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Bistable motoneurons of the spinal cord exhibit warmth-activated plateau potential driven by Na+ and triggered by a brief excitation. The thermoregulating molecular mechanisms of bistability and their role in motor functions remain unknown. Here, we identify thermosensitive Na+-permeable Trpm5 channels as the main molecular players for bistability in mouse motoneurons. Pharmacological, genetic or computational inhibition of Trpm5 occlude bistable-related properties (slow afterdepolarization, windup, plateau potentials) and reduce spinal locomotor outputs while central pattern generators for locomotion operate normally. At cellular level, Trpm5 is activated by a ryanodine-mediated Ca2+ release and turned off by Ca2+ reuptake through the sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) pump. Mice in which Trpm5 is genetically silenced in most lumbar motoneurons develop hindlimb paresis and show difficulties in executing high-demanding locomotor tasks. Overall, by encoding bistability in motoneurons, Trpm5 appears indispensable for producing a postural tone in hindlimbs and amplifying the locomotor output., The authors show that Trpm5, but not Trpm4, is the main Na+ -permeant channel mediating the warmth-activated ICaN in lumbar motoneurons. Trpm5 is also critical in generating plateau potentials in bistable motoneurons that are essential for producing a postural tone in hindlimbs and amplifying the locomotor output.

In the malaria vector Anopheles gambiae, two point mutations in the acetylcholinesterase (ace-1R) and the sodium channel (kdrR) genes confer resistance to organophosphate/carbamate and pyrethroid insecticides, respectively. The mechanisms of compensation that recover the functional alterations associated with these mutations and their role in the modulation of insecticide efficacy are unknown. Using multidisciplinary approaches adapted to neurons isolated from resistant Anopheles gambiae AcerKis and KdrKis strains together with larval bioassays, we demonstrate that nAChRs, and the intracellular calcium concentration represent the key components of an adaptation strategy ensuring neuronal functions maintenance. In AcerKis neurons, the increased effect of acetylcholine related to the reduced acetylcholinesterase activity is compensated by expressing higher density of nAChRs permeable to calcium. In KdrKis neurons, changes in the biophysical properties of the L1014F mutant sodium channel, leading to enhance overlap between activation and inactivation relationships, diminish the resting membrane potential and reduce the fraction of calcium channels available involved in acetylcholine release. Together with the lower intracellular basal calcium concentration observed, these factors increase nAChRs sensitivity to maintain the effect of low concentration of acetylcholine. These results explain the opposite effects of the insecticide clothianidin observed in AcerKis and KdrKis neurons in vitro and in vivo., Perrier et al. examine the neurons of mosquitoes following the development of insecticide resistance-associated point mutations in the voltage-gated sodium channels and AChE1 genes for two resistant strains, KdrKis and AcerKis. Their results show that nAChRs and the intracellular calcium concentration provide a compensatory mechanism for AcerKis and KdrKis neurons to insecticide exposure.

GABAA receptors mediate most inhibitory synaptic transmission in the brain of vertebrates. Following GABA binding and fast activation, these receptors undergo a slower desensitization, the conformational pathway of which remains largely elusive. To explore the mechanism of desensitization, we used concatemeric α1β2γ2 GABAA receptors to selectively introduce gain-of-desensitization mutations one subunit at a time. A library of twenty-six mutant combinations was generated and their bi-exponential macroscopic desensitization rates measured. Introducing mutations at the different subunits shows a strongly asymmetric pattern with a key contribution of the γ2 subunit, and combining mutations results in marked synergistic effects indicating a non-concerted mechanism. Kinetic modelling indeed suggests a pathway where subunits move independently, the desensitization of two subunits being required to occlude the pore. Our work thus hints towards a very diverse and labile conformational landscape during desensitization, with potential implications in physiology and pharmacology., GABAA receptors mediate most inhibitory synaptic transmission in the brain. Here authors used concatemeric α1β2γ2 GABAA receptors to introduce gain-of-desensitization mutations one subunit at a time, revealing non-concerted rearrangements with a key contribution of the γ2 subunit during desensitization.

N-arachidonoyl glycine (NAGly) is an endogenous lipid deriving from the endocannabinoid anandamide (AEA). Identified as a ligand of several G-protein coupled receptors (GPCRs), it can however exert biological responses independently of GPCRs. NAGly was recently shown to depress store-operated Ca2+ entry (SOCE) but its mechanism of action remains elusive. The major aim of this study was to gain a better knowledge on the NAGly-dependent impairment of SOCE in neurons of the central nervous system (CNS) from mice. First, we examined the expression of genes encoding for putative lipid sensing GPCRs using transcriptomic data publicly available. This analysis showed that the most abundant GPCRs transcripts present in the cerebral cortices of embryonic brains were coding for lysophosphatidic acid (LPA) and sphingosine-1 phosphate (S1P) receptors. Next, the presence of functional receptors was assessed with live-cell calcium imaging experiments. In primary cortical cells S1P and LPA mobilize Ca2+ from internal stores via a mechanism sensitive to the S1P and LPA receptor antagonists Ex26, H2L5186303, or Ki16425. However, none of these compounds prevented or attenuated the NAGly-dependent impairment of SOCE. We found no evidence for the requirement of lipid sensing GPCRs in this inhibitory process, indicating that NAGly is an endogenous modulator interfering with the core machinery of SOCE. Moreover, these data also raise the intriguing possibility that the depression of SOCE could play a role in the central effects of NAGly.

The excitability of neurons is tightly dependent on their ion channel repertoire. Among these channels, the leak sodium channel NALCN plays a crucial role in the maintenance of the resting membrane potential. Importantly, NALCN mutations lead to complex neurodevelopmental syndromes, including infantile hypotonia with psychomotor retardation and characteristic facies (IHPRF) and congenital contractures of limbs and face, hypotonia and developmental delay (CLIFAHDD), which are recessively and dominantly inherited, respectively. Unfortunately, the biophysical properties of NALCN are still largely unknown to date, as well as the functional consequences of both IHPRF and CLIFAHDD mutations on NALCN current. Here we have set-up the heterologous expression of NALCN in the neuronal cell line NG108-15 to investigate the electrophysiological properties of NALCN carrying representative IHPRF and CLIFAHDD mutations. Several original properties of the wild-type (wt) NALCN current were retrieved: mainly carried by external Na+, blocked by Gd3+, insensitive to TTX and potentiated by low external Ca2+ concentration. However, we found that this current displays a time-dependent inactivation in the −80/−40 mV range of membrane potential, and a non linear current-voltage relationship indicative of voltage sensitivity. Importantly, no detectable current was recorded with the IHPRF missense mutation p.Trp1287Leu (W1287L), while the CLIFAHDD mutants, p.Leu509Ser (L509S) and p.Tyr578Ser (Y578S), showed higher current densities and slower inactivation, compared to wt NALCN current. This study reveals that heterologous expression of NALCN channel can be achieved in the neuronal cell line NG108-15 to study the electrophysiological properties of wt and mutants. From our results, we conclude that IHPRF and CLIFAHDD missense mutations are loss- and gain-of-function variants, respectively.

Piezo1 and Piezo2 encode mechanically activated cation channels that function as mechanotransducers involved in vascular system development and touch sensing, respectively. Structural features of Piezos remain unknown. Mouse Piezo1 is bioinformatically predicted to have 30–40 transmembrane (TM) domains. Here, we find that nine of the putative inter-transmembrane regions are accessible from the extracellular side. We use chimeras between mPiezo1 and dPiezo to show that ion-permeation properties are conferred by C-terminal region. We further identify a glutamate residue within a conserved region adjacent to the last two putative TM domains of the protein, that when mutated, affects unitary conductance and ion selectivity, and modulates pore block. We propose that this amino acid is either in the pore or closely associates with the pore. Our results describe important structural motifs of this channel family and lay the groundwork for a mechanistic understanding of how Piezos are mechanically gated and conduct ions., Piezo ion channels function as mechanotransducers involved in vascular development and touch sensing, but their structural features remain unknown. Here the authors find that the C-terminal region of Piezo protein encompasses the pore and identify a glutamate residue within this region involved in ion conduction properties.