RESUMO
BACKGROUND: α-Amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) and N-methyl-D-aspartate (NMDA) receptors mediate excitatory neurotransmission in the brain and may be targeted by autoantibodies, leading to autoimmune synaptic encephalitis (AE). AE can be associated with other autoimmune diseases. However, the cooccurrence of anti-AMPA and NMDA receptor AE together with myasthenia gravis (MG) is unusual. CASE PRESENTATION: A 24-year-old previously healthy male presented with seronegative ocular MG, the diagnosis of which was supported by single-fiber electrophysiology findings. Three months later, he developed AE, initially being positive for AMPA receptor antibodies and subsequently for NMDA receptor antibodies. No underlying malignancy was found. In response to aggressive immunosuppressive treatment, he recovered (modified Rankin Scale (mRS) score change from 5 to 1). Despite some cognitive problems at the 1-year follow-up, which were not revealed using the mRS, he was able to return to his studies. CONCLUSIONS: AE may coexist with other autoimmune disorders. Patients with seronegative MG, including ocular MG, may develop autoimmune encephalitis with more than one cell-surface antibody.
Assuntos
Encefalite Antirreceptor de N-Metil-D-Aspartato , Encefalite , Miastenia Gravis , Humanos , Masculino , Adulto Jovem , Adulto , Receptores de N-Metil-D-Aspartato , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico , Encefalite/diagnóstico , Autoanticorpos , Encefalite Antirreceptor de N-Metil-D-Aspartato/diagnósticoRESUMO
Prenatal stress is believed to increase the risk of developing neuropsychiatric disorders, including major depression. Adverse genetic and environmental impacts during early development, such as glucocorticoid hyper-exposure, can lead to changes in the foetal brain, linked to mental illnesses developed in later life. Dysfunction in the GABAergic inhibitory system is associated with depressive disorders. However, the pathophysiology of GABAergic signalling is poorly understood in mood disorders. Here, we investigated GABAergic neurotransmission in the low birth weight (LBW) rat model of depression. Pregnant rats, exposed to dexamethasone, a synthetic glucocorticoid, during the last week of gestation, yielded LBW offspring showing anxiety- and depressive-like behaviour in adulthood. Patch-clamp recordings from dentate gyrus granule cells in brain slices were used to examine phasic and tonic GABAA receptor-mediated currents. The transcriptional levels of selected genes associated with synaptic vesicle proteins and GABAergic neurotransmission were investigated. The frequency of spontaneous inhibitory postsynaptic currents (sIPSC) was similar in control and LBW rats. Using a paired-pulse protocol to stimulate GABAergic fibres impinging onto granule cells, we found indications of decreased probability of GABA release in LBW rats. However, tonic GABAergic currents and miniature IPSCs, reflecting quantal vesicle release, appeared normal. Additionally, we found elevated expression levels of two presynaptic proteins, Snap-25 and Scamp2, components of the vesicle release machinery. The results suggest that altered GABA release may be an essential feature in the depressive-like phenotype of LBW rats.
Assuntos
Depressão , Ácido gama-Aminobutírico , Gravidez , Feminino , Ratos , Animais , Ácido gama-Aminobutírico/metabolismo , Peso ao Nascer , Glucocorticoides/metabolismo , Hipocampo/metabolismo , Receptores de GABA-A/metabolismoRESUMO
Neural activity regulates local increases in cerebral blood flow (ΔCBF) and the cortical metabolic rate of oxygen (ΔCMRO2) that constitutes the basis of BOLD functional neuroimaging signals. Glutamate signaling plays a key role in brain vascular and metabolic control; however, the modulatory effect of GABA is incompletely understood. Here we performed in vivo studies in mice to investigate how THIP (which tonically activates extrasynaptic GABAARs) and Zolpidem (a positive allosteric modulator of synaptic GABAARs) impact stimulation-induced ΔCBF, ΔCMRO2, local field potentials (LFPs), and fluorescent cytosolic Ca(2+) transients in neurons and astrocytes. Low concentrations of THIP increased ΔCBF and ΔCMRO2 at low stimulation frequencies. These responses were coupled to increased synaptic activity as indicated by LFP responses, and to Ca(2+) activities in neurons and astrocytes. Intermediate and high concentrations of THIP suppressed ΔCBF and ΔCMRO2 at high stimulation frequencies. Zolpidem had similar but less-pronounced effects, with similar dependence on drug concentration and stimulation frequency. Our present findings suggest that slight increases in both synaptic and extrasynaptic GABAAR activity might selectively gate and amplify transient low-frequency somatosensory inputs, filter out high-frequency inputs, and enhance vascular and metabolic responses that are likely to be reflected in BOLD functional neuroimaging signals.
Assuntos
Cálcio/metabolismo , Circulação Cerebrovascular/fisiologia , Consumo de Oxigênio/fisiologia , Receptores de GABA-A/metabolismo , Córtex Somatossensorial/citologia , Córtex Somatossensorial/fisiologia , Potenciais de Ação/fisiologia , Animais , Biofísica , Circulação Cerebrovascular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ácido Egtázico/análogos & derivados , Ácido Egtázico/metabolismo , Estimulação Elétrica , Lateralidade Funcional , Agonistas GABAérgicos/farmacologia , Isoxazóis/farmacologia , Camundongos , Consumo de Oxigênio/efeitos dos fármacos , Pressão Parcial , Piridinas/farmacologia , Sulfonamidas/metabolismo , Tiazóis/metabolismo , Vibrissas/inervação , Zolpidem , Ácido gama-Aminobutírico/farmacologiaRESUMO
Amyotrophic lateral sclerosis (ALS) is a progressive degenerative disease of the central nervous system. Symptomatic and presymptomatic ALS patients demonstrate cortical hyperexcitability, which raises the possibility that alterations in inhibitory gamma-aminobutyric acid (GABA)ergic system could underlie this dysfunction. Here, we studied the GABAergic system in cortex using patch-clamp recordings in the wobbler mouse, a model of ALS. In layer 5 pyramidal neurons of motor cortex, the frequency of GABA(A) receptor-mediated spontaneous inhibitory postsynaptic currents was reduced by 72% in wobbler mice. Also, miniature inhibitory postsynaptic currents recorded under blockade of action potentials were decreased by 64%. Tonic inhibition mediated by extrasynaptic GABA(A) receptors was reduced by 87%. In agreement, we found a decreased density of parvalbumin- and somatostatin-positive inhibitory interneurons and reduced vesicular GABA transporter immunoreactivity in the neuropil. Finally, we observed an increased input resistance and excitability of wobbler excitatory neurons, which could be explained by lack of GABA(A) receptor-mediated influences. In conclusion, we demonstrate decreases in GABAergic inhibition, which might explain the cortical hyperexcitability in wobbler mice.
Assuntos
Esclerose Lateral Amiotrófica/fisiopatologia , Potenciais Pós-Sinápticos Inibidores/fisiologia , Córtex Motor/fisiopatologia , Inibição Neural/fisiologia , Ácido gama-Aminobutírico/metabolismo , Esclerose Lateral Amiotrófica/metabolismo , Animais , Modelos Animais de Doenças , Imuno-Histoquímica , Camundongos , Potenciais Pós-Sinápticos em Miniatura/fisiologia , Córtex Motor/metabolismo , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Transmissão Sináptica/fisiologiaRESUMO
In major depression, one line of research indicates that a dysfunctional GABAergic inhibitory system is linked to the appearance of depressive symptoms. However, as the mechanistic details of such GABAergic deficit are largely unknown, we undertook a functional investigation of the GABAergic system in the rat chronic mild stress model of depression. Adult rats were exposed to an eight-week long stress protocol leading to anhedonic-like behavior. In hippocampal brain slices, phasic, and tonic GABA(A) receptor-mediated currents in dentate gyrus granule cells were examined using patch-clamp recordings. In granule cells, the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) was reduced to 41% in anhedonic-like rats, which was associated with a reduced probability of evoked GABA release. Using immunohistochemical analysis, there was no change in the number of parvalbumin-positive interneurons in the dentate gyrus. Notably, we observed a 60% increase in THIP-activated tonic GABA(A) mediated current in anhedonic-like rats, suggesting an upregulation of extrasynaptic GABA(A) receptors. Finally, five weeks treatment with the antidepressant escitalopram partially reversed the sIPSCs frequency. In summary, we have revealed a hippocampal dysfunction in the GABAergic system in the chronic mild stress model of depression in rats, caused by a reduction in action potential-dependent GABA release. Since the function of the GABAergic system was improved by antidepressant treatment, in parallel with behavioral read outs, it suggests a role of the GABAergic system in the pathophysiology of depression.
Assuntos
Giro Denteado/metabolismo , Agonistas de Receptores de GABA-A/farmacologia , Potenciais Pós-Sinápticos Inibidores/fisiologia , Isoxazóis/farmacologia , Receptores de GABA-A , Ácido gama-Aminobutírico/metabolismo , Potenciais de Ação/efeitos dos fármacos , Animais , Antidepressivos de Segunda Geração/farmacologia , Citalopram/farmacologia , Depressão/tratamento farmacológico , Modelos Animais de Doenças , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hipocampo/fisiopatologia , Imuno-Histoquímica , Interneurônios/metabolismo , Masculino , Parvalbuminas/metabolismo , Técnicas de Patch-Clamp , Ratos , Ratos Wistar , Receptores de GABA-A/efeitos dos fármacos , Receptores de GABA-A/metabolismo , Inibidores Seletivos de Recaptação de Serotonina/farmacologiaRESUMO
Mature BDNF and its precursor proBDNF may both be secreted to exert opposite effects on synaptic plasticity in the hippocampus. However, it is unknown how proBDNF and mature BDNF affect the excitability of GABAergic interneurons and thereby regulate GABAergic inhibition. We made recordings of GABAergic spontaneous IPSCs (sIPSCs) in mouse dentate gyrus granule cells and found that chronic or acute BDNF reductions led to large increases in the sIPSC frequencies, which were TTX (tetrodotoxin) sensitive and therefore action-potential driven. Conversely, addition of mature BDNF, but not proBDNF, within minutes led to a decrease in the sIPSC frequency to 44%. Direct recordings from fast-spiking GABAergic interneurons revealed that mature BDNF reduced their excitability and depressed their action potential firing, whereas proBDNF had no effect. Using the TrkB inhibitor K-252a, or mice deficient for the common neurotrophin receptor p75(NTR), the regulation of GABAergic activity was shown specifically to be mediated by BDNF binding to the neurotrophin receptor TrkB. In agreement, immunohistochemistry demonstrated that TrkB, but not p75(NTR), was expressed in parvalbumin-positive interneurons. Our results suggest that mature BDNF decreases the excitability of GABAergic interneurons via activation of TrkB, while proBDNF does not impact on GABAergic activity. Thus, by affecting the firing of GABAergic interneurons, mature BDNF may play an important role in regulating network oscillations in the hippocampus.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Giro Denteado/metabolismo , Interneurônios/metabolismo , Potenciais de Ação , Animais , Giro Denteado/citologia , Masculino , Camundongos , Camundongos Mutantes , Inibição Neural/fisiologia , Plasticidade Neuronal/fisiologia , Técnicas de Patch-Clamp , Receptor trkB/metabolismo , Regulação para CimaRESUMO
Tonic inhibitory currents, mediated by extrasynaptic GABAA receptors, are elevated at a delay following stroke. Flavonoids minimise the extent of cellular damage following stroke, but little is known about their mode of action. We demonstrate that the flavonoid, 2'-methoxy-6-methylflavone (0.1-10 µM; 2'MeO6MF), increases GABAA receptor tonic currents presumably via δ-containing GABAA receptors. Treatment with 2'MeO6MF 1-6 h post focal ischaemia dose dependently decreases infarct volume and improves functional recovery. The effect of 2'MeO6MF was attenuated in δ-/- mice, indicating that the effects of the flavonoid were mediated via δ-containing GABAA receptors. Further, as flavonoids have been shown to have multiple modes of action, we investigated the anti-inflammatory effects of 2'MeO6MF. Using a macrophage cell line, we show that 2'MeO6MF can dampen an LPS-induced elevation in NFkB activity. Assessment of vehicle-treated stroke animals revealed a significant increase in circulating IL1ß, TNFα and IFγ levels. Treatment with 2'MeO6MF dampened the stroke-induced increase in circulating cytokines, which was blocked in the presence of the pan-AKT inhibitor, GSK690693. These studies support the hypothesis that compounds that potentiate tonic inhibition via δ-containing GABAA receptors soon after stroke can afford neuroprotection.
Assuntos
Isquemia Encefálica/tratamento farmacológico , Flavonas/administração & dosagem , Moduladores GABAérgicos/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Animais , Encéfalo/metabolismo , Modelos Animais de Doenças , Flavonas/farmacocinética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Receptores de GABA-A/efeitos dos fármacos , Receptores de GABA-A/genética , Receptores de GABA-A/fisiologia , Acidente Vascular Cerebral/tratamento farmacológico , Potenciais Sinápticos/efeitos dos fármacos , Potenciais Sinápticos/fisiologiaRESUMO
Clinical and experimental data suggest that fronto-cortical GABAergic deficits contribute to the pathophysiology of major depressive disorder (MDD). To further test this hypothesis, we used a well characterized rat model for depression and examined the effect of stress on GABAergic neuron numbers and GABA-mediated synaptic transmission in the medial prefrontal cortex (mPFC) of rats. Adult male Wistar rats were subjected to 9-weeks of chronic mild stress (CMS) and based on their hedonic-anhedonic behavior they were behaviorally phenotyped as being stress-susceptible (anhedonic) or stress-resilient. Post mortem quantitative histopathology was used to examine the effect of stress on parvalbumin (PV)-, calretinin- (CR), calbindin- (CB), cholecystokinin- (CCK), somatostatin-(SST) and neuropeptide Y-positive (NPY+) GABAergic neuron numbers in all cortical subareas of the mPFC (anterior cingulate (Cg1), prelimbic (PrL) and infralimbic (IL) cortexes). In vitro, whole-cell patch-clamp recordings from layer II-III pyramidal neurons of the ventral mPFC was used to examine GABAergic neurotransmission. The cognitive performance of the animals was assessed in a hippocampal-prefrontal-cortical circuit dependent learning task. Stress exposure reduced the number of CCK-, CR- and PV-positive GABAergic neurons in the mPFC, most prominently in the IL cortex. Interestingly, in the stress-resilient animals, we found higher number of neuropeptide Y-positive neurons in the entire mPFC. The electrophysiological analysis revealed reduced frequencies of spontaneous and miniature IPSCs in the anhedonic rats and decreased release probability of perisomatic-targeting GABAergic synapses and alterations in GABAB receptor mediated signaling. In turn, pyramidal neurons showed higher excitability. Anhedonic rats were also significantly impaired in the object-place paired-associate learning task. These data demonstrate that long-term stress results in functional and structural deficits of prefrontal GABAergic networks. Our findings support the concept that fronto-limbic GABAergic dysfunctions may contribute to emotional and cognitive symptoms of MDD.
RESUMO
BACKGROUND: The schizophrenia-associated BRD1 gene encodes a transcriptional regulator whose comprehensive chromatin interactome is enriched with schizophrenia risk genes. However, the biology underlying the disease association of BRD1 remains speculative. METHODS: This study assessed the transcriptional drive of a schizophrenia-associated BRD1 risk variant in vitro. Accordingly, to examine the effects of reduced Brd1 expression, we generated a genetically modified Brd1+/- mouse and subjected it to behavioral, electrophysiological, molecular, and integrative genomic analyses with focus on schizophrenia-relevant parameters. RESULTS: Brd1+/- mice displayed cerebral histone H3K14 hypoacetylation and a broad range of behavioral changes with translational relevance to schizophrenia. These behaviors were accompanied by striatal dopamine/serotonin abnormalities and cortical excitation-inhibition imbalances involving loss of parvalbumin immunoreactive interneurons. RNA-sequencing analyses of cortical and striatal micropunches from Brd1+/- and wild-type mice revealed differential expression of genes enriched for schizophrenia risk, including several schizophrenia genome-wide association study risk genes (e.g., calcium channel subunits [Cacna1c and Cacnb2], cholinergic muscarinic receptor 4 [Chrm4)], dopamine receptor D2 [Drd2], and transcription factor 4 [Tcf4]). Integrative analyses further found differentially expressed genes to cluster in functional networks and canonical pathways associated with mental illness and molecular signaling processes (e.g., glutamatergic, monoaminergic, calcium, cyclic adenosine monophosphate [cAMP], dopamine- and cAMP-regulated neuronal phosphoprotein 32 kDa [DARPP-32], and cAMP responsive element binding protein signaling [CREB]). CONCLUSIONS: Our study bridges the gap between genetic association and pathogenic effects and yields novel insights into the unfolding molecular changes in the brain of a new schizophrenia model that incorporates genetic risk at three levels: allelic, chromatin interactomic, and brain transcriptomic.
Assuntos
Comportamento Animal/fisiologia , Expressão Gênica/genética , Histona Acetiltransferases/fisiologia , Esquizofrenia/genética , Transmissão Sináptica/genética , Acetilação , Animais , Animais Geneticamente Modificados/genética , Corpo Estriado/metabolismo , Dopamina/metabolismo , Histona Acetiltransferases/genética , Histonas/metabolismo , Interneurônios/fisiologia , Camundongos , Serotonina/metabolismoRESUMO
GABA transporter subtype 1 (GAT1) knock-out (KO) mice display normal reproduction and life span but have reduced body weight (female, -10%; male, -20%) and higher body temperature fluctuations in the 0.2-1.5/h frequency range. Mouse GAT1 (mGAT1) KO mice exhibit motor disorders, including gait abnormality, constant 25-32 Hz tremor, which is aggravated by flunitrazepam, reduced rotarod performance, and reduced locomotor activity in the home cage. Open-field tests show delayed exploratory activity, reduced rearing, and reduced visits to the central area, with no change in the total distance traveled. The mGAT1 KO mice display no difference in acoustic startle response but exhibit a deficiency in prepulse inhibition. These open-field and prepulse inhibition results suggest that the mGAT1 KO mice display mild anxiety or nervousness. The compromised GABA uptake in mGAT1 KO mice results in an increased GABA(A) receptor-mediated tonic conductance in both cerebellar granule and Purkinje cells. The reduced rate of GABA clearance from the synaptic cleft is probably responsible for the slower decay of spontaneous IPSCs in cerebellar granule cells. There is little or no compensatory change in other proteins or structures related to GABA transmission in the mGAT1 KO mice, including GAT1-independent GABA uptake, number of GABAergic interneurons, and GABA(A)-, vesicular GABA transporter-, GAD65-, and GAT3-immunoreactive structures in cerebellum or hippocampus. Therefore, the excessive extracellular GABA present in mGAT1 KO mice results in behaviors that partially phenocopy the clinical side effects of tiagabine, suggesting that these side effects are inherent to a therapeutic strategy that targets the widely expressed GAT1 transporter system.
Assuntos
Ansiedade/genética , Ataxia/genética , Cerebelo/fisiopatologia , Proteínas da Membrana Plasmática de Transporte de GABA/deficiência , Tremor/genética , Animais , Comportamento Animal/fisiologia , Benzodiazepinas/uso terapêutico , Temperatura Corporal/genética , Peso Corporal/genética , Cerebelo/citologia , Estimulação Elétrica/métodos , Eletroencefalografia , Comportamento Exploratório/fisiologia , Antagonistas GABAérgicos/farmacologia , Proteínas da Membrana Plasmática de Transporte de GABA/metabolismo , Transtornos Neurológicos da Marcha/induzido quimicamente , Transtornos Neurológicos da Marcha/tratamento farmacológico , Transtornos Neurológicos da Marcha/genética , Glutamato Descarboxilase/metabolismo , Imuno-Histoquímica/métodos , Técnicas In Vitro , Inibição Psicológica , Isoenzimas/metabolismo , Aprendizagem em Labirinto/fisiologia , Potenciais da Membrana/genética , Potenciais da Membrana/efeitos da radiação , Camundongos , Camundongos Knockout , Atividade Motora/genética , Neurônios/fisiologia , Neurônios/efeitos da radiação , Pentilenotetrazol , Desempenho Psicomotor/fisiologia , Piridazinas/farmacologia , Receptores de GABA-A/metabolismo , Reflexo Acústico/genética , Teste de Desempenho do Rota-Rod/métodos , Convulsões/induzido quimicamente , Sinaptossomos/metabolismo , Tremor/tratamento farmacológico , Proteínas Vesiculares de Transporte de Aminoácidos Inibidores/metabolismo , Ácido gama-Aminobutírico/metabolismoRESUMO
GABA transporter subtype 1 (GAT1) molecules were counted near GABAergic synapses, to a resolution of approximately 0.5 microm. Fusions between GAT1 and green fluorescent protein (GFP) were tested in heterologous expression systems, and a construct was selected that shows function, expression level, and trafficking similar to that of wild-type (WT) GAT1. A strain of knock-in mice was constructed that expresses this mGAT1-GFP fusion in place of the WT GAT1 gene. The pattern of fluorescence in brain slices agreed with previous immunocytochemical observations. [3H]GABA uptake, synaptic electrophysiology, and subcellular localization of the mGAT1-GFP construct were also compared with WT mice. Quantitative fluorescence microscopy was used to measure the density of mGAT1-GFP at presynaptic structures in CNS preparations from the knock-in mice. Fluorescence measurements were calibrated with transparent beads and gels that have known GFP densities. Surface biotinylation defined the fraction of transporters on the surface versus those in the nearby cytoplasm. The data show that the presynaptic boutons of GABAergic interneurons in cerebellum and hippocampus have a membrane density of 800-1300 GAT1 molecules per square micrometer, and the axons that connect boutons have a linear density of 640 GAT1 molecules per micrometer. A cerebellar basket cell bouton, a pinceau surrounding a Purkinje cell axon, and a cortical chandelier cell cartridge carry 9000, 7.8 million, and 430,000 GAT1 molecules, respectively; 61-63% of these molecules are on the surface membrane. In cultures from hippocampus, the set of fluorescent cells equals the set of GABAergic interneurons. Knock-in mice carrying GFP fusions of membrane proteins provide quantitative data required for understanding the details of synaptic transmission in living neurons.
Assuntos
Proteínas de Transporte/biossíntese , Membrana Celular/metabolismo , Citoplasma/metabolismo , Proteínas de Membrana/biossíntese , Proteínas de Membrana Transportadoras , Transportadores de Ânions Orgânicos , Terminações Pré-Sinápticas/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Animais , Biotinilação , Proteínas de Transporte/genética , Linhagem Celular , Cerebelo/citologia , Cerebelo/metabolismo , DNA Complementar/genética , DNA Complementar/metabolismo , Proteínas da Membrana Plasmática de Transporte de GABA , Proteínas de Fluorescência Verde , Hipocampo/citologia , Hipocampo/metabolismo , Humanos , Rim/citologia , Rim/metabolismo , Proteínas Luminescentes/genética , Proteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Microscopia de Fluorescência , Neurônios/citologia , Neurônios/metabolismo , Especificidade de Órgãos , Técnicas de Patch-Clamp , Proteínas Recombinantes de Fusão/genética , Células-Tronco/citologia , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/metabolismoRESUMO
Two main questions are important for understanding and treating affective disorders: why are certain individuals susceptible or resilient to stress, and what are the features of treatment response and resistance? To address these questions, we used a chronic mild stress (CMS) rat model of depression. When exposed to stress, a fraction of rats develops anhedonic-like behavior, a core symptom of major depression, while another subgroup of rats is resilient to CMS. Furthermore, the anhedonic-like state is reversed in about half the animals in response to chronic escitalopram treatment (responders), while the remaining animals are resistant (non-responder animals). Electrophysiology in hippocampal brain slices was used to identify a synaptic hallmark characterizing these groups of animals. Presynaptic properties were investigated at GABAergic synapses onto single dentate gyrus granule cells. Stress-susceptible rats displayed a reduced probability of GABA release judged by an altered paired-pulse ratio of evoked inhibitory postsynaptic currents (IPSCs) (1.48 ± 0.25) compared with control (0.81 ± 0.05) and stress-resilient rats (0.78 ± 0.03). Spontaneous IPSCs (sIPSCs) occurred less frequently in stress-susceptible rats compared with control and resilient rats. Finally, a subset of stress-susceptible rats responding to selective serotonin reuptake inhibitor (SSRI) treatment showed a normalization of the paired-pulse ratio (0.73 ± 0.06) whereas non-responder rats showed no normalization (1.2 ± 0.2). No changes in the number of parvalbumin-positive interneurons were observed. Thus, we provide evidence for a distinct GABAergic synaptopathy which associates closely with stress-susceptibility and treatment-resistance in an animal model of depression.
Assuntos
Depressão/fisiopatologia , Plasticidade Neuronal , Sinapses/fisiologia , Animais , Masculino , Ratos , Ratos Wistar , Ácido gama-Aminobutírico/fisiologiaRESUMO
In addition to their classical roles in neuronal growth, survival and differentiation, neurotrophins are also rapid regulators of excitability, synaptic transmission and activity-dependent synaptic plasticity. We have recently shown that mature BDNF (Brain Derived Neurotrophic Factor), but not proBDNF, modulates the excitability of interneurons in dentate gyrus within minutes. Here, we used brain slice patch-clamp recordings to study the mechanisms through which BDNF modulates the firing of interneurons in rat dentate gyrus by binding to TrkB receptors. Bath application of BDNF (15 ng/ml) under current-clamp decreased the firing frequency (by 80%) and input resistance, blocking the delayed firing observed at near-threshold voltage ranges, with no changes in resting membrane potential or action potential waveform. Using TEA (tetraethylammonium), or XE991(a Kv7/KCNQ channel antagonist), the effect of BDNF was abolished, whereas application of retigabine (a Kv7/KCNQ channel opener) mimicked the effect of BDNF, suggesting that the M-current could be implicated in the modulation of the firing. In voltage-clamp experiments, BDNF increased the M-like current amplitude with no change in holding current. This effect was again blocked by XE991 and mimicked by retigabine, the latter accompanied with a change in holding current. In agreement with the electrophysiology, parvalbumin-positive interneurons co-expressed TrkB receptors and Kv7.2/KCNQ2 channels. In conclusion, BDNF depresses the excitability of interneurons by activating an M-like current and possibly blocking Kv1 channels, thereby controlling interneuron resting membrane potential and excitability.
Assuntos
Potenciais de Ação , Fator Neurotrófico Derivado do Encéfalo/fisiologia , Giro Denteado/fisiologia , Interneurônios/fisiologia , Animais , Giro Denteado/citologia , Giro Denteado/efeitos dos fármacos , Feminino , Proteínas de Ligação ao GTP/fisiologia , Técnicas In Vitro , Interneurônios/efeitos dos fármacos , Interneurônios/metabolismo , Masculino , Parvalbuminas/metabolismo , Técnicas de Patch-Clamp , Bloqueadores dos Canais de Potássio/farmacologia , Ratos , Ratos Wistar , Fosfolipases Tipo C/fisiologiaRESUMO
Amyotrophic lateral sclerosis (ALS) is the most common adult-onset motor neuron disease. It is a fatal degenerative disease, best recognized for its debilitating neuromuscular effects. ALS however also induces cognitive impairments in as many as 50% of affected individuals. Moreover, many ALS patients demonstrate cortical hyperexcitability, which has been shown to precede the onset of clinical symptoms. The wobbler mouse is a model of ALS, and like ALS patients the wobbler mouse displays cortical hyperexcitability. Here we investigated if the neocortical aberrations of the wobbler mouse also occur in the hippocampus. Consequently, we performed extracellular field excitatory postsynaptic potential recordings in the CA1 region of the hippocampus on acute brain slices from symptomatic (P45-P60) and presymptomatic (P17-P21) wobbler mice. Significant increased excitation of hippocampal synapses was revealed by leftward shifted input/output-curves in both symptomatic and presymptomatic wobbler mice, and substantiated by population spike occurrence analyses, demonstrating that the increased synaptic excitation precedes the onset of visible phenotypic symptoms in the mouse. Synaptic facilitation tested by paired-pulse facilitation and trains in wobbler and control mice showed no differences, suggesting the absence of presynaptic defects. Immunohistochemical staining revealed that symptomatic wobbler mice have a lower number of parvalbumin positive interneurons when compared to controls and presymptomatic mice. This study reveals that the wobbler mouse model of ALS exhibits hippocampal hyperexcitability. We suggest that the hyperexcitability could be caused by increased excitatory synaptic transmission and a concomitant reduced inhibition due to a decreased number of parvalbumin positive interneurons. Thus we substantiate that wobbler brain impairments are not confined to the motor cortex, but extend to the hippocampus. Importantly, we have revealed more details of the early pathophysiology in asymptomatic animals, and studies like the present may facilitate the development of novel treatment strategies for earlier intervention in ALS patients in the future.
Assuntos
Esclerose Lateral Amiotrófica/patologia , Esclerose Lateral Amiotrófica/fisiopatologia , Hipocampo/patologia , Hipocampo/fisiopatologia , Interneurônios/patologia , Terminações Pré-Sinápticas/metabolismo , Animais , Modelos Animais de Doenças , Camundongos , Células Piramidais/metabolismo , Células Piramidais/fisiopatologia , Potenciais SinápticosRESUMO
δ-subunit containing extrasynaptic GABA(A) receptors are potential targets for modifying neuronal activity in a range of brain disorders. With the aim of gaining more insight in synaptic and extrasynaptic inhibition, we used a new positive modulator, AA29504, of δ-subunit containing GABA(A) receptors in mouse neurons in vitro and in vivo. Whole-cell patch-clamp recordings were carried out in the dentate gyrus in mouse brain slices. In granule cells, AA29504 (1 µM) caused a 4.2-fold potentiation of a tonic current induced by THIP (1 µM), while interneurons showed a potentiation of 2.6-fold. Moreover, AA29504 (1 µM) increased the amplitude and prolonged the decay of miniature inhibitory postsynaptic currents (mIPSCs) in granule cells, and this effect was abolished by Zn²âº (15 µM). AA29504 (1 µM) also induced a small tonic current (12.7 ± 3.2 pA) per se, and when evaluated in a nominally GABA-free environment using Ca²âº imaging in cultured neurons, AA29504 showed GABA(A) receptor agonism in the absence of agonist. Finally, AA29504 exerted dose-dependent stress-reducing and anxiolytic effects in mice in vivo. We propose that AA29504 potentiates δ-containing GABA(A) receptors to enhance tonic inhibition, and possibly recruits perisynaptic δ-containing receptors to participate in synaptic phasic inhibition in dentate gyrus.
Assuntos
GABAérgicos/farmacologia , Agonistas GABAérgicos/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Receptores de GABA-A/fisiologia , Animais , Ansiedade/tratamento farmacológico , Ansiedade/psicologia , Encéfalo/metabolismo , Cálcio/metabolismo , Interpretação Estatística de Dados , Giro Denteado/citologia , Giro Denteado/efeitos dos fármacos , Fenômenos Eletrofisiológicos , Febre/etiologia , GABAérgicos/metabolismo , Isoxazóis/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Técnicas de Patch-Clamp , Células Piramidais/efeitos dos fármacos , Ensaio Radioligante , Receptores de GABA-A/efeitos dos fármacos , Estresse Psicológico/fisiopatologia , Transmissão Sináptica/efeitos dos fármacosRESUMO
Succinic semialdehyde dehydrogenase (SSADH; aldehyde dehydrogenase 5a1, ALDH5A1; E.C. 1.2.1.24; OMIM 610045, 271980) deficiency is a rare heritable disorder that disrupts the metabolism of the inhibitory neurotransmitter 4-aminobutyric acid (GABA). Identified in conjunction with increased urinary excretion of the GABA analog gamma-hydroxybutyric acid (GHB), numerous patients have been identified worldwide and the autosomal-recessive disorder has been modeled in mice. The phenotype is one of nonprogressive neurological dysfunction in which seizures may be prominently displayed. The murine model is a reasonable phenocopy of the human disorder, yet the severity of the seizure disorder in the mouse exceeds that observed in SSADH-deficient patients. Abnormalities in GABAergic and GHBergic neurotransmission, documented in patients and mice, form a component of disease pathophysiology, although numerous other disturbances (metabolite accumulations, myelin abnormalities, oxidant stress, neurosteroid depletion, altered bioenergetics, etc.) are also likely to be involved in developing the disease phenotype. Most recently, the demonstration of a redox control system in the SSADH protein active site has provided new insights into the regulation of SSADH by the cellular oxidation/reduction potential. The current review summarizes some 30 years of research on this protein and disease, addressing pathological mechanisms in human and mouse at the protein, metabolic, molecular, and whole-animal level.
Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/enzimologia , Succinato-Semialdeído Desidrogenase/metabolismo , Ácido gama-Aminobutírico/metabolismo , Aldeídos/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/patologia , Animais , Domínio Catalítico , Deficiências do Desenvolvimento , Modelos Animais de Doenças , Humanos , Hidroxibutiratos/urina , Camundongos , Oxirredução , Succinato-Semialdeído Desidrogenase/química , Succinato-Semialdeído Desidrogenase/deficiência , Succinato-Semialdeído Desidrogenase/genéticaRESUMO
Succinic semialdehyde dehydrogenase (SSADH) deficiency is an autosomal-recessively inherited disorder of gamma-aminobutyrate (GABA) catabolism characterized by ataxia and epilepsy. Since SSADH is responsible for GABA break-down downstream of GABA transaminase, patients manifest high extracellular levels of GABA, as well as the GABA(B) receptor (GABA(B)R) agonist gamma-hydroxybutyrate (GHB). SSADH knockout (KO) mice display absence seizures, which progress into lethal tonic-clonic seizures at around 3weeks of age. It is hypothesized that desensitization of GABA(B)Rs plays an important role in the disease, although detailed studies of pre- and postsynaptic GABA(B)Rs are not available. We performed patch-clamp recordings from layer 2/3 pyramidal neurons in neocortical brain slices of wild-type (WT) and SSADH KO mice. Electrical stimulation of GABAergic fibers during wash in of the GABA(B)R agonist baclofen revealed no difference in presynaptic GABA(B)R mediated inhibition of GABA release between WT and SSADH KO mice. In contrast, a significant decrease in postsynaptic baclofen-induced potassium currents was seen in SSADH KO mice. This reduction was unlikely to be caused by accumulation of potassium, GABA or GHB in the brain slices, or an altered expression of regulators of G-protein signaling (RGS) proteins. Finally, adenosine-induced potassium currents were also reduced in SSADH KO mice, which could suggest heterologous desensitization of the G-protein dependent effectors, leading to a reduction in G-protein coupled inwardly rectifying potassium (GIRK) channel responses. Our findings indicate that high GABA and GHB levels desensitize postsynaptic, but not certain presynaptic, GABA(B)Rs, promoting a decrease in GIRK channel function. These changes could contribute to the development of seizures in SSADH KO mice and potentially also in affected patients.
Assuntos
Plasticidade Neuronal/fisiologia , Terminações Pré-Sinápticas/enzimologia , Receptores de GABA-B/fisiologia , Succinato-Semialdeído Desidrogenase/deficiência , Succinato-Semialdeído Desidrogenase/genética , Transmissão Sináptica/genética , Adenosina/farmacologia , Animais , Baclofeno/farmacologia , Modelos Animais de Doenças , Antagonistas GABAérgicos/farmacologia , Camundongos , Camundongos Knockout , Plasticidade Neuronal/efeitos dos fármacos , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/metabolismo , Receptores de GABA-B/metabolismo , Transmissão Sináptica/efeitos dos fármacos , Ácido gama-Aminobutírico/metabolismoRESUMO
Human and murine succinic semialdehyde dehydrogenase (SSADH; gamma-hydroxybutyric (GHB) aciduria) deficiency represents an epileptic disorder associated with hyperGABA- and hyperGHB-ergic states. Despite significant neurotransmitters alterations, well-defined single-cell electrophysiological studies, aimed to provide insight into regional neuropathology, have been lacking. In this study, we characterized the effect of residual SSADH enzyme function/increased GABA levels on single-cell hippocampal electrophysiology in SSADH+/+ (wild-type; WT), SSADH+/- (heterozygous; HET), and SSADH-/- (knock-out; KO) mice. Tonic extrasynaptic GABAA receptor (GABAAR)-mediated currents were elevated in HET and KO mice, whereas phasic synaptic GABAAR currents were unaltered in dentate gyrus granule cells. Similarly, tonic GABAAR-mediated currents were increased in dentate gyrus interneurons of KO animals, while phasic GABAergic neurotransmission was unaffected in the same cells. Our results indicate global disruption of cortical networks in SSADH KO mice, affecting both excitatory and inhibitory neurons. Our findings provide new clues concerning seizure evolution in the murine model (absence-->tonic-clonic-->status epilepticus), and extend pathophysiological insight into human SSADH deficiency.
Assuntos
Dosagem de Genes/genética , Hipocampo/patologia , Potenciais da Membrana/genética , Neurônios/fisiologia , Convulsões , Succinato-Semialdeído Desidrogenase/deficiência , Animais , Animais Recém-Nascidos , Biofísica , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Estimulação Elétrica/métodos , Feminino , Antagonistas GABAérgicos/farmacologia , Humanos , Técnicas In Vitro , Modelos Lineares , Lisina/análogos & derivados , Masculino , Potenciais da Membrana/efeitos dos fármacos , Camundongos , Camundongos Knockout , Neurônios/classificação , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp/métodos , Piridazinas/farmacologia , Convulsões/genética , Convulsões/patologia , Convulsões/fisiopatologia , Ácido gama-Aminobutírico/metabolismoRESUMO
We have previously demonstrated that L-type Ca(2+) channels are involved in post-tetanic potentiation (PTP) of GABAergic IPSCs in cultured hippocampal neurons. Here we have used intracellular Fluo-3 to detect [Ca(2+)](i) in single GABAergic boutons in response to stimulation that evokes PTP. During control stimulation of the presynaptic GABAergic neuron at 40 Hz for 1-2 s, DeltaF/F(0) increased rapidly to a peak value and started to decline shortly after the train ended, returning to baseline within 10-20 s. The L-type channel blocker, isradipine (5 microM), had no significant effect on the amplitude or kinetics of the Ca(2+) signal. Following blockade of N- and P/Q-type Ca(2+)-channels, the amplitude was reduced by 52.9+/-3%. Isradipine caused a reduction of the remaining response (by 26.6+/-5%, P<0.01), that was fully reversible on washing. The L-type channel "agonist", BayK 8644 (8 microM), caused a significant enhancement of the peak (by 18.7%+/-7%, P<0.05). The rising phase of the Ca(2+) signal, which is related to the rate of entry of Ca(2+) into the bouton, was decreased by isradipine (by 25.5+/-6%, P<0.05) and enhanced by BayK 8644 (by 45.2%+/-16%, P<0.05). These Ca(2+) imaging experiments support the putative role of L-type channels in PTP of GABAergic synapses on cultured hippocampal neurons. We expect L-channels to be few in number, although they may couple strongly to intracellular signalling cascades that could amplify a signal that regulates synaptic vesicle turnover in the GABAergic boutons.
Assuntos
Canais de Cálcio Tipo L/metabolismo , Cálcio/metabolismo , Hipocampo/fisiologia , Neurônios/fisiologia , Terminações Pré-Sinápticas/fisiologia , Receptores Pré-Sinápticos/metabolismo , Ácido gama-Aminobutírico/fisiologia , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil)/farmacologia , Animais , Canais de Cálcio Tipo N/metabolismo , Células Cultivadas , Hipocampo/citologia , Isradipino/farmacologia , Potenciais da Membrana , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Ratos WistarRESUMO
Activity of extrasynaptic GABA A receptors mediating tonic inhibition is thought to play an important role for the excitability of the mammalian cerebral cortex. However, little is known about the cell type-specific expression of tonic inhibition in particular types of cortical interneurons. Here, we used transgenic mice expressing green fluorescent protein (GFP) in somatostatin-positive (SOM) interneurons and investigated tonic inhibition in SOM interneurons versus pyramidal cells in neocortical layers 2/3. In brain slices, pyramidal cells showed a tonic current of 66 +/- 19 pA in response to the delta-subunit selective GABA A agonist THIP (1 microM). On the other hand, tonic inhibition was absent in SOM interneurons (8 +/- 1 pA) in response to THIP. As opposed to pyramidal cells, SOM interneurons were also insensitive to the delta-subunit preferring neurosteroid allotetrahydrodeoxycorticosterone (THDOC) (100 nM) and to elevated endogenous GABA levels in the slice. Finally, SOM interneurons received only 45% of the phasic charge transfer during GABA A receptor-mediated synaptic activity compared with pyramidal cells. Altogether, our study indicates that SOM interneurons receive relatively weak inhibitory input and cannot be brought under the influence of tonic inhibition.