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The transient receptor potential melastatin 4 (TRPM4) channel contributes extensively to cardiac electrical activity, especially cardiomyocyte action potential formation. Mechanical stretch can induce changes in heart rate and rhythm, and the mechanosensitive channel Piezo1 is expressed in many cell types within the myocardium. Our previous study showed that TRPM4 and Piezo1 are closely co-localized in the t-tubules of ventricular cardiomyocytes and contribute to the Ca 2+ -dependent signalling cascade that underlies hypertrophy in response to mechanical pressure overload. However, there was no direct evidence showing that Piezo1 activation was related to TRPM4 activation in situ. In the present study, we employed the HL-1 mouse atrial myocyte-like cell line as an in vitro model to investigate whether Piezo1-TRPM4 coupling can affect action potential properties. We used the small molecule Piezo1 agonist, Yoda1, as a surrogate for mechanical stretch to activate Piezo1 and detected the action potential changes in HL-1 cells using FluoVolt, a fluorescent voltage sensitive dye. Our results demonstrate that Yoda1-induced activation of Piezo1 changes the action potential frequency in HL-1 cells. This change in action potential frequency is reduced by Piezo1 knockdown using small intefering RNA. Importantly knockdown or pharmacological inhibition of TRPM4 significantly affected the degree to which Yoda1-evoked Piezo1 activation influenced action potential frequency. Thus, the present study provides in vitro evidence of a functional coupling between Piezo1 and TRPM4 in a cardiomyocyte-like cell line. The coupling of a mechanosensitive Ca 2+ permeable channel and a Ca 2+ -activated TRP channel probably represents a ubiquitous model for the role of TRP channels in mechanosensory transduction. KEY POINTS: The transient receptor potential melastatin 4 (TRPM4) and Piezo1 channels have been confirmed to contribute to the Ca 2+ -dependent signalling cascade that underlies cardiac hypertrophy in response to mechanical pressure overload. However, there was no direct evidence showing that Piezo1 activation was related to TRPM4 activation in situ. We employed the HL-1 mouse atrial myocyte-like cell line as an in vitro model to investigate the effect of Piezo1-TRPM4 coupling on cardiac electrical properties. The results show that both pharmacological and genetic inhibition of TRPM4 significantly affected the degree to which Piezo1 activation influenced action potential frequency in HL-1 cells. Our findings provide in vitro evidence of a functional coupling between Piezo1 and TRPM4 in a cardiomyocyte-like cell line. The coupling of a mechanosensitive Ca 2+ permeable channel and a Ca 2+ -activated TRP channel probably represents a ubiquitous model for the role of TRP channels in mechanosensory transduction in various (patho)physiological processes., (© 2023 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Borneol has been used successfully for the treatment of itchy skin in traditional Chinese medicine. However, the antipruritic effect of borneol has rarely been studied, and the mechanism is unclear. Here, we showed that topical application of borneol on skin substantially suppressed pruritogen chloroquine- and compound 48/80-induced itching in mice. The potential targets of borneol, including transient receptor potential cation channel subfamily V member 3 (TRPV3), transient receptor potential cation channel subfamily A member 1 (TRPA1), transient receptor potential cation channel subfamily M member 8 (TRPM8), and gamma-aminobutyric acid type A (GABA A ) receptor were pharmacologically inhibited or genetically knocked out one by one in mouse. Itching behavior studies demonstrated that the antipruritic effect of borneol is largely independent of TRPV3 and GABA A receptor, and TRPA1 and TRPM8 channels are responsible for a major portion of the effect of borneol on chloroquine-induced nonhistaminergic itching. Borneol activates TRPM8 and inhibits TRPA1 in sensory neurons of mice. Topical co-application of TRPA1 antagonist and TRPM8 agonist mimicked the effect of borneol on chloroquine-induced itching. Intrathecal injection of a group II metabotropic glutamate receptor antagonist partially attenuated the effect of borneol and completely abolished the effect of TRPM8 agonist on chloroquine-induced itching, suggesting that a spinal glutamatergic mechanism is involved. In contrast, the effect of borneol on compound 48/80-induced histaminergic itching occurs through TRPA1-and TRPM8-independent mechanisms. Our work demonstrates that borneol is an effective topical itch reliever, and TRPA1 inhibition and TRPM8 activation in peripheral nerve terminals account for its antipruritic effect., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Transient receptor potential (TRP) family play critical roles in cardiovascular system. TRPM family as largest TRP subfamily is non-voltage Ca2+-activated selective channels which has 8 members. This study aimed to discuss the role of TRPM family in cardiovascular system and diseases. Systematic search was performed covering PubMed, ISI Web of Science, and Google Scholar from inception until June 2021 using related keywords and Mesh terms for English studies with human, animal and in-vitro subjects. Finally 10 studies were selected for data extraction. Reviewing the articles showed that TRPM2, TRPM4, TRPM5, TRPM6 and TRPM7 play important roles in cardiovascular system and diseases. TRPM2 could be activated by reactive oxygen species (ROS) and effects on cardiac injury and cardiac fibrosis. TRPM7 and TRPM6 also have been reported to be associated with cardiac fibrosis and atrial fibrosis development respectively. TRPM4 channels contributed to resting membrane potential of cerebral artery smooth muscle cells and atrial contraction. TRPM5 channels are bitter taste sensors and prevent high salt intake and consequently high blood pressure due to the high salt intake. In conclusion based on the proof of the effectiveness of some members of TRPM family in the cardiovascular system, research on other members of this channel group seems to be useful and necessary to find their possible connection to the cardiovascular system., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Ion channels are ubiquitously expressed in almost all living cells, and are the third-largest category of drug targets, following enzymes and receptors. The transient receptor potential melastatin (TRPM) subfamily of ion channels are important to cell function and survival. Studies have shown upregulation of the TRPM family of ion channels in various brain tumours. Gliomas are the most prevalent form of primary malignant brain tumours with no effective treatment; thus, drug development is eagerly needed. TRPM2 is an essential ion channel for cell function and has important roles in oxidative stress and inflammation. In response to oxidative stress, ADP-ribose (ADPR) is produced, and in turn activates TRPM2 by binding to the NUDT9-H domain on the C-terminal. TRPM2 has been implicated in various cancers and is significantly upregulated in brain tumours. This article reviews the current understanding of TRPM2 in the context of brain tumours and overviews the effects of potential drug therapies targeting TRPM2 including hydrogen peroxide (H 2 O 2 ), curcumin, docetaxel and selenium, paclitaxel and resveratrol, and botulinum toxin. It is long withstanding knowledge that gliomas are difficult to treat effectively, therefore investigating TRPM2 as a potential therapeutic target for brain tumours may be of considerable interest in the fields of ion channels and pharmacology., (© 2021. The Author(s), under exclusive licence to CPS and SIMM.)

D. Julius was awarded the 2021 Medicine Nobel prize for the discovery of new cationic channels that detect temperatures either over 40 °C (TRPV1) or cold (TRPM8) ranging from 8-15 °C, followed by the latter identification of other channels that sense temperatures within other ranges. On the other hand, A. Patapoutian shared the 2021 Nobel prize for the independent and simultaneous co-discovery of the TRPM8 cationic channel. Furthermore, Patapoutian iden-tified piezo 1 and 2 channels previously referred to as the cell mechanosensors related to the sense of touch and proprioception. These experimental findings indicate that these novel cationic channels localized in nerve endings of the skin, mouth, lips, bronchial tree, the nephron, plus a variety of tissues transduce phy-sical stimuli into electrical activity that reach the brain sensory cortex to process these stimuli and elicit animal behavior.

In the lower urinary tract, transient receptor potential (TRP) channels are primarily involved in physiological function, especially in cellular sensors responding to chemical and physical stimuli. Among TRP channels, TRP melastatin 8 (TRPM8) channels, responding to cold temperature and/or chemical agents, such as menthol or icilin, are mainly expressed in the nerve endings of the primary afferent neurons and in the cell bodies of dorsal root ganglia innervating the urinary bladder (via Aδ- and C-fibers); this suggests that TRPM8 channels primarily contribute to bladder sensory (afferent) function. Storage symptoms of overactive bladder, benign prostatic hyperplasia, and interstitial cystitis are commonly related to sensory function (bladder hypersensitivity); thus, TRPM8 channels may also contribute to the pathophysiology of bladder hypersensitivity. Indeed, it has been reported in a pharmacological investigation using rodents that TRPM8 channels contribute to the pathophysiological bladder afferent hypersensitivity of mechanosensitive C-fibers. Similar findings have also been reported in humans. Therefore, a TRPM8 antagonist would be a promising therapeutic target for bladder hypersensitive disorders, including urinary urgency or nociceptive pain. In this review article, the functional role of the TRPM8 channel in the lower urinary tract and the potential of its antagonist for the treatment of bladder disorders was described.

Magnesium is essential for cellular life, but how it is homeostatically controlled still remains poorly understood. Here, we report that members of CNNM family, which have been controversially implicated in both cellular Mg2+ influx and efflux, selectively bind to the TRPM7 channel to stimulate divalent cation entry into cells. Coexpression of CNNMs with the channel markedly increased uptake of divalent cations, which is prevented by an inactivating mutation to the channel's pore. Knockout (KO) of TRPM7 in cells or application of the TRPM7 channel inhibitor NS8593 also interfered with CNNM-stimulated divalent cation uptake. Conversely, KO of CNNM3 and CNNM4 in HEK-293 cells significantly reduced TRPM7-mediated divalent cation entry, without affecting TRPM7 protein expression or its cell surface levels. Furthermore, we found that cellular overexpression of phosphatases of regenerating liver (PRLs), known CNNMs binding partners, stimulated TRPM7-dependent divalent cation entry and that CNNMs were required for this activity. Whole-cell electrophysiological recordings demonstrated that deletion of CNNM3 and CNNM4 from HEK-293 cells interfered with heterologously expressed and native TRPM7 channel function. We conclude that CNNMs employ the TRPM7 channel to mediate divalent cation influx and that CNNMs also possess separate TRPM7-independent Mg2+ efflux activities that contribute to CNNMs' control of cellular Mg2+ homeostasis., Competing Interests: The authors have declared that no competing interests exist.

Transient receptor potential melastatin 2 (TRPM2), a Ca 2+ -permeable cation channel, is gated by intracellular adenosine diphosphate ribose (ADPR), Ca 2+ , warm temperature, and oxidative stress. It is critically involved in physiological and pathological processes ranging from inflammation to stroke to neurodegeneration. At present, the channel's gating and ion permeation mechanisms, such as the location and identity of the selectivity filter, remain ambiguous. Here, we report the cryo-electron microscopy (cryo-EM) structure of human TRPM2 in nanodisc in the ligand-free state. Cryo-EM map-guided computational modeling and patch-clamp recording further identify a quadruple-residue motif as the ion selectivity filter, which adopts a restrictive conformation in the closed state and acts as a gate, profoundly contrasting with its widely open conformation in the Nematostella vectensis TRPM2. Our study reveals the gating of human TRPM2 by the filter and demonstrates the feasibility of using cryo-EM in conjunction with computational modeling and functional studies to garner structural information for intrinsically dynamic but functionally important domains., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Excessive Ca 2+ influx and mitochondrial oxidative stress (OS) of trigeminal ganglia (TG) have essential roles in the etiology of migraine headache and aura. The stimulation of TRPM2 channel via the generation of OS and ADP-ribose (ADPR) induces pain, inflammatory, and oxidative neurotoxicity, although its inhibition reduces the intensity of pain and neurotoxicity in several neurons. However, the cellular and molecular effects of TRPM2 in the TG of migraine model (glyceryl trinitrate, GTN) on the induction of pain, OS, apoptosis, and inflammation remain elusive. GTN-mediated increases of pain intensity, apoptosis, death, cytosolic reactive oxygen species (ROS), mitochondrial ROS, caspase -3, caspase -9, cytosolic Ca 2+ levels, and cytokine generations (TNF-α, IL-1β, and IL-6) in the TG of TRPM2 wild-type mouse were further increased by the TRPM2 activation, although they were modulated by the treatments of GSH, PARP-1 inhibitors (PJ34 and DPQ), and TRPM2 blockers (ACA and 2APB). However, the effects of GTN were not observed in the TG of TRPM2 knockout mice. The current data indicate that the maintaining activation of TRPM2 is not only important for the quenching OS, inflammation, and neurotoxicity in the TG neurons of mice with experimental migraine but also equally critical to the modulation of GTN-induced pain., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Hippocampal cell death and cognitive dysfunction are common following global cerebral ischemia across all ages, including children. Most research has focused on preventing neuronal death. Restoration of neuronal function after cell death is an alternative approach (neurorestoration). We previously identified transient receptor potential M2 (TRPM2) ion channels as a potential target for acute neuroprotection and delayed neurorestoration in an adult CA/CPR mouse model. Cardiac arrest/cardiopulmonary resuscitation (CA/CPR) in juvenile (p20-25) mice was used to investigate the role of ion TRPM2 channels in neuroprotection and ischemia-induced synaptic dysfunction in the developing brain. Our novel TRPM2 inhibitor, tatM2NX, did not confer protection against CA1 pyramidal cell death but attenuated synaptic plasticity (long-term plasticity (LTP)) deficits in both sexes. Further, in vivo administration of tatM2NX two weeks after CA/CPR reduced LTP impairments and restored memory function. These data provide evidence that pharmacological synaptic restoration of the surviving hippocampal network can occur independent of neuroprotection via inhibition of TRPM2 channels, providing a novel strategy to improve cognitive recovery in children following cerebral ischemia. Importantly, these data underscore the importance of age-appropriate models in disease research., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2021 Robert M. Dietz et al.)

Magnesium is an essential cofactor in many cellular processes, and aberrations in magnesium homeostasis can have life-threatening consequences. The kidney plays a central role in maintaining serum magnesium within a narrow range (0.70-1.10 mmol/L). Along the proximal tubule and thick ascending limb, magnesium reabsorption occurs via paracellular pathways. Members of the claudin family form the magnesium pores in these segments, and also regulate magnesium reabsorption by adjusting the transepithelial voltage that drives it. Along the distal convoluted tubule transcellular reabsorption via heteromeric TRPM6/7 channels predominates, although paracellular reabsorption may also occur. In this segment, the NaCl cotransporter plays a critical role in determining transcellular magnesium reabsorption. Although the general machinery involved in renal magnesium reabsorption has been identified by studying genetic forms of magnesium imbalance, the mechanisms regulating it are poorly understood. This review discusses pathways of renal magnesium reabsorption by different segments of the nephron, emphasizing newer findings that provide insight into regulatory process, and outlining critical unanswered questions., (Copyright © 2021 by the American Society of Nephrology.)

Transient receptor potential melastatin 4 (TRPM4) plays an important role in many tissues, including pacemaker and conductive tissues of the heart, but much less is known about its electrophysiological role in ventricular myocytes. Our earlier results showed the lack of selectivity of 9-phenanthrol, so CBA ((4-chloro-2-(2-chlorophenoxy)acetamido) benzoic acid) was chosen as a new, potentially selective inhibitor. Goal: Our aim was to elucidate the effect and selectivity of CBA in canine left ventricular cardiomyocytes and to study the expression of TRPM4 in the canine heart. Experiments were carried out in enzymatically isolated canine left ventricular cardiomyocytes. Ionic currents were recorded with an action potential (AP) voltage-clamp technique in whole-cell configuration at 37 °C. An amount of 10 mM BAPTA was used in the pipette solution to exclude the potential activation of TRPM4 channels. AP was recorded with conventional sharp microelectrodes. CBA was used in 10 µM concentrations. Expression of TRPM4 protein in the heart was studied by Western blot. TRPM4 protein was expressed in the wall of all four chambers of the canine heart as well as in samples prepared from isolated left ventricular cells. CBA induced an approximately 9% reduction in AP duration measured at 75% and 90% of repolarization and decreased the short-term variability of APD 90 . Moreover, AP amplitude was increased and the maximal rates of phase 0 and 1 were reduced by the drug. In AP clamp measurements, CBA-sensitive current contained a short, early outward and mainly a long, inward current. Transient outward potassium current (I to ) and late sodium current (I Na,L ) were reduced by approximately 20% and 47%, respectively, in the presence of CBA, while L-type calcium and inward rectifier potassium currents were not affected. These effects of CBA were largely reversible upon washout. Based on our results, the CBA induced reduction of phase-1 slope and the slight increase of AP amplitude could have been due to the inhibition of I to . The tendency for AP shortening can be explained by the inhibition of inward currents seen in AP-clamp recordings during the plateau phase. This inward current reduced by CBA is possibly I Na,L , therefore, CBA is not entirely selective for TRPM4 channels. As a consequence, similarly to 9-phenanthrol, it cannot be used to test the contribution of TRPM4 channels to cardiac electrophysiology in ventricular cells, or at least caution must be applied.

The cardiac Mg 2+ -sensitive, TRPM6, and TRPM7-like channels remain undefined, especially with the uncertainty regarding TRPM6 expression in cardiomyocytes. Additionally, their contribution to the cardiac action potential (AP) profile is unclear. Immunofluorescence assays showed the expression of the TRPM6 and TRPM7 proteins in isolated pig atrial and ventricular cardiomyocytes, of which the expression was modulated by incubation in extracellular divalent cation-free conditions. In patch clamp studies of cells dialyzed with solutions containing zero intracellular Mg 2+ concentration ([Mg 2+ ] i ) to activate the Mg 2+ -sensitive channels, raising extracellular [Mg 2+ ] ([Mg 2+ ] o ) from the 0.9-mM baseline to 7.2 mM prolonged the AP duration (APD). In contrast, no such effect was observed in cells dialyzed with physiological [Mg 2+ ] i . Under voltage clamp, in cells dialyzed with zero [Mg 2+ ] i , depolarizing ramps induced an outward-rectifying current, which was suppressed by raising [Mg 2+ ] o and was absent in cells dialyzed with physiological [Mg 2+ ] i . In cells dialyzed with physiological [Mg 2+ ] i , raising [Mg 2+ ] o decreased the L-type Ca 2+ current and the total delayed-rectifier current but had no effect on the APD. These results suggest a co-expression of the TRPM6 and TRPM7 proteins in cardiomyocytes, which are therefore the molecular candidates for the native cardiac Mg 2+ -sensitive channels, and also suggest that the cardiac Mg 2+ -sensitive current shortens the APD, with potential implications in arrhythmogenesis.

Cerebral ischemia-reperfusion (I-R) transiently increased autophagy by producing excessively reactive oxygen species (ROS); on the other hand, activated autophagy would remove ROS-damaged mitochondria and proteins, which led to cell survival. However, the regulation mechanism of autophagy activity during cerebral I-R is still unclear. In this study, we found that deficiency of the TRPM2 channel which is a ROS sensor significantly decreased I-R-induced neuronal damage. I-R transiently increased autophagy activity both in vitro and in vivo . More importantly, TRPM2 deficiency decreased I-R-induced neurological deficit score and infarct volume. Interestingly, our results indicated that TRPM2 deficiency could further activate AMPK rather than Beclin1 activity, suggesting that TRPM2 inhibits autophagy by regulating the AMPK/mTOR pathway in I-R. In conclusion, our study reveals that ROS-activated TRPM2 inhibits autophagy by downregulating the AMPK/mTOR pathway, which results in neuronal death induced by cerebral I-R, further supporting that TRPM2 might be a potential drug target for cerebral ischemic injury therapy., Competing Interests: The authors declare no conflict of interest., (Copyright © 2021 Xupang Hu et al.)

The transient receptor potential (TRP) melastatin-like subfamily member 2 (TRPM2) is a non-selective calcium-permeable cation channel. It is expressed by many mammalian tissues, including bone marrow, spleen, lungs, heart, liver, neutrophils, and endothelial cells. The best-known mechanism of TRPM2 activation is related to the binding of ADP-ribose to the nudix-box sequence motif (NUDT9-H) in the C-terminal domain of the channel. In cells, the production of ADP-ribose is a result of increased oxidative stress. In the context of endothelial function, TRPM2-dependent calcium influx seems to be particularly interesting as it participates in the regulation of barrier function, cell death, cell migration, and angiogenesis. Any impairments of these functions may result in endothelial dysfunction observed in such conditions as atherosclerosis or hypertension. Thus, TRPM2 seems to be an attractive therapeutic target for the conditions connected with the increased production of reactive oxygen species. However, before the application of TRPM2 inhibitors will be possible, some issues need to be resolved. The main issues are the lack of specificity, poor membrane permeabilization, and low stability in in vivo conditions. The article aims to summarize the latest findings on a role of TRPM2 in endothelial cells. We also show some future perspectives for the application of TRPM2 inhibitors in cardiovascular system diseases.

Ophiocordyceps sinensis (OCS), an entomopathogenic fungus, is known to exert antiproliferative and antitissue remodeling effects. Vascular remodeling and vasoconstriction play critical roles in the development of pulmonary hypertension (PH). The therapeutic potential of OCS for PH was investigated using rodent PH models, and cultured pulmonary artery endothelial and smooth muscle cells (PAECs and PASMCs), with a focus on the involvement of TRPM7. OCS ameliorated the development of PH, right ventricular hypertrophy and dysfunction in the monocrotaline-induced PH rats. The genetic knockout of TRPM7 attenuated the development of PH in mice with monocrotaline pyrrole-induced PH. TRPM7 was associated with medial hypertrophy and the plexiform lesions in rats and humans with PH. OCS suppressed proliferation of PASMCs derived from the PH patients. Ethanol extracts of OCS inhibited TRPM7-like current, TGF-β2-induced endothelial-mesenchymal transition, IL-6-induced STAT3 phosphorylation, and PDGF-induced Akt phosphorylation in PAECs or PASMCs. These inhibitory effects were recapitulated by either siRNA-mediated TRPM7 knockdown or treatment with TRPM7 antagonist FTY-720. OCS and FTY-720 induced vasorelaxation in the isolated normal human pulmonary artery. As a result, the present study proposes the therapeutic potential of OCS for the treatment of PH. The inhibition of TRPM7 is suggested to underlie the therapeutic effect of OCS., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Upon exposure to harmful microorganisms, hosts engage in protective molecular and behavioral immune responses, both of which are ultimately regulated by the nervous system. Using the nematode Caenorhabditis elegans , we show that ingestion of Enterococcus faecalis leads to a fast pathogen avoidance behavior that results in aversive learning. We have identified multiple sensory mechanisms involved in the regulation of avoidance of E. faecalis. The G-protein coupled receptor NPR-1-dependent oxygen-sensing pathway opposes this avoidance behavior, while an ASE neuron-dependent pathway and an AWB and AWC neuron-dependent pathway are directly required for avoidance. Colonization of the anterior part of the intestine by E. faecalis leads to AWB and AWC mediated olfactory aversive learning. Finally, two transient receptor potential melastatin (TRPM) channels, GON-2 and GTL-2, mediate this newly described rapid pathogen avoidance. These results suggest a mechanism by which TRPM channels may sense the intestinal distension caused by bacterial colonization to elicit pathogen avoidance and aversive learning by detecting changes in host physiology., Competing Interests: AF, JL, AA No competing interests declared, (© 2021, Filipowicz et al.)

Pyramidal neurons in the medial prefrontal cortical layer 2/3 are an essential contributor to the cellular basis of working memory; thus, changes in their intrinsic excitability critically affect medial prefrontal cortex (mPFC) functional properties. Transient Receptor Potential Melastatin 4 (TRPM4), a calcium-activated nonselective cation channel (CAN), regulates the membrane potential in a calcium-dependent manner. In this study, we uncovered the role of TRPM4 in regulating the intrinsic excitability plasticity of pyramidal neurons in the mouse mPFC layer of 2/3 using a combination of conventional and nystatin perforated whole-cell recordings. Interestingly, we found that TRPM4 is open at resting membrane potential, and its inhibition increases input resistance and hyperpolarizes membrane potential. After high-frequency stimulation, pyramidal neurons increase a calcium-activated non-selective cation current, increase the action potential firing, and the amplitude of the afterdepolarization, these effects depend on intracellular calcium. Furthermore, pharmacological inhibition or genetic silencing of TRPM4 reduces the firing rate and the afterdepolarization after high frequency stimulation. Together, these results show that TRPM4 plays a significant role in the excitability of mPFC layer 2/3 pyramidal neurons by modulating neuronal excitability in a calcium-dependent manner.

Neutrophils must navigate accurately towards pathogens in order to destroy invaders and thus defend our bodies against infection. Here we show that hydrogen peroxide, a potent neutrophil chemoattractant, guides chemotaxis by activating calcium-permeable TRPM2 ion channels and generating an intracellular leading-edge calcium "pulse". The thermal sensitivity of TRPM2 activation means that chemotaxis towards hydrogen peroxide is strongly promoted by small temperature elevations, suggesting that an important function of fever may be to enhance neutrophil chemotaxis by facilitating calcium influx through TRPM2. Chemotaxis towards conventional chemoattractants such as LPS, CXCL2 and C5a does not depend on TRPM2 but is driven in a similar way by leading-edge calcium pulses. Other proposed initiators of neutrophil movement, such as PI3K, Rac and lyn, influence chemotaxis by modulating the amplitude of calcium pulses. We propose that intracellular leading-edge calcium pulses are universal drivers of the motile machinery involved in neutrophil chemotaxis.

Resistance to targeted therapy and immunotherapy remains a major obstacle in improving care for patients with advanced melanoma. MicroRNAs play important roles in regulating gene networks involved in disease progression and resistance to therapy in cancers such as melanoma. MicroRNA miR-211 contributes to melanocyte and melanoma biology and has been implicated in targeted therapy resistance. Lee et al. (2020) report a novel mechanism by which miR-211 promotes resistance to BRAF V600E inhibitor therapy via the upregulation of the extracellular signal-regulated kinase 5 signaling pathway., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: To explore the role of transient receptor potential melastatin 8 (TRPM8) in the occurrence and development of bladder pain in interstitial cystitis/bladder pain syndrome (IC/BPS) patients. The differences in the content and location distribution of TRPM8 in bladder were compared between IC/BPS and control group., Methods: All enrolled patients answered questionnaire such as O'leary-Sant symptom index, visual analog scale (VAS), quality of life (QOL), and pelvic pain and urinary urgency frequency (PUF) score, then bladder specimens were collected. Analyses such as qRT-PCR, western blot, and immunofluorescence were performed to determine the changes in TRPM8 content and expression in neurons and sensory nerves between the IC/BPS and control group, and the relationships between TRPM8 and various clinical scores were also analyzed., Results: There were significant differences in the O'leary-Sant score, PUF score, VAS, and QOL score between IC/BPS and the control group (P < .05). Compared with the control group, the expression levels of TRPM8 mRNA and protein were significantly increased in the IC/BPS bladder tissues (P < .01). Immunofluorescence examination also revealed that (1) the number of neurons and sensory nerves displayed a significant upward trend in the bladder tissue of IC/BPS patients (2) the expression levels of TRPM8 on neurons and sensory nerves also increased significantly in IC/BPS group., Conclusion: In IC/BPS patients, TRPM8 content increased significantly and mainly expressed on increased neurons and sensory nerves in bladder tissue. These results may indicate a mechanism by which bladder pain is more easily to spread in IC/BPS patients, and may also indicate an important mechanism for pain sensitization in such patients., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The transient receptor potential melastatin-subfamily member 7 (TRPM7) is a ubiquitously expressed protein that contains both an ion channel and an active kinase. TRPM7 has involved in a variety of cellular functions and critically participates in various diseases mainly including cancer and neurodegenerative disorders. However, the theme trends and knowledge structures for TRPM7 have not yet been studied bibliometrically. The main purposes of this research are to compare the scientific production in the research field of TRPM7 among countries and to evaluate the publication trend between 2004 and 2019. All publications were extracted from the Web of Science Core Collection (WoSCC) database from 2004 to 2019. Microsoft Excel 2018, Prism 6, and CiteSpace V were applied to analyze the scientific research outputs including journals, countries, territories, institutions, authors, and research hotspots. In this report, a total of 860 publications related to TRPM7 were analyzed. Biophysical Journal ranked top for publishing 31 papers. The United States of America had the largest number of publications (320) with a high citation frequency (11,298) and H-index (58). Chubanov V (38 publications) and Gudermann T (38 citations), who from Ludwig Maximilian University of Munich, were the most productive authors and had the greatest co-citation counts. Our study also combined the bibliometric study with a systematic review on TRPM7, highlighting the four research frontiers of TRPM7. This is the first study that demonstrated the trends and future development in TRPM7 publications, providing a clear and intuitive profile for the contributions in this field.

Transient receptor potential melastatin 2 (TRPM2) is a Ca 2+ - permeable nonselective cation channel that is involved in diverse biological functions as a cellular sensor for oxidative stress and temperature. It has been considered a promising therapeutic target for the treatment of ischemia/reperfusion (IR) injury, inflammation, cancer, and neurodegenerative diseases. Development of highly potent and selective TRPM2 inhibitors and validation of their use in relevant disease models will advance drug discovery. In this review, we describe the molecular structures and gating mechanism of the TRPM2 channel, and offer a comprehensive review of advances in the discovery of TRPM2 inhibitors. Furthermore, we analyze the properties of reported TRPM2 inhibitors with an emphasis on how specific inhibitors targeting this channel could be better developed., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Aims: Vascular calcification is a common complication in patients with chronic kidney disease and associated with increased morbidity and mortality. The role of TRPM7 in vascular smooth muscle cell (VSMC) transformation during vascular calcification is not clear. We aim to investigate the effects of phosphate and indoxyl sulphate on the expression of TRPM7 and calcification-related molecules in VSMC., Main Methods: Human aortic smooth muscle cells (HASMC) were treated with phosphate (3.3 mM) or indoxyl sulphate (500 μM and 1000 μM). 2-APB, a channel blocker of TRPM7 was added simultaneously in blocking experiment. Cells were then examined grossly and alizarin red solution was employed for calcification assessment. Lastly, cells were harvested for gene expression and protein abundance analysis., Key Findings: Phosphate treatment induced significant increase in BMP2, RUNX2, BMP7, vitamin D receptor (VDR), calcium sensing receptor (CaSR) and TRPM7, but 1-alpha hydroxylase, klotho, DKK1 and sclerostin were not changed. The addition of 2-APB prevented increase of BMP2, RUNX2, BMP7, VDR, CaSR and TRPM7. Indoxyl sulphate treatment was associated with decrease in TRPM7 and DKK1, but increase in RUNX2, BMP2 and VDR were noted. There were no significant alterations in BMP7, CaSR, klotho,1-alpha hydroxylase and sclerostin. Co-treatment with 2-APB reversed the increase in VDR., Significance: Both phosphate and indoxyl sulphate induced calcification in VSMC but it was more prominent in phosphate. TRPM7 was upregulated by phosphate but downregulated in indoxyl sulphate treatment. Vascular calcification was reduced by blocking TRPM7 with 2-APB and there was partial anti-calcification effect in indoxyl sulphate., (Copyright © 2020. Published by Elsevier Inc.)

Background: Calcium is an essential signal transduction element that has been associated with aggressive behaviours in several cancers. Cell motility is a prerequisite for metastasis, the major cause of lung cancer death, yet its association with calcium signalling and underlying regulatory axis remains an unexplored area., Methods: Bioinformatics database analyses were employed to assess correlations between calcium influx channels and clinical outcomes in non-small cell lung cancer (NSCLC). Functional and regulatory roles of influx channels in cell migration and invasion were conducted and experimental lung metastasis was examined using in vivo live imaging., Results: High expression of TRPM7 channel correlates well with the low survival rate of patients and high metastatic potential. Inhibition of TRPM7 suppresses cell motility in various NSCLC cell lines and patient-derived primary cells and attenuates experimental lung metastases. Mechanistically, TRPM7 acts upstream of O-GlcNAcylation, a post-translational modification and a crucial sensor for metabolic changes. We reveal for the first time that caveolin-1 and c-Myc are favourable molecular targets of TRPM7/O-GlcNAc that regulates NSCLC motility. O-GlcNAcylation of caveolin-1 and c-Myc promotes protein stability by interfering with their ubiquitination and proteasomal degradation., Conclusions: TRPM7/O-GlcNAc axis represents a potential novel target for lung cancer therapy that may overcome metastasis.

The importance of the intracellular Ca 2+ concentration ([Ca 2+ ] i ) in neutrophil function has been intensely studied. However, the role of the intracellular Na + concentration ([Na + ] i ) which is closely linked to the intracellular Ca 2+ regulation has been largely overlooked. The [Na + ] i is regulated by Na + transport proteins such as the Na + /Ca 2+ -exchanger (NCX1), Na + /K + -ATPase, and Na + -permeable, transient receptor potential melastatin 2 (TRPM2) channel. Stimulating with either N-formylmethionine-leucyl-phenylalanine (fMLF) or complement protein C5a causes distinct changes of the [Na + ] i . fMLF induces a sustained increase of [Na + ] i , surprisingly, reaching higher values in TRPM2 -/- neutrophils. This outcome is unexpected and remains unexplained. In both genotypes, C5a elicits only a transient rise of the [Na + ] i . The difference in [Na + ] i measured at t = 10 min after stimulation is inversely related to neutrophil chemotaxis. Neutrophil chemotaxis is more efficient in C5a than in an fMLF gradient. Moreover, lowering the extracellular Na + concentration from 140 to 72 mM improves chemotaxis of WT but not of TRPM2 -/- neutrophils. Increasing the [Na + ] i by inhibiting the Na + /K + -ATPase results in disrupted chemotaxis. This is most likely due to the impact of the altered Na + homeostasis and presumably NCX1 function whose expression was shown by means of qPCR and which critically relies on proper extra- to intracellular Na + concentration gradients. Increasing the [Na + ] i by a few mmol/l may suffice to switch its transport mode from forward (Ca 2+ -efflux) to reverse (Ca 2+ -influx) mode. The role of NCX1 in neutrophil chemotaxis is corroborated by its blocker, which also causes a complete inhibition of chemotaxis., (Copyright © 2020 Najder, Rugi, Lebel, Schröder, Oster, Schimmelpfennig, Sargin, Pethő, Bulk and Schwab.)

Oxidative stress-induced Ca 2+ permeable transient receptor potential melastatin 2 (TRPM2) channels are expressed at high levels in the brain, appear to link neuronal excitability to cellular metabolism, and are involved in the pathogenesis of neurodegenerative disorders. We aimed to study the electrophysiological properties of TRPM2 channels in stellate cells of the mouse ventral cochlear nucleus (VCN) using molecular, immunohistochemical and electrophysiological approaches. In the present study, the real time PCR analysis revealed the presence of the TRPM2 mRNA in the mouse VCN tissue. Cell bodies of stellate cells were moderately labeled with TRPM2 antibodies using immunohistochemical staining. Stellate cells were sensitive to intracellular ADP-ribose (ADPR), a TRPM2 agonist. Upon the application of ADPR, the resting membrane potential of the stellate cells was significantly depolarized, shifting from -61.2 ± 0.9 mV to -57.0 ± 0.8 mV (P < 0.001; n = 21), and the firing rate significantly increased (P < 0.001, n = 6). When the pipette solution contained ADPR (300 μM) and the TRPM2 antagonists flufenamic acid (FFA) (100 μM), N-(p-amylcinnamoyl) anthranilic acid (ACA) (50 μM) and 8-bromo-cADP-Ribose (8-Br-cADPR) (50 μM), the membrane potential shifted in a hyperpolarizing direction. ADPR did not significantly change the resting membrane potential and action potential firing rate of stellate cells from TRPM2-/- mice. In conclusion, the results obtained using these molecular, immunohistochemical and electrophysiological approaches reveal the expression of functional TRPM2 channels in stellate neurons of the mouse VCN. TRPM2 might exert a significant modulatory effect on setting the level of resting excitability., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Ion channels are important therapeutic targets due to their plethoric involvement in physiological and pathological consequences. The transient receptor potential cation channel subfamily M member 8 (TRPM8) is a nonselective cation channel that controls Ca 2+ homeostasis. It has been proposed to be the predominant thermoreceptor for cellular and behavioral responses to cold stimuli in the transient receptor potential (TRP) channel subfamilies and exploited so far to reach the clinical-stage of drug development. TRPM8 channels can be found in multiple organs and tissues, regulating several important processes such as cell proliferation, migration and apoptosis, inflammatory reactions, immunomodulatory effects, pain, and vascular muscle tension. The related disorders have been expanded to new fields ranging from cancer and migraine to dry eye disease, pruritus, irritable bowel syndrome (IBS), and chronic cough. This review is aimed to summarize the distribution of TRPM8 and disorders related to it from a clinical perspective, so as to broaden the scope of knowledge of researchers to conduct more studies on this subject., (Copyright © 2020 Elsevier B.V. All rights reserved.)

When in a particular scientific field, major progress is rapidly reached after a long period of relative stand-still, this is often achieved by the development or exploitation of new techniques and methods. A striking example is the new insights brought into the understanding of the gating mechanism of the transient receptor potential melastatin type 2 cation channel (TRPM2) by cryogenic electron microscopy structure analysis. When conventional methods are complemented by new ones, it is quite natural that established researchers are not fully familiar with the possibilities and limitations of the new method. On the other hand, newcomers may need some assistance in perceiving the previous knowledge in detail; they may not realize that some of their interpretations are at odds with previous results and need refinement. This may in turn trigger further studies with new and promising perspectives, combining the promises of several methodological approaches. With this review, I aim to give a comprehensive overview on functional data of several orthologous of TRPM2 that are nicely explained by structural studies. Moreover, I wish to point out some functional contradictions raised by the structural data. Finally, some open questions and some lines of possible future experimental approaches shall be discussed.

Chronic cerebral hypoperfusion (CCH) been well characterized as a common pathological status contributing to neurodegenerative diseases such as Alzheimer's disease and vascular dementia. CCH is an important factor that leads to cognitive impairment, but the underlying neurobiological mechanism is poorly understood and no effective treatment is available. Recently, transient receptor potential melastatin 4 (TRPM4) cation channel has been identified as an important molecular element in focal cerebral ischemia. Over activation of the channel is a major molecular mechanism of oncotic cell death. However, the role of TRPM4 in CCH that propagates global brain hypoxia have not been explored. Therefore, the present study is designed to investigate the effect of TRPM4 inhibition on the cognitive functions of the rats following CCH via permanent bilateral occlusion of common carotid arteries (PBOCCA) model. In this model, treatment with siRNA suppressed TRPM4 expression at both the mRNA and protein levels and improved cognitive deficits of the CCH rats without affecting their motor function. Furthermore, treatment with siRNA rescued the LTP impairment in CCH-induced rats. Consistent with the restored of LTP, western blot analysis revealed that siRNA treatment prevented the reduction of synaptic proteins, including calcium/calmodulin-dependent kinase II alpha (CaMKIIα) and brain-derived neurotrophic factor (BDNF) in brain regions of CCH rats. The present findings provide a novel role of TRPM4 in restricting cognitive functions in CCH and suggest inhibiting TRPM4 may represent a promising therapeutic strategy in targeting ion channels to prevent the progression of cognitive deficits induced by ischemia., Competing Interests: Declaration of Competing Interest The authors declare no potential conflicts of interest with respect to the research, authorship, and/or publication of this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

A common theme in organogenesis is how the final structure of organs emerge from epithelial tube structures, with the formation of the neural tube being one of the best examples. Two types of cell movements co-occur during neural tube closure involving the migration of cells toward the midline of the embryo (mediolateral intercalation or convergent extension) as well as the deep movement of cells from inside the embryo to the outside of the lateral side of the neural plate (radial intercalation). Failure of either type of cell movement will prevent neural tube closure, which can produce a range of neural tube defects (NTDs), a common congenital disease in humans. Numerous studies have identified signaling pathways that regulate mediolateral intercalation during neural tube closure. Less understood are the pathways that govern radial intercalation. Using the Xenopus laevis system, our group reported the identification of transient receptor potential (TRP) channels, TRPM6 and TRPM7, and the Mg 2+ ion they conduct, as novel and key factors regulating both mediolateral and radial intercalation during neural tube closure. Here we broadly discuss tubulogenesis and cell intercalation from the perspective of neural tube closure and the respective roles of TRPM7 and TRPM6 in this critical embryonic process., (© 2020 Wiley Periodicals, Inc.)

Transient receptor potential (TRP) melastatin 8 (TRPM8) is a temperature-sensing ion channel mainly expressed in primary sensory neurons (A δ -fibers and C-fibers in the dorsal root ganglion). In this report, we characterized KPR-5714 ( N -[( R )-3,3-difluoro-4-hydroxy-1-(2 H -1,2,3-triazol-2-yl)butan-2-yl]-3-fluoro-2-[5-(4-fluorophenyl)-1 H -pyrazol-3-yl]benzamide), a novel and selective TRPM8 antagonist, to assess its therapeutic potential against frequent urination in rat models with overactive bladder (OAB). In calcium influx assays with HEK293T cells transiently expressing various TRP channels, KPR-5714 showed a potent TRPM8 antagonistic effect and high selectivity against other TRP channels. Intravenously administered KPR-5714 inhibited the hyperactivity of mechanosensitive C-fibers of bladder afferents and dose-dependently increased the intercontraction interval shortened by intravesical instillation of acetic acid in anesthetized rats. Furthermore, we examined the effects of KPR-5714 on voiding behavior in conscious rats with cerebral infarction and in those exposed to cold in metabolic cage experiments. Cerebral infarction and cold exposure induced a significant decrease in the mean voided volume and increase in voiding frequency in rats. Orally administered KPR-5714 dose-dependently increased the mean voided volume and decreased voiding frequency without affecting total voided volume in these models. This study demonstrates that KPR-5714 improves OAB in three different models by inhibiting exaggerated activity of mechanosensitive bladder C-fibers and suggests that KPR-5714 may provide a new and useful approach to the treatment of OAB. SIGNIFICANCE STATEMENT: TRPM8 is involved in bladder sensory transduction and plays a role in the abnormal activation in hypersensitive bladder disorders. KPR-5714, as a novel and selective TRPM8 antagonist, may provide a useful treatment for the disorders related to the hyperactivity of bladder afferent nerves, particularly in overactive bladder., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Hippocampal injury and cognitive impairments are common after cardiac arrest and stroke and do not have an effective intervention despite much effort. Therefore, we developed a new approach aimed at reversing synaptic dysfunction by targeting TRPM2 channels. Cardiac arrest/cardiopulmonary resuscitation (CA/CPR) in mice was used to investigate cognitive deficits and the role of the calcium-permeable ion channel transient receptor potential-M2 (TRPM2) in ischemia-induced synaptic dysfunction. Our data indicates that absence (TRPM2 -/- ) or acute inhibition of TRPM2 channels with tatM2NX reduced hippocampal cell death in males only, but prevented synaptic plasticity deficits in both sexes. Remarkably, administration of tatM2NX weeks after injury reversed hippocampal plasticity and memory deficits. Finally, TRPM2-dependent activation of calcineurin-GSK3β pathway contributes to synaptic plasticity impairments. These data suggest persistent TRPM2 activity following ischemia contributes to impairments of the surviving hippocampal network and that inhibition of TRPM2 channels at chronic time points may represent a novel strategy to improve functional recovery following cerebral ischemia that is independent of neuroprotection.

Aging is a crucial cause of cognitive decline and a major risk factor for Alzheimer's disease (AD); however, AD's underlying molecular mechanisms remain unclear. Recently, tetraspanins have emerged as important modulators of synaptic function and memory. We demonstrate that the level of tetraspanin CD82 is upregulated in the brains of AD patients and middle-aged mice. In young adult mice, injection of AAV-CD82 to the hippocampus induced AD-like cognitive deficits and impairments in neuronal spine density. CD82 overexpression increased TRPM7 α-kinase cleavage via caspase-3 activation and induced Numb phosphorylation at Thr346 and Ser348 residues. CD82 overexpression promoted beta-amyloid peptide (Aβ) secretion which could be reversed by Numb T346S348 mutants. Importantly, hippocampus-related memory functions were improved in Cd82 -/- mice. Taken together, our findings provide the evidence that links the elevated CD82-TRPM7-Numb signaling to age-related cognitive impairment.

During infection, phagocytic cells pursue homeostasis in the host via multiple mechanisms that control microbial invasion. Neutrophils respond to infection by exerting a variety of cellular processes, including chemotaxis, activation, phagocytosis, degranulation and the generation of reactive oxygen species (ROS). Calcium (Ca 2+ ) signaling and the activation of specific Ca 2+ channels are required for most antimicrobial effector functions of neutrophils. The transient receptor potential melastatin-2 (TRPM2) cation channel has been proposed to play important roles in modulating Ca 2+ mobilization and oxidative stress in neutrophils. In the present study, we use a mouse model of Listeria monocytogenes infection to define the role of TRPM2 in the regulation of neutrophils' functions during infection. We show that the susceptibility of Trpm2 -/- mice to L. monocytogenes infection is characterized by increased migration rates of neutrophils and monocytes to the liver and spleen in the first 24 h. During the acute phase of L. monocytogenes infection, Trpm2 -/- mice developed septic shock, characterized by increased serum levels of TNF-α, IL-6, and IL-10. Furthermore, in vivo depletion of neutrophils demonstrated a critical role of these immune cells in regulating acute inflammation in Trpm2 -/- infected mice. Gene expression and inflammatory cytokine analyses of infected tissues further confirmed the hyperinflammatory profile of Trpm2 -/- neutrophils. Finally, the increased inflammatory properties of Trpm2 -/- neutrophils correlated with the dysregulated cytoplasmic concentration of Ca 2+ and potentiated membrane depolarization, in response to L. monocytogenes . In conclusion, our findings suggest that the TRPM2 channel plays critical functional roles in regulating the inflammatory properties of neutrophils and preventing tissue damage during Listeria infection., (Copyright © 2020 Robledo-Avila, Ruiz-Rosado, Brockman and Partida-Sánchez.)

Epilepsy is one of the most common neurological conditions. Recent findings suggest that one of the mechanisms promoting its existence is calcium influx. The transient receptor potential melastatin type 2 channel (TRPM2) is a Ca 2+ -permeable cation channel that contributes to cell apoptosis; its possible signaling pathway is the PARP1/BNIP3/AIF/Endo G pathway that may be related to epilepsy. The aim of this study was to investigate the TRPM2 channel's involvement in epilepsy and how it works. We also explored the possible role of the TRPM2 channel on cognitive ability and emotion in epilepsy. To accomplish our goals, we used different animal epilepsy models to study the effect of the TRPM2 channel on epilepsy. The results showed that the knockout (KO) of the TRPM2 gene might play a protective role in epilepsy. Considering the advantages attributed to pentylenetetrazole (PTZ)-induced kindling mouse model, we used the model for the following assessments: 1. to observe changes in cognition and anxiety between wild type (WT) mice and TRPM2-KO mice with the recognition of new things trial and elevated plus-maze; 2. to determine the expression of apoptosis-associated proteins (PARP1, BNIP3, AIF, and Endo G) using Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot; 3. to observe neurons pathologic damages and astrocyte activation in each group. The main findings of our study were: (a) TRPM2-KO had a protective effect on epilepsy; (b) TRPM2-KO improved spatial memory deficits overtime during epilepsy, but it did not improve anxiety; (c) the protective effect probably occurred via the PARP1 downstream signaling pathway; (d) TRPM2-KO could ameliorate epilepsy-induced hippocampal pathological damages and weaken astrocyte activation. These findings may provide a new approach for the treatment of epilepsy and early intervention., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Hyperosmolar sweet foods onto exposed tooth dentin evoke sudden and intense dental pain, called dentin hypersensitivity . However, it remains unclear how hyperosmolar stimuli excite dental primary afferent (DPA) neurons and thereby lead to dentin hypersensitivity. This study elucidated whether TRPM8, which is well known as a cold temperature- or menthol-activated receptor, additionally mediates nociception in response to hyperosmolar stimuli in adult mouse DPA neurons, which are identified by a fluorescent retrograde tracer: DiI. Single-cell reverse transcription polymerase chain reaction revealed that TRPM8 was expressed in subsets of DPA neurons and that TRPM8 was highly colocalized with TRPV1 and Piezo2. Immunohistochemical analysis also confirmed TRPM8 expression in DPA neurons. By using Fura-2-based calcium imaging, application of hyperosmolar sucrose solutions elicited calcium transients in subsets of the trigeminal ganglion neurons, which was significantly abolished by a selective TRPM8 antagonist: N-(3-Aminopropyl)-2-[(3-methylphenyl)methoxy]-N-(2-thienylmethyl)benzamide (AMTB) hydrochloride. When we further examined changes of c-fos expression (a neuronal activation marker) in the spinal trigeminal nucleus after hyperosmolar stimulation onto exposed tooth dentin, c-fos mRNA and protein expression were increased and were also significantly reduced by AMTB, especially in the spinal trigeminal interpolaris-caudalis transition zone (Vi/Vc). Taken together, our results provide strong evidence that TRPM8 expressed in DPA neurons might mediate dental pain as a hyperosmosensor in adult mice.

For protection from inhaled pathogens many strategies have evolved in the airways such as mucociliary clearance and cough. We have previously shown that protective respiratory reflexes to locally released bacterial bitter "taste" substances are most probably initiated by tracheal brush cells (BC). Our single-cell RNA-seq analysis of murine BC revealed high expression levels of cholinergic and bitter taste signaling transcripts (Tas2r108, Gnat3, Trpm5). We directly demonstrate the secretion of acetylcholine (ACh) from BC upon stimulation with the Tas2R agonist denatonium. Inhibition of the taste transduction cascade abolished the increase in [Ca 2+ ] i in BC and subsequent ACh-release. ACh-release is regulated in an autocrine manner. While the muscarinic ACh-receptors M3R and M1R are activating, M2R is inhibitory. Paracrine effects of ACh released in response to denatonium included increased [Ca 2+ ] i in ciliated cells. Stimulation by denatonium or with Pseudomonas quinolone signaling molecules led to an increase in mucociliary clearance in explanted tracheae that was Trpm5- and M3R-mediated. We show that ACh-release from BC via the bitter taste cascade leads to immediate paracrine protective responses that can be boosted in an autocrine manner. This mechanism represents the initial step for the activation of innate immune responses against pathogens in the airways., (© 2019 The Authors. The FASEB Journal published by Wiley Periodicals, Inc. on behalf of Federation of American Societies for Experimental Biology.)

As a crucial signaling molecule, calcium plays a critical role in many physiological and pathological processes by regulating ion channel activity. Recently, one study resolved the structure of the transient receptor potential melastatin 2 (TRPM2) channel from Nematostella vectensis (nvTRPM2). This identified a calcium-binding site in the S2-S3 loop, while its effect on channel gating remains unclear. Here, we investigated the role of this calcium-binding site in both nvTRPM2 and human TRPM2 (hTRPM2) by mutagenesis and patch-clamp recording. Unlike hTRPM2, nvTRPM2 cannot be activated by calcium alone. Moreover, the inactivation rate of nvTRPM2 was decreased as intracellular calcium concentration was increased. In addition, our results showed that the four key residues in the calcium-binding site of S2-S3 loop have similar effects on the gating processes of nvTRPM2 and hTRPM2. Among them, the mutations at negatively charged residues (glutamate and aspartate) substantially decreased the currents of nvTRPM2 and hTRPM2. This suggests that these sites are essential for calcium-dependent channel gating. For the charge-neutralizing residues (glutamine and asparagine) in the calcium-binding site, our data showed that glutamine mutating to alanine or glutamate did not affect the channel activity, but glutamine mutating to lysine caused loss of function. Asparagine mutating to aspartate still remained functional, while asparagine mutating to alanine or lysine led to little channel activity. These results suggest that the side chain of glutamine has a less contribution to channel gating than does asparagine. However, our data indicated that both glutamine mutating to alanine or glutamate and asparagine mutating to aspartate accelerated the channel inactivation rate, suggesting that the calcium-binding site in the S2-S3 loop is important for calcium-dependent channel inactivation. Taken together, our results uncovered the effect of four key residues in the S2-S3 loop of TRPM2 on the TRPM2 gating process.

The cornea is extensively innervated by trigeminal ganglion cold thermoreceptor neurons expressing TRPM8 (transient receptor potential cation channel subfamily M member 8). These neurons respond to cooling, hyperosmolarity and wetness of the corneal surface. Surgical injury of corneal nerve fibers alters tear production and often causes dry eye sensation. The contribution of TRPM8-expressing corneal cold-sensitive neurons (CCSNs) to these symptoms is unclear. Using extracellular recording of CCSNs nerve terminals combined with in vivo confocal tracking of reinnervation, Ca 2+ imaging and patch-clamp recordings of fluorescent retrogradely labeled corneal neurons in culture, we analyzed the functional modifications of CCSNs induced by peripheral axonal damage in male mice. After injury, the percentage of CCSNs, the cold- and menthol-evoked intracellular [Ca 2+ ] rises and the TRPM8 current density in CCSNs were larger than in sham animals, with no differences in the brake K + current I KD Active and passive membrane properties of CCSNs from both groups were alike and corresponded mainly to those of canonical low- and high-threshold cold thermoreceptor neurons. Ongoing firing activity and menthol sensitivity were higher in CCSN terminals of injured mice, an observation accounted for by mathematical modeling. These functional changes developed in parallel with a partial reinnervation of the cornea by TRPM8(+) fibers and with an increase in basal tearing in injured animals compared with sham mice. Our results unveil key TRPM8-dependent functional changes in CCSNs in response to injury, suggesting that increased tearing rate and ocular dryness sensation derived from deep surgical ablation of corneal nerves are due to enhanced functional expression of TRPM8 channels in these injured trigeminal primary sensory neurons. SIGNIFICANCE STATEMENT We unveil a key role of TRPM8 channels in the sensory and autonomic disturbances associated with surgical damage of eye surface nerves. We studied the damage-induced functional alterations of corneal cold-sensitive neurons using confocal tracking of reinnervation, extracellular corneal nerve terminal recordings, tearing measurements in vivo , Ca 2+ imaging and patch-clamp recordings of cultured corneal neurons, and mathematical modeling. Corneal nerve ablation upregulates TRPM8 mainly in canonical cold thermoreceptors, enhancing their cold and menthol sensitivity, inducing a rise in the ongoing firing activity of TRPM8(+) nerve endings and an increase in basal tearing. Our results suggest that unpleasant dryness sensations, together with augmented tearing rate after corneal nerve injury, are largely due to upregulation of TRPM8 in cold thermoreceptor neurons., (Copyright © 2019 the authors.)

Transient receptor potential melastatin 3 (TRPM3) is a nonselective cation channel that is inhibited by Gβγ subunits liberated following activation of Gα i/o protein-coupled receptors. Here, we demonstrate that TRPM3 channels are also inhibited by Gβγ released from Gα s and Gα q Activation of the G s -coupled adenosine 2B receptor and the G q -coupled muscarinic acetylcholine M1 receptor inhibited the activity of TRPM3 heterologously expressed in HEK293 cells. This inhibition was prevented when the Gβγ sink βARK1-ct (C terminus of β-adrenergic receptor kinase-1) was coexpressed with TRPM3. In neurons isolated from mouse dorsal root ganglion (DRG), native TRPM3 channels were inhibited by activating G s -coupled prostaglandin-EP2 and G q -coupled bradykinin B2 (BK2) receptors. The G i/o inhibitor pertussis toxin and inhibitors of PKA and PKC had no effect on EP2- and BK2-mediated inhibition of TRPM3, demonstrating that the receptors did not act through Gα i/o or through the major protein kinases activated downstream of G-protein-coupled receptor (GPCR) activation. When DRG neurons were dialyzed with GRK2i, which sequesters free Gβγ protein, TRPM3 inhibition by EP2 and BK2 was significantly reduced. Intraplantar injections of EP2 or BK2 agonists inhibited both the nocifensive response evoked by TRPM3 agonists, and the heat hypersensitivity produced by Freund's Complete Adjuvant (FCA). Furthermore, FCA-induced heat hypersensitivity was completely reversed by the selective TRPM3 antagonist ononetin in WT mice and did not develop in Trpm3 -/- mice. Our results demonstrate that TRPM3 is subject to promiscuous inhibition by Gβγ protein in heterologous expression systems, primary neurons and in vivo , and suggest a critical role for this ion channel in inflammatory heat hypersensitivity. SIGNIFICANCE STATEMENT The ion channel TRPM3 is widely expressed in the nervous system. Recent studies showed that Gα i/o -coupled GPCRs inhibit TRPM3 through a direct interaction between Gβγ subunits and TRPM3. Since Gβγ proteins can be liberated from other Gα subunits than Gα i/o , we examined whether activation of G s - and G q -coupled receptors also influence TRPM3 via Gβγ. Our results demonstrate that activation of G s - and G q -coupled GPCRs in recombinant cells and sensory neurons inhibits TRPM3 via Gβγ liberation. We also demonstrated that G s - and G q -coupled receptors inhibit TRPM3 in vivo , thereby reducing pain produced by activation of TRPM3, and inflammatory heat hypersensitivity. Our results identify Gβγ inhibition of TRPM3 as an effector mechanism shared by the major Gα subunits., (Copyright © 2019 the authors.)

Introduction: Erectile dysfunction (ED) is frequently encountered in patients with arterial hypertension and there is a recent functional correlation between the expression of thermoreceptor channels TRPM8 (melastatin 8) and alterations in blood pressure in hypertension. The aim of this study was to investigate the function of cold-sensing TRPM8 channel in internal pudendal artery (IPA) in both normotensive and hypertensive rats., Methods: We performed experiments integrating physiological, pharmacological, biochemical and cellular techniques., Results: TRPM8 channels are expressed in the IPA and in vascular smooth muscle cells from IPA. In addition, TRPM8 activation, by both a cooling compound icilin (82.1 ± 3.0%, n = 6) and cold temperature [thermal stimulus, basal tone (25 °C, 41.2 ± 3.4%, n = 5) or pre-contracted tone induced by phenylephrine (25 °C, 87.0 ± 3.6%, n = 7)], induced relaxation in IPA. Furthermore, the results showed that the concentration-response curve to icilin was significantly shifted to the right in different conditions, such as: the absence of the vascular endothelium, in the presence of L-NAME (10 -4 M), or indomethacin (10 -5 M) or by a combination of charybdotoxin (10 -7 M) and apamin (5 × 10 -6 M), and Y27632 (10 -6 M). Interestingly, icilin-induced vasodilation was significantly higher in IPA from spontaneously hypertensive (SHR, E 10 -4 M = 75.3 ± 1.7%) compared to wistar rats (E 10 -4 M = 56.4 ± 2.6%), despite no changes in the TRPM8 expression in IPA between the strains, suggesting that the sensitivity of TRPM8 channels is higher in SHR., Conclusions: These data demonstrate for the first time, the expression and function of TRPM8 channels in the IPA involving, at least in part, endothelium-derived relaxing factors and ROCK inhibition. Overall, this channel could potentially be a new target for the treatment of hypertension associated-ED., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

There was no clear evidence of the TRPA1 ion channel involvement in the formation of thermoregulatory responses. The present results convincingly show that the skin TRPA1 ion channel activation has significant influence on the formation of thermoregulatory responses of the body to cooling; it is especially strongly manifested for the metabolic component. At the TRPA1 activation by its agonist AITC (0.04%), an enhancement in thermoregulatory responses is observed: the temperature thresholds for the first phase and the second one of the metabolic response decrease, the values of all components of the metabolic response considerably increase: the increment of oxygen consumption in the first phase increases from 1.8 ± 0.24 in the control to 2.9 ± 0.35 ml/min*kg under AITC, P = 0.04; the increment of oxygen consumption in the second phase increases from 6.2 ± 2.06 to 17.4 ± 1.20 ml/min*kg, P = 0.002, as well as shivering rises from 7.8 ± 1.79 to 15.4 ± 1.87 mV, P = 0.011. In consideration of our previous results on the influence of TRPM8 ion channel activation on thermoregulatory responses (Kozyreva et al., J. Therm.Biol., 2010) it is obvious that the TRPM8 and TRPA1 ion channels have a pronounced, but unequal effects on the values of different phases of the metabolic response to cold. The TRPM8 activation manifests itself in an increase of value only the urgent first phase, this phase is associated with carbohydrate metabolism. As the recent results have shown the influence of the TRPA1 activation is realized predominantly in the clearly marked increase in the second phase of the metabolic response associated with lipid metabolism, as well as in evident shivering gain. The ability to predominantly control different parameters of thermoregulatory responses to cold may indicate the importance of both the TRPM8 and the TRPA1 ion channels in the processes of maintaining temperature homeostasis. The obtained data testify to the joint sequential operation of these thermosensitive ion channels., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The ability to safely control transgene expression with simple synthetic gene switches is critical for effective gene- and cell-based therapies. In the present study, the signaling pathway controlled by human transient receptor potential (TRP) melastatin 8 (hTRPM8), a TRP channel family member 1 , is harnessed to control transgene expression. Human TRPM8 signaling is stimulated by menthol, an innocuous, natural, cooling compound, or by exposure to a cool environment (15-18 °C). By functionally linking hTRPM8-induced signaling to a synthetic promoter containing elements that bind nuclear factor of activated T cells, a synthetic gene circuit was designed that can be adjusted by exposure to either a cool environment or menthol. It was shown that this gene switch is functional in various cell types and human primary cells, as well as in mice implanted with engineered cells. In response to transdermal delivery of menthol, microencapsulated cell implants harboring this gene circuit, coupled to expression of either of two therapeutic proteins, insulin or a modified, activin type IIB, receptor ligand trap protein (mActRIIB ECD -hFc), could alleviate hyperglycemia in alloxan-treated mice (a model of type 1 diabetes) or reverse muscle atrophy in dexamethasone-treated mice (a model of muscle wasting), respectively. This fully human-derived orthogonal transgene switch should be amenable to a wide range of clinical applications.

Potential harmful stimuli like heat, mechanical pressure or chemicals are detected by specialized cutaneous nerve fiber endings of nociceptor neurons in a process called nociception. Acute stimulation results in immediate protective reflexes and pain sensation as a normal, physiological behavior. However, ongoing (chronic) pain is a severe pathophysiological condition with diverse pathogeneses that is clinically challenging because of limited therapeutic options. Therefore, an urgent need exists for new potent and specific analgesics without afflicting adverse effects. Recently, TRPM3, a member of the superfamily of transient receptor potential (TRP) ion channels, has been shown to be expressed in nociceptors and to be involved in the detection of noxious heat (acute pain) as well as inflammatory hyperalgesia (acute and chronic pain). Current results in TRPM3 research indicate that this ion channel might not only be part of yet unraveled mechanisms underlying chronic pain but also has the potential to become a clinically relevant pharmacological target of future analgesic strategies. The aim of this review is to summarize and present the basic features of TRPM3 proteins and channels, to highlight recent findings and developments and to provide an outlook on emerging directions of TRPM3 research in the field of chronic pain.

External thermosensation is crucial to regulate animal behavior and homeostasis, but the development of the mammalian thermosensory system is not well known. We investigated whether temperature could play a role in the control of movements in a mammalian model born very immature, the opossum ( Monodelphis domestica ). Like other marsupials, at birth the opossum performs alternate and rhythmic movements with its forelimbs (FLs) to reach a teat where it attaches in order to continue its development. It was shown that FL movements can be induced by mechanical stimulation of the snout in in vitro preparations of newborns consisting of the neuraxis with skin and FLs intact. In the present study, we used puff ejections of cold, neutral (bath temperature) and hot liquid directed toward the snout to induce FL responses in such preparations. Either the responses were visually observed under a microscope or triceps muscle activity was recorded. Cold liquid systematically induced FL movements and triceps contractions, but neutral and hot temperatures were less potent to do so. Sections of the trigeminal nerves and removal of the facial skin diminished responses to cold and nearly abolished those to hot and neutral stimulations. Transient receptor potential melastatin 8 (TRPM8) being the major cold receptor cation channel in adult mammals, we employed immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) to test for its expression, but found that it is not expressed before 13 postnatal days. Overall our results indicate that cold thermosensation exerts a strong influence on motor behaviors in newborn opossums., (Copyright © 2019 Corriveau-Parenteau et al.)

The role of TRP channels in the ventilatory response to CO 2 was investigated in vivo. To this end, the respiration of unrestrained adult TRPM8-, TRPV1- and TRPV4-channel knockout mice was measured using whole-body plethysmography. Under control conditions and hyperoxic hypercapnia, no difference in respiratory parameters was observed between adult wild-type mice and TRPV1- and TRPV4-channel knockout mice. However, TRPM8-channel knockout mice showed decreased tidal volume under both hypercapnia and resting conditions. In addition, the expression of TRPM8, TRPV1 and TRPV4 mRNAs was detected in EGFP-positive glial cells in the medulla of GFAP promoter-EGFP transgenic mice by real-time PCR. Furthermore, we measured intracellular Ca 2+ responses of TRPM8-overexpressing HEK-293 cells to hypercapnic acidosis. Subpopulations of cells that exhibited hypercapnic acidosis-induced Ca 2+ response also responded to the application of menthol. These results suggest that TRPM8 partially mediates the ventilatory response to CO 2 via changes in intracellular Ca 2+ and is a chemosensing protein that may be involved in detecting endogenous CO 2 production., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Electrical stimulation of right Schaffer collateral in Trpm4 -/- knockout and wild type rats were used to study the role of Trpm4 channels for signal processing in the hippocampal formation. Stimulation induced neuronal activity was simultaneously monitored in the CA1 region by in vivo extracellular field recordings and in the entire brain by BOLD fMRI measurements. In wild type and Trpm4 -/- knockout rats, consecutive 5 Hz pulse trains elicited similar neuronal responses in the CA1 region and similar BOLD responses in the stimulated right hippocampus. Stimulus-related positive BOLD responses were also found in the left dorsal hippocampus. In contrast to the right dorsal hippocampus, baseline BOLD signals in the left hippocampus significantly decreased during consecutive stimulation trains. Similarly, slowly developing significant declines in baseline BOLD signals, in absence of any positive BOLD responses, were also observed in the right entorhinal, right piriform cortex, right basolateral amygdala and right dorsal striatum whereas baseline BOLD signals remained almost stable in the corresponding left regions. Furthermore, significant declines in baseline BOLD signals were found in the prefrontal cortex and prelimbic/infralimbic cortex. Because significant baseline BOLD declines were only observed in target regions of the right dorsal hippocampus, it might reflect functional connectivity between these regions. In all observed regions the decline in baseline BOLD signals was significantly delayed and less pronounced in Trpm4 -/- knockout rats when compared to wild type rats. Thus, either Trpm4 channels are involved in mediating these baseline BOLD shifts or functional connectivity of the hippocampus is impaired in Trpm4 -/- knockout rats., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Here, we investigated the effects of 9-hydroxyphenanthrene (9-phenanthrol), a potent and selective transient receptor potential melastatin 4 (TRPM4) channel blocker, on the resting membrane potential and cholinergic contractile responses to elucidate the functional role of TRPM4 channels in the contractile activities of mouse detrusor and ileal longitudinal smooth muscles. We observed that, 9-phenanthrol (3-30 µM) did not significantly inhibit high K + -induced contractions in both preparations; however, 9-phenanthrol (10 µM) strongly inhibited cholinergic contractions evoked by electrical field stimulation in detrusor preparations compared to inhibitions in ileal preparations. 9-Phenanthrol (10 µM) significantly inhibited the muscarinic agonist, carbachol-induced contractile responses and slowed the maximum upstroke velocities of the contraction in detrusor preparations. However, the agent (10 µM) did not inhibit the contractions due to intracellular Ca 2+ release evoked by carbachol, suggesting that the inhibitory effect of 9-phenanthrol may primarily be due to the inhibition of the membrane depolarization process incurred by TRPM4 channels. On the other hand, 9-phenanthrol (10 µM) did not affect carbachol-induced contractile responses in ileal preparations. Further, 9-phenanthrol (10 µM) significantly hyperpolarized the resting membrane potential and decreased the basal tone in both detrusor and ileal muscle preparations. Taken together, our results suggest that TRPM4 channels are constitutively active and are involved in setting of the resting membrane potential, thereby regulating the basal tone in detrusor and ileal smooth muscles. Thus, TRPM4 channels play a significant role in cholinergic signaling in detrusor, but not ileal, smooth muscles.

Pain is the leading cause for hospitalization in patients with sickle cell disease (SCD). While the characteristics of SCD pain can vary widely between patients and between phases of the disease (e.g. vasoocclusive crisis pain vs. chronic pain), similar neuronal mechanisms likely underlie the various aspects of nociceptive processing. In the peripheral nervous system, small unmyelinated C fibers and lightly-myelinated Aδ fibers detect and transmit noxious stimuli. Both classes of neurons express members of the transient receptor potential (TRP) family, a group of ligand gated ion-channels that are activated by thermal, chemical, and mechanical stimuli. Promiscuous TRP channel family members are activated by a wide range of stimuli, many of which are dysregulated in patients with SCD and transgenic SCD mouse models. In 2011, our lab published the first report of TRP channel contributions to rodent SCD pain. Since that time, additional basic and clinical research efforts have investigated the genetic and biochemical status of TRP channels in SCD, placing particular focus on TRPV1. This review will discuss these advances and highlight the clinical SCD presentations that have not yet been studied, but which may be mediated by TRP channel activity., (Copyright © 2018 Elsevier B.V. All rights reserved.)