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Communications Biology Oct 2023Pyroptosis is a cell death process that causes inflammation and contributes to numerous diseases. Pyroptosis is mediated by caspase-1 family proteases that cleave the...
Pyroptosis is a cell death process that causes inflammation and contributes to numerous diseases. Pyroptosis is mediated by caspase-1 family proteases that cleave the pore-forming protein gasdermin D, causing plasma membrane rupture and release of pathogenic cellular contents. We previously identified muscimol as a small molecule that prevents plasma membrane rupture during pyroptosis via an unidentified mechanism. Here, we show that muscimol has reversible activity to prevent cellular lysis without affecting earlier pyroptotic events. Although muscimol is a well-characterized agonist for neuronal GABA receptors, muscimol protection is not altered by GABA receptor antagonists or recapitulated by other GABA agonists, suggesting that muscimol acts via a novel mechanism. We find that muscimol blocks oligomerization of ninjurin-1, which is required for plasma membrane rupture downstream of gasdermin D pore formation. Our structure-activity relationship studies reveal distinct molecular determinants defining inhibition of pyroptotic lysis compared to GABA binding. In addition, we demonstrate that muscimol reduces lethality during LPS-induced septic shock. Together, these findings demonstrate that ninjurin-1-mediated plasma membrane rupture can be pharmacologically modulated and pave the way toward identification of therapeutic strategies for pathologic conditions associated with pyroptosis.
Topics: Pyroptosis; Muscimol; Gasdermins; Intracellular Signaling Peptides and Proteins; Cell Membrane; Receptors, GABA-A; gamma-Aminobutyric Acid
PubMed: 37798443
DOI: 10.1038/s42003-023-05354-4 -
Reviews in the Neurosciences Aug 2020In this review, a series of experiments is presented, in which γ-amino butyric acid (GABA)ergic and glutamatergic effects on dopamine function in the rat nigrostriatal... (Review)
Review
In this review, a series of experiments is presented, in which γ-amino butyric acid (GABA)ergic and glutamatergic effects on dopamine function in the rat nigrostriatal and mesolimbic system was systematically assessed after pharmacological challenge with GABAA receptor (R) and and N-methyl d-aspartate (NMDA)R agonists and antagonists. In these studies, [123I]iodobenzamide binding to the D2/3R was mesured in nucleus accumbens (NAC), caudateputamen (CP), substantia nigra/ventral tegmental area (SN/VTA), frontal (FC), motor (MC) and parietal cortex (PC) as well as anterior (aHIPP) and posterior hippocampus (pHIPP) with small animal SPECT in baseline and after injection of either the GABAAR agonist muscimol (1 mg/kg), the GABAAR antagonist bicuculline (1 mg/kg), the NMDAR agonist d-cycloserine (20 mg/kg) or the NMDAR antagonist amantadine (40 mg/kg). Muscimol reduced D2/3R binding in NAC, CP, SN/VTA, THAL and pHIPP, while, after amantadine, decreases were confined to NAC, CP and THAL. In contrast, d-cycloserine elevated D2/3R binding in NAC, SN/VTA, THAL, frontal cortex, motor cortex, PC, aHIPP and pHIPP, while, after bicuculline, increases were confined to CP and THAL. Taken together, similar actions on regional dopamine levels were exterted by the GABAAR agonist and the NMDAR antagonist on the one side and by the GABAAR antagonist and the NMDAR agonist on the other, with agonistic action, however, affecting more brain regions. Thereby, network analysis suggests different roles of GABAARs and NMDARs in the mediation of nigrostriatal, nigrothalamocortical and mesolimbocortical dopamine function.
Topics: Animals; Bicuculline; Dopamine; Humans; Muscimol; Nucleus Accumbens; Rats; Receptors, GABA-A; Receptors, N-Methyl-D-Aspartate
PubMed: 32619197
DOI: 10.1515/revneuro-2019-0112 -
Molecular Pharmacology Oct 2020Muscimol is a psychoactive isoxazole derived from the mushroom and a potent orthosteric agonist of the GABA receptor. The binding of [H]muscimol has been used to...
Muscimol is a psychoactive isoxazole derived from the mushroom and a potent orthosteric agonist of the GABA receptor. The binding of [H]muscimol has been used to evaluate the distribution of GABA receptors in the brain, and studies of modulation of [H]muscimol binding by allosteric GABAergic modulators such as barbiturates and steroid anesthetics have provided insight into the modes of action of these drugs on the GABA receptor. It has, however, not been feasible to directly apply interaction parameters derived from functional studies to describe the binding of muscimol to the receptor. Here, we employed the Monod-Wyman-Changeux concerted transition model to analyze muscimol binding isotherms. We show that the binding isotherms from recombinant 13 GABA receptors can be qualitatively predicted using electrophysiological data pertaining to properties of receptor activation and desensitization in the presence of muscimol. The model predicts enhancement of [H]muscimol binding in the presence of the steroids allopregnanolone and pregnenolone sulfate, although the steroids interact with distinct sites and either enhance (allopregnanolone) or reduce (pregnenolone sulfate) receptor function. We infer that the concerted transition model can be used to link radioligand binding and electrophysiological data. SIGNIFICANCE STATEMENT: The study employs a three-state resting-active-desensitized model to link radioligand binding and electrophysiological data. We show that the binding isotherms can be qualitatively predicted using parameters estimated in electrophysiological experiments and that the model accurately predicts the enhancement of [H]muscimol binding in the presence of the potentiating steroid allopregnanolone and the inhibitory steroid pregnenolone sulfate.
Topics: Allosteric Regulation; Binding Sites; GABA-A Receptor Agonists; HEK293 Cells; Humans; Multiprotein Complexes; Muscimol; Pregnanolone; Pregnenolone; Receptors, GABA-A; Recombinant Proteins; Steroids; Tritium
PubMed: 32873746
DOI: 10.1124/molpharm.120.000066 -
Brain and Behavior May 2020Due to side effects of medications used for chronic pain, combination therapy seems to be an appropriate solution for alleviation of chronic pain and reducing the side...
INTRODUCTION
Due to side effects of medications used for chronic pain, combination therapy seems to be an appropriate solution for alleviation of chronic pain and reducing the side effects. The role of inhibitory GABA system is well proven in reducing neuropathic pain. Also, special attention has been focused on endogenous morphine (endomorphins) in reducing chronic pain originates from damage to the nervous system. The purpose of this study is to investigate the analgesic effect of simultaneous administration of GABA agonist and endomorphin-1 on neuropathic pain in rat model of spinal cord injury (SCI). The role of oxidative stress, NR1 subunits of NMDA receptors, and α subunits of GABA receptors in the spinal cord has also been investigated.
METHODS
Spinal cord at level of T6-T8 was compressed. Three weeks after spinal cord injury, muscimol and endomorphin-1 were injected (intrathecally once a day for 7 days) individually or in combination. Mechanical and cold allodynia, thermal and mechanical hyperalgesia were evaluated before injection and 15 and 60 min after injection. At the end of behavioral experiments, histological and biochemical evaluations were done on prepared spinal cord samples.
RESULTS
Isobologram results showed that combination therapy significantly increased the pain threshold comparing to injection of endomorphin-1 (EM) or muscimol alone. Histological studies indicated the increased expression of α2 subunits of GABA receptors, and NR1 subunits of NMDA receptors in the spinal cord. The combination therapy also increased the glutathione (GSH) and superoxide dismutase (SOD) level and decreased the malondialdehyde (MDA) levels in the spinal cord.
CONCLUSION
Simultaneous administration of muscimol and endomorphine-1 could be a new candidate for alleviation of pain resulting from spinal cord injury.
Topics: Animals; Hyperalgesia; Injections, Spinal; Muscimol; Neuralgia; Oligopeptides; Rats; Spinal Cord; Spinal Cord Injuries
PubMed: 32189472
DOI: 10.1002/brb3.1576 -
ACS Chemical Neuroscience Mar 2013GABAA receptors are ligand-gated ion channels that mediate inhibitory synaptic signaling in the CNS. Fluorescent probes with the ability to target these receptors can...
GABAA receptors are ligand-gated ion channels that mediate inhibitory synaptic signaling in the CNS. Fluorescent probes with the ability to target these receptors can provide insights into receptor location, distribution and dynamics in live cells, while revealing abnormalities in their distribution and dynamics that could occur in a variety of diseases. We have developed fluorescent probes of GABAA receptors that are composed of a CdSe/ZnS core-shell nanocrystal (quantum dot; qdot) conjugated to pegylated derivatives of the GABA receptor agonists GABA and muscimol (GABA-qdots and muscimol-qdots, respectively). Quantitative fluorescence imaging was used to analyze the binding activity of these conjugates to α1β2γ2 GABAA and ρ1 GABAA receptors expressed in Xenopus oocytes. The selectivity of these conjugates for α1β2γ2 GABAA and ρ1 GABAA receptors was determined by their ability to compete with the antagonists bicuculline and methyl-(1,2,3,6-tetrahydropyridin-4-yl)phosphinic acid (TPMPA). Both GABA- and muscimol-qdots exhibited robust binding to both α1β2γ2 and ρ1 GABAA receptors. At α1β2γ2 receptors, pretreatment with bicuculline reduced conjugate binding by ≥8-fold on average, an extent far exceeding the reduction produced by TPMPA (~30%). Conversely, at ρ1 receptors, pretreatment with TPMPA inhibited binding by ~10-fold, an extent greatly exceeding the change produced by bicuculline (~50% or less). These results indicate specific binding of muscimol-qdots and GABA-qdots to α1β2γ2 GABAA and ρ1 GABAA receptors in a manner that preserves the respective pharmacological sensitivities of these receptors to TPMPA and bicuculline, and encourage the use of qdot-conjugated neurotransmitter analogs as labeling agents at GABAA receptors.
Topics: Animals; Binding Sites; Female; Humans; Muscimol; Protein Binding; Protein Subunits; Quantum Dots; Receptors, GABA-A; Receptors, GABA-B; Xenopus laevis; gamma-Aminobutyric Acid
PubMed: 23509979
DOI: 10.1021/cn300144v -
The Journal of Biological Chemistry Sep 2022Anthelmintics are used to treat human and veterinary parasitic diseases and to reduce crop and livestock production loss associated with parasitosis. The free-living...
Anthelmintics are used to treat human and veterinary parasitic diseases and to reduce crop and livestock production loss associated with parasitosis. The free-living nematode Caenorhabditis elegans, a model system for anthelmintic drug discovery, has a serotonin (5-HT)-gated chloride channel, MOD-1, which belongs to the Cys-loop receptor family and modulates locomotory and behavioral functions. Since MOD-1 is unique to nematodes, it is emerging as an attractive anthelmintic drug target, but details of MOD-1 function are unclear. Here, we revealed novel aspects of MOD-1 function from the molecular level to the organism level and identified compounds targeting this receptor, which may provide new directions for anthelmintic drug discovery. We used whole-cell current recordings from heterologously expressed MOD-1 to show that tryptamine (Tryp), a weak partial agonist of vertebrate serotonin type 3 (5-HT) receptors, efficaciously activates MOD-1. A screen for modulators revealed that GABAergic ligands piperazine (PZE) and muscimol reduce 5-HT-elicited currents, thus identifying novel MOD-1 allosteric inhibitors. Next, we performed locomotor activity assays, and we found 5-HT and Tryp rapidly decrease worm motility, which is reversible only at low 5-HT concentrations. Mutants lacking MOD-1 are partially resistant to both drugs, demonstrating its role in locomotion. Acting as an antagonist of MOD-1, we showed PZE reduces the locomotor effects of exogenous 5-HT. Therefore, Tryp- and PZE-derived compounds, acting at MOD-1 through different molecular mechanisms, emerge as promising anthelmintic agents. This study enhances our knowledge of the function and drug selectivity of Cys-loop receptors and postulates MOD-1 as a potential target for anthelmintic therapy.
Topics: Animals; Anthelmintics; Caenorhabditis elegans; Chloride Channels; Cysteine Loop Ligand-Gated Ion Channel Receptors; Humans; Muscimol; Nematoda; Piperazines; Serotonin
PubMed: 35952761
DOI: 10.1016/j.jbc.2022.102356 -
PloS One 2022Previous studies have shown that spontaneously active cultured networks of cortical neuron grown planar microelectrode arrays are sensitive to radiofrequency (RF) fields...
Previous studies have shown that spontaneously active cultured networks of cortical neuron grown planar microelectrode arrays are sensitive to radiofrequency (RF) fields and exhibit an inhibitory response more pronounced as the exposure time and power increase. To better understand the mechanism behind the observed effects, we aimed at identifying similarities and differences between the inhibitory effect of RF fields (continuous wave, 1800 MHz) to the γ-aminobutyric acid type A (GABAA) receptor agonist muscimol (MU). Inhibition of the network bursting activity in response to RF exposure became apparent at an SAR level of 28.6 W/kg and co-occurred with an elevation of the culture medium temperature of ~1°C. Exposure to RF fields preferentially inhibits bursting over spiking activity and exerts fewer constraints on neural network bursting synchrony, differentiating it from a pharmacological inhibition with MU. Network rebound excitation, a phenomenon relying on the intrinsic properties of cortical neurons, was observed following the removal of tonic hyperpolarization after washout of MU but not in response to cessation of RF exposure. This implies that hyperpolarization is not the main driving force mediating the inhibitory effects of RF fields. At the level of single neurons, network inhibition induced by MU and RF fields occurred with reduced action potential (AP) half-width. As changes in AP waveform strongly influence efficacy of synaptic transmission, the narrowing effect on AP seen under RF exposure might contribute to reducing network bursting activity. By pointing only to a partial overlap between the inhibitory hallmarks of these two forms of inhibition, our data suggest that the inhibitory mechanisms of the action of RF fields differ from the ones mediated by the activation of GABAA receptors.
Topics: Action Potentials; Muscimol; Neurons; Radio Waves; Synaptic Transmission
PubMed: 36044461
DOI: 10.1371/journal.pone.0268605 -
Epilepsy Research Feb 2023Intracerebral drug delivery is an emerging treatment strategy aiming to manage seizures in patients with systemic drug-resistant epilepsies. In rat seizure and epilepsy...
Intracerebral drug delivery is an emerging treatment strategy aiming to manage seizures in patients with systemic drug-resistant epilepsies. In rat seizure and epilepsy models, the GABA receptor agonist muscimol has shown powerful antiseizure potential when injected acutely into the subthalamic nucleus (STN), known for its capacity to provide remote control of different seizure types. However, chronic intrasubthalamic muscimol delivery required for long-term seizure suppression has not yet been investigated. We tested the hypothesis that chronic convection-enhanced delivery (CED) of muscimol into the STN produces long-lasting antiseizure effects in the intravenous pentylenetetrazole seizure threshold test in female rats. Acute microinjection was included to verify efficacy of intrasubthalamic muscimol delivery in this seizure model and caused significant antiseizure effects at 30 and 60 ng per hemisphere with a dose-dependent increase of responders and efficacy and only mild adverse effects compared to controls. For the chronic study, muscimol was bilaterally infused into the STN over three weeks at daily doses of 60, 300, or 600 ng per hemisphere using an implantable pump and cannula system. Chronic intrasubthalamic CED of muscimol caused significant long-lasting antiseizure effects for up to three weeks at 300 and 600 ng daily. Drug responder rate increased dose-dependently, as did drug tolerance rates. Transient ataxia and body weight loss were the main adverse effects. Drug distribution was comparable (about 2-3 mm) between acute and chronic delivery. This is the first study providing proof-of-concept that not only acute, but also chronic, continuous CED of muscimol into the STN raises seizure thresholds.
Topics: Rats; Female; Animals; Subthalamic Nucleus; Muscimol; Convection; Epilepsy; Seizures
PubMed: 36736200
DOI: 10.1016/j.eplepsyres.2023.107097 -
Convection-Enhanced Delivery of Muscimol Into the Bilateral Subthalamic Nuclei of Nonhuman Primates.Neurosurgery Jun 2019Muscimol is a gamma-aminobutyric acid receptor agonist that selectively and temporarily inhibits neurons. Local bolus injection of muscimol has been used experimentally...
BACKGROUND
Muscimol is a gamma-aminobutyric acid receptor agonist that selectively and temporarily inhibits neurons. Local bolus injection of muscimol has been used experimentally to inhibit neuronal populations within discrete anatomical structures and discern their physiological function.
OBJECTIVE
To determine the safety and behavioral effects of convection-enhanced delivery of muscimol into the bilateral subthalamic nuclei (STN) of nonhuman primate rhesus macaques (NHPs).
METHODS
Six awake NHPs underwent co-infusion of gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA), a surrogate magnetic resonance imaging (MRI) tracer, with increasing concentrations of muscimol for behavioral and histological assessment. Three other NHPs were co-infused with Gd-DTPA and 3H-muscimol into the STN to determine muscimol distribution by MRI and autoradiography. Two NHPs underwent microcatheter implantation without muscimol infusion for control comparison.
RESULTS
MRI revealed selective and complete perfusion of the bilateral STN in animals infused with Gd-DTPA and muscimol. No abnormal movements occurred at 0.125 mM. Muscimol doses between 0.25 and 4.4 mM resulted in transient, dose-dependent hyperkinesia. Muscimol (8.8 mM) resulted in severe bilateral dyskinesias, ballistic movements, and sedation. An 88.8 mM dose produced unresponsiveness in 1 animal. Infusion-related pathological abnormities or toxicity was not present on histological examination. MRI distribution of co-infused Gd-DTPA was similar to autoradiographic distribution of 3H-muscimol (Vd; R = 0.94). Mean Vd of infused animals was 37.9 mm3 ± 11.7 mm3 and mean Vd: Vi 7.6 ± 2.3.
CONCLUSION
Bilateral convection-enhanced delivery of muscimol into the primate STN resulted in dose-related hyperkinetic movements that resolved after stopping the infusion. Muscimol was not toxic to brain tissue. Gd-DTPA accurately tracked muscimol distribution.
Topics: Animals; Contrast Media; Convection; Female; GABA-A Receptor Agonists; Gadolinium DTPA; Macaca mulatta; Magnetic Resonance Imaging; Male; Muscimol; Subthalamic Nucleus
PubMed: 29931364
DOI: 10.1093/neuros/nyy279 -
International Journal of Molecular... Aug 2021The two-pore domain K (K) channel, which is involved in setting the resting membrane potential in neurons, is an essential target for receptor agonists. Activation of...
The two-pore domain K (K) channel, which is involved in setting the resting membrane potential in neurons, is an essential target for receptor agonists. Activation of the γ-aminobutyric acid (GABA) receptors (GABAR and GABAR) reduces cellular excitability through Cl influx and K efflux in neurons. Relatively little is known about the link between GABAR and the K channel. The present study was performed to identify the effect of GABAR agonists on K channel expression and activity in the neuroblastic B35 cells that maintain glutamic acid decarboxylase (GAD) activity and express GABA. TASK and TREK/TRAAK mRNA were expressed in B35 cells with a high level of TREK-2 and TRAAK. In addition, TREK/TRAAK proteins were detected in the GABAergic neurons obtained from GABA transgenic mice. Furthermore, TREK-2 mRNA and protein expression levels were markedly upregulated in B35 cells by GABAR and GABAR agonists. In particular, muscimol, a GABAR agonist, significantly increased TREK-2 expression and activity, but the effect was reduced in the presence of the GABAR antagonist bicuculine or TREK-2 inhibitor norfluoxetine. In the whole-cell and single-channel patch configurations, muscimol increased TREK-2 activity, but the muscimol effect disappeared in the N-terminal deletion mutant. These results indicate that muscimol directly induces TREK-2 activation through the N-terminus and suggest that muscimol can reduce cellular excitability by activating the TREK-2 channel and by inducing Cl influx in GABAergic neurons.
Topics: Animals; Cells, Cultured; GABA-A Receptor Agonists; GABAergic Neurons; HEK293 Cells; Humans; Male; Membrane Potentials; Mice; Muscimol; Potassium Channels, Tandem Pore Domain; Rats; Receptors, GABA
PubMed: 34502229
DOI: 10.3390/ijms22179320