TRPV1 Expression Research Articles

Complement Receptor C3aR1 Contributes to Paclitaxel-Induced Peripheral Neuropathic Pain in Mice and Rats.

Cancer chemotherapy-induced neuropathic pain is a devastating pain syndrome without effective therapies. We previously reported that rats deficient in complement C3, the central component of complement activation cascade, showed a reduced degree of paclitaxel-induced mechanical allodynia (PIMA), suggesting that complement is integrally involved in the pathogenesis of this model. However, the underlying mechanism was unclear. Complement activation leads to the production of C3a, which mediates inflammation through its receptor C3aR1. In this article, we report that the administration of paclitaxel induced a significantly higher expression level of C3aR1 on dorsal root ganglion (DRG) macrophages and expansion of these macrophages in DRGs in wild-type (WT) compared with in C3aR1 knockout (KO) mice. We also found that paclitaxel induced less severe PIMA, along with a reduced DRG expression of transient receptor potential channels of the vanilloid subtype 4 (TRPV4), an essential mediator for PIMA, in C3aR1 KO than in WT mice. Treating WT mice or rats with a C3aR1 antagonist markedly attenuated PIMA in association with downregulated DRG TRPV4 expression, reduced DRG macrophages expansion, suppressed DRG neuron hyperexcitability, and alleviated peripheral intraepidermal nerve fiber loss. Administration of C3aR1 antagonist to TRPV4 KO mice further protected them from PIMA. These results suggest that complement regulates PIMA development through C3aR1 to upregulate TRPV4 on DRG neurons and promote DRG macrophage expansion. Targeting C3aR1 could be a novel therapeutic approach to alleviate this debilitating pain syndrome.

The Potential of Berberine to Target Telocytes in Rabbit Heart.

A brand-new class of interstitial cells, called telocytes, has been detected in the heart. Telocytes can connect and transmit signals to almost all cardiomyocytes; this is highly interrelated with the occurrence and development of heart diseases. Modern studies have shown that berberine has a therapeutic effect on cardiovascular health. However, berberine's mechanism of action on the cardiovascular system through cardiac telocytes is unclear. Interestingly, 5 µm of berberine remarkably decreased the concentration of intracellular calcium and membrane depolarization in cultured telocytes, upregulated the expression of CX43 and β-catenin, and downregulated the expressions of TRPV4 and TRPV1. Here, telocytes were identified in the vascular adventitia and intima, endocardium, myocardium, adventitia, and heart valves. Moreover, telocytes were broadly dispersed around cardiac vessels and interacted directly through gap junctions and indirectly through extracellular vesicles. Together, cardiac telocytes interact with berberine and then deliver drug information to the heart. Telocytes may be an essential cellular target for drug therapy of the cardiovascular system.

Transient receptor potential vanilloid 4 promotes cutaneous wound healing by regulating keratinocytes and fibroblasts migration and collagen production in fibroblasts in a mouse model

BackgroundTransient receptor potential vanilloid 4 (TRPV4), a cation ion channel, is expressed in different cells, and it regulates the development of different diseases. We recently found a high TRPV4 expression in the wounded skin area. However, the role of TRPV4 in cutaneous wound healing is unknown. ObjectiveTo investigate the role of TRPV4 in cutaneous wound healing in a mouse model. MethodsSkin wound healing experiment and histopathological studies were performed between WT and TRPV4 KO mice. The effect of TRPV4 antagonist and agonist on cell migration, proliferation, and differentiation were examined in vitro. ResultsTRPV4 expression was enhanced in wounded area in the skin. TRPV4 KO mice had impaired cutaneous wound healing compared with the WT mice. Further, they had significantly suppressed re-epithelialization and formation of granulation tissue, amount of collagen deposition, and number of α-SMA-positive myofibroblasts in skin wounds. qPCR revealed that the KO mice had decreased mRNA expression of COL1A1 and ACTA2 in skin wounds. In vitro, treatment with selective TRPV4 antagonist suppressed migrating capacity, scratch stimulation enhanced the expression of phospho-ERK in keratinocytes, and TGF-β stimulation enhanced the mRNA expression of COL1A1 and ACTA2 in fibroblasts. Selective TRPV4 agonist suppressed cell migration in keratinocytes, and did not enhance proliferation and migration, but promoted differentiation in fibroblasts. ConclusionTRPV4 mediates keratinocytes and fibroblasts migration and increases collagen deposition in the wound area, thereby promoting cutaneous wound healing.

The state of mitochondrial membrane potential in peripheral blood leukocytes of patients with chronic obstructive pulmonary disease

Introduction. Mitochondria provide energy homeostasis of the cell by maintaining an optimal transmembrane electrochemical gradient (ΔΨm), which does not allow excessive formation of reactive oxygen species (ROS). However, under conditions of pathology, the normal functioning of mitochondria is disrupted, which can lead to ATP deficiency and/or increased production of ROS.Aim. The aim of this study was to investigate the ΔΨm parameters and their relationship with the expression of TRP channels in peripheral blood leukocytes of patients with chronic obstructive pulmonary disease (COPD).Materials and methods. The study included 23 patients with COPD of varying severity, 8 smokers without signs of bronchial obstruction and 9 healthy volunteers who had never smoked. All subjects underwent spirometry to assess the lung function. ΔΨm was determined by staining the cells with tetramethylrhodamine ethyl ester (TMRE) and measuring the fluorescent signal by flow cytometry, under basal conditions and pro-inflammatory stimulation with phorbol-12-myristate-13-acetate (PMA).Results. We found that COPD patients were characterized by a significant increase in basal ΔΨm of monocytes (161.8 [153.8; 206.8] vs. 129.3 [75.5; 161.8], p=0.03) and lymphocytes (209,7 [184.7; 257.8] vs. 122.5 [67.9; 164.3], p=0.003) as compared with the control group. Stimulation of cells with PMA led to multidirectional changes in ΔΨm, while its increased level was still preserved in COPD. In monocytes of COPD patients, a decrease in ΔΨm in response to PMA stimulation was prevalent (75%), while in the majority (53.9%) of individuals in the control group ΔΨm, on the contrary, increased (p=0.08). In addition, among COPD patients, an increase in ΔΨm in monocytes was accompanied by an enhanced expression of TRPV4, while in the control group, among individuals with positive dynamics of ΔΨm, TRPV4 expression was, on the contrary, reduced.Conclusion. The increased level of ΔΨm in the mononuclears of COPD patients is consistent with previously detected enhanced ROS production, but does not support the assumption about energy deficit in the cells. The revealed differences in the relationship between TRPV4 expression and ΔΨm dynamics may indicate the presence of pathological features in TRP signaling in COPD patients.

Neuroinflammation in the Dorsal Root Ganglia and Dorsal Horn Contributes to Persistence of Nociceptor Sensitization in SIV-Infected Antiretroviral Therapy-Treated Macaques

Despite the development of antiretroviral therapy (ART), HIV-associated distal sensory polyneuropathy remains prevalent. Using SIV-infected rhesus macaques, this study examined molecular mechanisms of peripheral and central sensitization to infer chronic pain from HIV infection. Previous studies identified atrophy in nociceptive neurons during SIV infection, which was associated with monocyte infiltration into the dorsal root ganglia (DRG). However, the sensory signaling mechanism connecting this pathology to symptoms remains unclear, especially because pain persists after resolution of high viremia and inflammation with ART. We hypothesized that residual DRG and dorsal horn neuroinflammation contributes to nociceptive sensitization. Using three cohorts of macaques [uninfected (SIV-), SIV-infected (SIV+), and SIV infected with ART (SIV+/ART)], this study showed an increase in the cellular and cytokine inflammatory profiles in the DRG of SIV+/ART macaques compared with uninfected animals. It found significant increase in the expression of nociceptive ion channels, TRPV1, and TRPA1 among DRG neurons in SIV+/ART compared with uninfected animals. SIV-infected and SIV+/ART animals showed reduced innervation of the nonpeptidergic nociceptors into the dorsal horn compared with uninfected animals. Finally, there were a significantly higher number of CD68+ cells in the dorsal horn of SIV+/ART macaques compared with uninfected animals. In summary, these data demonstrate that neuroinflammation, characteristics of nociceptor sensitization, and central terminal atrophy persists in SIV+/ART animals.

Maresin-1 prevents blood-spinal cord barrier disruption associated with TRPV4 elevation in the experimental model of spinal cord injury

Recently, we reported the TRPV4 ion channel activation and its association with secondary damage after spinal cord injury (SCI). TRPV4 activation is linked with blood-spinal cord barrier (BSCB) disruption, endothelial damage, and inflammation after SCI. Specialized pro-resolving mediators (SPM) are endogenous lipid mediators released for inflammation resolution. Studies suggest that SPM could act as an endogenous antagonist of ion channels directly or indirectly at the plasma membrane. Herein, we studied the effect of maresin-1, a docosahexaenoic acid (DHA)-derived SPM, in SCI-induced TRPV4 expression and subsequent associated damage. First, employing a particular agonist (4αPDD) in endothelial and neuronal cell lines, we examined the potential of maresin-1 to block TRPV4 activation. Then we quantify the DHA levels in plasma and epicenter of the spinal cord in sham and at 1, 3, 7, 14, 21, and 28-days post-injury (DPI) using LC-MS. Then, we exogenously administered maresin-1 using two dosing regimens i.e., single-dose (1 μg) and multiple-dose (1 μg/day for seven days), to confirm its role in the TRPV4 inhibition and its linked damage. After SCI, DHA levels decrease in the spinal cord epicenter area as well as in the plasma. Treatment with maresin-1 attenuates TRPV4 expression, inflammatory cytokines, and chemokines and impedes neutrophil infiltration. Furthermore, treatment with maresin-1 prevents BSCB disruption, alleviates glial scar formation, and improves functional recovery. Thus, our results suggest that maresin-1 could modulate TRPV4 expression and could be a safe and promising approach to target inflammation and BSCB damage after SCI.

Differential outcomes of high-fat diet on age-related rescaling of cochlear frequency place coding.

Auditory frequency coding is place-specific, which depends on the mechanical coupling of the basilar membrane-outer hair cell (OHC)-tectorial membrane network. Prestin-based OHC electromotility improves cochlear frequency selectivity and sensitivity. Cochlear amplification determines the frequency coding wherein discrete sound frequencies find a 'best' place along the cochlear length. Loss of OHC is the leading cause of age-related hearing loss (ARHL) and is the most common cause of sensorineural hearing loss and compromised speech perception. Lipid interaction with Prestin impacts OHC function. It has been established that high-fat diet (HFD) is associated with ARHL. To determine whether genetic background and metabolism preserve cochlear frequency place coding, we examined the effect of HFD in C57BL/6J (B6) and CBA/CaJ (CBA) on ARHL.We found a significant rescuing effect on ARHL in aged B6 HFD cohort. Prestin levels and cell sizes were better maintained in the experimental B6-HFD group. We also found that distortion product otoacoustic emission (DPOAE) group delay measurement was preserved, which suggested stable frequency place coding. In contrast, the response to HFD in the CBA cohort was modest with no appreciable benefit to hearing threshold. Notably, group delay was shortened with age along with the control. In addition, the frequency dependent OHC nonlinear capacitance gradient was most pronounced at young age but decreased with age. Cochlear RNA-seq analysis revealed differential TRPV1 expression and lipid homeostasis. Activation of TRPV1 and downregulation of arachidonic acid led to downregulation of inflammatory response in B6 HFD, which protects the cochlea from ARHL. The genetic background and metabolic state-derived changes in OHC morphology and function collectively contribute to a redefined cochlear frequency place coding and improved age-related pitch perception.

Capsaicin alleviates the intestinal oxidative stress via activation of TRPV1/PKA/UCP2 and Keap1/Nrf2 pathways in heat-stressed mice

Heat stress (HS) can lead to oxidative stress (OS), which jeopards human health and animal production. Capsaicin (CAP) has antioxidant activities. This study aimed to investigate the impacts of CAP on the alleviation of intestinal OS in heat-stressed mice. All 40 male C57BL/6J mice were randomly allotted to 4 groups in a 2 × 2 factorial arrangement with 2 levels of CAP (0 and 0.4 mg/d for 15 days) and 2 ambient temperatures (normal and HS (39.5 °C per day for 2 h for the last 6 days) conditions). CAP treatment enhanced the villus height under HS conditions. CAP supplementation increased serum superoxide dismutase (SOD) activity, jejunal glutathione peroxidase (GPX) activity, and HO-1, SOD1, SOD2, and GPX1 gene abundance, also upregulated TRPV1 and UCP2 expressions in heat-stressed mice. Meanwhile, CAP administration reversed the HS-induced elevation of the TNF-α and IFN-γ abundance. Collectively, CAP could enhance intestinal antioxidant capacity in heat-stressed mice.

Knockdown of TRPV2 inhibits the migration of RAW264.7 cells toward low fluid shear stress region.

Our previous studies have demonstrated that macrophages (RAW264.7) have a special ability for sensing the gradient of fluid shear stress (FSS) and migrate toward the low-FSS region. However, the molecular mechanism regulating this phenomenon is still unclear. In this study, we examined the transcriptome genes in RAW264.7 cells, MC3T3-E1 osteoblasts, mesenchymal stem cells, canine renal epithelial cells, and periodontal ligament cells. The expression levels of genes related to cell migration, force transfer, and force sensitivity in the Ca2+ signaling pathway were analyzed. We observed that the transient receptor potential cation channel type 2 (TRPV2) was highly expressed in RAW264.7 cells. Furthermore, we used lentiviral transfection to knockdown TRPV2 expression in RAW264.7 cells and studied the effect of TRPV2 on the migration of RAW264.7 cells under a gradient FSS field. The results showed that compared with normal cells, TRPV2-knockdown cells had impaired ability for sensing FSS gradient to migrate toward the low-FSS region and lower intracellular calcium response to FSS stimulation. This study may reveal the molecular mechanism of regulating the directional migration of macrophages under a gradient FSS field.

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.2

Transient Receptor Potential channels (TRP) in GtoPdb v.2023.2

Effect of hypoandrogenism on expression of transient receptor potential vanilloid channels in rat penile corpus cavernosum and erectile function.

Hypoandrogenism is a cause of erectile dysfunction (ED). Vascular smooth muscle cell contraction and relaxation are regulated by TRPV1-4 channels. However, the influence of hypoandrogenism on TRPV1-4 and its relationship with erectile function remain unclear. To reveal whether hypoandrogenism affects erectile function by influencing TRPV1-4 expression in the corpus cavernosum of rats. Male Sprague-Dawley rats (N = 36) aged 8 weeks were assigned to 6 groups at random (n = 6): sham operation, castrated, castrated + testosterone replacement, sham operation + transfection, castrated + transfection, and castrated + empty transfection. Four weeks after castration, 20μL of lentiviral vector (1 × 108 TU/mL) carrying the TRPV4 gene was injected into the penile cavernous tissue of the transfection groups. One week after transfection, the maximum intracavernous pressure (ICPmax)/mean arterial pressure (MAP) and the content of TRPV1-4, phosphorylated eNOS (p-eNOS)/eNOS, and nitric oxide (NO) in penile cavernous tissue of each group were measured. Under low androgen conditions, TRPV4 expression in endothelial cells in the rat penile cavernosum was sharply reduced, resulting in a decrease in p-eNOS/eNOS and NO content, which could inhibit erectile function. In rat penile cavernous tissue, TRPV1-4 was expressed in the cell membranes of endothelial cells and smooth muscle cells. The ICPmax/MAP and the content of TRPV4, p-eNOS/eNOS, and NO end product nitrite level in rat penile cavernous tissue was markedly reduced in the castrated group as compared with the sham group (P < .05). The ICPmax/MAP and the content of TRPV4, p-eNOS/eNOS, and NO end product nitrite level in rat penile cavernous tissue were markedly improved in the castrated + transfection group vs the castrated group (P < .01). Upregulation of TRPV4 expression in penile cavernosum tissue might be a viable therapeutic for ED caused by hypoandrogenism. The specific mechanism of TRPV4 in ED needs to be further verified by androgen receptor or TRPV4 gene knockout experiments. Hypoandrogenism may cause ED by reducing the expression of TRPV4 in rat penile cavernous tissue. Upregulation of TRPV4 expression in penile cavernous tissue can increase the ratio of p-eNOS/eNOS and NO levels and ameliorate the erectile function of castrated rats.

Transient receptor potential M2 channel in the hypothalamic preoptic area and its impact on thermoregulation during menopause

BackgroundDecreased estrogen levels can cause abnormal thermosensitivity of the preoptic area (POA) in the hypothalamus during menopause, which may cause hot flashes. Thermosensitive transient receptors (ThermoTRPs) affect the thermosensitivity of neurons. It is worth exploring whether ThermoTRPs change under low estrogen state and participate in the abnormal thermoregulation of POA. MethodsAdult female Sprague–Dawley rats were randomly divided into sham operation (SHAM), ovariectomy (OVX) and estrogen treatment after ovariectomy (OVX+E) groups. Under 10 ℃, 18 ℃, 25 ℃, 37 ℃ and 45 ℃ incubations, their skin temperature was monitored and the expression of TRPA1, TRPM8, TRPM2, and TRPV1 in POA were investigated. ResultsThe skin temperature of ovariectomized rats changed faster and more dramatically under different incubation temperatures. The results at mRNA level show that only the expression of TRPM2 decreased in POA of OVX group compared with the other two groups at 25 ℃, TRPA1 expression in POA of the three groups increased at 10 ℃, TRPM8 increased at 10 ℃ and 18 ℃, TRPV1 increased at 10 ℃ and 45 ℃, while the expression of TRPM2 decreased at 10 ℃ and 18 ℃ and increased at 37 ℃ and 45 ℃. In all these cases, the magnitudes of the changes were less in the OVX group relative to the other two groups. The further immunohistochemical and Western blot results of TRPM2 and the activated TRPM2 positive cells labeled by c-Fos were consistent with the results of mRNA level. ConclusionsThe expression and thermosensitivity of TRPM2 in POA changed greatly under different incubation temperatures, but the changes in ovariectomized rats were less. This may be the key factor triggering thermoregulation dysfunction under low estrogen and may cause hot flashes.

Mechanisms of peripheral sensitization in endometriosis patients with peritoneal lesions and acyclical pain

PurposeEndometriosis (EM) is one of the most frequent differential diagnoses concerning chronic pelvic pain. Women under hormonal therapy (HT) often benefit from it but sometimes suffer a setback and develop acyclical pelvic pain. Due to the assumption that mechanisms of neurogenic inflammation are involved in the generation of chronic pelvic pain, we aimed to investigate the expression of sensory nerve markers in EM-associated nerve fibers of patients with/without HT.MethodsLaparoscopically excised peritoneal samples from 45 EM and 10 control women were immunohistochemically stained for: PGP9.5, Substance P (SP), NK1R, NGFp75, TRPV-1, and TrkA. Demographics and severity of pain were documented.ResultsEM patients showed a higher nerve fiber density (PGP9.5 and SP) and increased expression of NGFp75, TRPV1, TrkA, and NK1R in blood vessels and immune cells compared with controls. Patients with HT have cycle-dependent pelvic pain but suffer from acyclical pelvic pain. Interestingly, reducing NK1R expression in blood vessels under HT was observed. A correlation between dyspareunia severity and nerve fibers density and between NGFRp75 expression in blood vessels and cycle-dependent pelvic pain severity was observed.ConclusionPatients under HT have no ovulation and no (menstrual) bleeding, which correlate with inflammation and cyclical pain. However, acyclical pain seems to be due to peripheral sensitization once it is present under treatment. Neurotransmitters, like SP and their receptors, are involved in mechanisms of neurogenic inflammation, which are relevant for pain initiation. These findings indicate that in both groups (EM with/without HT), neurogenic inflammation is present and responsible for acyclical pain.

Vitexin inhibits pain and itch behavior via modulating TRPV4 activity in mice

Itching and pain are distinct unpleasant sensations. The transient receptor potential cation channel subfamily V member 4 (TRPV4) pathway is regarded as a shared pathway that mediates pain and itching. Vitexin (Mujingsu, MJS), a C-glycosylflavonoid, is an effective analgesic. This study aimed to explore the antinociceptive and anti-pruritic effects of MJS and whether its effects are mediated via the TRPV4 pathway. Mice were treated with MJS (7.5 mg/kg) 0.5 h prior to the initiation of the pain or itch modeling process. The results showed that MJS suppressed pain-like behavior in hot plate, thermal infiltration, glacial acetic acid twisting, and formalin tests. Administration of MJS decreased the pruritus response induced by histamine, C48/80, chloroquine and BAM8-22 within 30 min. MJS reduced scratching bouts and lessened the wiping reaction of mice under TRPV4 activation by GSK101 (10 µg/5 μl). MJS inhibited scratching behavior in acetone–ether–water (AEW)-treated mice within 60 min. An H1 receptor antagonist—chlorpheniramine (CLP, 400 mg/kg)—and a TRPV4 antagonist—HC067047 (250 ng/kg), exhibited similar effects to those of MJS. Moreover, MJS ameliorated dry skin itch-associated cutaneous barrier disruption in mice. MJS did not inhibit the expression of TRPV4 in the dorsal root ganglion neurons at L2–L3 in AEW mice. These results indicate that the analgesic and anti-pruritic effects of MJS in acute and chronic pain and itching, as well as itching caused by TRPV4 activation, could be attributed to the TRPV4 pathway modulation.

Protein Arginine Methyltransferase 5 Contributes to Paclitaxel-Induced Neuropathic Pain by Activating Transient Receptor Potential Vanilloid 1 Epigenetic Modification in Dorsal Root Ganglion.

Paclitaxel (PTX), which is a first-line chemotherapy drug used to treat various types of cancers, exhibits peripheral neuropathy as a common side effect that is difficult to treat. Protein arginine methyltransferase 5 (PRMT 5) is a key regulator of the chemotherapy response, as chemotherapy drugs induce PRMT5 expression. However, little is known about the PRMT5-mediated epigenetic mechanisms involved in PTX-induced neuropathic allodynia. Sprague-Dawley rats were intraperitoneally given PTX to induce neuropathic pain. Biochemical analyses were conducted to measure the protein expression levels in the dorsal root ganglion (DRG) of the animals. The von Frey test and hot plate test were used to evaluate nociceptive behaviors. PTX increased the PRMT5 (mean difference [MD]: 0.68, 95% confidence interval [CI], 0.88-0.48; P < .001 for vehicle)-mediated deposition of histone H3R2 dimethyl symmetric (H3R2me2s) at the transient receptor potential vanilloid 1 ( Trpv1 ) promoter in the DRG. PRMT5-induced H3R2me2s recruited WD repeat domain 5 (WDR5) to increase trimethylation of lysine 4 on histone H3 (H3K4me3) at Trpv1 promoters, thus resulting in TRPV1 transcriptional activation (MD: 0.65, 95% CI, 0.82-0.49; P < .001 for vehicle) in DRG in PTX-induced neuropathic pain. Moreover, PTX increased the activity of NADPH oxidase 4 (NOX4) (MD: 0.66, 95% CI, 0.81-0.51; P < .001 for vehicle), PRMT5-induced H3R2me2s, and WDR5-mediated H3K4me3 in the DRG in PTX-induced neuropathic pain. Pharmacological antagonism and the selective knockdown of PRMT5 in DRG neurons completely blocked PRMT5-mediated H3R2me2s, WDR5-mediated H3K4me3, or TRPV1 expression and neuropathic pain development after PTX injection. Remarkably, NOX4 inhibition not only attenuated allodynia behavior and reversed the above-mentioned signaling but also reversed NOX4 upregulation via PTX. Thus, the NOX4/PRMT5-associated epigenetic mechanism in DRG has a dominant function in the transcriptional activation of TRPV1 in PTX-induced neuropathic pain.

Nerve terminals in the tumor microenvironment as targets for local infiltration analgesia

Nerve terminals within the tumor microenvironment as potential pain-mitigating targets for local infiltration analgesia is relatively less explored. In this study, we examine the role of key analgesics administered as local infiltration analgesia in a model of cancer-induced bone pain (CIBP). CIBP was induced by administration of allogenic MRMT1 breast cancer cells in the proximal tibia of rats, and tumor mass characterized using radiogram, micro-CT, and histological analysis. In vitro responsiveness to key analgesics δ-opioid receptor agonist (DOPr), Ca2+ channel and TRPV1 antagonists was assessed using ratiometric Ca2+ imaging in sensory neurons innervating the tumor site. Effectiveness of locally infiltrated analgesics administered independently or in combination was assessed by quantifying evoked limb withdrawal thresholds at two distinct sites for up to 14 days. CIBP animals demonstrated DOPr, N-, and L-type and TRPV1 expression in lumbar dorsal root ganglion neurons (DRG), comparable to controls. Evoked Ca2+ transients in DRG neurons from CIBP animals were significantly reduced in response to treatment with compounds targeting DOPr, N-, L-type Ca2+ channels and TRPV1 proteins. Behaviourally, evoked hyperalgesia at the tumor site was strongly mitigated by peritumoral injection of the DOPr agonist and T-type calcium antagonist, via its activity on bone afferents. Results from this study suggest that nerve terminals at tumor site could be utilized as targets for specific analgesics, using local infiltration analgesia.

Functional expression of the thermally activated transient receptor potential channels TRPA1 and TRPM8 in human myotubes

Transient potential (TRP) ion channels expressed in primary sensory neurons act as the initial detectors of environmental cold and heat, information which controls muscle energy expenditure. We hypothesize that non-neuronal TRPs have direct cellular responses to thermal exposure, also affecting cellular metabolism. In the present study we show expression of TRPA1, TRPM8 and TRPV1 in rat skeletal muscle and human primary myotubes by qPCR. Effects of TRP activity on metabolism in human myotubes were studied using radiolabeled glucose. FURA-2 was used for Ca2+ imaging.TRPA1, TRPM8 and TRPV1 were expressed at low levels in primary human myotubes and in m. gastrocnemius, m. soleus, and m. trapezius from rat. Activation of TRPA1 by ligustilide resulted in an increased glucose uptake and oxidation in human myotubes, whereas activation of TRPM8 by menthol and icilin significantly decreased glucose uptake and oxidation. Activation of heat sensing TRPV1 by capsaicin had no effect on glucose metabolism. Agonist-induced increases in intracellular Ca2+ levels by ligustilide and icilin in human myotubes confirmed a direct activation of TRPA1 and TRPM8, respectively. The mRNA expression of some genes involved in thermogenesis, i.e. peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), uncoupling protein (UCP) 1 and UCP3, were downregulated in human myotubes following TRPA1 activation, while the mRNA expression of TRPM8 and TRPA1 were downregulated following TRPM8 activation by menthol and icilin, respectively. Cold exposure (18 °C) of cultured myotubes followed by a short recovery period had no effect on glucose uptake and oxidation in the basal situation, however when TRPA1 and TRPM8 channels were chemically inhibited a temperature-induced difference in glucose metabolism was found.In conclusion, mRNA of TRPA1, TRPM8 and TRPV1 are expressed in rat skeletal muscle and human skeletal muscle cells. Modulation of TRPA1 and TRPM8 by chemical agents induced changes in Ca2+ levels and glucose metabolism in human skeletal muscle cells, indicating functional receptors.

Capsaicin Changes the Pattern of Brain Rhythms in Sleeping Rats.

The heat and capsaicin sensor TRPV1 ion channels were originally discovered in sensory neurons of dorsal root ganglia, and later found in many other tissues and organs. However, whether TRPV1 channels are present in brain regions other than the hypothalamus has been a subject of debate. Here, we addressed this issue with an unbiased functional test by recording electroencephalograms (EEGs) to examine whether capsaicin injection directly into the rat lateral ventricle could alter brain electrical activity. We observed that EEGs during the sleep stage could be significantly perturbed by capsaicin, whereas EEGs during the awake stage did not show a detectable change. Our results are consistent with TRPV1 expression in selective brain regions whose activities are dominative during the sleep stage.

Human Osteoarthritic Chondrocytes Express Nineteen Different TRP-Genes-TRPA1 and TRPM8 as Potential Drug Targets.

Transient receptor potential (TRP) ion channels are expressed in neuronal and some non-neuronal cells and are involved particularly in pain and thermosensation. We previously showed that TRPA1 is functionally expressed in human osteoarthritic (OA) chondrocytes and mediates inflammation, cartilage degradation, and pain in monosodium-iodoacetate-induced experimental OA. In the present study, we explored the expression of TRP-channels in primary human OA chondrocytes and investigated whether drugs used in the treatment of OA, ibuprofen and glucocorticoids, have effects on TRP-channel expression. OA cartilage was obtained from knee replacement surgery and chondrocytes were isolated with enzyme digestion. NGS analysis showed the expression of 19 TRP-genes in OA chondrocytes, with TRPM7, TRPV4, TRPC1, and TRPM8 having the highest counts in unstimulated cells. These results were verified with RT-PCR in samples from a different group of patients. Interleukin-1β (IL-1β) significantly increased TRPA1 expression, while TRPM8 and TRPC1 expression was decreased, and TRPM7 and TRPV4 expression remained unaffected. Furthermore, dexamethasone attenuated the effect of IL-1β on TRPA1 and TRPM8 expression. The TRPM8 and TRPA1 agonist menthol increased the expression of the cartilage-degrading enzymes MMP-1, MMP-3, and MMP-13 and the inflammatory factors iNOS and IL-6 in OA chondrocytes. In conclusion, human OA chondrocytes express 19 different TRP-genes, of which the significant TRPM8 expression is a novel finding. Dexamethasone attenuated IL-1β-induced TRPA1 expression. Interestingly, the TRPM8 and TRPA1 agonist menthol increased MMP expression. These results support the concept of TRPA1 and TRMP8 as potential novel drug targets in arthritis.

Endothelial basement membrane laminins - new players in mouse and human myoendothelial junctions and shear stress communication

Basement membranes (BMs) are critical but frequently ignored components of the vascular system. Using high-resolution confocal imaging of whole-mount-stained mesenteric arteries, we identify integrins, vinculin, focal adhesion kinase (FAK) and several BM proteins including laminins as novel components of myoendothelial junctions (MEJs), anatomical microdomains that are emerging as regulators of cross-talk between endothelium and smooth muscle cells (SMCs). Electron microscopy revealed multiple layers of the endothelial BM that surround endothelial projections into the smooth muscle layer as structural characteristics of MEJs. The shear-responsive calcium channel TRPV4 is broadly distributed in endothelial cells and occurs in a proportion of MEJs where it localizes to the tips of the endothelial projections that are in contact with the underlying SMCs. In mice lacking the major endothelial laminin isoform, laminin 411 (Lama4−/−), which we have previously shown over-dilate in response to shear and exhibit a compensatory laminin 511 upregulation, localization of TRPV4 at the endothelial-SMC interface in MEJs was increased. Endothelial laminins do not affect TRPV4 expression, rather in vitro electrophysiology studies using human umbilical cord arterial endothelial cells revealed enhanced TRPV4 signalling upon culturing on an RGD-motif containing domain of laminin 511. Hence, integrin-mediated interactions with laminin 511 in MEJ structures unique to resistance arteries modulate TRPV4 localization at the endothelial-smooth muscle interface in MEJs and signalling over this shear-response molecule.