Unraveling the molecular reason of opposing effects of α-mangostin and norfluoxetine on TREK-2 at the same binding site [[EQUATION]].

TWIK-related K(+) channel (TREK)-2, expressed in sensory neurons, is involved in setting membrane potential, and its modulations contributes to the generation of nociceptive signals. Although acute and chronic pain is a common symptom experienced by patients with various conditions, most existing analgesics exhibit low efficacy and are associated with adverse effects. For this reason, finding the novel modulator of TREK-2 is of significance for the development of new analgesics. Recent studies have shown that α-Mangostin (α-MG) activates TREK-2, facilitating analgesic effects, yet the underlying molecular mechanisms remain elusive. Intriguingly, even though norfluoxetine (NFx) is known to inhibit TREK-2, α-MG is also observed to share the same binding site with NFx, and this implies that TREK-2 might be modulated in a highly complicated manner. Therefore, we examine the mechanism of how TREK-2 is activated by α-MG using computational methods and patch clamp experiments in the present study. Based on these results, we offer an explanation of how α-MG and NFx exhibit opposing effects at the same binding site of TREK-2. These findings will broaden our understanding of TREK-2 modulation, providing clues for designing novel analgesic drugs.

Sortilin-Mediated Inhibition of TREK1/2 Channels in Primary Sensory Neurons Promotes Prediabetic Neuropathic Pain.

Neuropathic pain can occur during the prediabetic stage, even in the absence of hyperglycemia. The presence of prediabetic neuropathic pain (PDNP) poses challenges to the management of individuals with prediabetes. However, the mechanisms underlying this pain remain unclear. This study aims to investigate the underlying mechanism and identify potential therapeutic targets of PDNP. A prediabetic animal model induced by a high-energy diet exhibits both mechanical allodynia and thermal hyperalgesia. Furthermore, hyperexcitability and decreased potassium currents are observed in the dorsal root ganglion (DRG) neurons of these rats. TREK1 and TREK2 channels, which belong to the two-pore-domain K+ channel (K2P) family and play an important role in controlling cellular excitability, are downregulated in DRG neurons. Moreover, this alteration is modulated by Sortilin, a molecular partner that modulates the expression of TREK1. The overexpression of Sortilin negatively affects the expression of TREK1 and TREK2, leading to increased neuronal excitability in the DRG and enhanced peripheral pain sensitivity in rats. Moreover, the downregulation of Sortilin or activation of TREK1 and TREK2 channels by genetic or pharmacological approaches can alleviate PDNP. Therefore, targeting the Sortilin-mediated TREK1/2 pathway may provide a therapeutic approach for ameliorating PDNP.

ß-COP Suppresses the Surface Expression of the TREK2.

K2P channels, also known as two-pore domain K+ channels, play a crucial role in maintaining the cell membrane potential and contributing to potassium homeostasis due to their leaky nature. The TREK, or tandem of pore domains in a weak inward rectifying K+ channel (TWIK)-related K+ channel, subfamily within the K2P family consists of mechanical channels regulated by various stimuli and binding proteins. Although TREK1 and TREK2 within the TREK subfamily share many similarities, ß-COP, which was previously known to bind to TREK1, exhibits a distinct binding pattern to other members of the TREK subfamily, including TREK2 and the TRAAK (TWIK-related acid-arachidonic activated K+ channel). In contrast to TREK1, ß-COP binds to the C-terminus of TREK2 and reduces its cell surface expression but does not bind to TRAAK. Furthermore, ß-COP cannot bind to TREK2 mutants with deletions or point mutations in the C-terminus and does not affect the surface expression of these TREK2 mutants. These results emphasize the unique role of ß-COP in regulating the surface expression of the TREK family.

Evaluation of the Ion Channel Assembly in a Eukaryotic Cell-Free System Focusing on Two-Pore Domain Potassium Channels K2P.

Oligomeric ion channels are abundant in nature. However, the recombinant expression in cell culture-based systems remains tedious and challenging due to negative side effects, limiting the understanding of their role in health and disease. Accordingly, in this work, we demonstrate the cell-free synthesis (CFS) as an alternative platform to study the assembly of two-pore domain potassium channels (K2P) within endogenous endoplasmic reticulum-derived microsomes. Exploiting the open nature of CFS, we investigate the cotranslational translocation of TREK-2 into the microsomes and suggest a cotranslational assembly with typical single-channel behavior in planar lipid-bilayer electrophysiology. The heteromeric assembly of K2P channels is a contentious matter, accordingly we prove the successful assembly of TREK-2 with TWIK-1 using a biomolecular fluorescence complementation assay, Western blot analysis and autoradiography. The results demonstrate that TREK-2 homodimer assembly is the initial step, followed by heterodimer formation with the nascent TWIK-1, providing evidence of the intergroup heterodimerization of TREK-2 and TWIK-1 in eukaryotic CFS. Since K2P channels are involved in various pathophysiological conditions, including pain and nociception, CFS paves the way for in-depth functional studies and related pharmacological interventions. This study highlights the versatility of the eukaryotic CFS platform for investigating ion channel assembly in a native-like environment.

Antiarrhythmic calcium channel blocker verapamil inhibits trek currents in sympathetic neurons.

Background and Purpose: Verapamil, a drug widely used in certain cardiac pathologies, exert its therapeutic effect mainly through the blockade of cardiac L-type calcium channels. However, we also know that both voltage-dependent and certain potassium channels are blocked by verapamil. Because sympathetic neurons of the superior cervical ganglion (SCG) are known to express a good variety of potassium currents, and to finely tune cardiac activity, we speculated that the effect of verapamil on these SCG potassium channels could explain part of the therapeutic action of this drug. To address this question, we decided to study, the effects of verapamil on three different potassium currents observed in SCG neurons: delayed rectifier, A-type and TREK (a subfamily of K2P channels) currents. We also investigated the effect of verapamil on the electrical behavior of sympathetic SCG neurons. Experimental Approach: We employed the Patch-Clamp technique to mouse SCG neurons in culture. Key Results: We found that verapamil depolarizes of the resting membrane potential of SCG neurons. Moreover, we demonstrated that this drug also inhibits A-type potassium currents. Finally, and most importantly, we revealed that the current driven through TREK channels is also inhibited in the presence of verapamil. Conclusion and Implications: We have shown that verapamil causes a clear alteration of excitability in sympathetic nerve cells. This fact undoubtedly leads to an alteration of the sympathetic-parasympathetic balance which may affect cardiac function. Therefore, we propose that these possible peripheral alterations in the autonomic system should be taken into consideration in the prescription of this drug.

Glial-derived neurotrophic factor regulates the expression of TREK2 in rat primary sensory neurons leading to attenuation of axotomy-induced neuropathic pain.

TREK2 is a member of the 2-pore domain family of K+ channels (K2P) preferentially expressed by unmyelinated, slow-conducting and non-peptidergic isolectin B4-binding (IB4+) primary sensory neurons of the dorsal root ganglia (DRG). IB4+ neurons depend on the glial-derived neurotrophic factor (GDNF) family of ligands (GFL's) to maintain their phenotype. In our previous work, we demonstrated that 7 days after spinal nerve axotomy (SNA) of the L5 DRG, TREK2 moves away from the cell membrane resulting in a more depolarised resting membrane potential (Em). Given that axotomy deprives DRG neurons from peripherally-derived GFL's, we hypothesized that they might control the expression of TREK2. Using a combination of immunohistochemistry, immunocytochemistry, western blotting, in vivo pharmacological manipulation and behavioral tests we examined the ability of the GFL's (GDNF, neurturin and artemin) and their selective receptors (GFRα1, GFRα2 and GFRα3) to regulate the expression and function of TREK2 in the DRG. We found that TREK2 correlated strongly with the three receptors normally and ipsilaterally for all GFR's after SNA. GDNF, but not NGF, neurturin or artemin up-regulated the expression of TREK2 in cultured DRG neurons. In vivo continuous, subcutaneous administration of GDNF restored the subcellular distribution of TREK2 ipsilaterally and reversed mechanical and cold allodynia 7 days after SNA. This is the first demonstration that GDNF controls the expression of a K2P channel in nociceptors. As TREK2 controls the Em of C-nociceptors affecting their excitability, our finding has therapeutic potential in the treatment of chronic pain.

Piezo1 and Piezo2 foster mechanical gating of K2P channels.

Mechanoelectrical transduction is mediated by the opening of different types of force-sensitive ion channels, including Piezo1/2 and the TREK/TRAAK K2P channels. Piezo1 curves the membrane locally into an inverted dome that reversibly flattens in response to force application. Moreover, Piezo1 forms numerous preferential interactions with various membrane lipids, including cholesterol. Whether this structural architecture influences the functionality of neighboring membrane proteins is unknown. Here, we show that Piezo1/2 increase TREK/TRAAK current amplitude, slow down activation/deactivation, and remove inactivation upon mechanical stimulation. These findings are consistent with a mechanism whereby Piezo1/2 cause a local depletion of membrane cholesterol associated with a prestress of TREK/TRAAK channels. This regulation occurs in mouse fibroblasts between endogenous Piezo1 and TREK-1/2, both channel types acting in concert to delay wound healing. In conclusion, we demonstrate a community effect between different structural and functional classes of mechanosensitive ion channels.

Activity of TREK-2-like Channels in the Pyramidal Neurons of Rat Medial Prefrontal Cortex Depends on Cytoplasmic Calcium.

TREK-2-like channels in the pyramidal neurons of rat prefrontal cortex are characterized by a wide range of spontaneous activity-from very low to very high-independent of the membrane potential and the stimuli that are known to activate TREK-2 channels, such as temperature or membrane stretching. The aim of this study was to discover what factors are involved in high levels of TREK-2-like channel activity in these cells. Our research focused on the PI(4,5)P2-dependent mechanism of channel activity. Single-channel patch clamp recordings were performed on freshly dissociated pyramidal neurons of rat prefrontal cortexes in both the cell-attached and inside-out configurations. To evaluate the role of endogenous stimulants, the activity of the channels was recorded in the presence of a PI(4,5)P2 analogue (PI(4,5)P2DiC8) and Ca2+. Our research revealed that calcium ions are an important factor affecting TREK-2-like channel activity and kinetics. The observation that calcium participates in the activation of TREK-2-like channels is a new finding. We showed that PI(4,5)P2-dependent TREK-2 activity occurs when the conditions for PI(4,5)P2/Ca2+ nanocluster formation are met. We present a possible model explaining the mechanism of calcium action.

Effects of GABAB receptor activation on excitability of IB4-positive maxillary trigeminal ganglion neurons: Possible involvement of TREK2 activation.

IB4-positive maxillary trigeminal ganglion (TG) neurons are a subtype of afferent neurons involving nociception in orofacial regions, and excitability of these neurons is associated with orofacial nociceptive sensitivity. TREK-2 channel is a member of two-pore domain potassium (K2P) channel family mediating leak K+ currents. It has been shown previously that TREK-2 channel activity can be enhanced following GABAB receptor activation, leading to a reduction of cortical neuron excitability. In the present study, we have characterized TREK-2 channel expression on maxillary TG neurons and investigated the effect of the GABAB agonist baclofen on electrophysiological properties of small-sized maxillary TG neurons of rats. We show with immunohistochemistry that TREK-2 channels are predominantly expressed in small-sized IB4-positive maxillary TG neurons. Patch-clamp recordings on neurons in ex vivo TG preparations show that baclofen hyperpolarizes resting membrane potentials, increases outward leak currents, and decreases input resistances in IB4-positive maxillary TG neurons. Moreover, baclofen significantly reduces action potential (AP) firing in IB4-positive maxillary TG neurons. In contrast, baclofen shows no significant effect on electrophysiological properties of small-sized nociceptive-like and non-nociceptive-like maxillary trigeminal neurons that are IB4-negatve. Our results suggest that TREK-2 channel activity can be enhanced by baclofen, leading to reduced excitability of IB4-positive maxillary TG neurons. This finding provides new insights into the role of TREK-2 and GABAB receptors in controlling nociceptive sensitivity in orofacial regions, which may have therapeutic implications.

Negative Influence by the Force: Mechanically Induced Hyperpolarization via K2P Background Potassium Channels.

The two-pore domain K2P subunits form background (leak) potassium channels, which are characterized by constitutive, although not necessarily constant activity, at all membrane potential values. Among the fifteen pore-forming K2P subunits encoded by the KCNK genes, the three members of the TREK subfamily, TREK-1, TREK-2, and TRAAK are mechanosensitive ion channels. Mechanically induced opening of these channels generally results in outward K+ current under physiological conditions, with consequent hyperpolarization and inhibition of membrane potential-dependent cellular functions. In the past decade, great advances have been made in the investigation of the molecular determinants of mechanosensation, and members of the TREK subfamily have emerged among the best-understood examples of mammalian ion channels directly influenced by the tension of the phospholipid bilayer. In parallel, the crucial contribution of mechano-gated TREK channels to the regulation of membrane potential in several cell types has been reported. In this review, we summarize the general principles underlying the mechanical activation of K2P channels, and focus on the physiological roles of mechanically induced hyperpolarization.

Pharmacological Approaches to Studying Potassium Channels.

In this review, we consider the pharmacology of potassium channels from the perspective of these channels as therapeutic targets. Firstly, we describe the three main families of potassium channels in humans and disease states where they are implicated. Secondly, we describe the existing therapeutic agents which act on potassium channels and outline why these channels represent an under-exploited therapeutic target with potential for future drug development. Thirdly, we consider the evidence desired in order to embark on a drug discovery programme targeting a particular potassium channel. We have chosen two "case studies": activators of the two-pore domain potassium (K2P) channel TREK-2 (K2P10.1), for the treatment of pain and inhibitors of the voltage-gated potassium channel KV1.3, for use in autoimmune diseases such as multiple sclerosis. We describe the evidence base to suggest why these are viable therapeutic targets. Finally, we detail the main technical approaches available to characterise the pharmacology of potassium channels and identify novel regulatory compounds. We draw particular attention to the Comprehensive in vitro Proarrhythmia Assay initiative (CiPA, https://cipaproject.org ) project for cardiac safety, as an example of what might be both desirable and possible in the future, for ion channel regulator discovery projects.

The Prostacyclin Analogue, Treprostinil, Used in the Treatment of Pulmonary Arterial Hypertension, is a Potent Antagonist of TREK-1 and TREK-2 Potassium Channels.

Pulmonary arterial hypertension (PAH) is an aggressive vascular remodeling disease that carries a high morbidity and mortality rate. Treprostinil (Remodulin) is a stable prostacyclin analogue with potent vasodilatory and anti-proliferative activity, approved by the FDA and WHO as a treatment for PAH. A limitation of this therapy is the severe subcutaneous site pain and other forms of pain experienced by some patients, which can lead to significant non-compliance. TWIK-related potassium channels (TREK-1 and TREK-2) are highly expressed in sensory neurons, where they play a role in regulating sensory neuron excitability. Downregulation, inhibition or mutation of these channels leads to enhanced pain sensitivity. Using whole-cell patch-clamp electrophysiological recordings, we show, for the first time, that treprostinil is a potent antagonist of human TREK-1 and TREK-2 channels but not of TASK-1 channels. An increase in TASK-1 channel current was observed with prolonged incubation, consistent with its therapeutic role in PAH. To investigate treprostinil-induced inhibition of TREK, site-directed mutagenesis of a number of amino acids, identified as important for the action of other regulatory compounds, was carried out. We found that a gain of function mutation of TREK-1 (Y284A) attenuated treprostinil inhibition, while a selective activator of TREK channels, BL-1249, overcame the inhibitory effect of treprostinil. Our data suggests that subcutaneous site pain experienced during treprostinil therapy may result from inhibition of TREK channels near the injection site and that pre-activation of these channels prior to treatment has the potential to alleviate this nociceptive activity.

Higher expression of KCNK10 (TREK-2) K+ channels and their functional upregulation by lipopolysaccharide treatment in mouse peritoneal B1a cells.

Innate-like CD5+ B1a cells localized in serous cavities are activated by innate stimuli, such as lipopolysaccharide (LPS), leading to T cell-independent antibody responses. Although ion channels play crucial roles in the homeostasis and activation of immune cells, the electrophysiological properties of B1a cells have not been investigated to date. Previously, in the mouse B cell lymphoma cells, we found that the voltage-independent two-pore-domain potassium (K2P) channels generate a negative membrane potential and drive Ca2+ influx. Here, we newly compared the expression and activities of K2P channels in mouse splenic follicular B (FoB), marginal zone B (MZB), and peritoneal B1a cells. Next-generation sequencing analysis showed higher levels of transcripts for TREK-2 and TWIK-2 in B1a cells than those in FoB or MZB cells. Electrophysiological analysis, using patch clamp technique, revealed higher activity of TREK-2 with the characteristic large unitary conductance (~ 250 pS) in B1a than that in FoB or MZB cells. TREK-2 activity was further increased by LPS treatment (>2 h), which was more prominent in B1a than that in MZB or FoB cells. The cytosolic Ca2+ concentration of B cells was decreased by high-K+-induced depolarization (ΔRKCl (%)), suggesting the basal Ca2+ influx to be driven by negative membrane potential. The LPS treatment significantly increased the ΔRKCl (%) in B1a, though not in FoB and MZB cells. Our study was the first to compare the K2P channels in mouse primary B cell subsets, elucidating the functional upregulation of TREK-2 and augmentation of Ca2+ influx by the stimulation of Toll-like receptor 4 in B1a cells.

Dual regulatory effects of PI(4,5)P2 on TREK-2 K+ channel through antagonizing interaction between the alkaline residues (K330 and R355-357) in the cytosolic C-terminal helix.

TWIK-related two-pore domain K+ channel-2 (TREK-2) has voltageindependent activity and shows additional activation by acidic intracellular pH (pHi) via neutralizing the E332 in the cytoplasmic C terminal (Ct). We reported opposite regulations of TREK-2 by phosphatidylinositol 4,5-bisphosphate (PIP2) via the alkaline K330 and triple Arg residues (R355-357); inhibition and activation, respectively. The G334 between them appeared critical because its mutation (G334A) endowed hTREK-2 with tonic activity, similar to the mutation of the inhibitory K330 (K330A). To further elucidate the role of putative bent conformation at G334, we compared the dual mutation forms, K330A/G334A and G334A/R355-7A, showing higher and lower basal activity, respectively. The results suggested that the tonic activity of G334A owes to a dominant influence from R355-7. Since there are additional triple Arg residues (R377-9) distal to R355-7, we also examined the triple mutant (G334A/R355-7A/R377-9A) that showed tonic inhibition same with G334A/R355-7A. Despite the state of tonic inhibition, the activation by acidic pHi was preserved in both G334A/R355-7A and G334A/R355-7A/R377-9A, similar to the R355-7A. Also, the inhibitory effect of ATP could be commonly demonstrated under the activation by acidic pHi in R355-7A, G334A/R355-7A, and G334A/R355-7A/R377-9A. These results suggest that the putative bent conformation at G334 is important to set the tug-of-war between K330 and R355-7 in the PIP2-dependent regulation of TREK-2.

Spadin Selectively Antagonizes Arachidonic Acid Activation of TREK-1 Channels.

TREK-1 channel activity is a critical regulator of neuronal, cardiac, and smooth muscle physiology and pathology. The antidepressant peptide, spadin, has been proposed to be a TREK-1-specific blocker. Here we sought to examine the mechanism of action underlying spadin inhibition of TREK-1 channels. Heterologous expression in Xenopus laevis oocytes and electrophysiological analysis using two-electrode voltage clamp in standard bath solutions was used to characterize the pharmacological profile of wild-type and mutant murine TREK-1 and TREK-2 channels using previously established human K2P activators; arachidonic acid (AA), cis-4,7,10,13,16,19-docosahexaenoic acid (DHA), BL-1249, and cinnamyl-3,4-dihydroxy-α-cyanocinnamate (CDC) and inhibitors; spadin and barium (Ba2+). Mouse TREK-1 and TREK-2 channel currents were both significantly increased by AA, BL-1249, and CDC, similar to their human homologs. Under basal conditions, both TREK-1 and TREK-2 currents were insensitive to application of spadin, but could be blocked by Ba2+. Spadin did not significantly inhibit either TREK-1 or TREK-2 currents either chemically activated by AA, BL-1249, or CDC, or structurally activated via a gating mutation. However, pre-exposure to spadin significantly perturbed the subsequent activation of TREK-1 currents by AA, but not TREK-2. Furthermore, spadin was unable to prevent activation of TREK-1 by BL-1249, CDC, or the related bioactive lipid, DHA. Spadin specifically antagonizes the activation of TREK-1 channels by AA, likely via an allosteric mechanism. Lack of intrinsic activity may explain the absence of clinical side effects during antidepressant therapy.

Neuronal microRNAs modulate TREK two-pore domain K+ channel expression and current density.

The TREK family of leak potassium channels has been found to play critical roles in nociception, sensitivity to general anaesthetics, neuroprotection, and memory. The three members of the family, TREK1, TREK2 and TRAAK establish the resting potential and modify the duration, frequency and amplitude of action potentials. Despite their apparent importance, the repertoire of regulatory interactions utilized by cells to control their expression is poorly understood. Herein, the contribution of miRNAs in the regulation of their post-transcriptional gene expression has been examined. Using different assays, miR-124 and to a lesser extent miR-128 and miR-183 were found to reduce TREK1 and TREK2 levels through specific binding to their 3'UTRs. In contrast, miR-9 which was predicted to bind to TRAAK 3'UTR, did not alter its expression. Expression of miR-124, miR-128 and miR-183 was found to mirror that of Trek1 and Trek2 mRNAs during brain development. Moreover, application of proinflammatory mediators in dorsal root ganglion (DRG) neurons revealed an inverse correlation between miR-124 and Trek1 and Trek2 mRNA expression. Voltage clamp recordings of TREK2-mediated currents showed that miR-124 reduced the sensitivity of TREK2-expressing cells to non-aversive warmth stimulation. Overall, these findings reveal a significant regulatory mechanism by which TREK1 and TREK2 expression and hence activity are controlled in neurons and uncover new druggable targets for analgesia and neuroprotection.Abbreviations: microRNA: miRNA; UTR: untranslated region; K2p channels: two-pore domain K+channels; DRG: dorsal root ganglion; CNS: central nervous system; FBS: fetal bovine serum; TuD: Tough Decoy; TREK: tandem P-domain weak inward rectifying K+ (TWIK)-related K+ channel 1; TRAAK: TWIK-related arachidonic acid K+.

Pranlukast is a novel small molecule activator of the two-pore domain potassium channel TREK2.

TREK2 (KCNK10, K2P10.1) is a two-pore domain potassium (K2P) channel and a potential target for the treatment of pain. Like the majority of the K2P superfamily, there is currently a lack of useful pharmacological tools to study TREK2. Here we present a strategy for identifying novel TREK2 activators. A cell-based thallium flux assay was developed and used to screen a library of drug-like molecules, from which we identified the CysLT1 antagonist Pranlukast as a novel activator of TREK2. This compound was selective for TREK2 versus TREK1 and showed no activity at TRAAK. Pranlukast was also screened against other members of the K2P superfamily. Several close analogues of Pranlukast and other CysLT1 antagonists were also tested for their ability to activate K2P channels. Consistent with previous work, structure activity relationships showed that subtle structural changes to these analogues completely attenuated the activation of TREK2, whereas for TREK1, analogues moved from activators to inhibitors. Pranlukast's activity was also confirmed using whole-cell patch clamp electrophysiology. Studies using mutant forms of TREK2 suggest Pranlukast does not bind in the K2P modulator pocket or the BL-1249 binding site. Pranlukast therefore represents a novel tool by which to study the mechanism of TREK2 activation.

Diurnal Rhythm of trek2a Expression is Associated with Diurnal Rhythm of gnrh3 Expression in Zebrafish.

The two-pore domain potassium ion (K + ) channel-related K + (TREK) channel and melatonin receptors play roles in the regulation of reproduction in zebrafish. Since reproduction is regulated by diurnal rhythms, the TREK family and melatonin receptors may exhibit diurnal rhythms in expression. In this study, we aimed to investigate diurnal variations of the gene expressions of TREK family and melatonin receptors and their associations with kisspeptin and gonadotrophin-releasing hormone (GnRH). Diurnal variations of trek1b, trek2a, trek2b, mt1, mt2, mel1a, kiss2 and gnrh3 expressions were examined by real-time PCR. For reproduction-related genes, kiss2 and gnrh3 exhibited diurnal rhythms. trek2a revealed a diurnal rhythm in the TREK family. mt2 and mel1c exhibited diurnal rhythms in the melatonin receptors. Since Trek2a regulates gnrh3 expression, the diurnal rhythm of gnrh3 expression suggests to be regulated by the diurnal rhythm of trek2a expression.

Towards a TREK-1/2 (TWIK-Related K+ Channel 1 and 2) dual activator tool compound: Multi-dimensional optimization of BL-1249.

This letter describes a focused, multi-dimensional optimization campaign around BL-1249, a fenamate class non-steroidal anti-inflammatory and a known activator of the K2P potassium channels TREK-1 (K2P2.1) and TREK-2 (K2P10.1). While BL-1249 has been widely profiled in vitro as a dual TREK-1/2 activator, poor physicochemical and DMPK properties have precluded a deeper understanding of the therapeutic potential of these key K2P channels across a broad spectrum of peripheral and central human disease. Here, we report multi-dimensional SAR that led to a novel TREK-1/2 dual activator chemotype, exemplified by ONO-2960632/VU6011992, with improved DMPK properties, representing a new lead for further optimization towards robust in vivo tool compounds.

Tandem pore TWIK-related potassium channels and neuroprotection.

TWIK-related potassium channels (TREK) belong to a subfamily of the two-pore domain potassium channels family with three members, TREK1, TREK2 and TWIK-related arachidonic acid-activated potassium channels. The two-pore domain potassium channels is the last big family of channels being discovered, therefore it is not surprising that most of the information we know about TREK channels predominantly comes from the study of heterologously expressed channels. Notwithstanding, in this review we pay special attention to the limited amount of information available on native TREK-like channels and real neurons in relation to neuroprotection. Mainly we focus on the role of free fatty acids, lysophospholipids and other neuroprotective agents like riluzole in the modulation of TREK channels, emphasizing on how important this modulation may be for the development of new therapies against neuropathic pain, depression, schizophrenia, epilepsy, ischemia and cardiac complications.