayed. On the other hand, systemic administration of TRPV1 antagonist capsazepine (CPZ, 0.05 and 0.5 mg/kg) and TRPV1 knockout mice exhibited delayed latency to TCS and reduced mortality. Furthermore, hippocampal administration of CPZ (10 and 33 nmol/μL/side) was firstly reported to increase the latency to TCS, decrease the maximal grade of seizure and mortality. It is worth noting that decreased susceptibility of PTZ-induced seizures was observed in hippocampal TRPV1 overexpression mice and hippocampal CAP administration (33 nmol/μL/side), which is opposite from results of systemic agonist CAP. Our findings suggest that the systemic administration of TRPV1 antagonist may be a novel therapeutic target for epilepsy, and alteration of hippocampal TRPV1 function exerts a critical role in seizure susceptibility.

Molecular Mechanism of TRPV1 Activation by Capsaicin

Molecular Mechanism of TRPV1 Activation by Capsaicin

Dietary Capsaicin and Exercise: Analysis of a Two-Pronged Approach to Counteract Obesity

Obesity contributes to diabetes, hypertension and myocardial infarction. Exercise is an effective measure to counteract obesity. Recent research demonstrates a regulatory role of transient receptor potential vanilloid 1 (TRPV1) in high fat diet (HFD)-induced obesity. Here, we evaluated the effects of exercise ± dietary capsaicin (CAP, 0.01% of total HFD), an active ingredient in natural chili peppers and a TRPV1 agonist, on HFD-fed wild type and TRPV1-/- (TRPV1 knockout) mice. We evaluated the performance of normal chow diet (NCD) or HFD (±CAP)-fed mice on computer-controlled rotarod. Trained mice were exercised for 12 min./day for five days a week. HFD+CAP-fed mice walked on the rotarod for a longer duration of the exercise regimen (630 ± 69 sec.) and showed lesser weight gain after 25 weeks of feeding (11.5 ± 2.1 g) compared to exercised HFD-fed mice (440 ± 215 sec.; 27.5 ± 2.1 g). Both sedentary and exercised HFD-fed groups exhibited similar weight gain, albeit an increase in food consumption shown by exercised HFD-fed group. Also, exercised HFD + CAP-fed mice showed an increased metabolic activity compared to exercised HFD-fed group. Further, NCD-fed WT mice walked for longer duration on the rotarod (704 ± 14 sec) and gained lesser weight at 20 weeks of feeding (4.5 ± 0.7 g) than NCD-fed TRPV1-/- mice (665 ± 50 sec.; 7.7 ± 2.1 g). CAP prevented weight gain to a similar extent in both sedentary and exercised wild type mice. Also, Dietary CAP improved the endurance of mice on rotarod and counteracted HFD-induced suppression of muscle coordination. This suggests a novel role of TRPV1 in metabolism and muscle coordination function. Collectively, our data provide evidence for the role of TRPV1 and its activation by dietary CAP and exercise to inhibit HFD-induced obesity.

Transient Receptor Potential Vanilloid 4-Induced Modulation of Voltage-Gated Sodium Channels in Hippocampal Neurons.

Transient receptor potential vanilloid 4 (TRPV4) is reported to control the resting membrane potential and increase excitability in many types of cells. Voltage-gated sodium channels (VGSCs) play an important role in initiating action potentials in neurons. However, whether VGSCs can be modulated by the activation of TRPV4 in hippocampal pyramidal neurons remains unknown. In this study, we tested the effect of TRPV4 agonists (GSK1016790A and 4α-PDD) on voltage-gated sodium current (I Na) in hippocampal CA1 pyramidal neurons and the protein levels of α/β-subunit of VGSCs in the hippocampus of mice subjected to intracerebroventricular (icv.) injection of GSK1016790A (GSK-injected mice). Herein, we report that I Na was inhibited by acute application of GSK1016790A or 4α-PDD. In the presence of TRPV4 agonists, the voltage-dependent inactivation curve shifted to the hyperpolarization, whereas the voltage-dependent activation curve remained unchanged. The TRPV4 agonist-induced inhibition of I Na was blocked by the TRPV4 antagonist or tetrodotoxin. Moreover, blocking protein kinase A (PKA) markedly attenuated the GSK1016790A-induced inhibition of I Na, whereas antagonism of protein kinase C or p38 mitogen-activated protein kinase did not change GSK1016790A action. Finally, the protein levels of Nav1.1, Nav1.2, and Nav1.6 in the hippocampus increased in GSK-injected mice, whereas those of Nav1.3 and Navβ1 remained nearly unchanged. We conclude that I Na is inhibited by the acute activation of TRPV4 through PKA signaling pathway in hippocampal pyramidal neurons, but protein expression of α-subunit of VGSCs is increased by sustained TRPV4 activation, which may compensate for the acute inhibition of I Na and provide a possibility for hyper-excitability upon sustained TRPV4 activation.

Activation of Transient Receptor Potential Vanilloid 4 is Involved in Neuronal Injury in Middle Cerebral Artery Occlusion in Mice.

Transient receptor potential vanilloid 4 (TRPV4) is widely expressed in the central nervous system and can be activated by multiple stimuli during cerebral ischemia. Recently, we reported that intracerebroventricular (icv.) injection of HC-067047, a specific TRPV4 antagonist, reduced brain infarction following 60-min of middle cerebral artery occlusion (MCAO). This study was undertaken to investigate the molecular mechanisms underlying TRPV4-mediated neuronal injury in cerebral ischemia. We demonstrated that TRPV4 expression was upregulated in the ipsilateral hippocampus at 4 to 48h post-MCAO, peaking at 18h post-MCAO. Treatment with TRPV4 antagonists (HC-067047 and ruthenium red) dose-dependently reduced brain infarction at 24h post-MCAO. Phosphorylation of protein kinase B (p-Akt) was downregulated and that of extracellular signal-related kinase (p-ERK) was upregulated at 8 to 24h post-MCAO, which was markedly blocked by treatment with HC-067047. Icv. injection of GSK1016790A (a TRPV4 agonist), dose-dependently induced hippocampal neuronal death, accompanied by an increase in phosphorylation of the NR2B subunit of the N-methyl-D-aspartate receptor (NMDAR). In addition, the level of p-Akt was decreased and that of p-ERK was increased by GSK1016790A-injection, which was sensitive to an NR2B antagonist. The neuronal toxicity of GSK1016790A was blocked by treatment with an NR2B antagonist and a phosphatidylinositol-3-kinase (PI3K) agonist but not by administration of a MAPK/ERK kinase antagonist. We conclude that the activation of TRPV4 is upregulated and involved in neuronal injury during cerebral ischemia and that the neurotoxicity associated with TRPV4-activation is mediated through NR2B-NMDAR and the related downregulation of the Akt signaling pathway.

Transient receptor potential vanilloid 1 is expressed in human cytotrophoblasts: Induction of cell apoptosis and impairment of syncytialization

The normal development of placenta relies essentially on a balanced proliferation, differentiation and apoptosis of cytotrophoblasts. These processes are tightly regulated by several hormones, cytokines, lipids and other molecules and anomalies in these events are associated with gestational complications. The cation channel transient receptor potential vanilloid 1 (TRPV1) is expressed in several organs and tissues and it participates in cellular events like nociception, inflammation and cell death. However, the expression and importance of this receptor in human placenta still remains unknown. In this work, we found that TRPV1 is expressed in human cytotrophoblasts and syncytiotrophoblasts. Furthermore, the TRPV1 agonists capsaicin and anandamide decreased cytotrophoblast viability and induced morphological alterations, such as chromatin condensation and fragmentation, which suggest the occurrence of apoptosis. Also, both TRPV1 agonists induced a loss of mitochondrial membrane potential and an increase of caspase 3/7 activity and production of reactive species of oxygen and nitrogen. Furthermore, capsaicin (10μM) impaired the spontaneous in vitro differentiation of cytotrophoblasts into syncytiotrophoblasts by triggering TRPV1, as observed by the decrease in placental alkaline phosphatase activity and in human chorionic gonadotropin secretion. On the other hand, anandamide decreased placental alkaline phosphatase activity via a TRPV1-independent mechanism but did not influence the secretion of human chorionic gonadotropin. In conclusion, we showed that TRPV1 is expressed in human cytotrophoblasts and syncytiotrophoblasts and also reported the involvement of this receptor in cytotrophoblast apoptosis and differentiation.

Abstract 084: Activation Of Trpv1-expressing Renal Sensory Nerves With N-oleoyldopamine Attenuates High Fat Diet-induced Impairment In Renal Function

Renal injury occurs in obesity. Accumulating evidence indicates that activation of the transient receptor potential vanilloid 1 (TRPV1) protects tissues from injury albeit the mechanisms are largely unknown. We test the hypothesis that high fat diet (HFD) intake impairs afferent renal nerves expressing TRPV1 channels, leading to increased renal sympathetic nerve activity (RSNA), decreased renal function, and hypertension, and that chronic activation of TRPV1 positive afferent renal nerves attenuates HFD-induced impairment. N-oleoyldopamine (OLDA, 1 ng/kg, daily), a selective TRPV1 agonist, was administrated intrathecally (T8-L3) via a indwelled catheter to chronically activate TRPV1 positive afferent renal nerves in rats fed a HFD or normal fat diet (Con) for 8 weeks. HFD decreased renal TRPV1 expression and afferent renal nerve activity (ARNA, Con: 133±8, Con+OLDA: 127±15, HFD: 84±5, HFD+OLDA: 115±7, p&lt;0.05) in response to intra-pelvis perfusion of capsaicin (4 μM, 3 min, 20 ml/min), a selective TRPV1 agonist, which were prevented by OLDA. HFD increased RSNA responses to intrathecal injection of muscimol (3 nmol/kg), a GABA-A receptor agonist, and urinary norepinephrine levels, which were prevented by OLDA. HFD decreased creatinine clearance and increased urinary albumin levels, which were prevented by OLDA (Creatinine clearance, Con: 0.52±0.09, Con+OLDA: 0.54±0.12, HFD: 0.24±0.04, HFD+OLDA: 0.40±0.06 ml/min/100 gbwt, p&lt;0.05). HFD increased levels of collagen deposition, connective tissue growth factor (CTGF), and matrix metalloproteinase-2 (MMP-2) in the kidney, which were prevented by OLDA. HFD increased systolic blood pressure by the week of 6 after HFD, which was prevented by OLDA. Thus, HFD impairs TRPV1-positive afferent renal nerves, increases renal sympathetic nerve activity, and leads to renal injury and hypertension. Segment-specific intrathecal injection of OLDA protects against HFD-induced impairment in afferent renal nerves and prevents HFD-induced renal injury and hypertension. Our data illustrate that preservation of TRPV1 positive afferent renal nerves may be a therapeutic strategy in preventing obesity- or type 2 diabetes-induced renal injury and hypertension.

Functional transient receptor potential vanilloid 1 and transient receptor potential vanilloid 4 channels along different segments of the renal vasculature.

Transient receptor potential vanilloid 1 (TRPV1) and vanilloid 4 (TRPV4) cation channels have been recently identified to promote endothelium-dependent relaxation of mouse mesenteric arteries. However, the role of TRPV1 and TRPV4 in the renal vasculature is largely unknown. We hypothesized that TRPV1/4 plays a role in endothelium-dependent vasodilation of renal blood vessels. We studied the distribution of functional TRPV1/4 along different segments of the renal vasculature. Mesenteric arteries were studied as control vessels. The TRPV1 agonist capsaicin relaxed mouse mesenteric arteries with an EC50 of 25 nm, but large mouse renal arteries or rat descending vasa recta only at >100-fold higher concentrations. The vasodilatory effect of capsaicin in the low-nanomolar concentration range was endothelium-dependent and absent in vessels of Trpv1 -/- mice. The TRPV4 agonist GSK1016790A relaxed large conducting renal arteries, mesenteric arteries and vasa recta with EC50 of 18, 63 nm and ~10 nm respectively. These effects were endothelium-dependent and inhibited by a TRPV4 antagonist, AB159908 (10 μm). Capsaicin and GSK1016790A produced vascular dilation in isolated mouse perfused kidneys with EC50 of 23 and 3 nm respectively. The capsaicin effects were largely reduced in Trpv1 -/- kidneys, and the effects of GSK1016790A were inhibited in Trpv4 -/- kidneys. Our results demonstrate that two TRPV channels have unique sites of vasoregulatory function in the kidney with functional TRPV1 having a narrow, discrete distribution in the resistance vasculature and TRPV4 having more universal, widespread distribution along different vascular segments. We suggest that TRPV1/4 channels are potent therapeutic targets for site-specific vasodilation in the kidney.

Nonpsychotropic plant cannabinoids, cannabidivarin (CBDV) and cannabidiol (CBD), activate and desensitize transient receptor potential vanilloid 1 (TRPV1) channels in vitro: potential for the treatment of neuronal hyperexcitability.

Epilepsy is the most common neurological disorder, with over 50 million people worldwide affected. Recent evidence suggests that the transient receptor potential cation channel subfamily V member 1 (TRPV1) may contribute to the onset and progression of some forms of epilepsy. Since the two nonpsychotropic cannabinoids cannabidivarin (CBDV) and cannabidiol (CBD) exert anticonvulsant activity in vivo and produce TRPV1-mediated intracellular calcium elevation in vitro, we evaluated the effects of these two compounds on TRPV1 channel activation and desensitization and in an in vitro model of epileptiform activity. Patch clamp analysis in transfected HEK293 cells demonstrated that CBD and CBDV dose-dependently activate and rapidly desensitize TRPV1, as well as TRP channels of subfamily V type 2 (TRPV2) and subfamily A type 1 (TRPA1). TRPV1 and TRPV2 transcripts were shown to be expressed in rat hippocampal tissue. When tested on epileptiform neuronal spike activity in hippocampal brain slices exposed to a Mg(2+)-free solution using multielectrode arrays (MEAs), CBDV reduced both epileptiform burst amplitude and duration. The prototypical TRPV1 agonist, capsaicin, produced similar, although not identical effects. Capsaicin, but not CBDV, effects on burst amplitude were reversed by IRTX, a selective TRPV1 antagonist. These data suggest that CBDV antiepileptiform effects in the Mg(2+)-free model are not uniquely mediated via activation of TRPV1. However, TRPV1 was strongly phosphorylated (and hence likely sensitized) in Mg(2+)-free solution-treated hippocampal tissue, and both capsaicin and CBDV caused TRPV1 dephosphorylation, consistent with TRPV1 desensitization. We propose that CBDV effects on TRP channels should be studied further in different in vitro and in vivo models of epilepsy.

Study on relations between transient receptor potential vanilloid 1 and pungent property of traditional Chinese medicines

The five-flavor theory of traditional Chinese medicines (TCM) and the flavor efficacy generation mechanism has long been focuses and difficulties in studies on traditional Chinese medicinal properties. In this paper, by using the pharmacophore-based virtual screening technique, the authors discussed the relations between the pungent property and transient receptor potential vanilloid 1 (TRPV1) by studying the TCM components' role in regulating TRPV1 ion channel. The results showed that the matching relationship between TRPV1 agonist pharmacophore model and TCM chemical components could identify the active ingredients from pungent herbs. Therefore, the authors proposed that TRPV1 is one of the potential targets for efficient pungent herbs. The pungent property of TCMs is decided by its chemical components, and consistent with the inherited and additive characteristics.

Effects of neonatal treatment with the TRPV1 agonist, capsaicin, on adult rat brain and behaviour

Treatment of neonatal rats with the transient receptor potential vanilloid 1 (TRPV1) channel agonist, capsaicin, produces life-long loss of sensory neurons expressing TRPV1 channels. Previously it was shown that rats treated on day 2 of life with capsaicin had behavioural hyperactivity in a novel environment at 5–7 weeks of age and brain changes reminiscent of those found in subjects with schizophrenia. The objective of the present study was to investigate brain and behavioural responses of adult rats treated as neonates with capsaicin. It was found that the brain changes found at 5–7 weeks in rats treated as neonates with capsaicin persisted into adulthood (12 weeks) but were less in older rats (16–18 weeks). Increased prepulse inhibition (PPI) of acoustic startle was found in these rats at 8 and 12 weeks of age rather than the deficit commonly found in animal models of schizophrenia. Subjects with schizophrenia also have reduced flare responses to niacin and methylnicotinate proposed to be mediated by prostaglandin D2 (PGD2). Flare responses are accompanied by cutaneous plasma extravasation. It was found that the cutaneous plasma extravasation responses to methylnicotinate and PGD2 were reduced in capsaicin-treated rats. In conclusion, several neuroanatomical changes observed in capsaicin-treated rats, as well as the reduced cutaneous plasma extravasation responses, indicate that the role of TRPV1 channels in schizophrenia is worthy of investigation.

TRPV4 activation mediates flow-induced nitric oxide production in the rat thick ascending limb

Nitric oxide (NO) regulates renal function. Luminal flow stimulates NO production in the thick ascending limb (TAL). Transient receptor potential vanilloid 4 (TRPV4) is a mechano-sensitive channel activated by luminal flow in different types of cells. We hypothesized that TRPV4 mediates flow-induced NO production in the rat TAL. We measured NO production in isolated, perfused rat TALs using the fluorescent dye DAF FM. Increasing luminal flow from 0 to 20 nl/min stimulated NO from 8 ± 3 to 45 ± 12 arbitrary units (AU)/min (n = 5; P < 0.05). The TRPV4 antagonists, ruthenium red (15 μmol/l) and RN 1734 (10 μmol/l), blocked flow-induced NO production. Also, luminal flow did not increase NO production in the absence of extracellular calcium. We also studied the effect of luminal flow on NO production in TALs transduced with a TRPV4shRNA. In nontransduced TALs luminal flow increased NO production by 47 ± 17 AU/min (P < 0.05; n = 5). Similar to nontransduced TALs, luminal flow increased NO production by 39 ± 11 AU/min (P < 0.03; n = 5) in TALs transduced with a control negative sequence-shRNA while in TRPV4shRNA-transduced TALs, luminal flow did not increase NO production (Δ10 ± 15 AU/min; n = 5). We then tested the effect of two different TRPV4 agonists on NO production in the absence of luminal flow. 4α-Phorbol 12,13-didecanoate (1 μmol/l) enhanced NO production by 60 ± 11 AU/min (P < 0.002; n = 7) and GSK1016790A (10 ηmol/l) increased NO production by 52 ± 15 AU/min (P < 0.03; n = 5). GSK1016790A (10 ηmol/l) did not stimulate NO production in TRPV4shRNA-transduced TALs. We conclude that activation of TRPV4 channels mediates flow-induced NO production in the rat TAL.

S3-4 H2S targets transient receptor potential receptors to relieve from inflammation and pain

Transient Receptor Potential Vanilloid 4 (TRPV4) and Transient Receptor Potential Ankyrin 1 (TRPA1) are present both in sensory neurons and in intestinal epithelial cells, where their activation causes intracellular calcium mobilization. Activation of each receptor in sensory neurons conveys pro-nociceptive signals, while in intestinal epithelial cells, their activation confers a pro-inflammatory phenotype with chemokine release. Further, TRPV4 or TRPA1 activation in mouse intestine induced visceral hyperalgesia and colitis, confirming that these channels could participate to intestinal pain and inflammation. We hypothesized that H 2 S is able to regulate TRPV4 and TRPA1functions in intestinal epithelial cells and sensory neurons. Methods TRPV4 and TRPA1 responses were evaluated by measuring calcium flux with a Fluo-8 probe in HEK cells transfected with TRPV4 or with TRPA1, but also in normal derived Colon Mucosa (NCM460) cells, Caco-2 cells (an intestinal epithelium cell line) and mouse DRG (dorsal ganglion roots) primary neurons. Cells were pre-exposed to Sodium Hydrogen Sulfide (NaHS) a H 2 S donor, or their vehicle, 3-min before a second stimulation with the specific agonist of TRPV4, 4 α PDD (4 α -phorbol-12,13-didenoate), the agonist of TRPA1 allyl isothiocyanate (AITC) or vehicle. IL-8 expression was also evaluated after 6 h of treatment with NaHS and TRPV4 or TRPA1 stimulation. Results 4 α PDD or AITC but not NaHS increased the intracellular calcium concentration respectively in HEK TRPV4 and HEK TRPA1. TRPV4 or TRPA1 agonist induced a dose-dependent calcium flux in NCM460 cells, Caco-2 cells and neurons. Incubation of NCM460 and Caco-2 cells with NaHS (200–1000 μM) dose-dependently decreased the calcium response to a pro-inflammatory dose of TRPV4 or TRPA1 agonist (50 μM). In neurons, the calcium response to 25 μM of 4 α PDD or AITC was also significantly reduced with a NaHS pre-treatment (1 mM). Furthermore, IL-8 expression was decreased when NCM460 and Caco-2 cells were pre-treated with NaHS. Conclusions NaHS exposure decreased cellular response to TRPV4 or TRPA1 agonist. Considering the pro-inflammatory and pro-nociceptive effects of TRPV4 or TRPA1 activation in the gut, H 2 S could exert there, anti-inflammatory effects through the inhibition of TRPV4 and TRPA1. Hydrogen sulfide efficiently reduces visceral inflammation and pain as previously demonstrated, our results suggest that these protective effects of H2S donnors could be mediated by TRPV4 and TRPA1 inhibition.