Transient Receptor Potential Melastatin Subtype Research Articles

Inhibition of TRPM8 function by prostacyclin receptor agonists requires coupling to Gq/11 proteins.

The TRPM8 ion channel is involved in innocuous cold sensing and has a potent anti-inflammatory action. Its activation by lower temperature or chemical agonists such as menthol and icilin induces analgesic effects, reversing hypersensitivity and reducing chronic pain. On the other hand, prostacyclin (PGI2) enhances pain and inflammation by activating the IP receptors. Due to the critical roles of TRPM8 and IP receptors in the regulation of inflammatory pain, and considering their overlapping expression pattern, we analysed the functional interaction between human TRPM8 and IP receptors. We transiently expressed human TRPM8 channels and IP receptors in HEK293T cells and carried out intracellular calcium and cAMP measurements. Additionally, we cultured neurons from the dorsal root ganglia (DRGs) of mice and determined the increase in intracellular calcium triggered by the TRPM8 agonist, icilin, in the presence of the IP receptor agonist cicaprost, the IP receptor antagonist Cay10441, and the Gq/11 inhibitor YM254890. Activation of IP receptors by selective agonists (cicaprost, beraprost, and iloprost) inhibited TRPM8 channel function, independently of the Gs-cAMP pathway. The potent inhibition of TRPM8 channels by IP receptor agonists involved Gq/11 coupling. These effects were also observed in neurons isolated from murine DRGs. Our results demonstrate an unusual signalling pathway of IP receptors by coupling to Gq/11 proteins to inhibit TRPM8 channel function. This pathway may contribute to a better understanding of the role of TRPM8 channels and IP receptors in regulating pain and inflammation.

Tegaserod maleate exhibits antileukemic activity by targeting TRPM8

To identify therapeutic targets in acute myeloid leukemia (AML), we conducted growth inhibition screens of 2040 small molecules from a library of FDA-approved drugs using a panel of 12 AML cell lines. Tegaserod maleate, a 5-hydroxytryptamine 4 receptor partial agonist, elicits strong anti-AML effects in vitro and in vivo by targeting transient receptor potential melastatin subtype 8 (TRPM8), which plays critical roles in several important processes. However, the role of TRPM8 remains incompletely described in AML, whose treatment is based mostly on antimitotic chemotherapy. Here, we report an unexpected role of TRPM8 in leukemogenesis. Strikingly, TRPM8 knockout inhibits AML cell survival/proliferation by promoting apoptosis. Mechanistically, TRPM8 exerts its oncogenic effect by regulating the ERK–CREB/c-Fos signaling axis. Hyperactivation of ERK signaling can be reversed by TRPM8 inhibition. Importantly, TRPM8 is overexpressed in AML patients, indicating that it is a new prognostic factor in AML. Collectively, our work demonstrates the anti-AML effects of tegaserod maleate via targeting TRPM8 and indicates that TRPM8 is a regulator of leukemogenesis with therapeutic potential in AML.

Correlation between nasal mucosal temperature change and nasal airflow perception

The mechanism of nasal airflow perception remains little known. It is currently believed that the main mechanism for perceiving nasal patency is to activate transient receptor potential melastatin subtype 8. Computer fluent dynamics show that increased airflow and heat flux are associated with higher subjective scores. Similarly, physical measurements of the nasal cavity using a temperature probe show a correlation between the lower nasal mucosa temperature and better results. Trigeminal function detection also indirectly confirms this. This literature review aimed to explore the role of nasal mucosal temperature change in the subjective perception of nasal patency and the secondary aim was to appraise the relevant evidence about the mechanism.

β-Lactam TRPM8 Antagonist RGM8-51 Displays Antinociceptive Activity in Different Animal Models.

Transient receptor potential melastatin subtype 8 (TRPM8) is a cation channel extensively expressed in sensory neurons and implicated in different painful states. However, the effectiveness of TRPM8 modulators for pain relief is still a matter of discussion, since structurally diverse modulators lead to different results, depending on the animal pain model. In this work, we described the antinociceptive activity of a β–lactam derivative, RGM8-51, showing good TRPM8 antagonist activity, and selectivity against related thermoTRP channels and other pain-mediating receptors. In primary cultures of rat dorsal root ganglion (DRG) neurons, RGM8-51 potently reduced menthol-evoked neuronal firing without affecting the major ion conductances responsible for action potential generation. This compound has in vivo antinociceptive activity in response to cold, in a mouse model of oxaliplatin-induced peripheral neuropathy. In addition, it reduces cold, mechanical and heat hypersensitivity in a rat model of neuropathic pain arising after chronic constriction of the sciatic nerve. Furthermore, RGM8-51 exhibits mechanical hypersensitivity-relieving activity, in a mouse model of NTG-induced hyperesthesia. Taken together, these preclinical results substantiate that this TRPM8 antagonist is a promising pharmacological tool to study TRPM8-related diseases.

TRPM8 Channels: Advances in Structural Studies and Pharmacological Modulation.

The transient receptor potential melastatin subtype 8 (TRPM8) is a cold sensor in humans, activated by low temperatures (>10, <28 °C), but also a polymodal ion channel, stimulated by voltage, pressure, cooling compounds (menthol, icilin), and hyperosmolarity. An increased number of experimental results indicate the implication of TRPM8 channels in cold thermal transduction and pain detection, transmission, and maintenance in different tissues and organs. These channels also have a repercussion on different kinds of life-threatening tumors and other pathologies, which include urinary and respiratory tract dysfunctions, dry eye disease, and obesity. This compendium firstly covers newly described papers on the expression of TRPM8 channels and their correlation with pathological states. An overview on the structural knowledge, after cryo-electron microscopy success in solving different TRPM8 structures, as well as some insights obtained from mutagenesis studies, will follow. Most recently described families of TRPM8 modulators are also covered, along with a section of molecules that have reached clinical trials. To finalize, authors provide an outline of the potential prospects in the TRPM8 field.

Diminished Cold Avoidance Behaviours after Chronic Cold Exposure – Potential Involvement of TRPM8

Ambient temperature changes trigger plastic biological responses. Cold temperature is detected by the somatosensory system and evokes perception of cold together with adaptive physiological responses. We addressed whether chronic cold exposure induces adaptive adjustments of (1) thermosensory behaviours, and (2) the principle molecular cold sensor in the transduction machinery, transient receptor potential melastatin subtype 8 (TRPM8). Mice in two groups were exposed to either cold (6 °C) or thermoneutral (27 °C) ambient temperatures for 4 weeks and subjected to thermosensory behavioural testing. Cold group mice behaved different from Thermoneutral group in the Thermal Gradient Test: the former occupied a wider temperature range and was less cold avoidant. Furthermore, subcutaneous injection of the TRPM8 agonist icilin, enhanced cold avoidance in both groups in the Thermal Gradient Test, but Cold group mice were significantly less affected by icilin. Primary sensory neuron soma are located in dorsal root ganglia (DRGs), and western blotting showed diminished TRPM8 levels in DRGs of Cold group mice, as compared to the Thermoneutral group. We conclude that acclimation to chronic cold altered thermosensory behaviours, so that mice appeared less cold sensitive, and potentially, TRPM8 is involved.

Recent Progress in TRPM8 Modulation: An Update.

The transient receptor potential melastatin subtype 8 (TRPM8) is a nonselective, multimodal ion channel, activated by low temperatures (<28 °C), pressure, and cooling compounds (menthol, icilin). Experimental evidences indicated a role of TRPM8 in cold thermal transduction, different life-threatening tumors, and other pathologies, including migraine, urinary tract dysfunction, dry eye disease, and obesity. Hence, the modulation of the TRPM8 channel could be essential in order to understand its implications in these pathologies and for therapeutic intervention. This short review will cover recent progress on the TRPM8 agonists and antagonists, describing newly reported chemotypes, and their application in the pharmacological characterization of TRPM8 in health and disease. The recently described structures of the TRPM8 channel alone or complexed with known agonists and PIP2 are also discussed.

A1954 TRPM8 activation blunts mitochondrial dysfunction-mediated inflammation in monocytes during cold-induced hypertension

Objectives: Inflammation and oxidative stress in mononuclear cells (MCs) are enhanced in cold-induced hypertension (CIH). Transient receptor potential melastatin subtype 8 (TRPM8) is a mild cold-sensing nonselective cation channel activated by menthol. Little is known about the effect of TRPM8 activation by menthol on inflammation and oxidative stress in MCs during CIH. Methods: TRPM8 knockout (KO) mice and wild-type (WT) littermates were maintained at 23–25°C, whereas mice treated with cold stimulation were exposed to 4°C in a temperature-controlled and ventilated chamber for 4 h every day for 24 weeks. Mice were given normal chow or normal chow +0.5% menthol. TRPM8 expression and mitochondrial respiratory function were determined. Intracellular and mitochondrial free calcium as well as reactive oxygen species (ROS) were measured. Results: Dietary menthol supplementation lowered systolic blood pressure in WT mice in a TRPM8-dependent manner. TRPM8 activation by dietary menthol blunted inflammatory responses in CIH. Dietary menthol restored cold stimulation-induced vasodilatation of mesenteric arterial rings in WT but not in TRPM8−/− mice. This beneficial effect of TRPM8 activation by dietary menthol was associated with reduction in expression of the NLRP3 inflammasome in MCs from CIH mice. TRPM8 activation attenuated the dysfunction and ROS generation in mitochondria mediated by activation of the NLRP3 inflammasomein THP-1 MCs. Conclusion: Chronic dietary menthol antagonizes hypertension and cellular inflammation through improvement of mitochondrial dysfunction in CIH.

Nociceptive Roles of TRPM2 Ion Channel in Pathologic Pain.

Pain is a protective mechanism that enables us to avoid potentially harmful environments. However, when pathologically persisted and aggravated under severely injured or inflamed conditions, pain often reduces the quality of life and thus is considered as a disease to eliminate. Inflammatory and/or neuropathic mechanisms may exaggerate interactions between damaged tissues and neural pathways for pain mediation. Similar mechanisms also promote the communication among cellular participants in synapses at spinal or higher levels, which may amplify nociceptive firing and subsequent signal transmission, deteriorating the pain sensation. In this pathology, important cellular players are afferent sensory neurons, peripheral immune cells, and spinal glial cells. Arising from damage of injury, overloaded interstitial and intracellular reactive oxygen species (ROS) and intracellular Ca2+ are key messengers in the development and maintenance of pathologic pain. Thus, an ROS-sensitive and Ca2+-permeable ion channel that is highly expressed in the participant cells might play a critical role in the pathogenesis. Transient receptor potential melastatin subtype 2 (TRPM2) is the unique molecule that satisfies all of the requirements: the sensitivity, permeability, and its expressing cells. Notable progress in delineating the role of TRPM2 in pain has been achieved during the past decade. In the present review, we summarize the important findings in the key cellular components that are involved in pathologic pain. This overview will help to understand TRPM2-mediated pain mechanisms and speculate therapeutic strategies by utilizing this updated information.

Activation of Transient Receptor Potential Melastatin Subtype 8 Attenuates Cold-Induced Hypertension Through Ameliorating Vascular Mitochondrial Dysfunction.

BackgroundEnvironmental cold‐induced hypertension is common, but how to treat cold‐induced hypertension remains an obstacle. Transient receptor potential melastatin subtype 8 (TRPM8) is a mild cold‐sensing nonselective cation channel that is activated by menthol. Little is known about the effect of TRPM8 activation by menthol on mitochondrial Ca2+ homeostasis and the vascular function in cold‐induced hypertension.Methods and ResultsPrimary vascular smooth muscle cells from wild‐type or Trpm8−/− mice were cultured. In vitro, we confirmed that sarcoplasmic reticulum–resident TRPM8 participated in the regulation of cellular and mitochondrial Ca2+ homeostasis in the vascular smooth muscle cells. TRPM8 activation by menthol antagonized angiotensin II induced mitochondrial respiratory dysfunction and excess reactive oxygen species generation by preserving pyruvate dehydrogenase activity, which hindered reactive oxygen species–triggered Ca2+ influx and the activation of RhoA/Rho kinase pathway. In vivo, long‐term noxious cold stimulation dramatically increased vasoconstriction and blood pressure. The activation of TRPM8 by dietary menthol inhibited vascular reactive oxygen species generation, vasoconstriction, and lowered blood pressure through attenuating excessive mitochondrial reactive oxygen species mediated the activation of RhoA/Rho kinase in a TRPM8‐dependent manner. These effects of menthol were further validated in angiotensin II–induced hypertensive mice.ConclusionsLong‐term dietary menthol treatment targeting and preserving mitochondrial function may represent a nonpharmaceutical measure for environmental noxious cold–induced hypertension.

Segmental transport of Ca²⁺ and Mg²⁺ along the gastrointestinal tract.

Calcium (Ca(2+)) and magnesium (Mg(2+)) ions are involved in many vital physiological functions. Since dietary intake is the only source of minerals for the body, intestinal absorption is essential for normal homeostatic levels. The aim of this study was to characterize the absorption of Ca(2+) as well as Mg(2+) along the gastrointestinal tract at a molecular and functional level. In both humans and mice the Ca(2+) channel transient receptor potential vanilloid subtype 6 (TRPV6) is expressed in the proximal intestinal segments, whereas Mg(2+) channel transient receptor potential melastatin subtype 6 (TRPM6) is expressed in the distal parts of the intestine. A method was established to measure the rate of Mg(2+) absorption from the intestine in a time-dependent manner by use of (25)Mg(2+). In addition, local absorption of Ca(2+) and Mg(2+) in different segments of the intestine of mice was determined by using surgically implanted intestinal cannulas. By these methods, it was demonstrated that intestinal absorption of Mg(2+) is regulated by dietary needs in a vitamin D-independent manner. Also, it was shown that at low luminal concentrations, favoring transcellular absorption, Ca(2+) transport mainly takes place in the proximal segments of the intestine, whereas Mg(2+) absorption predominantly occurs in the distal part of the gastrointestinal tract. Vitamin D treatment of mice increased serum Mg(2+) levels and 24-h urinary Mg(2+) excretion, but not intestinal absorption of (25)Mg(2+). Segmental cannulation of the intestine and time-dependent absorption studies using (25)Mg(2+) provide new ways to study intestinal Mg(2+) absorption.

Activation of Cold-Sensing Transient Receptor Potential Melastatin Subtype 8 Antagonizes Vasoconstriction and Hypertension Through Attenuating RhoA/Rho Kinase Pathway

Environmental cold is a nonmodifiable hypertension risk factor. Transient receptor potential melastatin subtype 8 (TRPM8) is a cold-sensing cation channel that can be activated by menthol, a compound with a naturally cold sensation in mint. Little is known about the effect of TRPM8 activation on vascular function and blood pressure. Here, we report that TRPM8 is abundantly expressed in the vasculature. TRPM8 activation by menthol attenuated vasoconstriction via RhoA/Rho kinase pathway inhibition in wild-type mice, but the effect was absent in TRPM8(-/-) mice. Chronic dietary menthol blunted mesenteric arterial constriction and lowered blood pressure in genetic hypertensive rats via inhibition of RhoA/Rho kinase expression and activity in the vivo study. TRPM8 effect was associated with inhibition of intracellular calcium release from the sarcoplasmic reticulum, RhoA/Rho kinase activity, and sustained arterial contraction in the vitro study. Importantly, 8-week chronic menthol capsule treatment moderately lowered systolic blood pressure and diastolic blood pressure in prehypertensive individuals compared with the placebo group. Furthermore, chronic menthol capsule administration also improved flow-mediated dilatation in prehypertensive individuals, but not in the placebo group. In conclusion, our study demonstrates that TRPM8 activation by menthol benefits vascular function and blood pressure by inhibiting calcium signaling-mediated RhoA/Rho kinase activation in the vasculature. These findings add to the evidence that long-term dietary menthol treatment had favorable effects on hypertension treatment.

Involvement of transient receptor potential melastatin-8 (TRPM8) in menthol-induced calcium entry, reactive oxygen species production and cell death in rheumatoid arthritis rat synovial fibroblasts

Rheumatoid arthritis is most prominently characterized by synoviocyte hyperplasia which therefore serves as an important target for clinical therapy. In the present study, it was observed that menthol, the specific agonist of transient receptor potential melastatin subtype 8 (TRPM8), could induce sustained increases of cytosolic calcium concentration ([Ca2+]c) in synoviocytes isolated from collagen-induced arthritis rats in dose-dependent manner, which was evidently blocked by applying an extracellular Ca2+-free buffer. Menthol-induced [Ca2+]c increase was also significantly inhibited by potent TRPM8 antagonist capsazepine (CZP), indicating that this [Ca2+]c elevation was mostly attributed to TRPM8-mediated Ca2+ entry. Besides, RT-PCR indeed demonstrated presence of TRPM8 in the synoviocytes. Meanwhile, it was found that menthol evoked production of intracellular reactive oxygen species, which could be abolished by Ca2+ free solutions or CZP. Further experiments showed that menthol reduced the cell numbers and survival of synoviocytes. This reduction was associated with apoptosis as suggested by mitochondrial membrane depolarization, nuclear condensation and a caspase 3/7 apoptotic assay. Menthol-induced death and apoptosis of synoviocytes both were obviously inhibited by CZP, intracellular calcium chelator BAPTA-AM, and reactive oxygen species inhibitor diphenylene iodonium, respectively. Taken together, our data indicated that menthol resulted in synoviocyte death associated with apoptosis via calcium entry and reactive oxygen species production depending on TRPM8 activation.

Epithelial Mg2+ channel TRPM6: insight into the molecular regulation

Our understanding of the molecular mechanisms of renal magnesium (Mg2+) handling has greatly enhanced over recent years. This review highlights the regulatory factors controlling Mg2+ homeostasis through its effects on the epithelial Mg2+ channel TRPM6 (Transient Receptor Potential Melastatin subtype 6), the gatekeeper of the body's Mg2+ balance. Drug treatment, acid-base status, and several hormones have been shown to regulate TRPM6 expression, while its channel activity is modified by intracellular Mg2+, pH, and ATP. Recently, epidermal growth factor (EGF) and estrogen have been implicated as magnesiotropic hormones. The stimulation of the EGF receptor (EGFR) leads to an intracellular cascade involving Rac1 that promotes trafficking of TRPM6 to the plasma membrane. Furthermore, long-term EGF treatment upregulates the expression of TRPM6. Estrogen has also been shown to stimulate TRPM6 activity upon short-term treatment, next to its long-term regulatory effect on TRPM6 transcription. TRPM6, and its closest homologue TRPM7, are composed of a Mg2+ -permeable channel fused to an alpha-kinase domain. In the intracellular compartment, the receptor for activated C-kinase (RACK1), the repressor for estrogen receptor activity (REA), and ATP were identified as negative modulators of TRPM6 activity through its alpha-kinase domain. Therefore, the a-kinase domain acts as an indirect player involved in Mg2+ homeostasis by its feedback function in the TRPM6-mediated Mg2+ influx.

Identification of Pore Residues Engaged in Determining Divalent Cationic Permeation in Transient Receptor Potential Melastatin Subtype Channel 2

The molecular basis for divalent cationic permeability in transient receptor potential melastatin subtype (TRPM) channels is not fully understood. Here we studied the roles of all eight acidic residues, glutamate or aspartate, and also the glutamine residue between pore helix and selectivity filter in the pore of TRPM2 channel. Mutants with alanine substitution in each of the acidic residues, except Glu-960 and Asp-987, formed functional channels. These channels exhibited similar Ca2+ and Mg2+ permeability to wild type channel, with the exception of the E1022A mutant, which displayed increased Mg2+ permeability. More conservative E960Q, E960D, and D987N mutations also led to loss of function. The D987E mutant was functional and showed greater Ca2+ permeability along with concentration-dependent inhibition of Na+-carrying currents by Ca2+. Incorporation of negative charge in place of Gln-981 between the pore helix and selectivity filter by changing it to glutamate, which is present in the more Ca2+-permeable TRPM channels, substantially increased Ca2+ permeability. Expression of concatemers linking wild type and E960D mutant subunits resulted in functional channels that exhibited reduced Ca2+ permeability. These data taken together suggest that Glu-960, Gln-981, Asp-987, and Glu-1022 residues are engaged in determining divalent cationic permeation properties of the TRPM2 channel.