Volatile anaesthetics inhibit the thermosensitive nociceptor ion channel transient receptor potential melastatin 3 (TRPM3)

BACKGROUND: Volatile anaesthetics (VAs) are the most widely used compounds to induce reversible loss of consciousness and maintain general anaesthesia during surgical interventions. Although the mechanism of their action is not yet fully understood, it is generally believed, that VAs depress central nervous system functions mainly through modulation of ion channels in the neuronal membrane, including 2-pore-domain K+ channels, GABA and NMDA receptors. Recent research also reported their action on nociceptive and thermosensitive TRP channels expressed in the peripheral nervous system, including TRPV1, TRPA1, and TRPM8. Here, we investigated the effect of VAs on TRPM3, a less characterized member of the thermosensitive TRP channels playing a central role in noxious heat sensation. METHODS: We investigated the effect of VAs on the activity of recombinant and native TRPM3, by monitoring changes in the intracellular Ca2+ concentration and measuring TRPM3-mediated transmembrane currents. RESULTS: All the investigated VAs (chloroform, halothane, isoflurane, sevoflurane) inhibited both the agonist-induced (pregnenolone sulfate, CIM0216) and heat-activated Ca2+ signals and transmembrane currents in a concentration dependent way in HEK293T cells overexpressing recombinant TRPM3. Among the tested VAs, halothane was the most potent blocker (IC50 = 0.52 ± 0.05 mM). We also investigated the effect of VAs on native TRPM3 channels expressed in sensory neurons of the dorsal root ganglia. While VAs activated certain sensory neurons independently of TRPM3, they strongly and reversibly inhibited the agonist-induced TRPM3 activity. CONCLUSIONS: These data provide a better insight into the molecular mechanism beyond the analgesic effect of VAs and propose novel strategies to attenuate TRPM3 dependent nociception.status: publishe

Transcriptome-based screening of ion channels and transporters in a migratory chondroprogenitor cell line isolated from late-stage osteoarthritic cartilage

Abstract Chondrogenic progenitor cells (CPCs) may be used as an alternative source of cells with potentially superior chondrogenic potential compared to mesenchymal stem cells (MSCs), and could be exploited for future regenerative therapies targeting articular cartilage in degenerative diseases such as osteoarthritis (OA). In this study, we hypothesised that CPCs derived from OA cartilage may be characterised by a distinct channelome. First, a global transcriptomic analysis using Affymetrix microarrays was performed. We studied the profiles of those ion channels and transporter families that may be relevant to chondroprogenitor cell physiology. Following validation of the microarray data with quantitative reverse transcription-polymerase chain reaction, we examined the role of calcium-dependent potassium channels in CPCs and observed functional large-conductance calcium-activated potassium (BK) channels involved in the maintenance of the chondroprogenitor phenotype. In line with our very recent results, we found that the KCNMA1 gene was upregulated in CPCs and observed currents that could be attributed to the BK channel. The BK channel inhibitor paxilline significantly inhibited proliferation, increased the expression of the osteogenic transcription factor RUNX2, enhanced the migration parameters, and completely abolished spontaneous Ca2+ events in CPCs. Through characterisation of their channelome we demonstrate that CPCs are a distinct cell population but are highly similar to MSCs in many respects. This study adds key mechanistic data to the in-depth characterisation of CPCs and their phenotype in the context of cartilage regeneration

Maurocalcine interacts with the cardiac ryanodine receptor without inducing channel modification

We have previously shown that MCa (maurocalcine), a toxin from the venom of the scorpion Maurus palmatus, binds to RyR1 (type 1 ryanodine receptor) and induces strong modifications of its gating behaviour. In the present study, we investigated the ability of MCa to bind to and modify the gating process of cardiac RyR2. By performing pull-down experiments we show that MCa interacts directly with RyR2 with an apparent affinity of 150 nM. By expressing different domains of RyR2 in vitro, we show that MCa binds to two domains of RyR2, which are homologous with those previously identified on RyR1. The effect of MCa binding to RyR2 was then evaluated by three different approaches: (i) [3H]ryanodine binding experiments, showing a very weak effect of MCa (up to 1 μM), (ii) Ca2+ release measurements from cardiac sarcoplasmic reticulum vesicles, showing that MCa up to 1 μM is unable to induce Ca2+ release, and (iii) single-channel recordings, showing that MCa has no effect on the open probability or on the RyR2 channel conductance level. Long-lasting opening events of RyR2 were observed in the presence of MCa only when the ionic current direction was opposite to the physiological direction, i.e. from the cytoplasmic face of RyR2 to its luminal face. Therefore, despite the conserved MCa binding ability of RyR1 and RyR2, functional studies show that, in contrast with what is observed with RyR1, MCa does not affect the gating properties of RyR2. These results highlight a different role of the MCa-binding domains in the gating process of RyR1 and RyR2