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PloS One 2024Gulf War Illness (GWI) is a chronic condition characterized by multisystem symptoms that still affect up to one-third of veterans who engaged in combat in the Gulf War...
Gulf War Illness (GWI) is a chronic condition characterized by multisystem symptoms that still affect up to one-third of veterans who engaged in combat in the Gulf War three decades ago. The aetiology of GWI is mainly explained by exposure to multiple toxic agents, vaccines, and medications. As there is a significant overlap in symptoms between GWI and Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS), the objective of this study was to investigate a biomarker widely reported in Natural Killer (NK) cells from ME/CFS patients, the Transient Receptor Potential Melastatin 3 (TRPM3) ion channel. NK cells from 6 healthy controls (HC) and 6 GWI participants were isolated, and TRPM3 function was assessed through whole-cell patch-clamp. As demonstrated by prior studies, NK cells from HC expressed typical TRPM3 function after pharmacomodulation. In contrast, this pilot investigation demonstrates a dysfunctional TRPM3 in NK cells from GWI participants through application of a TRPM3 agonist and confirmed by a TRPM3 antagonist. There was a significant reduction in TRPM3 function from GWI than results measured in HC. This study provides an unprecedented research field to investigate the involvement of TRP ion channels in the pathomechanism and potential medical interventions to improve GWI quality of life.
Topics: Humans; TRPM Cation Channels; Persian Gulf Syndrome; Killer Cells, Natural; Male; Middle Aged; Adult; Female; Case-Control Studies; Patch-Clamp Techniques
PubMed: 38917121
DOI: 10.1371/journal.pone.0305704 -
Proceedings of the National Academy of... Jul 2024HCN1-4 channels are the molecular determinants of the I/I current that crucially regulates cardiac and neuronal cell excitability. HCN dysfunctions lead to sinoatrial...
HCN1-4 channels are the molecular determinants of the I/I current that crucially regulates cardiac and neuronal cell excitability. HCN dysfunctions lead to sinoatrial block (HCN4), epilepsy (HCN1), and chronic pain (HCN2), widespread medical conditions awaiting subtype-specific treatments. Here, we address the problem by solving the cryo-EM structure of HCN4 in complex with ivabradine, to date the only HCN-specific drug on the market. Our data show ivabradine bound inside the open pore at 3 Å resolution. The structure unambiguously proves that Y507 and I511 on S6 are the molecular determinants of ivabradine binding to the inner cavity, while F510, pointing outside the pore, indirectly contributes to the block by controlling Y507. Cysteine 479, unique to the HCN selectivity filter (SF), accelerates the kinetics of block. Molecular dynamics simulations further reveal that ivabradine blocks the permeating ion inside the SF by electrostatic repulsion, a mechanism previously proposed for quaternary ammonium ions.
Topics: Ivabradine; Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; Molecular Dynamics Simulation; Humans; Cryoelectron Microscopy; Animals; Potassium Channels; Muscle Proteins
PubMed: 38917012
DOI: 10.1073/pnas.2402259121 -
Proceedings of the National Academy of... Jul 2024S100A1, a small homodimeric EF-hand Ca-binding protein (~21 kDa), plays an important regulatory role in Ca signaling pathways involved in various biological functions...
S100A1, a small homodimeric EF-hand Ca-binding protein (~21 kDa), plays an important regulatory role in Ca signaling pathways involved in various biological functions including Ca cycling and contractile performance in skeletal and cardiac myocytes. One key target of the S100A1 interactome is the ryanodine receptor (RyR), a huge homotetrameric Ca release channel (~2.3 MDa) of the sarcoplasmic reticulum. Here, we report cryoelectron microscopy structures of S100A1 bound to RyR1, the skeletal muscle isoform, in absence and presence of Ca. Ca-free apo-S100A1 binds beneath the bridging solenoid (BSol) and forms contacts with the junctional solenoid and the shell-core linker of RyR1. Upon Ca-binding, S100A1 undergoes a conformational change resulting in the exposure of the hydrophobic pocket known to serve as a major interaction site of S100A1. Through interactions of the hydrophobic pocket with RyR1, Ca-bound S100A1 intrudes deeper into the RyR1 structure beneath BSol than the apo-form and induces sideways motions of the C-terminal BSol region toward the adjacent RyR1 protomer resulting in tighter interprotomer contacts. Interestingly, the second hydrophobic pocket of the S100A1-dimer is largely exposed at the hydrophilic surface making it prone to interactions with the local environment, suggesting that S100A1 could be involved in forming larger heterocomplexes of RyRs with other protein partners. Since S100A1 interactions stabilizing BSol are implicated in the regulation of RyR-mediated Ca release, the characterization of the S100A1 binding site conserved between RyR isoforms may provide the structural basis for the development of therapeutic strategies regarding treatments of RyR-related disorders.
Topics: Ryanodine Receptor Calcium Release Channel; S100 Proteins; Calcium; Cryoelectron Microscopy; Animals; Protein Binding; Binding Sites; Models, Molecular; Protein Conformation; Humans
PubMed: 38917010
DOI: 10.1073/pnas.2400497121 -
Proceedings of the National Academy of... Jul 2024Spinal cord dorsal horn inhibition is critical to the processing of sensory inputs, and its impairment leads to mechanical allodynia. How this decreased inhibition...
Spinal cord dorsal horn inhibition is critical to the processing of sensory inputs, and its impairment leads to mechanical allodynia. How this decreased inhibition occurs and whether its restoration alleviates allodynic pain are poorly understood. Here, we show that a critical step in the loss of inhibitory tone is the change in the firing pattern of inhibitory parvalbumin (PV)-expressing neurons (PVNs). Our results show that PV, a calcium-binding protein, controls the firing activity of PVNs by enabling them to sustain high-frequency tonic firing patterns. Upon nerve injury, PVNs transition to adaptive firing and decrease their PV expression. Interestingly, decreased PV is necessary and sufficient for the development of mechanical allodynia and the transition of PVNs to adaptive firing. This transition of the firing pattern is due to the recruitment of calcium-activated potassium (SK) channels, and blocking them during chronic pain restores normal tonic firing and alleviates chronic pain. Our findings indicate that PV is essential for controlling the firing pattern of PVNs and for preventing allodynia. Developing approaches to manipulate these mechanisms may lead to different strategies for chronic pain relief.
Topics: Parvalbumins; Animals; Chronic Pain; Mice; Neurons; Hyperalgesia; Male; Action Potentials; Small-Conductance Calcium-Activated Potassium Channels
PubMed: 38916998
DOI: 10.1073/pnas.2403777121 -
Drug Design, Development and Therapy 2024This study probed the mechanism of action of Xinfeng Capsule (XFC) in myocardial injury in rats with adjuvant arthritis (AA) via the growth arrest-specific transcript 5...
PURPOSE
This study probed the mechanism of action of Xinfeng Capsule (XFC) in myocardial injury in rats with adjuvant arthritis (AA) via the growth arrest-specific transcript 5 (GAS5)/microRNA-21 (miR-21)/Toll-like receptor 4 (TLR4) axis.
METHODS
Rats were injected with Freund's complete adjuvant to establish a rat model of AA. Then, some modeled rats were given normal saline or drugs only, and some modeled rats were injected with adeno-associated viruses or necrosulfonamide (NSA; a pyroptosis inhibitor) before drug administration. Toe swelling and arthritis index (AI) were calculated. Pathological and morphological changes in synovial and myocardial tissues were analyzed with hematoxylin-eosin staining, and pyroptotic vesicles and the ultrastructural changes of myocardial tissues were observed with transmission electron microscopy. The serum levels of interleukin (IL)-1β, IL-18, IL-6, and tumor necrosis factor (TNF)-α were detected, and lactate dehydrogenase (LDH) release was measured in myocardial tissues, accompanied by the examination of GAS5, miR-21, TLR4, nuclear factor-kB (NF-κB) p65, nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), Caspase-1, and Gasdermin D (GSDMD) expression in myocardial tissues.
RESULTS
After AA modeling, rats presented with significantly increased toe swelling and AI scores, synovial and myocardial tissue damage, elevated pyroptotic vesicles, and markedly enhanced serum levels of IL-1β, IL-18, IL-6, and TNF-α, accompanied by significantly diminished GAS5 expression, substantially augmented miR-21, TLR4, NF-κB p65, NLRP3, Caspase-1, and GSDMD expression, greatly increased LDH release in myocardial tissues. XFC treatment significantly declined toe swelling, AI scores, synovial and myocardial tissue damage, and the serum levels of IL-1β, IL-18, IL-6, and TNF-α in AA rats. Additionally, XFC treatment markedly elevated GAS5 expression and substantially lowered LDH release and miR-21, TLR4, NF-κB p65, NLRP3, Caspase-1, and GSDMD expression in myocardial tissues of AA rats. Moreover, the above effects of XFC in AA rats were further promoted by GAS5 overexpression or NSA treatment.
CONCLUSION
XFC alleviated myocardial injury in AA rats by regulating the GAS5/miR-21/TLR4 axis and inhibiting pyroptosis and pro-inflammatory cytokine secretion.
Topics: Animals; Toll-Like Receptor 4; Pyroptosis; Rats; Arthritis, Experimental; MicroRNAs; Drugs, Chinese Herbal; Rats, Sprague-Dawley; Male; Phosphate-Binding Proteins; Freund's Adjuvant; Gasdermins
PubMed: 38915862
DOI: 10.2147/DDDT.S456783 -
BioRxiv : the Preprint Server For... Jun 2024Toxin-antidote systems are selfish genetic elements composed of a linked toxin and antidote. The toxin-antidote system in consists of a transmembrane toxin protein...
Toxin-antidote systems are selfish genetic elements composed of a linked toxin and antidote. The toxin-antidote system in consists of a transmembrane toxin protein PEEL-1 which acts cell autonomously to kill cells. Here we investigate the molecular mechanism of PEEL-1 toxicity. We find that PEEL-1 requires a small membrane protein, PMPL-1, for toxicity. Together, PEEL-1 and PMPL-1 are sufficient for toxicity in a heterologous system, HEK293T cells, and cause cell swelling and increased cell permeability to monovalent cations. Using purified proteins, we show that PEEL-1 and PMPL-1 allow ion flux through lipid bilayers and generate currents which resemble ion channel gating. Our work suggests that PEEL-1 kills cells by co-opting PMPL-1 and creating a cation channel.
PubMed: 38915716
DOI: 10.1101/2024.06.11.598564 -
BioRxiv : the Preprint Server For... Jun 2024The vestibular nerve is comprised of neuron sub-groups with diverse functions related to their intrinsic biophysical properties. This diversity is partly due to...
The vestibular nerve is comprised of neuron sub-groups with diverse functions related to their intrinsic biophysical properties. This diversity is partly due to differences in the types and numbers of low-voltage-gated potassium channels found in the neurons' membranes. Expression for some low-voltage gated ion channels like KCNQ4 is upregulated during early post-natal development; suggesting that ion channel composition and neuronal diversity may be shaped by hair cell activity. This idea is consistent with recent work showing that glutamatergic input from hair cells is necessary for the normal diversification auditory neurons. To test if biophysical diversity is similarly dependent on glutamatergic input in vestibular neurons, we examined the maturation of the vestibular epithelium and ganglion neurons in mice whose hair cell synapses lack glutamate. Despite lacking glutamatergic input, the knockout mice showed no notable balance deficits and crossed challenging balance beams with little difficulty. Immunolabeling of the vestibular epithelia showed normal development as indicated by an identifiable striolar zone with calyceal terminals labeled by molecular marker calretinin, and normal expression of KCNQ4 by the end of the second post-natal week. We found similar numbers of Type I and Type II hair cells in the knockout and wildtype animals, regardless of epithelial zone. Thus, the presumably quiescent Type II hair cells are not cleared from the epithelium. Patch-clamp recordings showed that biophysical diversity of vestibular ganglion neurons in the mice is comparable to that found in wildtype controls, with a similar range firing patterns at both immature and juvenile ages. However, our results suggest a subtle biophysical alteration to the largest ganglion cells (putative somata of central zone afferents); those in the knockout had smaller net conductance and were more excitable than those in the wild type. Thus, unlike in the auditory nerve, glutamatergic signaling is unnecessary for producing biophysical diversity in vestibular ganglion neurons. And yet, because the input signals from vestibular hair cells are complex and not solely reliant on quantal release of glutamate, whether diversity of vestibular ganglion neurons is simply hardwired or regulated by a more complex set of input signals remains to be determined.
PubMed: 38915604
DOI: 10.1101/2024.06.12.597464 -
BioRxiv : the Preprint Server For... Jun 2024Hypothalamic kisspeptin (Kiss1) neurons are vital for pubertal development and reproduction. Arcuate nucleus Kiss1 (Kiss1) neurons are responsible for the pulsatile...
Hypothalamic kisspeptin (Kiss1) neurons are vital for pubertal development and reproduction. Arcuate nucleus Kiss1 (Kiss1) neurons are responsible for the pulsatile release of Gonadotropin-releasing Hormone (GnRH). In females, the behavior of Kiss1 neurons, expressing Kiss1, Neurokinin B (NKB), and Dynorphin (Dyn), varies throughout the ovarian cycle. Studies indicate that 17β-estradiol (E2) reduces peptide expression but increases mRNA and glutamate neurotransmission in these neurons, suggesting a shift from peptidergic to glutamatergic signaling. To investigate this shift, we combined transcriptomics, electrophysiology, and mathematical modeling. Our results demonstrate that E2 treatment upregulates the mRNA expression of voltage-activated calcium channels, elevating the whole-cell calcium current and that contribute to high-frequency burst firing. Additionally, E2 treatment decreased the mRNA levels of Canonical Transient Receptor Potential (TPRC) 5 and G protein-coupled K (GIRK) channels. When TRPC5 channels in Kiss1 neurons were deleted using CRISPR, the slow excitatory postsynaptic potential (sEPSP) was eliminated. Our data enabled us to formulate a biophysically realistic mathematical model of the Kiss1 neuron, suggesting that E2 modifies ionic conductances in Kiss1 neurons, enabling the transition from high frequency synchronous firing through NKB-driven activation of TRPC5 channels to a short bursting mode facilitating glutamate release. In a low E2 milieu, synchronous firing of Kiss1 neurons drives pulsatile release of GnRH, while the transition to burst firing with high, preovulatory levels of E2 would facilitate the GnRH surge through its glutamatergic synaptic connection to preoptic Kiss1 neurons.
PubMed: 38915596
DOI: 10.1101/2024.02.20.581121 -
BioRxiv : the Preprint Server For... Jun 2024Purinergic P2X3 receptors form trimeric cation-gated channels, which are activated by extracellular ATP. P2X3 plays a crucial role in chronic cough and affects over 10%...
Purinergic P2X3 receptors form trimeric cation-gated channels, which are activated by extracellular ATP. P2X3 plays a crucial role in chronic cough and affects over 10% of the population. Despite considerable efforts to develop drugs targeting P2X3, the highly conserved structure within the P2X receptor family presents obstacles for achieving selectivity. Camlipixant, a potent and selective P2X3 antagonist, is currently in phase III clinical trials. However, the mechanisms underlying receptor desensitization, ion permeation, principles governing antagonism, and the structure of P2X3 when bound to camlipixant remain elusive. In this study, we established a stable cell line expressing homotrimeric P2X3 and utilized a peptide scaffold to purify the complex and determine its structure using cryo-electron microscopy (cryo-EM). P2X3 binds to camlipixant at a previously unidentified drug-binding site and functions as an allosteric inhibitor. Structure-activity studies combined with modeling and simulations have shed light on the mechanisms underlying the selective targeting and inhibition of P2X3 by camlipixant, distinguishing it from other members of the P2X receptor family.
PubMed: 38915546
DOI: 10.1101/2024.06.10.598171 -
BioRxiv : the Preprint Server For... Jun 2024Ion channels are essential for proper morphogenesis of the craniofacial skeleton. However, the molecular mechanisms underlying this phenomenon are unknown. Loss of the...
UNLABELLED
Ion channels are essential for proper morphogenesis of the craniofacial skeleton. However, the molecular mechanisms underlying this phenomenon are unknown. Loss of the potassium channel disrupts Bone Morphogenetic Protein (BMP) signaling within the developing palate. BMP signaling is essential for the correct development of several skeletal structures, including the palate, though little is known about the mechanisms that govern BMP secretion. We introduce a tool to image the release of bone morphogenetic protein 4 (BMP4) from mammalian cells. Using this tool, we show that depolarization induces BMP4 release from mouse embryonic palate mesenchyme cells in a calcium-dependent manner. We show native transient changes in intracellular calcium occur in cranial neural crest cells, the cells from which embryonic palate mesenchyme derives. Waves of transient changes in intracellular calcium suggest that these cells are electrically coupled and may temporally coordinate BMP release. These transient changes in intracellular calcium persist in palate mesenchyme cells from embryonic day (E) 9.5 to 13.5 mice. Disruption of significantly decreases the amplitude of calcium transients and the ability of cells to secrete BMP. Together, these data suggest that temporal control of developmental cues is regulated by ion channels, depolarization, and changes in intracellular calcium for mammalian craniofacial morphogenesis.
SUMMARY
We show that embryonic palate mesenchyme cells undergo transient changes in intracellular calcium. Depolarization of these cells induces BMP4 release suggesting that ion channels are a node in BMP4 signaling.
PubMed: 38915514
DOI: 10.1101/2024.06.11.598333