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1.
Exp Neurol ; 347: 113881, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34597681

RESUMO

Hypo-excitability was reported in the peri-infarct tissue following stroke, an effect counteracted by a blockage of α5-GABAA receptors in adult rodents. Our present study aims to evaluate the effect of a selective α5-GABAA receptor antagonist, S 44819, in stroke in juvenile animals. We have set up and characterized an original model of transient ischemic stroke in 28 day-old Sprague-Dawley rats (45-min occlusion of the middle cerebral artery by intraluminal suture). In this model, S 44819 (1, 3 and 10 mg/kg, b.i.d) was orally administered from day 3 to day 16 after stroke onset. Sensorimotor recovery was assessed on day 1, day 9 and day 16 after stroke onset. Results show that rats treated with S 44819 at the doses of 3 and 10 mg/kg displayed a significant improvement of the neurological deficits (neuroscore) on day 9 and day 16, when compared with animals treated with vehicle. Grip-test data analysis reveals that rats treated with S 44819 at the dose of 3 mg/kg displayed a better recovery on day 9 and day 16. These results are in agreement with those previously observed in adult rats, demonstrating that targeting α5-GABAA receptors improves neurological recovery after stroke in juvenile rats.


Assuntos
Benzodiazepinas/farmacologia , Antagonistas de Receptores de GABA-A/farmacologia , Oxazóis/farmacologia , Recuperação de Função Fisiológica/efeitos dos fármacos , Acidente Vascular Cerebral/fisiopatologia , Animais , Masculino , Ratos , Ratos Sprague-Dawley
2.
Biomolecules ; 11(7)2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34356653

RESUMO

Rosmarinic acid, a major component of rosemary, is a polyphenolic compound with potential neuroprotective effects. Asreducing the synaptic release of glutamate is crucial to achieving neuroprotectant's pharmacotherapeutic effects, the effect of rosmarinic acid on glutamate release was investigated in rat cerebrocortical nerve terminals (synaptosomes). Rosmarinic acid depressed the 4-aminopyridine (4-AP)-induced glutamate release in a concentration-dependent manner. The removal of extracellular calcium and the blockade of vesicular transporters prevented the inhibition of glutamate release by rosmarinic acid. Rosmarinic acid reduced 4-AP-induced intrasynaptosomal Ca2+ elevation. The inhibition of N-, P/Q-type Ca2+ channels and the calcium/calmodulin-dependent kinase II (CaMKII) prevented rosmarinic acid from having effects on glutamate release. Rosmarinic acid also reduced the 4-AP-induced activation of CaMKII and the subsequent phosphorylation of synapsin I, the main presynaptic target of CaMKII. In addition, immunocytochemistry confirmed the presence of GABAA receptors. GABAA receptor agonist and antagonist blocked the inhibitory effect of rosmarinic acid on 4-AP-evoked glutamate release. Docking data also revealed that rosmarinic acid formed a hydrogen bond with the amino acid residues of GABAA receptor. These results suggested that rosmarinic acid activates GABAA receptors in cerebrocortical synaptosomes to decrease Ca2+ influx and CaMKII/synapsin I pathway to inhibit the evoked glutamate release.


Assuntos
Cinamatos/farmacologia , Depsídeos/farmacologia , Ácido Glutâmico/metabolismo , Sinaptossomos/efeitos dos fármacos , 4-Aminopiridina/farmacologia , Animais , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Cinamatos/química , Depsídeos/química , Agonistas de Receptores de GABA-A/farmacologia , Antagonistas de Receptores de GABA-A/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Simulação de Acoplamento Molecular , Fármacos Neuroprotetores/farmacologia , Fosforilação/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Ratos Sprague-Dawley , Receptores de GABA-A/química , Receptores de GABA-A/metabolismo , Sinaptossomos/metabolismo
3.
J Neurosci ; 41(39): 8103-8110, 2021 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-34385360

RESUMO

Entorhinal cortex neurons make monosynaptic connections onto distal apical dendrites of CA1 and CA2 pyramidal neurons through the perforant path (PP) projection. Previous studies show that differences in dendritic properties and synaptic input density enable the PP inputs to produce a much stronger excitation of CA2 compared with CA1 pyramidal neurons. Here, using mice of both sexes, we report that the difference in PP efficacy varies substantially as a function of presynaptic firing rate. Although a single PP stimulus evokes a 5- to 6-fold greater EPSP in CA2 compared with CA1, a brief high-frequency train of PP stimuli evokes a strongly facilitating postsynaptic response in CA1, with relatively little change in CA2. Furthermore, we demonstrate that blockade of NMDARs significantly reduces strong temporal summation in CA1 but has little impact on that in CA2. As a result of the differences in the frequency- and NMDAR-dependent temporal summation, naturalistic patterns of presynaptic activity evoke CA1 and CA2 responses with distinct dynamics, differentially tuning CA1 and CA2 responses to bursts of presynaptic firing versus single presynaptic spikes, respectively.SIGNIFICANCE STATEMENT Recent studies have demonstrated that abundant entorhinal cortical innervation and efficient dendritic propagation enable hippocampal CA2 pyramidal neurons to produce robust excitation evoked by single cortical stimuli, compared with CA1. Here we uncovered, unexpectedly, that the difference in efficacy of cortical excitation varies substantially as a function of presynaptic firing rate. A burst of stimuli evokes a strongly facilitating response in CA1, but not in CA2. As a result, the postsynaptic response of CA1 and CA2 to presynaptic naturalistic firing displays contrasting temporal dynamics, which depends on the activation of NMDARs. Thus, whereas CA2 responds to single stimuli, CA1 is selectively recruited by bursts of cortical input.


Assuntos
Região CA1 Hipocampal/fisiologia , Região CA2 Hipocampal/fisiologia , Córtex Cerebral/fisiologia , Potenciais Pós-Sinápticos Excitadores/fisiologia , Células Piramidais/fisiologia , Sinapses/fisiologia , Animais , Região CA1 Hipocampal/efeitos dos fármacos , Região CA2 Hipocampal/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Feminino , Antagonistas de Receptores de GABA-A/farmacologia , Antagonistas de Receptores de GABA-B/farmacologia , Masculino , Camundongos , Vias Neurais/efeitos dos fármacos , Vias Neurais/fisiologia , Técnicas de Patch-Clamp , Células Piramidais/efeitos dos fármacos , Sinapses/efeitos dos fármacos
4.
Int J Mol Sci ; 22(13)2021 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-34281255

RESUMO

Midazolam (MDZ) could affect lymphocyte immune functions. However, the influence of MDZ on cell's K+ currents has never been investigated. Thus, in the present study, the effects of MDZ on Jurkat T lymphocytes were studied using the patch-clamp technique. Results showed that MDZ suppressed the amplitude of delayed-rectifier K+ current (IK(DR)) in concentration-, time-, and state-dependent manners. The IC50 for MDZ-mediated reduction of IK(DR) density was 5.87 µM. Increasing MDZ concentration raised the rate of current-density inactivation and its inhibitory action on IK(DR) density was estimated with a dissociation constant of 5.14 µM. In addition, the inactivation curve of IK(DR) associated with MDZ was shifted to a hyperpolarized potential with no change on the slope factor. MDZ-induced inhibition of IK(DR) was not reversed by flumazenil. In addition, the activity of intermediate-conductance Ca2+-activated K+ (IKCa) channels was suppressed by MDZ. Furthermore, inhibition by MDZ on both IK(DR) and IKCa-channel activity appeared to be independent from GABAA receptors and affected immune-regulating cytokine expression in LPS/PMA-treated human T lymphocytes. In conclusion, MDZ suppressed current density of IK(DR) in concentration-, time-, and state-dependent manners in Jurkat T-lymphocytes and affected immune-regulating cytokine expression in LPS/PMA-treated human T lymphocytes.


Assuntos
Canais de Potássio de Retificação Tardia/antagonistas & inibidores , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/antagonistas & inibidores , Midazolam/farmacologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo , Animais , Citocinas/metabolismo , Canais de Potássio de Retificação Tardia/metabolismo , Relação Dose-Resposta a Droga , Flumazenil/farmacologia , Antagonistas de Receptores de GABA-A/farmacologia , Humanos , Hipnóticos e Sedativos/administração & dosagem , Hipnóticos e Sedativos/farmacologia , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/metabolismo , Células Jurkat , Cinética , Lipopolissacarídeos/farmacologia , Ativação Linfocitária , Microscopia Confocal , Midazolam/administração & dosagem , Técnicas de Patch-Clamp , Fito-Hemaglutininas/farmacologia , Linfócitos T/imunologia
5.
J Neurochem ; 159(1): 101-115, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34263932

RESUMO

Naturally occurring compounds such as sesquiterpenes and sesquiterpenoids (SQTs) have been shown to modulate GABAA receptors (GABAA Rs). In this study, the modulatory potential of 11 SQTs at GABAA Rs was analyzed to characterize their potential neurotropic activity. Transfected HEK293 cells and primary hippocampal neurons were functionally investigated using electrophysiological whole-cell recordings. Significantly different effects of ß-caryophyllene and α-humulene, as well as their respective derivatives ß-caryolanol and humulol, were observed in the HEK293 cell system. In neurons, the concomitant presence of phasic and tonic GABAA R configurations accounts for differences in receptor modulation by SQTs. The in vivo presence of the γ2 and δ subunits is important for SQT modulation. While phasic GABAA receptors in hippocampal neurons exhibited significantly altered GABA-evoked current amplitudes in the presence of humulol and guaiol, negative allosteric potential at recombinantly expressed α1 ß2 γ2 receptors was only verified for humolol. Modeling and docking studies provided support for the binding of SQTs to the neurosteroid-binding site of the GABAA R localized between transmembrane segments 1 and 3 at the (+ α)-(- α) interface. In sum, differences in the modulation of GABAA R isoforms between SQTs were identified. Another finding is that our results provide an indication that nutritional digestion affects the neurotropic potential of natural compounds.


Assuntos
Antagonistas de Receptores de GABA-A/farmacologia , Simulação de Acoplamento Molecular/métodos , Extratos Vegetais/farmacologia , Receptores de GABA-A/fisiologia , Sesquiterpenos/farmacologia , Regulação Alostérica/efeitos dos fármacos , Regulação Alostérica/fisiologia , Animais , Feminino , Antagonistas de Receptores de GABA-A/química , Antagonistas de Receptores de GABA-A/isolamento & purificação , Células HEK293 , Humanos , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Gravidez , Receptores de GABA-A/química , Sesquiterpenos/química , Sesquiterpenos/isolamento & purificação
6.
Front Immunol ; 12: 670153, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34135897

RESUMO

Background: Myocardial macrophages have key roles in cardiac remodeling and dysfunction. The gamma-aminobutyric acid subtype A (GABAA) receptor was recently found to be distributed in macrophages, allowing regulation of inflammatory responses to various diseases. This study aimed to clarify the role of GABAA receptor-mediated macrophage responses in pressure overload-induced heart failure. Methods and Results: C57BL/6J mice underwent transverse aortic constriction for pressure-overload hypertrophy (POH) and were intraperitoneally treated with a specific GABAA receptor agonist (topiramate) or antagonist (bicuculline). Echocardiography, histology, and flow cytometry were performed to evaluate the causes and effects of myocardial hypertrophy and fibrosis. Activation of the GABAA receptor by topiramate reduced ejection fraction and fractional shortening, enlarged the end-diastolic and end-systolic left ventricular internal diameter, aggravated myocardial hypertrophy and fibrosis, and accelerated heart failure in response to pressure overload. Mechanistically, topiramate increased the number of Ly6Clow macrophages in the heart during POH and circulating Ly6Chigh classic monocyte infiltration in late-phase POH. Further, topiramate drove Ly6Clow macrophages toward MHCIIhigh macrophage polarization. As a result, Ly6Clow macrophages activated the amphiregulin-induced AKT/mTOR signaling pathway, and Ly6ClowMHCIIhigh macrophage polarization increased expression levels of osteopontin and TGF-ß, which led to myocardial hypertrophy and fibrosis. Conversely, GABAA receptor blockage with bicuculline reversed these effects. Conclusions: Control of the GABAA receptor activity in monocytes/macrophages plays an important role in myocardial hypertrophy and fibrosis after POH. Blockade of the GABAA receptor has the potential to improve pressure overload-induced heart failure.


Assuntos
Antagonistas de Receptores de GABA-A/farmacologia , Insuficiência Cardíaca/tratamento farmacológico , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Macrófagos/efeitos dos fármacos , Receptores de GABA-A/efeitos dos fármacos , Animais , Aorta/fisiopatologia , Aorta/cirurgia , Pressão Arterial , Modelos Animais de Doenças , Fibrose , Insuficiência Cardíaca/etiologia , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Hipertrofia Ventricular Esquerda/etiologia , Hipertrofia Ventricular Esquerda/metabolismo , Hipertrofia Ventricular Esquerda/fisiopatologia , Ligadura , Macrófagos/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , Miocárdio/patologia , Receptores de GABA-A/metabolismo , Transdução de Sinais , Função Ventricular Esquerda/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacos
7.
World Neurosurg ; 153: e168-e178, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34166824

RESUMO

PURPOSE: To study the 24-hour expression of long noncoding RNAs (lncRNAs) in synaptic and extrasynaptic neurons expressing N-methyl-D-aspartate receptor (NMDAR), and normal neuronal cultures, via microarray analysis. MATERIALS AND METHODS: Cortical neurons from embryonic (day E18) Sprague-Dawley rats were used for primary neuronal culture. NMDAR activation was blocked and the cells were then incubated for 6 hours. Total RNA was extracted, quantified, and analyzed for purity and integrity. Double-stranded cDNA was synthesized, followed by quantile normalization, quantitative polymerase chain reaction validation, and data analysis. The interactions between transcription factors and lncRNAs were analyzed by Pearson correlation. RESULTS: The lncRNA profiles were obtained after synaptic and extrasynaptic NMDAR activation of rat cortical neuron cultures for 24 hours. In total, 251 lncRNAs were consistently upregulated, and 335 were downregulated, after extrasynaptic NMDAR activation compared with normal neurons. After synaptic NMDAR activation, only 9 lncRNAs were upregulated and 2 were downregulated. CONCLUSIONS: Differential expression of lncRNAs after synaptic and extrasynaptic NMDAR activation suggests that lncRNAs may be responsible for extrasynaptic NMDAR-induced neurodegeneration.


Assuntos
Neurônios/metabolismo , RNA Longo não Codificante/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapses/metabolismo , 4-Aminopiridina/farmacologia , Animais , Bicuculina/farmacologia , Córtex Cerebral/citologia , Regulação para Baixo , Agonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Receptores de GABA-A/farmacologia , Glicina/farmacologia , Glicinérgicos/farmacologia , Análise em Microsséries , N-Metilaspartato/farmacologia , Reação em Cadeia da Polimerase , Bloqueadores dos Canais de Potássio/farmacologia , Cultura Primária de Células , Ratos , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Sinapses/efeitos dos fármacos , Regulação para Cima
8.
Neuroscience ; 467: 1-15, 2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34033871

RESUMO

Stimulation with a low frequency electromagnetic field (LF-EMF) has proven to represent a powerful method for the suppression of seizures, as demonstrated in select clinical and laboratory studies. However, the mechanism by which LF-EMF suppresses seizures remains unclear. The purpose of the present study was to explore the modulatory effect of LF-EMF on epileptiform discharges (EDs) using rat hippocampal slices and investigate the underlying mechanisms that mediate these effects. EDs in hippocampal slices was induced by magnesium-free (zero-Mg2+) artificial cerebrospinal fluid (ACSF) and recorded using an in vitro micro-electrode array (MEA). A small sub-decimeter coil was designed and incorporated in a flexible magnetic stimulation device that allowed electromagnetic fields with different parameters to be delivered to slices. After a stable ED event was recorded, magnetic fields of 0.5 Hz (30 min) with a magnetic intensity of 0.13 mT (5 Vpp voltage input) and 0.25 mT (20 Vpp voltage input) were applied. The results indicated that a high-amplitude 0.5 Hz magnetic field could lead to persistent suppression of ictal discharges (IDs), while low-amplitude magnetic fields did not influence IDs. The persistent suppression of complex ED was prevented if the magnetic fields were applied in the presence of 10 µmol/L bicuculline (BIC), a γ-aminobutyric acid type A (GABAA) receptor antagonist, while the application of BIC subsequent to a magnetic field application led to the reappearance of ID. The addition of BIC resulted in EDs that had previously been inhibited by magnetic fields, reappearing. Low-frequency magnetic stimulation was able to inhibit the conversion from interictal discharges (IIDs) or preictal discharges (PIDs) to IDs. This suppression was attributed to the modulation of GABAA receptor activity.


Assuntos
Hipocampo , Convulsões , Animais , Bicuculina/farmacologia , Antagonistas de Receptores de GABA-A , Campos Magnéticos , Ratos
9.
Eur J Pharmacol ; 904: 174195, 2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34004209

RESUMO

Benzodiazepine receptor agonists are widely prescribed therapeutic agents that alter gamma-aminobutyric acid (GABA)A receptor activity and have anxiolytic effects. Post-operative use of benzodiazepines is a risk factor of delirium. Inflammatory conditions alter the anxiolytic effects of benzodiazepine. We investigated the effect of diazepam, a typical benzodiazepine anxiolytic, on changes in the emotional behavior of mice in a hole-board test after lipopolysaccharide (LPS) treatment. Diazepam dose-dependently increased the number of head-dips at doses that did not alter locomotor activity; however, diazepam dose-dependently significantly decreased the number of head-dips at doses that did not alter locomotor activity in LPS-treated mice. Flumazenil, a benzodiazepine receptor antagonist, normalized the decrease in head-dipping behavior caused by diazepam treatment in normal and LPS-treated mice. The decrease of the head-dipping effect caused by diazepam was attenuated by minocycline in LPS-treated mice. We further found that the decrease in head-dipping behavior caused by diazepam was blocked by bumetanide, a Na+-K+-2Cl- cotransporter isoform 1 (NKCC1) antagonist, in LPS-treated mice. These findings suggest that diazepam induces the anxiety-like behavior under inflammation conditions, and may cause the GABAA receptor dysfunction associated with the chloride plasticity mediated by NKCC1, which contributes to benzodiazepine-induced delirium after surgery.


Assuntos
Ansiolíticos/farmacologia , Ansiedade/prevenção & controle , Bumetanida/farmacologia , Diazepam/farmacologia , Agonistas de Receptores de GABA-A/farmacologia , Inibidores de Simportadores de Cloreto de Sódio e Potássio/farmacologia , Animais , Ansiolíticos/toxicidade , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Comportamento Animal/efeitos dos fármacos , Bicuculina/farmacologia , Bicuculina/uso terapêutico , Bumetanida/uso terapêutico , Diazepam/toxicidade , Emoções/efeitos dos fármacos , Flumazenil/farmacologia , Flumazenil/uso terapêutico , Agonistas de Receptores de GABA-A/efeitos adversos , Antagonistas de Receptores de GABA-A/farmacologia , Antagonistas de Receptores de GABA-A/uso terapêutico , Inflamação/induzido quimicamente , Inflamação/complicações , Lipopolissacarídeos/toxicidade , Masculino , Camundongos Endogâmicos ICR , Minociclina/farmacologia , Minociclina/uso terapêutico , Atividade Motora/efeitos dos fármacos , Inibidores de Simportadores de Cloreto de Sódio e Potássio/uso terapêutico
10.
Elife ; 102021 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-33904401

RESUMO

Output signals of neural circuits, including the retina, are shaped by a combination of excitatory and inhibitory signals. Inhibitory signals can act presynaptically on axon terminals to control neurotransmitter release and regulate circuit function. However, it has been difficult to study the role of presynaptic inhibition in most neural circuits due to lack of cell type-specific and receptor type-specific perturbations. In this study, we used a transgenic approach to selectively eliminate GABAA inhibitory receptors from select types of second-order neurons - bipolar cells - in mouse retina and examined how this affects the light response properties of the well-characterized ON alpha ganglion cell retinal circuit. Selective loss of GABAA receptor-mediated presynaptic inhibition causes an enhanced sensitivity and slower kinetics of light-evoked responses from ON alpha ganglion cells thus highlighting the role of presynaptic inhibition in gain control and temporal filtering of sensory signals in a key neural circuit in the mammalian retina.


Assuntos
Antagonistas de Receptores de GABA-A/farmacologia , Terminações Pré-Sinápticas/efeitos dos fármacos , Receptores de GABA-A/efeitos dos fármacos , Neurônios Retinianos/efeitos dos fármacos , Animais , Feminino , Cinética , Luz , Masculino , Camundongos , Camundongos Knockout , Terminações Pré-Sinápticas/fisiologia , Receptores de GABA-A/fisiologia , Neurônios Retinianos/fisiologia , Neurônios Retinianos/efeitos da radiação
11.
Sci Rep ; 11(1): 9300, 2021 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-33927244

RESUMO

Chronic pain is a significant public health problem that afflicts nearly 30% of the global population, but current pharmacotherapies are insufficient. Previous report indicated that N-demethylsinomenine, an active metabolite of sinomenine, is efficacious against postoperative pain. The present study investigated whether N-demethylsinomenine is effective for chronic painful conditions or whether repeated treatment alters its effect. Both chronic constriction injury (CCI) surgery and complete Freund's adjuvant (CFA) intraplantar injection induced significant and reliable mechanical allodynia at least for 7 days. Acute treatment with N-demethylsinomenine (10-40 mg/kg, i.p.) dose-dependently attenuated the mechanical allodynia both in CCI-induced neuropathic pain and CFA-induced inflammatory pain in mice. The potency of N-demethylsinomenine for reducing CFA-induced mechanical allodynia was slightly higher than sinomenine. During the period of repeated treatment, N-demethylsinomenine maintained its anti-allodynic effect against both neuropathic and inflammatory pain without producing carry-over effect. Pretreatment with bicuculline, a selective γ-aminobutyric acid type A (GABAA) receptor antagonist, almost completely blocked the anti-allodynia of N-demethylsinomenine (40 mg/kg) both in CCI and CFA-treated mice. Our findings indicated that N-demethylsinomenine exhibits GABAA receptor-mediated anti-allodynic effects in mouse models of neuropathic and inflammatory pain, suggesting it may be a useful novel pharmacotherapy for the control of chronic pain.


Assuntos
Analgésicos/uso terapêutico , Dor Crônica/tratamento farmacológico , Hiperalgesia/tratamento farmacológico , Morfinanos/uso terapêutico , Neuralgia/tratamento farmacológico , Analgésicos/administração & dosagem , Animais , Bicuculina/farmacologia , Modelos Animais de Doenças , Antagonistas de Receptores de GABA-A/farmacologia , Inflamação/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Morfinanos/administração & dosagem , Receptores de GABA-A/metabolismo
12.
Toxins (Basel) ; 13(2)2021 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-33672715

RESUMO

Cobra venoms contain three-finger toxins (TFT) including α-neurotoxins efficiently binding nicotinic acetylcholine receptors (nAChRs). As shown recently, several TFTs block GABAA receptors (GABAARs) with different efficacy, an important role of the TFTs central loop in binding to these receptors being demonstrated. We supposed that the positive charge (Arg36) in this loop of α-cobratoxin may explain its high affinity to GABAAR and here studied α-neurotoxins from African cobra N. melanoleuca venom for their ability to interact with GABAARs and nAChRs. Three α-neurotoxins, close homologues of the known N. melanoleuca long neurotoxins 1 and 2, were isolated and sequenced. Their analysis on Torpedocalifornica and α7 nAChRs, as well as on acetylcholine binding proteins and on several subtypes of GABAARs, showed that all toxins interacted with the GABAAR much weaker than with the nAChR: one neurotoxin was almost as active as α-cobratoxin, while others manifested lower activity. The earlier hypothesis about the essential role of Arg36 as the determinant of high affinity to GABAAR was not confirmed, but the results obtained suggest that the toxin loop III may contribute to the efficient interaction of some long-chain neurotoxins with GABAAR. One of isolated toxins manifested different affinity to two binding sites on Torpedo nAChR.


Assuntos
Colinérgicos/farmacologia , Proteínas Neurotóxicas de Elapídeos/farmacologia , Venenos Elapídicos/metabolismo , Antagonistas de Receptores de GABA-A/farmacologia , Naja , Receptores de GABA/efeitos dos fármacos , Receptor Nicotínico de Acetilcolina alfa7/efeitos dos fármacos , Animais , Sítios de Ligação , Ligação Competitiva , Linhagem Celular Tumoral , Colinérgicos/metabolismo , Proteínas Neurotóxicas de Elapídeos/metabolismo , Antagonistas de Receptores de GABA-A/metabolismo , Potenciais da Membrana , Camundongos , Ligação Proteica , Conformação Proteica , Receptores de GABA/genética , Receptores de GABA/metabolismo , Relação Estrutura-Atividade , Torpedo , Xenopus laevis , Receptor Nicotínico de Acetilcolina alfa7/metabolismo
13.
Psychopharmacology (Berl) ; 238(6): 1657-1669, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33715044

RESUMO

RATIONALE: Nucleus cuneiformis (NC), a reticular nucleus of the midbrain, is a part of the descending pain modulatory system and therefore has an important role in pain perception. OBJECTIVES: Considering the abundance of GABAA and cannabinoid receptors in the NC and also the bidirectional roles for GABA in controlling nociception, the present study examined the effects of bilateral intra-NC microinjection of different doses of the GABAA receptor agonist, muscimol, and the GABAA receptor antagonist, bicuculline, on pain modulation using formalin test. We also assessed interaction between canabinergic and GABAergic systems in the NC during this test. METHODS: Rats were exposed to intra-NC microinjection of bicuculline (50,100, and 200 ng/side) or muscimol (60, 120, and 240 ng/side) and then subjected to the formalin test. In another set of experiments, the effects of muscimol (60 ng/side) or bicuculline (50 ng/side) administration 5 min before a cannabinoid receptor agonist WIN 55,212-2 (5, 10, and 20 µg/side) microinjection into NC on the formalin test were evaluated. RESULTS: Microinjection of bicuculline and muscimol into the NC decreased and increased pain responses, respectively, in a dose-dependent manner during both phases of the test. Microinjection of WIN 55,212-2 into the NC significantly reduced pain responses in a dose-dependent manner. Microinjection of bicuculline or muscimol in combination with WIN 55,212-2 into the NC respectively potentiated and attenuated WIN 55,212-2-induced antinociception in the formalin test. CONCLUSIONS: This study shows that GABA in the NC is involved in pain modulation and suggests the existence of a GABAA-mediated inhibitory system in the NC on pain control. Furthermore, it seems that the antinociceptive effect of WIN 55,212-2 in the formalin test is mediated partly by the activity of local GABAA receptors in the NC.


Assuntos
Benzoxazinas/farmacologia , Bicuculina/farmacologia , Agonistas de Receptores de GABA-A/farmacologia , Morfolinas/farmacologia , Muscimol/farmacologia , Naftalenos/farmacologia , Animais , Bicuculina/administração & dosagem , Canabinoides/farmacologia , Antagonistas de Receptores de GABA-A/farmacologia , Masculino , Nociceptividade/efeitos dos fármacos , Dor/tratamento farmacológico , Medição da Dor , Ratos , Ratos Wistar , Receptores de GABA-A/efeitos dos fármacos , Ácido gama-Aminobutírico/metabolismo
14.
Phys Chem Chem Phys ; 23(6): 3993-4006, 2021 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-33554986

RESUMO

The activation of GABAA receptors by the neurotransmitter gamma-aminobutyric acid mediates the rapid inhibition response in the central nervous system of mammals. Many neurological and mental health disorders arise from alterations in the structure or function of these pentameric ion channels. GABAA receptors are targets for numerous drugs, including benzodiazepines, which bind to α1ß2γ2 GABAA receptors with high affinity to a site in the extracellular domain, between subunits α1 and γ2. It has been established experimentally that the binding of these drugs depends on the presence of one particular amino acid in the α1 subunit: histidine 102. However, the specific role it plays in the intermolecular interaction has not been elucidated. In this work, we applied in silico methods to understand whether certain protonation and rotamer states of α1His102 facilitate the binding of modulators. We analysed diazepam binding, a benzodiazepine, and the antagonist flumazenil to the GABAA receptor using molecular dynamics simulations and adaptive biasing force simulations. The binding free energy follows changes in the protonation state for both ligands, and rotameric states of α1His102 were specific for the different compounds, suggesting distinct preferences for positive allosteric modulators and antagonists. Moreover, in the presence of diazepam and favoured by a neutral tautomer, we identified a water molecule that links loops A, B, and C and may be relevant to the modulation mechanism.


Assuntos
Diazepam/metabolismo , Flumazenil/metabolismo , Moduladores GABAérgicos/metabolismo , Antagonistas de Receptores de GABA-A/metabolismo , Receptores de GABA-A/metabolismo , Histidina/química , Humanos , Simulação de Dinâmica Molecular , Conformação Proteica , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , Prótons , Receptores de GABA-A/química
15.
Br J Anaesth ; 126(3): 674-683, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33388140

RESUMO

BACKGROUND: Multiple cognitive and psychiatric disorders are associated with an increased tonic inhibitory conductance that is generated by α5 subunit-containing γ-aminobutyric acid type A (α5 GABAA) receptors. Negative allosteric modulators that inhibit α5 GABAA receptors (α5-NAMs) are being developed as treatments for these disorders. The effects of α5-NAMs have been studied on recombinant GABAA receptors expressed in non-neuronal cells; however, no study has compared drug effects on the tonic conductance generated by native GABAA receptors in neurones, which was the goal of this study. METHODS: The effects of five α5-NAMs (basmisanil, Ono-160, L-655,708, α5IA, and MRK-016) on tonic current evoked by a low concentration of GABA were studied using whole-cell recordings in cultured mouse hippocampal neurones. Drug effects on current evoked by a saturating concentration of GABA and on miniature inhibitory postsynaptic currents (mIPSCs) were also examined. RESULTS: The α5-NAMs caused a concentration-dependent decrease in tonic current. The potencies varied as the inhibitory concentration for 50% inhibition (IC50) of basmisanil (127 nM) was significantly higher than those of the other compounds (0.4-0.8 nM). In contrast, the maximal efficacies of the drugs were similar (35.5-51.3% inhibition). The α5-NAMs did not modify current evoked by a saturating GABA concentration or mIPSCs. CONCLUSIONS: Basmisanil was markedly less potent than the other α5-NAMs, an unexpected result based on studies of recombinant α5 GABAA receptors. Studying the effects of α5 GABAA receptor-selective drugs on the tonic inhibitory current in neurones could inform the selection of compounds for future clinical trials.


Assuntos
Disfunção Cognitiva/tratamento farmacológico , Antagonistas de Receptores de GABA-A/farmacologia , Hipocampo/efeitos dos fármacos , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Receptores de GABA-A/metabolismo , Regulação Alostérica , Animais , Células Cultivadas , Cognição/efeitos dos fármacos , Relação Dose-Resposta a Droga , Hipocampo/metabolismo , Camundongos , Neurônios/metabolismo , Técnicas de Patch-Clamp
16.
Neurosci Lett ; 744: 135619, 2021 01 23.
Artigo em Inglês | MEDLINE | ID: mdl-33421486

RESUMO

Progesterone acts on neurons directly by activating its receptor and through metabolic conversion to neurosteroids. There is emerging evidence that progesterone exerts excitatory effects by activating its cognate receptors (progesterone receptors, PRs) through enhanced expression of α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs). Progesterone metabolite 5α,3α-tetrahydro-progesterone (allopregnanolone, THP) mediates its anxiolytic and sedative actions through the potentiation of synaptic and extrasynaptic γ-aminobutyric acid type-A receptors (GABAARs). Here, we review progesterone's neuromodulatory actions exerted through PRs and THP and their opposing role in regulating seizures, catamenial epilepsy, and seizure exacerbation associated with progesterone withdrawal.


Assuntos
Anticonvulsivantes/metabolismo , Epilepsia/metabolismo , Neurônios/metabolismo , Progesterona/metabolismo , Receptores de GABA-A/metabolismo , Receptores de Progesterona/metabolismo , Animais , Anticonvulsivantes/uso terapêutico , Epilepsia/tratamento farmacológico , Antagonistas de Receptores de GABA-A/farmacologia , Antagonistas de Receptores de GABA-A/uso terapêutico , Humanos , Neurônios/efeitos dos fármacos , Progesterona/uso terapêutico
17.
Proc Natl Acad Sci U S A ; 118(1)2021 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-33443213

RESUMO

Diapause represents a major developmental switch in insects and is a seasonal adaptation that evolved as a specific subtype of dormancy in most insect species to ensure survival under unfavorable environmental conditions and synchronize populations. However, the hierarchical relationship of the molecular mechanisms involved in the perception of environmental signals to integration in morphological, physiological, behavioral, and reproductive responses remains unclear. In the bivoltine strain of the silkworm Bombyx mori, embryonic diapause is induced transgenerationally as a maternal effect. Progeny diapause is determined by the environmental temperature during embryonic development of the mother. Here, we show that the hierarchical pathway consists of a γ-aminobutyric acid (GABA)ergic and corazonin signaling system modulating progeny diapause induction via diapause hormone release, which may be finely tuned by the temperature-dependent expression of plasma membrane GABA transporter. Furthermore, this signaling pathway possesses similar features to the gonadotropin-releasing hormone (GnRH) signaling system for seasonal reproductive plasticity in vertebrates.


Assuntos
Bombyx/metabolismo , Diapausa/genética , Desenvolvimento Embrionário/genética , Proteínas da Membrana Plasmática de Transporte de GABA/metabolismo , Proteínas de Insetos/metabolismo , Neuropeptídeos/metabolismo , Ácido gama-Aminobutírico/metabolismo , Sequência de Aminoácidos , Animais , Bombyx/embriologia , Bombyx/genética , Membrana Celular/metabolismo , Proteínas da Membrana Plasmática de Transporte de GABA/genética , Antagonistas de Receptores de GABA-A/farmacologia , Neurônios GABAérgicos/efeitos dos fármacos , Neurônios GABAérgicos/metabolismo , Proteínas de Insetos/genética , Neuropeptídeos/genética , Óvulo/efeitos dos fármacos , Óvulo/metabolismo , Filogenia , Receptores de GABA-A/metabolismo , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
18.
Epilepsia ; 62(1): 238-249, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33417274

RESUMO

OBJECTIVE: LMR-101 is a bisphenol derivative of propofol, a short-acting general anesthetic, which is also used to manage status epilepticus (SE). We evaluated the sedative and anticonvulsant effects of LMR-101 to discover its potential to manage epilepsy and SE in the clinic. METHODS: Comparative studies between LMR-101 and propofol were performed in mice to elucidate an appropriate dose range for LMR-101 that produced anticonvulsant effects without significant sedation. Then, the anticonvulsive efficacy for LMR-101 was evaluated using seizure models induced by pentylenetetrazol and (+)-bicuculline. The ability of LMR-101 to inhibit SE was assessed using a rat model of SE induced by pilocarpine. Radioligand binding assay profiles for LMR-101 were performed to evaluate the potential mechanisms of action underlying its anticonvulsant properties. RESULTS: In the mouse study, LMR-101 exhibited greater anticonvulsant and lesser sedative effect compared with propofol. LMR-101 completely inhibited pentylenetetrazol-induced seizures at a dose of 50 mg/kg and exhibited heavy sedation at 300 mg/kg. Propofol anesthetized all mice and only decreased the seizure rate at 25 mg/kg. LMR-101 also suppressed seizure behaviors evoked by (+)-bicuculline in mice in a dose-dependent manner. In the pilocarpine-induced SE model, LMR-101 significantly decreased the maximum seizure score and seizure duration in a dose-dependent manner. The median effective dose for LMR-101 was 14.30 mg/kg and 121.87 mg/kg to prevent and inhibit sustained SE, respectively. In binding assays, LMR-101 primarily inhibited tert-[35 S] butylbicyclophosphorothionate binding to γ-aminobutyric acid type A (GABAA ) receptors (half-maximal inhibitory concentration = 2.06 µmol·L-1 ), but it did not affect [3 H] flunitrazepam or [3 H] muscimol binding. SIGNIFICANCE: It is anticipated that LMR-101 might play an essential role in the clinical management of epilepsy and SE. LMR-101 also might bind to a novel target site on the GABAA receptor that is different from existing antiepileptic drugs. Further study of the mechanisms of action of LMR-101 would be of considerable value in the search for new active drug sites on GABAA receptors.


Assuntos
Anticonvulsivantes/farmacologia , Propofol/farmacologia , Receptores de GABA-A/efeitos dos fármacos , Convulsões/tratamento farmacológico , Estado Epiléptico/tratamento farmacológico , Animais , Comportamento Animal/efeitos dos fármacos , Bicuculina/toxicidade , Eletroencefalografia , Antagonistas de Receptores de GABA-A/toxicidade , Hipnóticos e Sedativos/farmacologia , Camundongos , Agonistas Muscarínicos/toxicidade , Pentilenotetrazol/toxicidade , Fenóis/farmacologia , Pilocarpina/toxicidade , Propofol/análogos & derivados , Ratos , Receptores de GABA-A/metabolismo , Convulsões/induzido quimicamente , Estado Epiléptico/induzido quimicamente
19.
Med Chem ; 17(5): 453-461, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-31840612

RESUMO

BACKGROUND: Quinazolines and quinazolinones constitute a major class of biologically active molecules, both from natural and synthetic sources. The quinazolinone moiety is an important pharmacophore showing many types of pharmacological activities as shown in recent exhaustive review on the chemistry of 2-heteroaryl & heteroalkyl-4-quinazolinones4-quinazolinones that are the formal condensation products of anthranilic acid and amides. They can also be prepared in this fashion through the Niementowski quinazolinone synthesis, named after it's discoverer Stefan Niementowski. Quinazoline and condensed quinazoline exhibit potent central nervous system (CNS) activities like anti-anxiety, analgesic, anti-inflammatory and anticonvulsant. Quinazolin-4- ones with 2, 3-disubstitution is reported to possess significant analgesic, anti-inflammatory and anticonvulsant activities. METHODS: To expand these views and application profiles, efforts have been made for the synthesis of a new class of quinazolinone by incorporating different amines into synthesized benzoxazinone ring by replacing O atom in the ring. Up till now, a great number of various procedures have been proposed for the synthesis of quinazolin-4-ones in the past few years. Using microwave radiation, this reaction could be easily and rapidly performed in very good yields, providing a large number of various 3-substituted-2- propyl-quinazolin-4-one derivatives which can be employed as useful bioactive compounds. We report a facile and efficient method for the synthesis of 3-substituted-2- propyl-quinazolin-4-one by the condensation reaction of anthranilic acid or halogen substituted anthranilic acid or methyl anthranilate, butanoic anhydride with various amines. We also report a drug/ligand or receptor/protein interactions by identifying the suitable active sites in the human gamma-aminobutyric acid receptor, the gaba (a)r-beta3 homopentamer human gammaaminobutyric acid receptor, and the gaba (a)r-beta3 homopentamer protein. RESULTS: It was observed in the reaction, 3-alkyl/aryl-2-alkyl-quinazolin-4-one gave good yield as well as good quality of the product by using MW. All the synthesized compounds were subjected to grid-based molecular docking studies. The results show that compound 4t has good affinity to the active site residue of the human gamma-aminobutyric acid receptor, and the gaba (a)r-beta3 homopentamer. CONCLUSION: The Microwave irradiation for the synthesis of the title compounds offers a reduction in reaction time, operation simplicity, cleaner reaction, easy work-up and improved yields. The procedure clearly highlights the advantages of green chemistry. The data reported in this article may be helpful for the medicinal chemists who are working in this area. The protein-ligand interaction plays a significant role in structural based drug designing. In the present work, we have docked the ligand, 2, 3-disubstituted quinazolinone with the proteins that are used as the target for GABA-A receptor.


Assuntos
Antagonistas de Receptores de GABA-A/metabolismo , Quinazolinonas/metabolismo , Receptores de GABA-A/metabolismo , Domínio Catalítico , Antagonistas de Receptores de GABA-A/síntese química , Humanos , Micro-Ondas , Simulação de Acoplamento Molecular , Ligação Proteica , Quinazolinonas/síntese química , Receptores de GABA-A/química
20.
J Surg Res ; 259: 431-441, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33069391

RESUMO

BACKGROUND: Ivermectin (IVM) was first used as an antiparasitic agent; however, the role of this drug evolved into a broad spectrum. Many mechanisms have been proposed, including interaction with the GABAergic system. Considering the presence of GABA receptor in the skin tissue and its role in ischemia-reperfusion I/R injury, we aimed to evaluate the effect of IVM through GABA receptors on random-pattern skin flap survival. METHODS: Sixty Wistar male rats were used. Multiple doses of IVM (0.01, 0.05, 0.2, and 0.5 mg/kg) were injected intraperitoneally before the surgery. Baclofen (selective GABAB agonist) and bicuculline (selective GABAA antagonist) were administered in combination with IVM to assess the role of the GABAergic system. Histopathological evaluations, immunohistochemical staining, quantitative assessment of IL-1ß and TNFα, and the expression of GABAA α1 subunit and GABAB R1 receptors were evaluated in the skin tissue. RESULTS: IVM 0.05 mg/kg could significantly increase flap survival compared with the control group (P < 0.001). Subeffective dose of baclofen (0.1 mg/kg) had synergistic effect with the subeffective dose of IVM (0.01 mg/kg) (P < 0.001), whereas bicuculline 1 mg/kg reversed the effect of IVM (0.05 mg/kg) (P < 0.001). IVM 0.05 mg/kg could also decrease the IL-1ß and TNFα levels and increase the expression of GABAA α1 subunit and GABAB R1 receptors in the flap tissue compared with the control group. CONCLUSIONS: IVM could improve skin flap survival, probably mediated by the GABAergic pathway. Both GABAA and GABAB receptors are involved in this process. This finding may repurpose the use of old drug, "Ivermectin."


Assuntos
Sobrevivência de Enxerto/efeitos dos fármacos , Ivermectina/administração & dosagem , Retalhos Cirúrgicos/transplante , Ácido gama-Aminobutírico/metabolismo , Animais , Baclofeno/administração & dosagem , Bicuculina/administração & dosagem , Reposicionamento de Medicamentos , Antagonistas GABAérgicos/administração & dosagem , Antagonistas de Receptores de GABA-A/administração & dosagem , Humanos , Masculino , Modelos Animais , Ratos , Receptores de GABA-A/metabolismo , Receptores de GABA-B/metabolismo , Pele/efeitos dos fármacos , Pele/metabolismo , Retalhos Cirúrgicos/efeitos adversos
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