Transient Receptor Potential Melastatin Type Research Articles

Characterization of intestine-specific TRPM6 knockout C57BL/6J mice: effects of short-term omeprazole treatment.

The transient receptor potential melastatin type 6 (TRPM6) is a divalent cation channel pivotal for gatekeeping Mg2+ balance. Disturbance in Mg2+ balance has been associated with the chronic use of proton pump inhibitors (PPIs) such as omeprazole. In this study, we investigated if TRPM6 plays a role in mediating the effects of short-term (4days) omeprazole treatment on intestinal Mg2+ malabsorption using intestine-specific TRPM6 knockout (Vill1-TRPM6-/-) mice. To do this, forty-eight adult male C57BL/6J mice (50% TRPM6fl/fl and 50% Vill1-TRPM6-/-) were characterized, and the distal colon of these mice was subjected to RNA sequencing. Moreover, these mice were exposed to 20mg/kg bodyweight omeprazole or placebo for 4days. Vill1-TRPM6-/- mice had a significantly lower 25Mg2+ absorption compared to control TRPM6fl/fl mice, accompanied by lower Mg2+ serum levels, and urinary Mg2+ excretion. Furthermore, renal Slc41a3, Trpm6, and Trpm7 gene expressions were higher in these animals, indicating a compensatory mechanism via the kidney. RNA sequencing of the distal colon revealed a downregulation of the Mn2+ transporter Slc30a10. However, no changes in Mn2+ serum, urine, and feces levels were observed. Moreover, 4days omeprazole treatment did not affect Mg2+ homeostasis as no changes in serum 25Mg2+ and total Mg2+ were seen. In conclusion, we demonstrate here for the first time that Vill1-TRPM6-/- mice have a lower Mg2+ absorption in the intestines. Moreover, short-term omeprazole treatment does not alter Mg2+ absorption in both Vill1-TRPM6-/- and TRPM6fl/fl mice. This suggests that TRPM6-mediated Mg2+ absorption in the intestines is not affected by short-term PPI administration.

Pathogenic heterozygous TRPM7 variants and hypomagnesemia with developmental delay.

Heterozygous variants in Transient receptor potential melastatin type 7 (TRPM7), encoding an essential and ubiquitously expressed cation channel, may cause hypomagnesemia, but current evidence is insufficient to draw definite conclusions and it is unclear whether any other phenotypes can occur. Individuals with unexplained hypomagnesemia underwent whole-exome sequencing which identified TRPM7 variants. Pathogenicity of the identified variants was assessed by combining phenotypic, functional and in silico analyses. We report three new heterozygous missense variants in TRPM7 (p.Met1000Thr, p.Gly1046Arg, p.Leu1081Arg) in individuals with hypomagnesemia. Strikingly, autism spectrum disorder and developmental delay, mainly affecting speech and motor skills, was observed in all three individuals, while two out of three also presented with seizures. The three variants are predicted to be severely damaging by in silico prediction tools and structural modeling. Furthermore, these variants result in a clear loss-of-function of TRPM7-mediated magnesium uptake in vitro, while not affecting TRPM7 expression or insertion into the plasma membrane. This study provides additional evidence for the association between heterozygous TRPM7 variants and hypomagnesemia and adds developmental delay to the phenotypic spectrum of TRPM7-related disorders. Considering that the TRPM7 gene is relatively tolerant to loss-of-function variants, future research should aim to unravel by what mechanisms specific heterozygous TRPM7 variants can cause disease.

β-Lactam TRPM8 Antagonists Derived from Phe-Phenylalaninol Conjugates: Structure-Activity Relationships and Antiallodynic Activity.

The protein transient receptor potential melastatin type 8 (TRPM8), a non-selective, calcium (Ca2+)-permeable ion channel is implicated in several pathological conditions, including neuropathic pain states. In our previous research endeavors, we have identified β-lactam derivatives with high hydrophobic character that exhibit potent and selective TRPM8 antagonist activity. This work describes the synthesis of novel derivatives featuring C-terminal amides and diversely substituted N'-terminal monobenzyl groups in an attempt to increase the total polar surface area (TPSA) in this family of compounds. The primary goal was to assess the influence of these substituents on the inhibition of menthol-induced cellular Ca2+ entry, thereby establishing critical structure-activity relationships. While the substitution of the tert-butyl ester by isobutyl amide moieties improved the antagonist activity, none of the N'-monobencyl derivatives, regardless of the substituent on the phenyl ring, achieved the activity of the model dibenzyl compound. The antagonist potency of the most effective compounds was subsequently verified using Patch-Clamp electrophysiology experiments. Furthermore, we evaluated the selectivity of one of these compounds against other members of the transient receptor potential (TRP) ion channel family and some receptors connected to peripheral pain pathways. This compound demonstrated specificity for TRPM8 channels. To better comprehend the potential mode of interaction, we conducted docking experiments to uncover plausible binding sites on the functionally active tetrameric protein. While the four main populated poses are located by the pore zone, a similar location to that described for the N-(3-aminopropyl)-2-[(3-methylphenyl)methoxy]-N-(2-thienylmethyl)-benzamide (AMTB) antagonist cannot be discarded. Finally, in vivo experiments, involving a couple of selected compounds, revealed significant antinociceptive activity within a mice model of cold allodynia induced by oxaliplatin (OXA).

Targeting temperature-sensitive transient receptor potential channels in hypertension: far beyond the perception of hot and cold.

Transient receptor potential (TRP) channels are nonselective cation channels and participate in various physiological roles. Thus, changes in TRP channel function or expression have been linked to several disorders. Among the many TRP channel subtypes, the TRP ankyrin type 1 (TRPA1), TRP melastatin type 8 (TRPM8), and TRP vanilloid type 1 (TRPV1) channels are temperature-sensitive and recognized as thermo-TRPs, which are expressed in the primary afferent nerve. Thermal stimuli are converted into neuronal activity. Several studies have described the expression of TRPA1, TRPM8, and TRPV1 in the cardiovascular system, where these channels can modulate physiological and pathological conditions, including hypertension. This review provides a complete understanding of the functional role of the opposing thermo-receptors TRPA1/TRPM8/TRPV1 in hypertension and a more comprehensive appreciation of TRPA1/TRPM8/TRPV1-dependent mechanisms involved in hypertension. These channels varied activation and inactivation have revealed a signaling pathway that may lead to innovative future treatment options for hypertension and correlated vascular diseases.

Protein kinase C-mediated phosphorylation of transient receptor potential melastatin type 2 Thr738 counteracts the effect of cytosolic Ca2+ and elevates the temperature threshold.

The transient receptor potential melastatin type 2 (TRPM2) channel is a non-selective cation channel that has high Ca2+ permeability. TRPM2 is sensitive to warm temperatures and is expressed in cells and tissues that are maintained at core body temperature. TRPM2 activity is also regulated by endogenous factors including redox signalling, cytosolic Ca2+ and adenosine diphosphate ribose. As a result of its wide expression and function at core body temperature, these endogenous factors could regulate TRPM2 activity at body temperature under physiological and pathophysiological conditions. We previously reported that cellular redox signalling can lower TRPM2 temperature thresholds, although the mechanism that regulates these thresholds is unclear. Here, we used biochemical and electrophysiological techniques to explore another regulatory mechanism for TRPM2 temperature thresholds that is mediated by TRPM2 phosphorylation. Our results show that: (1) the temperature threshold for TRPM2 activation is lowered by cytosolic Ca2+ ; (2) protein kinase C-mediated phosphorylation of TRPM2 counteracts the effect of cytosolic Ca2+ ; and (3) Thr738 in mouse TRPM2 that lies near the Ca2+ binding site in the cytosolic cleft of the transmembrane domain is a potential phosphorylation site that may be involved in phosphorylation-mediated elevation of TRPM2 thresholds. These findings provide structure-based evidence to understand how temperature thresholds of thermo-sensitive TRP channels (thermo-TRPs) are determined and regulated. KEY POINTS: The transient receptor potential melastatin type 2 (TRPM2) ion channel is temperature-sensitive and Ca2+ -permeable. Endogenous factors and pathways such as redox signalling can regulate TRPM2 activity at body temperature under physiological and pathophysiological conditions. In the present study, we report the novel finding that cytosolic Ca2+ lowers the temperature threshold for TRPM2 activation in a concentration-dependent manner. Protein kinase C-mediated phosphorylation of TRPM2 at amino acid Thr782 elevates the temperature threshold for activation by counteracting the effects of cytosolic Ca2+ . These findings provide structure-based evidence to understand how temperature thresholds of thermo-sensitive TRP channels are determined and regulated.

Possible role for rare TRPM7 variants in patients with hypomagnesaemia with secondary hypocalcaemia.

Hypomagnesaemia with secondary hypocal-caemia (HSH) is a rare autosomal recessive disorder caused by pathogenic variants in TRPM6, encoding the channel-kinase transient receptor potential melastatin type 6. Patients have very low serum magnesium (Mg2+) levels and suffer from muscle cramps and seizures. Despite genetic testing, a subgroup of HSH patients remains without a diagnosis. In this study, two families with an HSH phenotype but negative for TRPM6 pathogenic variants were subjected to whole exome sequencing. Using a complementary combination of biochemical and functional analyses in overexpression systems and patient-derived fibroblasts, the effect of the TRPM7-identified variants on Mg2+ transport was examined. For the first time, variants in TRPM7 were identified in two families as a potential cause for hereditary HSH. Patients suffer from seizures and muscle cramps due to magnesium deficiency and episodes of hypocalcaemia. In the first family, a splice site variant caused the incorporation of intron 1 sequences into the TRPM7 messenger RNA and generated a premature stop codon. As a consequence, patient-derived fibroblasts exhibit decreased cell growth. In the second family, a heterozygous missense variant in the pore domain resulted in decreased TRPM7 channel activity. We establish TRPM7 as a prime candidate gene for autosomal dominant hypomagnesaemia and secondary hypocalcaemia. Screening of unresolved patients with hypocalcaemia and secondary hypocalcaemia may further establish TRPM7 pathogenic variants as a novel Mendelian disorder.

In Vitro and In Vivo Pharmacological Characterization of a Novel TRPM8 Inhibitor Chemotype Identified by Small-Scale Preclinical Screening.

Transient receptor potential melastatin type 8 (TRPM8) is a target for the treatment of different physio-pathological processes. While TRPM8 antagonists are reported as potential drugs for pain, cancer, and inflammation, to date only a limited number of chemotypes have been investigated and thus a limited number of compounds have reached clinical trials. Hence there is high value in searching for new TRPM8 antagonistic to broaden clues to structure-activity relationships, improve pharmacological properties and explore underlying molecular mechanisms. To address this, the EDASA Scientific in-house molecular library has been screened in silico, leading to identifying twenty-one potentially antagonist compounds of TRPM8. Calcium fluorometric assays were used to validate the in-silico hypothesis and assess compound selectivity. Four compounds were identified as selective TRPM8 antagonists, of which two were dual-acting TRPM8/TRPV1 modulators. The most potent TRPM8 antagonists (BB 0322703 and BB 0322720) underwent molecular modelling studies to highlight key structural features responsible for drug–protein interaction. The two compounds were also investigated by patch-clamp assays, confirming low micromolar potencies. The most potent compound (BB 0322703, IC50 1.25 ± 0.26 μM) was then profiled in vivo in a cold allodinya model, showing pharmacological efficacy at 30 μM dose. The new chemotypes identified showed remarkable pharmacological properties paving the way to further investigations for drug discovery and pharmacological purposes.

The Molecular Basis of COVID-19 Pathogenesis, Conventional and Nanomedicine Therapy.

In late 2019, a new member of the Coronaviridae family, officially designated as “severe acute respiratory syndrome coronavirus 2” (SARS-CoV-2), emerged and spread rapidly. The Coronavirus Disease-19 (COVID-19) outbreak was accompanied by a high rate of morbidity and mortality worldwide and was declared a pandemic by the World Health Organization in March 2020. Within the Coronaviridae family, SARS-CoV-2 is considered to be the third most highly pathogenic virus that infects humans, following the severe acute respiratory syndrome coronavirus (SARS-CoV) and the Middle East respiratory syndrome coronavirus (MERS-CoV). Four major mechanisms are thought to be involved in COVID-19 pathogenesis, including the activation of the renin-angiotensin system (RAS) signaling pathway, oxidative stress and cell death, cytokine storm, and endothelial dysfunction. Following virus entry and RAS activation, acute respiratory distress syndrome develops with an oxidative/nitrosative burst. The DNA damage induced by oxidative stress activates poly ADP-ribose polymerase-1 (PARP-1), viral macrodomain of non-structural protein 3, poly (ADP-ribose) glycohydrolase (PARG), and transient receptor potential melastatin type 2 (TRPM2) channel in a sequential manner which results in cell apoptosis or necrosis. In this review, blockers of angiotensin II receptor and/or PARP, PARG, and TRPM2, including vitamin D3, trehalose, tannins, flufenamic and mefenamic acid, and losartan, have been investigated for inhibiting RAS activation and quenching oxidative burst. Moreover, the application of organic and inorganic nanoparticles, including liposomes, dendrimers, quantum dots, and iron oxides, as therapeutic agents for SARS-CoV-2 were fully reviewed. In the present review, the clinical manifestations of COVID-19 are explained by focusing on molecular mechanisms. Potential therapeutic targets, including the RAS signaling pathway, PARP, PARG, and TRPM2, are also discussed in depth.

UCP1 and TRPM8 Expression in the Brown Fat Did Not Affect the Restriction of Menthol-Induced Hyperthermia by Estradiol in Ovariectomized Rats.

Estradiol (E2) modulates the central and peripheral thermoregulatory responses to cold. Menthol is an agonist of transient receptor potential melastatin type 8 (TRPM8), which is a peripheral cold receptor. E2 suppresses menthol-induced elevation of body temperature (Tb) in ovariectomized rats, but the mechanism is unknown. The aim of the present study is to investigate the effect of E2 on uncoupling protein 1 (UCP1), a thermogenic gene, and TRPM8 mRNA levels in ovariectomized rats applied menthol. A silastic tube was implanted in ovariectomized rats with and without E2 underneath the dorsal skin (E2(+) and E2(-) groups), and data loggers for Tb measurement into peritoneal cavity. After application of 10% L-menthol or vehicle to the skin of the whole trunk of rats, Tb was measured for 2 h. The interscapular brown adipose tissue (BAT) and spinal ganglia of cervical, thoracic, and lumbar parts were obtained for RT-qPCR assay. In the menthol application, Tb in the E2(+) group was lower than that in the E2(-) group. The UCP1 mRNA in the BAT, TRPM8 mRNA in the BAT and spinal ganglia in all areas did not differ between the E2(+) and E2(-) groups. In conclusion, the UCP1 and TRPM8 expression in the brown fat did not affect the restriction of the menthol-induced hyperthermia by estradiol in ovariectomized rats.

Genetic and drug-induced hypomagnesemia: different cause, same mechanism.

Magnesium (Mg2+) plays an essential role in many biological processes. Mg2+ deficiency is therefore associated with a wide range of clinical effects including muscle cramps, fatigue, seizures and arrhythmias. To maintain sufficient Mg2+ levels, (re)absorption of Mg2+ in the intestine and kidney is tightly regulated. Genetic defects that disturb Mg2+ uptake pathways, as well as drugs interfering with Mg2+ (re)absorption cause hypomagnesemia. The aim of this review is to provide an overview of the molecular mechanisms underlying genetic and drug-induced Mg2+ deficiencies. This leads to the identification of four main mechanisms that are affected by hypomagnesemia-causing mutations or drugs: luminal transient receptor potential melastatin type 6/7-mediated Mg2+ uptake, paracellular Mg2+ reabsorption in the thick ascending limb of Henle's loop, structural integrity of the distal convoluted tubule and Na+-dependent Mg2+ extrusion driven by the Na+/K+-ATPase. Our analysis demonstrates that genetic and drug-induced causes of hypomagnesemia share common molecular mechanisms. Targeting these shared pathways can lead to novel treatment options for patients with hypomagnesemia.

Nicotine and menthol independently exert neuroprotective effects against cisplatin- or amyloid- toxicity by upregulating Bcl-xl via JNK activation in SH-SY5Y cells

Nicotine and menthol, agonists of nicotinic acetylcholine receptor (nAChR) and transient receptor potential melastatin type 8 (TRPM8), serve important roles in the prevention of cell death-involved neurodegenerative diseases. However, the potential synergistic effects of nicotine and menthol on anti-apoptotic ability are still uncertain. In the present study, the potential synergistic effects of nicotine and menthol on cisplatin or amyloid β_1-42 induced cell model of the neurodegenerative diseases were explored by assessing cell viability, TNF-α expression, caspase-3 activation, and the collapse of mitochondrial membrane potential in human SH-SY5Y neuroblastoma cells. Statistical significance was tested using Student’s <i>t</i>-test or one-way ANOVA with <i>post hoc</i> Newman-Keuls test. The results showed that: Firstly, SH-SY5Y cell viability was obviously increased by the treatments with nicotine and menthol. Secondly, nicotine and menthol independently alleviated cisplatin or amyloid β_1-42 induced TNF-α up-regulation. Thirdly, nicotine and menthol abrogated the effect of cisplatin and amyloid β_25-35 on caspase-3 activation. Interestingly, the effect of cisplatin and amyloid β_1-42 on the collapse of mitochondrial membrane potential was efficiently attenuated by nicotine and menthol treatments. Most importantly, the inhibition of c-jun kinase (JNK) activation abolished the effect of cisplatin, and amyloid β_1-42 stimulated Bcl-xl expression. All these findings indicate that nicotine and menthol independently exert neuroprotective effects by upregulating Bcl-xl <i>via</i> JNK activation. Nicotine and menthol augmented Bcl-xl expression and JNK phosphorylation, and thus they are potential therapeutic targets for altering the progress of neurodegenerative diseases.

Structure-Function Relationship of TRPM2: Recent Advances, Contradictions, and Open Questions.

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.

TRPM2 ion channel is involved in the aggravation of cognitive impairment and down regulation of epilepsy threshold in pentylenetetrazole-induced kindling mice

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 Ca2+-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.

Regulation of TRPM8 channel activity by Src-mediated tyrosine phosphorylation.

The transient receptor potential melastatin type 8 (TRPM8) receptor channel is expressed in primary afferent neurons where it is the main transducer of innocuous cold temperatures and also in a variety of tumors, where it is involved in progression and metastasis. Modulation of this channel by intracellular signaling pathways has therefore important clinical implications. We investigated the modulation of recombinant and natively expressed TRPM8 by the Src kinase, which is known to be involved in cancer pathophysiology and inflammation. Human TRPM8 expressed in HEK293T cells is constitutively tyrosine phosphorylated by Src which is expressed either heterologously or endogenously. Src action on TRPM8 potentiates its activity, as treatment with PP2, a selective Src kinase inhibitor, reduces both TRPM8 tyrosine phosphorylation and cold-induced channel activation. RNA interference directed against the Src kinase diminished the extent of PP2-induced functional downregulation of TRPM8, confirming that PP2 acts mainly through Src inhibition. Finally, the effect of PP2 on TRPM8 cold activation was reproduced in cultured rat dorsal root ganglion neurons, and this action was antagonized by the protein tyrosine phosphatase inhibitor pervanadate, confirming that TRPM8 activity is sensitive to the cellular balance between tyrosine kinases and phosphatases. This positive modulation of TRPM8 by Src kinase may be relevant for inflammatory pain and cancer signaling.

Pharmacology of JNJ-28583113: A novel TRPM2 antagonist

Transient receptor potential melastatin type 2 (TRPM2) is a cation channel activated by free intracellular ADP-ribose and reactive oxygen species. TRPM2 signaling has been linked to the pathophysiology of CNS disorders such as neuropathic pain, bipolar disorder and Alzheimer's disease. In this manuscript, we describe the discovery of JNJ-28583113, a potent brain penetrant TRPM2 antagonist. Ca2+ flux assays in cells overexpressing TRPM2 and electrophysiological recordings were used to test the pharmacology of JNJ-28583113. JNJ-28583113 was assayed in vitro on GSK-3 phosphorylation levels, cell death, cytokine release in microglia and unbiased morphological phenotypic analysis. Finally, we dosed animals to evaluate its pharmacokinetic properties. Our results showed that JNJ-28583113 is a potent (126 ± 0.5 nM) TRPM2 antagonist. Blocking TRPM2 caused phosphorylation of GSK3α and β subunits. JNJ-28583113 also protected cells from oxidative stress induced cell death as well as morphological changes induced by non-cytotoxic concentrations of H2O2. In addition, inhibiting TRPM2 blunted cytokine release in response to pro-inflammatory stimuli in microglia. Lastly, we showed that JNJ-28583113 was brain penetrant but not suitable for systemic dosing as it was rapidly metabolized in vivo. While the in-vitro pharmacology of JNJ-28583113 is the best in class, its in-vivo properties would need optimization to assist in further probing key roles of TRPM2 in CNS pathophysiology.

Renal phospholipidosis and impaired magnesium handling in high-fat-diet-fed mice.

Hypomagnesemia (blood Mg2+ concentration <0.7 mM) is a common electrolyte disorder in patients with type 2 diabetes (T2D), but the etiology remains largely unknown. In patients with T2D, reduced blood Mg2+ levels are associated with an increased decline in renal function, independent of glycemic control and hypertension. To study the underlying mechanism of this phenomenon, we investigated the renal effects of hypomagnesemia in high-fat-diet (HFD)-fed mice. In mice fed a low dietary Mg2+, the HFD resulted in severe hypomagnesemia within 4 wk. Renal or intestinal Mg2+ wasting was not observed after 16 wk on the diets. Despite the absence of urinary or fecal Mg2+ loss, the HFD induced a reduction in the mRNA expression transient receptor potential melastatin type 6 in both the kidney and colon. mRNA expression of distal convoluted tubule (DCT)-specific genes was down-regulated by the LowMg-HFD, indicating atrophy of the DCT. The low dietary Mg2+ resulted in severe HFD-induced proximal tubule phospholipidosis, which was absent in mice on a NormalMg-HFD. This was accompanied by albuminuria, moderate renal damage, and alterations in renal energy metabolism, including enhanced gluconeogenesis and cholesterol synthesis. In conclusion, this study shows that hypomagnesemia is a consequence of diet-induced obesity and insulin resistance. Moreover, hypomagnesemia induces major structural changes in the diabetic kidney, including proximal tubular phospholipidosis, providing a novel mechanism for the increased renal decline in patients with hypomagnesemic T2D.-Kurstjens, S., Smeets, B., Overmars-Bos, C., Dijkman, H. B., den Braanker, D. J. W., de Bel, T., Bindels, R. J. M., Tack, C. J. J., Hoenderop, J. G. J., de Baaij, J. H. F. Renal phospholipidosis and impaired magnesium handling in high-fat-diet-fed mice.

Transient receptor potential melastatin 2 governs stress-induced depressive-like behaviors

Major depressive disorder (MDD) is a devastating disease that arises in a background of environmental risk factors, such as chronic stress, that produce reactive oxygen species (ROS) in the brain. The chronic stress-induced ROS production involves Ca2+ signals; however, the mechanism is poorly understood. Transient receptor potential melastatin type 2 (TRPM2) is a Ca2+-permeable cation channel that is highly expressed in the brain. Here we show that in animal models of chronic unpredictable stress (CUS), deletion of TRPM2 (Trpm2-/- ) produces antidepressant-like behaviors in mice. This phenotype correlates with reduced ROS, ROS-induced calpain activation, and enhanced phosphorylation of two Cdk5 targets including synapsin 1 and histone deacetylase 5 that are linked to synaptic function and gene expression, respectively. Moreover, TRPM2 mRNA expression is increased in hippocampal tissue samples from patients with MDD. Our findings suggest that TRPM2 is a key agent in stress-induced depression and a possible target for treating depression.

TRPM6 is Essential for Magnesium Uptake and Epithelial Cell Function in the Colon.

Intestinal magnesium (Mg) uptake is essential for systemic Mg homeostasis. Colon cells express the two highly homologous transient receptor potential melastatin type (TRPM) 6 and 7 Mg2+ channels, but their precise function and the consequences of their mutual interaction are not clear. To explore the functional role of TRPM6 and TRPM7 in the colon, we used human colon cell lines that innately express both channels and analyzed the functional consequences of genetic knocking-down, by RNA interference, or pharmacological inhibition, by NS8593, of either channel. TRPM7 silencing caused an increase in Mg2+ influx, and correspondingly enhanced cell proliferation and migration, while downregulation of TRPM6 did not affect significantly either Mg2+ influx or cell proliferation. Exposure to the specific TRPM6/7 inhibitor NS8593 reduced Mg2+ influx, and consequently cell proliferation and migration, but Mg supplementation rescued the inhibition. We propose a model whereby in colon cells the functional Mg2+ channel at the plasma membrane may consist of both TRPM7 homomers and TRPM6/7 heteromers. A different expression ratio between the two proteins may result in different functional properties. Altogether, our findings confirm that TRPM6 cannot be replaced by TRPM7, and that TRPM6/7 complexes and TRPM6/7-mediated Mg2+ influx are indispensable in human epithelial colon cells.

The anthelminthic drug praziquantel is a selective agonist of the sensory transient receptor potential melastatin type 8 channel

Praziquantel is the most effective anthelminthic drug for the treatment of schistosomiasis, an infectious disease caused by the platyhelminth Schistosoma mansoni. While praziquantel is known to trigger calcium influx into schisostomes, followed by spastic paralysis of the worms and tegumental disruption, the mechanism of action of the drug is not completely understood. Although relatively well tolerated, praziquantel has been reported to cause mild adverse effects, including nausea, abdominal pain and headaches. As a number of putative Transient Receptor Potential (TRP) channel genes have recently been predicted in S. mansoni, we sought to investigate the effect of praziquantel on three mammalian TRP channels, TRP melastatin type 8 (TRPM8), TRP vanilloid type 1 (TRPV1) and TRP ankyrin type 1 (TRPA1). Using calcium microfluorimetry and the patch clamp technique, we recorded the effect of praziquantel on HEK293T cells expressing recombinant TRPM8, TRPV1 or TRPA1, as well as on cultured dorsal root ganglion (DRG) neurons from wild type and TRPM8 null mutant mice. We discovered that praziquantel is a relatively potent and selective partial agonist of the mammalian and avian cold and menthol receptor TRPM8. The activation of cultured DRG neurons by clinically relevant concentrations of praziquantel is predominantly mediated by TRPM8. Our results may provide clues to a better understanding of praziquantel's mechanism of action and its adverse effects.

A Heterozygous de novo Mutation in SLC41A1 Causes Hypomagnesemia and Renal Magnesium Wasting

Over the last decades, the identification of hereditary hypomagnesemia‐causing genes has increased our understanding of the mechanisms underlying renal magnesium (Mg2+) reabsorption. In the kidney, urinary Mg2+ excretion is determined in the distal convoluted tubule, as no Mg2+ is reabsorbed beyond this segment. In the distal convoluted tubule, the epithelial Mg2+ channel transient receptor potential melastatin type 6 (TRPM6) mediates active Mg2+ reabsorption from the pro‐urine into the cell. However, to date the molecular identity of the proteins facilitating Mg2+ extrusion from the cell towards the blood remains elusive.In this study, a three‐year‐old girl was identified with tetanic convulsions caused by severe hypomagnesemia and hypocalcemia. A Mg2+‐loading test showed urinary Mg2+ wasting, demonstrating a renal origin of the disease. The aim of this study was to identify the gene mutation that is causative for hypomagnesemia in this patient. Moreover, we investigated the molecular mechanism of disease by cellular Mg2+ transport experiments and by gene expression knockdown experiments in the zebrafish model.Whole exome sequencing identified a heterozygous de novo p.Ile98Phe mutation in the SLC41A1 gene. 25Mg2+ transport assays in human embryonic kidney 293 cells overexpressing SLC41A1 demonstrated that this transporter mediates cellular Mg2+ extrusion. To examine the pathogenicity of the mutation, a slc41a1 zebrafish knockdown model was generated using two separate morpholino approaches. Slc41a1 knockdown resulted in a 20% reduction of the total magnesium content. Overexpression of SLC41A1 in slc41a1‐knockdown zebrafish restored the total magnesium content. Conversely, total magnesium levels were not normalized in slc41a1‐knockdown zebrafish overexpressing the patient's SLC41A1 mutant. Co‐expression of wild‐type and mutant SLC41A1 in zebrafish disturbed the Mg2+ balance, demonstrating that mutant SLC41A1 has a dominant negative effect over wild‐type SLC41A1.In conclusion, a heterozygous de novo mutation in SLC41A1 causes hypomagnesemia and renal Mg2+ wasting. These findings demonstrate the essential role of SLC41A1 in renal Mg2+ handling and maintenance of body Mg2+ balance. Based on our experiments, we propose that SLC41A1 may facilitate basolateral Mg2+ extrusion in the distal convoluted tubule.Support or Funding InformationThis work was supported by the Long‐Term Fellowship from the European Renal Association ‐ European Dialysis and Transplant Association (ERA‐EDTA, ERA LTF 175‐2014) and the Impulsion Grant from the ERA‐EDTA Working Group on Inherited Kidney Disorders (WGIKD) to dr. Francisco J. Arjona; and by the grants from the Netherlands Organization for Scientific Research (NWO VICI 016.130.668) and the EURenOmics project from the European Union seventh Framework Programme (FP7/2007–2013, agreement no. 305608) to prof. dr. Joost G.J. Hoenderop. Dr. Jeroen H.F. de Baaij is supported by grants from NWO (Rubicon 825.14.021), the Dutch Kidney Foundation (Kolff 14OKG17) and RIMLS.