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1.
Physiol Rev ; 103(2): 1095-1135, 2023 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-36302178

RESUMO

Synaptic inhibition plays a crucial role in regulating neuronal excitability, which is the foundation of nervous system function. This inhibition is largely mediated by the neurotransmitters GABA and glycine that activate Cl--permeable ion channels, which means that the strength of inhibition depends on the Cl- gradient across the membrane. In neurons, the Cl- gradient is primarily mediated by two secondarily active cation-chloride cotransporters (CCCs), NKCC1 and KCC2. CCC-mediated regulation of the neuronal Cl- gradient is critical for healthy brain function, as dysregulation of CCCs has emerged as a key mechanism underlying neurological disorders including epilepsy, neuropathic pain, and autism spectrum disorder. This review begins with an overview of neuronal chloride transporters before explaining the dependent relationship between these CCCs, Cl- regulation, and inhibitory synaptic transmission. We then discuss the evidence for how CCCs can be regulated, including by activity and their protein interactions, which underlie inhibitory synaptic plasticity. For readers who may be interested in conducting experiments on CCCs and neuronal excitability, we have included a section on techniques for estimating and recording intracellular Cl-, including their advantages and limitations. Although the focus of this review is on neurons, we also examine how Cl- is regulated in glial cells, which in turn regulate neuronal excitability through the tight relationship between this nonneuronal cell type and synapses. Finally, we discuss the relatively extensive and growing literature on how CCC-mediated neuronal excitability contributes to neurological disorders.


Assuntos
Transtorno do Espectro Autista , Doenças do Sistema Nervoso , Simportadores , Humanos , Cloretos/metabolismo , Simportadores/metabolismo , Neurônios/metabolismo , Doenças do Sistema Nervoso/metabolismo , Proteínas de Membrana Transportadoras
2.
Mol Cell ; 77(6): 1176-1192.e16, 2020 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-31999954

RESUMO

Microexons represent the most highly conserved class of alternative splicing, yet their functions are poorly understood. Here, we focus on closely related neuronal microexons overlapping prion-like domains in the translation initiation factors, eIF4G1 and eIF4G3, the splicing of which is activity dependent and frequently disrupted in autism. CRISPR-Cas9 deletion of these microexons selectively upregulates synaptic proteins that control neuronal activity and plasticity and further triggers a gene expression program mirroring that of activated neurons. Mice lacking the Eif4g1 microexon display social behavior, learning, and memory deficits, accompanied by altered hippocampal synaptic plasticity. We provide evidence that the eIF4G microexons function as a translational brake by causing ribosome stalling, through their propensity to promote the coalescence of cytoplasmic granule components associated with translation repression, including the fragile X mental retardation protein FMRP. The results thus reveal an autism-disrupted mechanism by which alternative splicing specializes neuronal translation to control higher order cognitive functioning.


Assuntos
Transtorno Autístico/fisiopatologia , Disfunção Cognitiva/patologia , Fator de Iniciação Eucariótico 4G/fisiologia , Éxons/genética , Proteína do X Frágil da Deficiência Intelectual/metabolismo , Neuroblastoma/patologia , Neurônios/patologia , Animais , Comportamento Animal , Disfunção Cognitiva/genética , Disfunção Cognitiva/metabolismo , Proteína do X Frágil da Deficiência Intelectual/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neuroblastoma/genética , Neuroblastoma/metabolismo , Neurogênese , Neurônios/metabolismo , Biossíntese de Proteínas , Splicing de RNA , Células Tumorais Cultivadas
3.
Mol Cell ; 64(6): 1023-1034, 2016 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-27984743

RESUMO

A key challenge in understanding and ultimately treating autism is to identify common molecular mechanisms underlying this genetically heterogeneous disorder. Transcriptomic profiling of autistic brains has revealed correlated misregulation of the neuronal splicing regulator nSR100/SRRM4 and its target microexon splicing program in more than one-third of analyzed individuals. To investigate whether nSR100 misregulation is causally linked to autism, we generated mutant mice with reduced levels of this protein and its target splicing program. Remarkably, these mice display multiple autistic-like features, including altered social behaviors, synaptic density, and signaling. Moreover, increased neuronal activity, which is often associated with autism, results in a rapid decrease in nSR100 and splicing of microexons that significantly overlap those misregulated in autistic brains. Collectively, our results provide evidence that misregulation of an nSR100-dependent splicing network controlled by changes in neuronal activity is causally linked to a substantial fraction of autism cases.


Assuntos
Processamento Alternativo , Transtorno do Espectro Autista/genética , Haploinsuficiência , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , Animais , Transtorno do Espectro Autista/metabolismo , Transtorno do Espectro Autista/fisiopatologia , Modelos Animais de Doenças , Embrião de Mamíferos , Éxons , Feminino , Expressão Gênica , Humanos , Masculino , Potenciais da Membrana , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/metabolismo , Neurônios/patologia , Reflexo de Sobressalto , Transmissão Sináptica
4.
J Physiol ; 599(2): 485-492, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32162694

RESUMO

Kainate receptors (KARs) are glutamate-type receptors that mediate both canonical ionotropic currents and non-canonical metabotropic signalling. While KARs are expressed widely throughout the brain, synaptic KAR currents have only been recorded at a limited set of synapses, and the KAR currents that have been recorded are relatively small and slow, which has led to the question, what is the functional significance of KARs? While the KAR current itself is relatively modest, its impact on inhibition in the hippocampus can be profound. In the CA1 region of the hippocampus, presynaptic KAR activation bidirectionally regulates γ-aminobutyric acid (GABA) release in a manner that depends on the glutamate concentration; lower levels of glutamate facilitate GABA release via an ionotropic pathway, while higher levels of glutamate depress GABA release via a metabotropic pathway. Postsynaptic interneuron KAR activation increases spike frequency through an ionotropic current, which in turn can strengthen inhibition. In the CA3 region, postsynaptic KAR activation in pyramidal neurons also strengthens inhibition, but in this case through a metabotropic pathway which regulates the neuronal chloride gradient and hyperpolarizes the reversal potential for GABA (EGABA ). Taken together, the evidence for KAR-mediated regulation of the strength of inhibition via pre- and postsynaptic mechanisms provides compelling evidence that KARs are ideally positioned to regulate excitation-inhibition balance - through sensing the excitatory tone and concomitantly tuning the strength of inhibition.


Assuntos
Hipocampo , Receptores de Ácido Caínico , Hipocampo/metabolismo , Interneurônios/metabolismo , Células Piramidais/metabolismo , Receptores de Ácido Caínico/metabolismo , Sinapses/metabolismo
5.
Brain ; 143(3): 800-810, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-32203578

RESUMO

Amyotrophic lateral sclerosis is a fatal disease resulting from motor neuron degeneration in the cortex and spinal cord. Cortical hyperexcitability is a hallmark feature of amyotrophic lateral sclerosis and is accompanied by decreased intracortical inhibition. Using electrophysiological patch-clamp recordings, we revealed parvalbumin interneurons to be hypoactive in the late pre-symptomatic SOD1*G93A mouse model of amyotrophic lateral sclerosis. We discovered that using adeno-associated virus-mediated delivery of chemogenetic technology targeted to increase the activity of the interneurons within layer 5 of the primary motor cortex, we were able to rescue intracortical inhibition and reduce pyramidal neuron hyperexcitability. Increasing the activity of interneurons in the layer 5 of the primary motor cortex was effective in delaying the onset of amyotrophic lateral sclerosis-associated motor deficits, slowing symptom progression, preserving neuronal populations, and increasing the lifespan of SOD1*G93A mice. Taken together, this study provides novel insights into the pathogenesis and treatment of amyotrophic lateral sclerosis.


Assuntos
Esclerose Lateral Amiotrófica/fisiopatologia , Interneurônios/fisiologia , Córtex Motor/fisiologia , Inibição Neural/fisiologia , Adenoviridae , Animais , Progressão da Doença , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Destreza Motora/fisiologia , Técnicas de Patch-Clamp , Células Piramidais/fisiologia , Superóxido Dismutase-1/genética , Transfecção
6.
Proc Natl Acad Sci U S A ; 115(7): E1618-E1626, 2018 02 13.
Artigo em Inglês | MEDLINE | ID: mdl-29382760

RESUMO

Huntington's disease (HD) is classically characterized as a movement disorder, however cognitive impairments precede the motor symptoms by ∼15 y. Based on proteomic and bioinformatic data linking the Huntingtin protein (Htt) and KCC2, which is required for hyperpolarizing GABAergic inhibition, and the important role of inhibition in learning and memory, we hypothesized that aberrant KCC2 function contributes to the hippocampal-associated learning and memory deficits in HD. We discovered that Htt and KCC2 interact in the hippocampi of wild-type and R6/2-HD mice, with a decrease in KCC2 expression in the hippocampus of R6/2 and YAC128 mice. The reduced expression of the Cl--extruding cotransporter KCC2 is accompanied by an increase in the Cl--importing cotransporter NKCC1, which together result in excitatory GABA in the hippocampi of HD mice. NKCC1 inhibition by the FDA-approved NKCC1 inhibitor bumetanide abolished the excitatory action of GABA and rescued the performance of R6/2 mice on hippocampal-associated behavioral tests.


Assuntos
Doença de Huntington/metabolismo , Doença de Huntington/psicologia , Transtornos da Memória/psicologia , Memória , Ácido gama-Aminobutírico/metabolismo , Animais , Bumetanida/administração & dosagem , Modelos Animais de Doenças , Feminino , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Humanos , Proteína Huntingtina/genética , Proteína Huntingtina/metabolismo , Doença de Huntington/tratamento farmacológico , Doença de Huntington/genética , Masculino , Memória/efeitos dos fármacos , Transtornos da Memória/etiologia , Transtornos da Memória/metabolismo , Camundongos , Camundongos Transgênicos , Membro 2 da Família 12 de Carreador de Soluto/genética , Membro 2 da Família 12 de Carreador de Soluto/metabolismo , Simportadores/genética , Simportadores/metabolismo , Cotransportadores de K e Cl-
7.
J Physiol ; 597(6): 1677-1690, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30570751

RESUMO

KEY POINTS: Potassium-chloride co-transporter 2 (KCC2) plays a critical role in regulating chloride homeostasis, which is essential for hyperpolarizing inhibition in the mature nervous system. KCC2 interacts with many proteins involved in excitatory neurotransmission, including the GluK2 subunit of the kainate receptor (KAR). We show that activation of KARs hyperpolarizes the reversal potential for GABA (EGABA ) via both ionotropic and metabotropic signalling mechanisms. KCC2 is required for the metabotropic KAR-mediated regulation of EGABA , although ionotropic KAR signalling can hyperpolarize EGABA independent of KCC2 transporter function. The KAR-mediated hyperpolarization of EGABA is absent in the GluK1/2-/- mouse and is independent of zinc release from mossy fibre terminals. The ability of KARs to regulate KCC2 function may have implications in diseases with disrupted excitation: inhibition balance, such as epilepsy, neuropathic pain, autism spectrum disorders and Down's syndrome. ABSTRACT: Potassium-chloride co-transporter 2 (KCC2) plays a critical role in the regulation of chloride (Cl- ) homeostasis within mature neurons. KCC2 is a secondarily active transporter that extrudes Cl- from the neuron, which maintains a low intracellular Cl- concentration [Cl- ]. This results in a hyperpolarized reversal potential of GABA (EGABA ), which is required for fast synaptic inhibition in the mature central nervous system. KCC2 also plays a structural role in dendritic spines and at excitatory synapses, and interacts with 'excitatory' proteins, including the GluK2 subunit of kainate receptors (KARs). KARs are glutamate receptors that display both ionotropic and metabotropic signalling. We show that activating KARs in the hippocampus hyperpolarizes EGABA , thus strengthening inhibition. This hyperpolarization occurs via both ionotropic and metabotropic KAR signalling in the CA3 region, whereas it is absent in the GluK1/2-/- mouse, and is independent of zinc release from mossy fibre terminals. The metabotropic signalling mechanism is dependent on KCC2, although the ionotropic signalling mechanism produces a hyperpolarization of EGABA even in the absence of KCC2 transporter function. These results demonstrate a novel functional interaction between a glutamate receptor and KCC2, a transporter critical for maintaining inhibition, suggesting that the KAR:KCC2 complex may play an important role in excitatory:inhibitory balance in the hippocampus. Additionally, the ability of KARs to regulate chloride homeostasis independently of KCC2 suggests that KAR signalling can regulate inhibition via multiple mechanisms. Activation of kainate-type glutamate receptors could serve as an important mechanism for increasing the strength of inhibition during periods of strong glutamatergic activity.


Assuntos
Cloretos/metabolismo , Potenciais Pós-Sinápticos Inibidores , Células Piramidais/metabolismo , Receptores de GABA/metabolismo , Receptores de Ácido Caínico/metabolismo , Animais , Região CA1 Hipocampal/citologia , Região CA1 Hipocampal/metabolismo , Região CA1 Hipocampal/fisiologia , Região CA3 Hipocampal/citologia , Região CA3 Hipocampal/metabolismo , Região CA3 Hipocampal/fisiologia , Células Cultivadas , Feminino , Homeostase , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fibras Musgosas Hipocampais/metabolismo , Fibras Musgosas Hipocampais/fisiologia , Células Piramidais/fisiologia , Simportadores/metabolismo , Cotransportadores de K e Cl- , Receptor de GluK2 Cainato
8.
J Neurosci ; 37(45): 10792-10799, 2017 11 08.
Artigo em Inglês | MEDLINE | ID: mdl-29118207

RESUMO

Inhibitory circuits are diverse, yet with a poorly understood cell biology. Functional characterization of distinct inhibitory neuron subtypes has not been sufficient to explain how GABAergic neurotransmission sculpts principal cell activity in a relevant fashion. Our Mini-Symposium brings together several emerging mechanisms that modulate GABAergic neurotransmission dynamically from either the presynaptic or the postsynaptic site. The first two talks discuss novel developmental and neuronal subtype-specific contributions to the excitatory/inhibitory balance and circuit maturation. The next three talks examine how interactions between cellular pathways, lateral diffusion of proteins between synapses, and chloride transporter function at excitatory and inhibitory synapses and facilitate inhibitory synapse adaptations. Finally, we address functional differences within GABAergic interneurons to highlight the importance of diverse, flexible, and versatile inputs that shape network function. Together, the selection of topics demonstrates how developmental and activity-dependent mechanisms coordinate inhibition in relation to the excitatory inputs and vice versa.


Assuntos
Sinapses/fisiologia , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/fisiologia , Animais , Humanos , Rede Nervosa/citologia , Rede Nervosa/fisiologia , Plasticidade Neuronal
9.
J Biol Chem ; 292(15): 6190-6201, 2017 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-28235805

RESUMO

Synaptic inhibition depends on a transmembrane gradient of chloride, which is set by the neuron-specific K+-Cl- co-transporter KCC2. Reduced KCC2 levels in the neuronal membrane contribute to the generation of epilepsy, neuropathic pain, and autism spectrum disorders; thus, it is important to characterize the mechanisms regulating KCC2 expression. In the present study, we determined the role of KCC2-protein interactions in regulating total and surface membrane KCC2 expression. Using quantitative immunofluorescence in cultured mouse hippocampal neurons, we discovered that the kainate receptor subunit GluK2 and the auxiliary subunit Neto2 significantly increase the total KCC2 abundance in neurons but that GluK2 exclusively increases the abundance of KCC2 in the surface membrane. Using a live cell imaging assay, we further determined that KCC2 recycling primarily occurs within 1-2 h and that GluK2 produces an ∼40% increase in the amount of KCC2 recycled to the membrane during this time period. This GluK2-mediated increase in surface recycling translated to a significant increase in KCC2 expression in the surface membrane. Moreover, we found that KCC2 recycling is enhanced by protein kinase C-mediated phosphorylation of the GluK2 C-terminal residues Ser-846 and Ser-868. Lastly, using gramicidin-perforated patch clamp recordings, we found that the GluK2-mediated increase in KCC2 recycling to the surface membrane translates to a hyperpolarization of the reversal potential for GABA (EGABA). In conclusion, our results have revealed a mechanism by which kainate receptors regulate KCC2 expression in the hippocampus.


Assuntos
Membrana Celular/metabolismo , Hipocampo/metabolismo , Potenciais da Membrana/fisiologia , Neurônios/metabolismo , Receptores de Ácido Caínico/metabolismo , Simportadores/metabolismo , Animais , Membrana Celular/genética , Células Cultivadas , Hipocampo/citologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Neurônios/citologia , Receptores de Ácido Caínico/genética , Simportadores/genética , Cotransportadores de K e Cl- , Receptor de GluK2 Cainato
10.
Proc Natl Acad Sci U S A ; 112(3): 851-6, 2015 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-25561528

RESUMO

Glioblastoma (GBM) is a cancer comprised of morphologically, genetically, and phenotypically diverse cells. However, an understanding of the functional significance of intratumoral heterogeneity is lacking. We devised a method to isolate and functionally profile tumorigenic clones from patient glioblastoma samples. Individual clones demonstrated unique proliferation and differentiation abilities. Importantly, naïve patient tumors included clones that were temozolomide resistant, indicating that resistance to conventional GBM therapy can preexist in untreated tumors at a clonal level. Further, candidate therapies for resistant clones were detected with clone-specific drug screening. Genomic analyses revealed genes and pathways that associate with specific functional behavior of single clones. Our results suggest that functional clonal profiling used to identify tumorigenic and drug-resistant tumor clones will lead to the discovery of new GBM clone-specific treatment strategies.


Assuntos
Neoplasias Encefálicas/patologia , Glioblastoma/patologia , Antineoplásicos/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Linhagem Celular Tumoral , Dacarbazina/análogos & derivados , Dacarbazina/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Humanos , Análise de Célula Única , Temozolomida
11.
J Physiol ; 594(10): 2593-605, 2016 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-26876607

RESUMO

KCC2 is the central regulator of neuronal Cl(-) homeostasis, and is critical for enabling strong hyperpolarizing synaptic inhibition in the mature brain. KCC2 hypofunction results in decreased inhibition and increased network hyperexcitability that underlies numerous disease states including epilepsy, neuropathic pain and neuropsychiatric disorders. The current holy grail of KCC2 biology is to identify how we can rescue KCC2 hypofunction in order to restore physiological levels of synaptic inhibition and neuronal network activity. It is becoming increasingly clear that diverse cellular signals regulate KCC2 surface expression and function including neurotransmitters and neuromodulators. In the present review we explore the existing evidence that G-protein-coupled receptor (GPCR) signalling can regulate KCC2 activity in numerous regions of the nervous system including the hypothalamus, hippocampus and spinal cord. We present key evidence from the literature suggesting that GPCR signalling is a conserved mechanism for regulating chloride homeostasis. This evidence includes: (1) the activation of group 1 metabotropic glutamate receptors and metabotropic Zn(2+) receptors strengthens GABAergic inhibition in CA3 pyramidal neurons through a regulation of KCC2; (2) activation of the 5-hydroxytryptamine type 2A serotonin receptors upregulates KCC2 cell surface expression and function, restores endogenous inhibition in motoneurons, and reduces spasticity in rats; and (3) activation of A3A-type adenosine receptors rescues KCC2 dysfunction and reverses allodynia in a model of neuropathic pain. We propose that GPCR-signals are novel endogenous Cl(-) extrusion enhancers that may regulate KCC2 function.


Assuntos
Cloretos/fisiologia , Homeostase/fisiologia , Neurônios/fisiologia , Neurotransmissores/fisiologia , Simportadores/fisiologia , Animais , Humanos , Transdução de Sinais/fisiologia
12.
Proc Natl Acad Sci U S A ; 110(9): 3561-6, 2013 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-23401525

RESUMO

KCC2 is a neuron-specific K(+)-Cl(-) cotransporter that is essential for Cl(-) homeostasis and fast inhibitory synaptic transmission in the mature CNS. Despite the critical role of KCC2 in neurons, the mechanisms regulating its function are not understood. Here, we show that KCC2 is critically regulated by the single-pass transmembrane protein neuropilin and tolloid like-2 (Neto2). Neto2 is required to maintain the normal abundance of KCC2 and specifically associates with the active oligomeric form of the transporter. Loss of the Neto2:KCC2 interaction reduced KCC2-mediated Cl(-) extrusion, resulting in decreased synaptic inhibition in hippocampal neurons.


Assuntos
Cloretos/metabolismo , Hipocampo/citologia , Proteínas de Membrana/deficiência , Neurônios/metabolismo , Simportadores/metabolismo , Potenciais de Ação/fisiologia , Sequência de Aminoácidos , Animais , Transporte Biológico , Espectrometria de Massas , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Neurônios/citologia , Ligação Proteica , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína , Relação Estrutura-Atividade , Simportadores/química , Ácido gama-Aminobutírico/metabolismo , Cotransportadores de K e Cl-
13.
J Neurosci ; 34(45): 14948-60, 2014 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-25378161

RESUMO

Hyperactivity within the ventral hippocampus (vHPC) has been linked to both psychosis in humans and behavioral deficits in animal models of schizophrenia. A local decrease in GABA-mediated inhibition, particularly involving parvalbumin (PV)-expressing GABA neurons, has been proposed as a key mechanism underlying this hyperactive state. However, direct evidence is lacking for a causal role of vHPC GABA neurons in behaviors associated with schizophrenia. Here, we probed the behavioral function of two different but overlapping populations of vHPC GABA neurons that express either PV or GAD65 by selectively inhibiting these neurons with the pharmacogenetic neuromodulator hM4D. We show that acute inhibition of vHPC GABA neurons in adult mice results in behavioral changes relevant to schizophrenia. Inhibiting either PV or GAD65 neurons produced distinct behavioral deficits. Inhibition of PV neurons, affecting ∼80% of the PV neuron population, robustly impaired prepulse inhibition of the acoustic startle reflex (PPI), startle reactivity, and spontaneous alternation, but did not affect locomotor activity. In contrast, inhibiting a heterogeneous population of GAD65 neurons, affecting ∼40% of PV neurons and 65% of cholecystokinin neurons, increased spontaneous and amphetamine-induced locomotor activity and reduced spontaneous alternation, but did not alter PPI. Inhibition of PV or GAD65 neurons also produced distinct changes in network oscillatory activity in the vHPC in vivo. Together, these findings establish a causal role for vHPC GABA neurons in controlling behaviors relevant to schizophrenia and suggest a functional dissociation between the GABAergic mechanisms involved in hippocampal modulation of sensorimotor processes.


Assuntos
Neurônios GABAérgicos/fisiologia , Hipocampo/fisiologia , Interneurônios/fisiologia , Aprendizagem em Labirinto , Inibição Neural , Reflexo de Sobressalto , Esquizofrenia/fisiopatologia , Potenciais de Ação , Animais , Clozapina/análogos & derivados , Clozapina/farmacologia , Neurônios GABAérgicos/metabolismo , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/metabolismo , Hipocampo/citologia , Hipocampo/metabolismo , Interneurônios/efeitos dos fármacos , Interneurônios/metabolismo , Locomoção , Camundongos , Parvalbuminas/genética , Parvalbuminas/metabolismo , Receptor Muscarínico M4/agonistas , Esquizofrenia/metabolismo , Potenciais Sinápticos
14.
Proc Natl Acad Sci U S A ; 108(27): 11274-9, 2011 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-21690381

RESUMO

Anoxic insults cause hyperexcitability and cell death in mammalian neurons. Conversely, in anoxia-tolerant turtle brain, spontaneous electrical activity is suppressed by anoxia (i.e., spike arrest; SA) and cell death does not occur. The mechanism(s) of SA is unknown but likely involves GABAergic synaptic transmission, because GABA concentration increases dramatically in anoxic turtle brain. We investigated this possibility in turtle cortical neurons exposed to anoxia and/or GABA(A/B) receptor (GABAR) modulators. Anoxia increased endogenous slow phasic GABAergic activity, and both anoxia and GABA reversibly induced SA by increasing GABA(A)R-mediated postsynaptic activity and Cl(-) conductance, which eliminated the Cl(-) driving force by depolarizing membrane potential (∼8 mV) to GABA receptor reversal potential (∼-81 mV), and dampened excitatory potentials via shunting inhibition. In addition, both anoxia and GABA decreased excitatory postsynaptic activity, likely via GABA(B)R-mediated inhibition of presynaptic glutamate release. In combination, these mechanisms increased the stimulation required to elicit an action potential >20-fold, and excitatory activity decreased >70% despite membrane potential depolarization. In contrast, anoxic neurons cotreated with GABA(A+B)R antagonists underwent seizure-like events, deleterious Ca(2+) influx, and cell death, a phenotype consistent with excitotoxic cell death in anoxic mammalian brain. We conclude that increased endogenous GABA release during anoxia mediates SA by activating an inhibitory postsynaptic shunt and inhibiting presynaptic glutamate release. This represents a natural adaptive mechanism in which to explore strategies to protect mammalian brain from low-oxygen insults.


Assuntos
Hipóxia Encefálica/fisiopatologia , Receptores de GABA-A/fisiologia , Receptores de GABA-B/fisiologia , Tartarugas/fisiologia , Potenciais de Ação , Adaptação Fisiológica , Animais , Fenômenos Eletrofisiológicos , Glutamina/fisiologia , Potenciais da Membrana , Neurônios/fisiologia , Transdução de Sinais , Ácido gama-Aminobutírico/fisiologia
15.
Neuroscience ; 537: 189-204, 2024 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-38036056

RESUMO

Rett syndrome (RTT) is a debilitating neurodevelopmental disorder caused by mutations in the X-linked methyl-CpG-binding protein 2 (MeCP2) gene, resulting in severe deficits in learning and memory. Alterations in synaptic plasticity have been reported in RTT, however most electrophysiological studies have been performed in male mice only, despite the fact that RTT is primarily found in females. In addition, most studies have focused on excitation, despite the emerging evidence for the important role of inhibition in learning and memory. Here, we performed an electrophysiological characterization in the CA1 region of the hippocampus in both males and females of RTT mouse models with a focus on neurogliaform (NGF) interneurons, given that they are the most abundant dendrite-targeting interneuron subtype in the hippocampus. We found that theta-burst stimulation (TBS) failed to induce long-term potentiation (LTP) in either pyramidal neurons or NGF interneurons in male or female RTT mice, with no apparent changes in short-term plasticity (STP). This failure to induce LTP was accompanied by excitation/inhibition (E/I) imbalances and altered excitability, in a sex- and cell-type specific manner. Specifically, NGF interneurons of male RTT mice displayed increased intrinsic excitability, a depolarized resting membrane potential, and decreased E/I balance, while in female RTT mice, the resting membrane potential was depolarized. Understanding the role of NGF interneurons in RTT animal models is crucial for developing targeted treatments to improve cognition in individuals with this disorder.


Assuntos
Síndrome de Rett , Masculino , Feminino , Camundongos , Animais , Síndrome de Rett/genética , Potenciação de Longa Duração , Proteína 2 de Ligação a Metil-CpG/metabolismo , Hipocampo/metabolismo , Plasticidade Neuronal/genética , Modelos Animais de Doenças
16.
J Neurosci ; 32(49): 17857-68, 2012 Dec 05.
Artigo em Inglês | MEDLINE | ID: mdl-23223304

RESUMO

Memory stabilization following encoding (synaptic consolidation) or memory reactivation (reconsolidation) requires gene expression and protein synthesis (Dudai and Eisenberg, 2004; Tronson and Taylor, 2007; Nader and Einarsson, 2010; Alberini, 2011). Although consolidation and reconsolidation may be mediated by distinct molecular mechanisms (Lee et al., 2004), disrupting the function of the transcription factor CREB impairs both processes (Kida et al., 2002; Mamiya et al., 2009). Phosphorylation of CREB at Ser133 recruits CREB binding protein (CBP)/p300 coactivators to activate transcription (Chrivia et al., 1993; Parker et al., 1996). In addition to this well known mechanism, CREB regulated transcription coactivators (CRTCs), previously called transducers of regulated CREB (TORC) activity, stimulate CREB-mediated transcription, even in the absence of CREB phosphorylation. Recently, CRTC1 has been shown to undergo activity-dependent trafficking from synapses and dendrites to the nucleus in excitatory hippocampal neurons (Ch'ng et al., 2012). Despite being a powerful and specific coactivator of CREB, the role of CRTC in memory is virtually unexplored. To examine the effects of increasing CRTC levels, we used viral vectors to locally and acutely increase CRTC1 in the dorsal hippocampus dentate gyrus region of mice before training or memory reactivation in context fear conditioning. Overexpressing CRTC1 enhanced both memory consolidation and reconsolidation; CRTC1-mediated memory facilitation was context specific (did not generalize to nontrained context) and long lasting (observed after virally expressed CRTC1 dissipated). CREB overexpression produced strikingly similar effects. Therefore, increasing CRTC1 or CREB function is sufficient to enhance the strength of new, as well as established reactivated, memories without compromising memory quality.


Assuntos
Giro Denteado/fisiologia , Memória/fisiologia , Fatores de Transcrição/fisiologia , Animais , Condicionamento Psicológico/fisiologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Giro Denteado/metabolismo , Medo/fisiologia , Medo/psicologia , Feminino , Genes fos/fisiologia , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Transgênicos , Neurônios/fisiologia , Cultura Primária de Células , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transfecção/métodos
17.
J Neurosci ; 32(25): 8746-51, 2012 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-22723714

RESUMO

KCC2 is the neuron-specific member of the of K(+)-Cl(-) cotransporter gene family. It is also the only member of its family that is active under physiologically normal conditions, in the absence of osmotic stress. By extruding Cl(-) from the neuron under isotonic conditions, this transporter maintains a low concentration of neuronal Cl(-), which is essential for fast inhibitory synaptic transmission by GABA and glycine in the mature nervous system. The other members of this K(+)-Cl(-) cotransporter gene family are exclusively swelling-activated. Here we demonstrate that a 15 aa region near the end of the C terminus, unique to KCC2 (termed the ISO domain), is required for KCC2 to cotransport K(+) and Cl(-) out of the neuron under isotonic conditions. We made this discovery by overexpressing chimeric KCC2-KCC4 cDNA constructs in cultured hippocampal neurons prepared from Sprague Dawley rat embryos and assaying neuronal Cl(-) through gramicidin perforated patch-clamp recordings. We found that when neurons had been transfected with a chimeric KCC2 that lacked the unique ISO domain, hyperpolarizing responses to GABA were abolished. This finding indicates that the ISO domain is required for neuronal Cl(-) regulation. Furthermore, we discovered that when KCC2 lacks the ISO domain, it still retains swelling-activated transport, which demonstrates that there are exclusive molecular determinants of isotonic and swelling-induced K(+)-Cl(-) cotransport in neurons.


Assuntos
Simportadores/fisiologia , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/fisiologia , Animais , Transporte Biológico Ativo/fisiologia , Tamanho Celular , Cloretos/metabolismo , DNA/genética , DNA/isolamento & purificação , Feminino , Hipocampo/citologia , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Transporte de Íons , Microscopia Confocal , Oócitos/metabolismo , Técnicas de Patch-Clamp , Potássio/metabolismo , Gravidez , Ratos , Ratos Sprague-Dawley , Xenopus , Cotransportadores de K e Cl-
18.
19.
Front Cell Neurosci ; 15: 817013, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35095429

RESUMO

Intracellular chloride (Cl-) levels in mature neurons must be tightly regulated for the maintenance of fast synaptic inhibition. In the mature central nervous system (CNS), synaptic inhibition is primarily mediated by gamma-amino butyric acid (GABA), which binds to Cl- permeable GABAA receptors (GABAARs). The intracellular Cl- concentration is primarily maintained by the antagonistic actions of two cation-chloride cotransporters (CCCs): Cl--importing Na+-K+-Cl- co-transporter-1 (NKCC1) and Cl- -exporting K+-Cl- co-transporter-2 (KCC2). In mature neurons in the healthy brain, KCC2 expression is higher than NKCC1, leading to lower levels of intracellular Cl-, and Cl- influx upon GABAAR activation. However, in neurons of the immature brain or in neurological disorders such as epilepsy and traumatic brain injury, impaired KCC2 function and/or enhanced NKCC1 expression lead to intracellular Cl- accumulation and GABA-mediated excitation. In Huntington's disease (HD), KCC2- and NKCC1-mediated Cl--regulation are also altered, which leads to GABA-mediated excitation and contributes to the development of cognitive and motor impairments. This review summarizes the role of Cl- (dys)regulation in the healthy and HD brain, with a focus on the basal ganglia (BG) circuitry and CCCs as potential therapeutic targets in the treatment of HD.

20.
Trends Pharmacol Sci ; 41(12): 897-899, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33097285

RESUMO

Finely tuned excitation-inhibition balance is essential for proper brain function, and loss of balance resulting from reduced synaptic inhibition is associated with neurological disorders. Savardi and colleagues have discovered a novel inhibitor of a cation-chloride transporter that is required for synaptic inhibition, and which restores behaviors associated with Down syndrome (DS) and autism spectrum disorder (ASD).

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