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1.
Elife ; 112022 12 19.
Artigo em Inglês | MEDLINE | ID: mdl-36534089

RESUMO

Spontaneous correlated activity is a universal hallmark of immature neural circuits. However, the cellular dynamics and intrinsic mechanisms underlying network burstiness in the intact developing brain are largely unknown. Here, we use two-photon Ca2+ imaging to comprehensively map the developmental trajectories of spontaneous network activity in the hippocampal area CA1 of mice in vivo. We unexpectedly find that network burstiness peaks after the developmental emergence of effective synaptic inhibition in the second postnatal week. We demonstrate that the enhanced network burstiness reflects an increased functional coupling of individual neurons to local population activity. However, pairwise neuronal correlations are low, and network bursts (NBs) recruit CA1 pyramidal cells in a virtually random manner. Using a dynamic systems modeling approach, we reconcile these experimental findings and identify network bi-stability as a potential regime underlying network burstiness at this age. Our analyses reveal an important role of synaptic input characteristics and network instability dynamics for NB generation. Collectively, our data suggest a mechanism, whereby developing CA1 performs extensive input-discrimination learning prior to the onset of environmental exploration.


Assuntos
Hipocampo , Células Piramidais , Camundongos , Animais , Hipocampo/fisiologia , Células Piramidais/fisiologia , Neurônios/fisiologia
2.
Cell Rep ; 38(13): 110568, 2022 03 29.
Artigo em Inglês | MEDLINE | ID: mdl-35354036

RESUMO

GABAergic signaling provides inhibitory stabilization and spatiotemporally coordinates the firing of recurrently connected excitatory neurons in mature cortical circuits. Inhibition thus enables self-generated neuronal activity patterns that underlie various aspects of sensation and cognition. In this review, we aim to provide a conceptual framework describing how and when GABA-releasing interneurons acquire their network functions during development. Focusing on the developing visual neocortex and hippocampus in mice and rats in vivo, we hypothesize that at the onset of patterned activity, glutamatergic neurons are stable by themselves and inhibitory stabilization is not yet functional. We review important milestones in the development of GABAergic signaling and illustrate how the cell-type-specific strengthening of synaptic inhibition toward eye opening shapes cortical network dynamics and allows the developing cortex to progressively disengage from extra-cortical synaptic drive. We translate this framework to human cortical development and discuss clinical implications for the treatment of neonatal seizures.


Assuntos
Interneurônios , Neocórtex , Animais , Hipocampo , Interneurônios/fisiologia , Camundongos , Neurônios/fisiologia , Ratos , Transdução de Sinais
4.
Proc Natl Acad Sci U S A ; 118(14)2021 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-33782119

RESUMO

NKCC1 is the primary transporter mediating chloride uptake in immature principal neurons, but its role in the development of in vivo network dynamics and cognitive abilities remains unknown. Here, we address the function of NKCC1 in developing mice using electrophysiological, optical, and behavioral approaches. We report that NKCC1 deletion from telencephalic glutamatergic neurons decreases in vitro excitatory actions of γ-aminobutyric acid (GABA) and impairs neuronal synchrony in neonatal hippocampal brain slices. In vivo, it has a minor impact on correlated spontaneous activity in the hippocampus and does not affect network activity in the intact visual cortex. Moreover, long-term effects of the developmental NKCC1 deletion on synaptic maturation, network dynamics, and behavioral performance are subtle. Our data reveal a neural network function of NKCC1 in hippocampal glutamatergic neurons in vivo, but challenge the hypothesis that NKCC1 is essential for major aspects of hippocampal development.


Assuntos
Hipocampo/crescimento & desenvolvimento , Membro 2 da Família 12 de Carreador de Soluto/fisiologia , Animais , Animais Recém-Nascidos , Ácido Glutâmico/metabolismo , Camundongos , Rede Nervosa , Neurônios/metabolismo , Sinapses/metabolismo , Córtex Visual/fisiologia , Ácido gama-Aminobutírico/metabolismo
5.
BMC Biol ; 17(1): 95, 2019 11 27.
Artigo em Inglês | MEDLINE | ID: mdl-31775747

RESUMO

BACKGROUND: Optogenetic silencing techniques have expanded the causal understanding of the functions of diverse neuronal cell types in both the healthy and diseased brain. A widely used inhibitory optogenetic actuator is eNpHR3.0, an improved version of the light-driven chloride pump halorhodopsin derived from Natronomonas pharaonis. A major drawback of eNpHR3.0 is related to its pronounced inactivation on a time-scale of seconds, which renders it unsuited for applications that require long-lasting silencing. RESULTS: Using transgenic mice and Xenopus laevis oocytes expressing an eNpHR3.0-EYFP fusion protein, we here report optimized photo-stimulation techniques that profoundly increase the stability of eNpHR3.0-mediated currents during long-term photo-stimulation. We demonstrate that optimized photo-stimulation enables prolonged hyperpolarization and suppression of action potential discharge on a time-scale of minutes. CONCLUSIONS: Collectively, our findings extend the utility of eNpHR3.0 to the long-lasting inhibition of excitable cells, thus facilitating the optogenetic dissection of neural circuits.


Assuntos
Potenciais de Ação/fisiologia , Proteínas de Bactérias/fisiologia , Halorrodopsinas/fisiologia , Neurônios/fisiologia , Optogenética/métodos , Animais , Animais Geneticamente Modificados , Encéfalo/fisiologia , Feminino , Halobacteriaceae/química , Masculino , Camundongos , Camundongos Transgênicos , Oócitos/fisiologia , Xenopus laevis
7.
Cell Rep ; 26(12): 3173-3182.e5, 2019 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-30893591

RESUMO

Synchronized activity is a universal characteristic of immature neural circuits that is essential for their developmental refinement and strongly depends on GABAergic neurotransmission. A major subpopulation of GABA-releasing interneurons (INs) expresses somatostatin (SOM) and proved critical for rhythm generation in adulthood. Here, we report a mechanism whereby SOM INs promote neuronal synchrony in the neonatal CA1 region. Combining imaging and electrophysiological approaches, we demonstrate that SOM INs and pyramidal cells (PCs) coactivate during spontaneous activity. Bidirectional optogenetic manipulations reveal excitatory GABAergic outputs to PCs that evoke correlated network events in an NKCC1-dependent manner and contribute to spontaneous synchrony. Using a dynamic systems modeling approach, we show that SOM INs affect network dynamics through a modulation of network instability and amplification threshold. Our study identifies a network function of SOM INs with implications for the activity-dependent construction of developing brain circuits.


Assuntos
Hipocampo/metabolismo , Interneurônios/metabolismo , Células Piramidais/metabolismo , Somatostatina/biossíntese , Transmissão Sináptica , Animais , Hipocampo/citologia , Interneurônios/citologia , Camundongos , Camundongos Transgênicos , Optogenética , Células Piramidais/citologia
8.
Brain Res ; 1701: 146-152, 2018 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-30205110

RESUMO

Diverse neuropsychiatric diseases were recently linked to specific anti-neuronal autoantibodies targeting synaptic proteins. Symptoms can range from epileptic seizures to cognitive impairment to movement disorders, commonly responding to treatment with immunotherapy. Several of these autoantibodies target inhibitory synapses that use GABA or glycine as neurotransmitters. Despite their relatively low abundance, inhibitory neurons are extraordinarily diverse in anatomical, electrophysiological and molecular terms, reflecting the variable clinical phenotypes of affected patients. Indeed, data on the antibody effects in neuronal cultures or animals models suggest that most of these antibodies are directly pathogenic by down-regulating synaptic proteins, activating complement or antagonizing ligand binding. The present review summarizes the current achievements in the field of humoral autoimmunity related to inhibitory networks, state-of-the-art diagnostics and clinical characterization of patients. In many instances, the phenotypic spectrum of patients with GABA receptor, glycine receptor, amphiphysin or GAD65 antibodies mirror the experimental findings, suggesting that ongoing work will markedly contribute to the better understanding of pathophysiology in this exciting patient group and might pave the way for disease-specific immunotherapy.


Assuntos
Autoanticorpos/fisiologia , Sinapses/imunologia , Sinapses/metabolismo , Animais , Autoanticorpos/metabolismo , Epilepsia/metabolismo , Humanos , Imunidade Humoral/fisiologia , Transtornos dos Movimentos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Neuropsiquiatria/métodos , Neurotransmissores , Receptores de GABA/metabolismo , Receptores de Glicina/metabolismo , Transmissão Sináptica/imunologia , Transmissão Sináptica/fisiologia
9.
Neuron ; 100(1): 91-105.e9, 2018 10 10.
Artigo em Inglês | MEDLINE | ID: mdl-30146304

RESUMO

AMPA receptors are essential for fast excitatory transmission in the CNS. Autoantibodies to AMPA receptors have been identified in humans with autoimmune encephalitis and severe defects of hippocampal function. Here, combining electrophysiology and high-resolution imaging with neuronal culture preparations and passive-transfer models in wild-type and GluA1-knockout mice, we analyze how specific human autoantibodies against the AMPA receptor subunit GluA2 affect receptor function and composition, synaptic transmission, and plasticity. Anti-GluA2 antibodies induce receptor internalization and a reduction of synaptic GluA2-containing AMPARs followed by compensatory ryanodine receptor-dependent incorporation of synaptic non-GluA2 AMPARs. Furthermore, application of human pathogenic anti-GluA2 antibodies to mice impairs long-term synaptic plasticity in vitro and affects learning and memory in vivo. Our results identify a specific immune-neuronal rearrangement of AMPA receptor subunits, providing a framework to explain disease symptoms.


Assuntos
Autoanticorpos/farmacologia , Encefalite/fisiopatologia , Doença de Hashimoto/fisiopatologia , Plasticidade Neuronal/efeitos dos fármacos , Receptores de AMPA/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Animais , Autoanticorpos/imunologia , Autoantígenos/imunologia , Encefalite/complicações , Encefalite/imunologia , Doença de Hashimoto/complicações , Doença de Hashimoto/imunologia , Hipocampo/efeitos dos fármacos , Humanos , Transtornos da Memória/etiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/efeitos dos fármacos , Receptores de AMPA/imunologia
10.
J Neurophysiol ; 119(5): 1863-1878, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29465325

RESUMO

Calcium imaging provides an indirect observation of the underlying neural dynamics and enables the functional analysis of neuronal populations. However, the recorded fluorescence traces are temporally smeared, thus making the reconstruction of exact spiking activity challenging. Most of the established methods to tackle this issue are limited in dealing with issues such as the variability in the kinetics of fluorescence transients, fast processing of long-term data, high firing rates, and measurement noise. We propose a novel, heuristic reconstruction method to overcome these limitations. By using both synthetic and experimental data, we demonstrate the four main features of this method: 1) it accurately reconstructs both isolated spikes and within-burst spikes, and the spike count per fluorescence transient, from a given noisy fluorescence trace; 2) it performs the reconstruction of a trace extracted from 1,000,000 frames in less than 2 s; 3) it adapts to transients with different rise and decay kinetics or amplitudes, both within and across single neurons; and 4) it has only one key parameter, which we will show can be set in a nearly automatic way to an approximately optimal value. Furthermore, we demonstrate the ability of the method to effectively correct for fast and rather complex, slowly varying drifts as frequently observed in in vivo data. NEW & NOTEWORTHY Reconstruction of spiking activities from calcium imaging data remains challenging. Most of the established reconstruction methods not only have limitations in adapting to systematic variations in the data and fast processing of large amounts of data, but their results also depend on the user's experience. To overcome these limitations, we present a novel, heuristic model-free-type method that enables an ultra-fast, accurate, near-automatic reconstruction from data recorded under a wide range of experimental conditions.


Assuntos
Potenciais de Ação/fisiologia , Cálcio/metabolismo , Córtex Cerebral/fisiologia , Processamento de Imagem Assistida por Computador/métodos , Modelos Teóricos , Neurônios/fisiologia , Imagem Óptica/métodos , Animais , Córtex Cerebral/diagnóstico por imagem , Córtex Cerebral/metabolismo , Simulação por Computador , Microscopia , Estudo de Prova de Conceito
11.
Neuroscientist ; 24(1): 36-53, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-28378628

RESUMO

In recent years, considerable progress has been achieved in deciphering the cellular and network functions of GABAergic transmission in the intact developing brain. First, in vivo studies in non-mammalian and mammalian species confirmed the long-held assumption that GABA acts as a mainly depolarizing neurotransmitter at early developmental stages. At the same time, GABAergic transmission was shown to spatiotemporally constrain spontaneous cortical activity, whereas firm evidence for GABAergic excitation in vivo is currently missing. Second, there is a growing body of evidence indicating that depolarizing GABA may contribute to the activity-dependent refinement of neural circuits. Third, alterations in GABA actions have been causally linked to developmental brain disorders and identified as potential targets of timed prophylactic interventions. In this article, we review these major recent findings and argue that both depolarizing and inhibitory GABA actions may be crucial for physiological brain maturation.


Assuntos
Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Transmissão Sináptica/fisiologia , Ácido gama-Aminobutírico/metabolismo , Animais , Humanos
12.
Sci Rep ; 7(1): 13015, 2017 10 12.
Artigo em Inglês | MEDLINE | ID: mdl-29026183

RESUMO

During neocortical development, network activity undergoes a dramatic transition from largely synchronized, so-called cluster activity, to a relatively sparse pattern around the time of eye-opening in rodents. Biophysical mechanisms underlying this sparsification phenomenon remain poorly understood. Here, we present a dynamic systems modeling study of a developing neural network that provides the first mechanistic insights into sparsification. We find that the rest state of immature networks is strongly affected by the dynamics of a transient, unstable state hidden in their firing activities, allowing these networks to either be silent or generate large cluster activity. We address how, and which, specific developmental changes in neuronal and synaptic parameters drive sparsification. We also reveal how these changes refine the information processing capabilities of an in vivo developing network, mainly by showing a developmental reduction in the instability of network's firing activity, an effective availability of inhibition-stabilized states, and an emergence of spontaneous attractors and state transition mechanisms. Furthermore, we demonstrate the key role of GABAergic transmission and depressing glutamatergic synapses in governing the spatiotemporal evolution of cluster activity. These results, by providing a strong link between experimental observations and model behavior, suggest how adult sparse coding networks may emerge developmentally.


Assuntos
Rede Nervosa/fisiologia , Sistema Nervoso/embriologia , Potenciais de Ação/fisiologia , Animais , Neurônios GABAérgicos/metabolismo , Modelos Neurológicos , Plasticidade Neuronal , Sinapses/fisiologia , Fatores de Tempo
13.
J Neurosci ; 36(18): 4959-75, 2016 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-27147650

RESUMO

UNLABELLED: This study evaluates single-cell indicators of glutamate transport in sulforhodamine 101-positive astrocytes of Q175 mice, a knock-in model of Huntington's disease (HD). Transport-related fluorescent ratio signals obtained with sodium-binding benzofuran isophtalate (SBFI) AM from unperturbed or voltage-clamped astrocytes and respective glutamate transporter currents (GTCs) were induced by photolytic or synaptic glutamate release and isolated pharmacologically. The HD-induced deficit ranged from -27% (GTC maximum at -100 mV in Ba(2+)) to -41% (sodium transients in astrocytes after loading SBFI-AM). Our specific aim was to clarify the mechanism(s) by which Kir4.1 channels can influence glutamate transport, as determined by either Na(+) imaging or transport-associated electrical signals. A decrease of Kir4.1 conductance was mimicked with Ba(2+) (200 µm), and an increase of Kir4.1 expression was obtained by intravenous administration of AAV9-gfaABC1D-Kir4.1-EGFP. The decrease of Kir4.1 conductance reduced the sodium transients but increased the amplitudes of somatic GTCs. Accordingly, after genetic upregulation of Kir4.1, somatic GTCs were found to be decreased. In individual cells, there was a negative correlation between Kir4.1 currents and GTCs. The relative effect of the Kir4.1 conductance was higher in the astrocyte periphery. These and other results suggest that the Kir4.1 conductance affects glutamate transporter activity in a dual manner: (1) by providing the driving force (voltage dependency of the transport itself) and (2) by limiting the lateral charge transfer (thereby reducing the interference with other electrogenic transporter functions). This leads to the testable prediction that restoring the high conductance state of passive astrocytes will not only normalize glutamate uptake but also restore other astrocytic transporter activities afflicted with HD. SIGNIFICANCE STATEMENT: Insufficiency of astrocytic glutamate uptake is a major element in the pathophysiology of neurodegenerative diseases. Considering the heterogeneity of astrocytes and their differential susceptibility to therapeutic interventions, it becomes necessary to evaluate the determinants of transport activity in individual astroglial cells. We have examined intracellular Na(+) transients and glutamate transporter currents as the most telling indicators of glutamate clearance after synaptic or photolytic release of glutamate in striatal slices. The results show that, in Huntington's disease, glutamate uptake activity critically depends on Kir4.1. These channels enable the high conductance state of the astrocytic plasma membrane, which ensures the driving force for glutamate transport and dumps the transport-associated depolarization along the astrocyte processes. This has significant implications for developing therapeutic targets.


Assuntos
Astrócitos/metabolismo , Ácido Glutâmico/metabolismo , Doença de Huntington/metabolismo , Neostriado/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Sistema X-AG de Transporte de Aminoácidos/metabolismo , Animais , Astrócitos/efeitos dos fármacos , Benzofuranos/farmacologia , Transportador 2 de Aminoácido Excitatório/genética , Transportador 2 de Aminoácido Excitatório/metabolismo , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Junções Comunicantes/efeitos dos fármacos , Junções Comunicantes/metabolismo , Técnicas de Introdução de Genes , Proteína Glial Fibrilar Ácida/metabolismo , Camundongos , Neostriado/citologia , Técnicas de Patch-Clamp , Ácidos Ftálicos/farmacologia , Canais de Potássio Corretores do Fluxo de Internalização/antagonistas & inibidores
14.
Neuroimage ; 138: 64-75, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27222218

RESUMO

Neuronal network activity in the developing brain is generated in a discontinuous manner. In the visual cortex during the period of physiological blindness of immaturity, this activity mainly comprises retinally triggered spindle bursts or Ca(2+) clusters thought to contribute to the activity-dependent construction of cortical circuits. In spite of potentially important developmental functions, the spatial structure of these activity patterns remains largely unclear. In order to address this issue, we here used three-dimensional two-photon Ca(2+) imaging in the visual cortex of neonatal mice at postnatal days (P) 3-4 in vivo. Large-scale voxel imaging covering a cortical depth of 200µm revealed that Ca(2+) clusters, identified as spindle bursts in simultaneous extracellular recordings, recruit cortical glutamatergic neurons of the upper cortical plate (CP) in a column-like manner. Specifically, the majority of Ca(2+) clusters exhibit prominent horizontal confinement and high intra-cluster density of activation involving the entire depth of the upper CP. Moreover, using simultaneous Ca(2+) imaging from hundreds of neurons at single-cellular resolution, we demonstrate that the degree of neuronal co-activation within Ca(2+) clusters displays substantial heterogeneity. We further provide evidence that co-activated cells within Ca(2+) clusters are spatially distributed in a non-stochastic manner. In summary, our data support the conclusion that dense coding in the form of column-like Ca(2+) clusters is a characteristic property of network activity in the developing visual neocortex. Such knowledge is expected to be relevant for a refined understanding of how specific spatiotemporal characteristics of early network activity instruct the development of cortical circuits.


Assuntos
Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Imageamento Tridimensional/métodos , Microscopia de Fluorescência por Excitação Multifotônica/métodos , Imagem Molecular/métodos , Neocórtex/metabolismo , Animais , Animais Recém-Nascidos , Feminino , Aumento da Imagem/métodos , Interpretação de Imagem Assistida por Computador/métodos , Masculino , Camundongos , Camundongos Transgênicos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Análise Espaço-Temporal , Distribuição Tecidual
15.
PLoS Comput Biol ; 12(2): e1004736, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26894748

RESUMO

Calcium imaging has been used as a promising technique to monitor the dynamic activity of neuronal populations. However, the calcium trace is temporally smeared which restricts the extraction of quantities of interest such as spike trains of individual neurons. To address this issue, spike reconstruction algorithms have been introduced. One limitation of such reconstructions is that the underlying models are not informed about the biophysics of spike and burst generations. Such existing prior knowledge might be useful for constraining the possible solutions of spikes. Here we describe, in a novel Bayesian approach, how principled knowledge about neuronal dynamics can be employed to infer biophysical variables and parameters from fluorescence traces. By using both synthetic and in vitro recorded fluorescence traces, we demonstrate that the new approach is able to reconstruct different repetitive spiking and/or bursting patterns with accurate single spike resolution. Furthermore, we show that the high inference precision of the new approach is preserved even if the fluorescence trace is rather noisy or if the fluorescence transients show slow rise kinetics lasting several hundred milliseconds, and inhomogeneous rise and decay times. In addition, we discuss the use of the new approach for inferring parameter changes, e.g. due to a pharmacological intervention, as well as for inferring complex characteristics of immature neuronal circuits.


Assuntos
Sinalização do Cálcio/fisiologia , Cálcio/metabolismo , Modelos Neurológicos , Neurônios/fisiologia , Potenciais de Ação/fisiologia , Animais , Teorema de Bayes , Região CA3 Hipocampal/citologia , Região CA3 Hipocampal/metabolismo , Células Cultivadas , Biologia Computacional , Camundongos , Camundongos Endogâmicos C57BL , Imagem Molecular
16.
Biomed Opt Express ; 6(10): 3678-93, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26504620

RESUMO

Two-photon laser-scanning microscopy enables to record neuronal network activity in three-dimensional space while maintaining single-cellular resolution. One of the proposed approaches combines galvanometric x-y scanning with piezo-driven objective movements and employs hardware feedback signals for position monitoring. However, readily applicable methods to quantify the accuracy of those feedback signals are currently lacking. Here we provide techniques based on contact-free laser reflection and laser triangulation for the quantification of positioning accuracy of each spatial axis. We found that the lateral feedback signals are sufficiently accurate (defined as <2.5 µm) for a wide range of scan trajectories and frequencies. We further show that axial positioning accuracy does not only depend on objective acceleration and mass but also its geometry. We conclude that the introduced methods allow a reliable quantification of position feedback signals in a cost-efficient, easy-to-install manner and should be applicable for a wide range of two-photon laser scanning microscopes.

17.
Nat Commun ; 6: 7750, 2015 Jul 16.
Artigo em Inglês | MEDLINE | ID: mdl-26177896

RESUMO

A large body of evidence from in vitro studies suggests that GABA is depolarizing during early postnatal development. However, the mode of GABA action in the intact developing brain is unknown. Here we examine the in vivo effects of GABA in cells of the upper cortical plate using a combination of electrophysiological and Ca(2+)-imaging techniques. We report that at postnatal days (P) 3-4, GABA depolarizes the majority of immature neurons in the occipital cortex of anaesthetized mice. At the same time, GABA does not efficiently activate voltage-gated Ca(2+) channels and fails to induce action potential firing. Blocking GABA(A) receptors disinhibits spontaneous network activity, whereas allosteric activation of GABA(A) receptors has the opposite effect. In summary, our data provide evidence that in vivo GABA acts as a depolarizing neurotransmitter imposing an inhibitory control on network activity in the neonatal (P3-4) neocortex.


Assuntos
GABAérgicos/metabolismo , Neocórtex/efeitos dos fármacos , Inibição Neural/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Lobo Occipital/efeitos dos fármacos , Ácido gama-Aminobutírico/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Animais Recém-Nascidos , GABAérgicos/farmacologia , Camundongos , Neocórtex/citologia , Neocórtex/metabolismo , Rede Nervosa/efeitos dos fármacos , Neurônios/metabolismo , Lobo Occipital/citologia , Lobo Occipital/metabolismo , Técnicas de Patch-Clamp , Receptores de GABA-A/metabolismo , Transmissão Sináptica/efeitos dos fármacos , Ácido gama-Aminobutírico/metabolismo
18.
J Am Heart Assoc ; 3(4)2014 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-25169792

RESUMO

BACKGROUND: Delayed cerebral vasospasm is the most common cause of mortality and severe neurological impairment in patients who survive subarachnoid hemorrhage. Despite improvements in the field of diagnostic imaging, options for prevention and medical treatment-primarily with the calcium channel antagonist nimodipine or hemodynamic manipulations-are insufficient. Previous studies have suggested that heme and bilirubin oxidation end products, originating from degraded hemoglobin around ruptured blood vessels, are involved in the development of vasospasm by inhibiting large conductance BKC a potassium channels in vascular smooth muscle cells. In this study, we identify individual heme degradation products regulating arteriolar diameter in dependence of BKC a channel activity. METHODS AND RESULTS: Using differential interference contrast video microscopy in acute brain slices, we determined diameter changes of intracerebral arterioles in mouse visual cortex. In preconstricted vessels, the specific BKC a channel blockers paxilline and iberiotoxin as well as iron-containing hemin caused vasoconstriction. In addition, the bilirubin oxidation end product Z-BOX A showed a stronger vasoconstrictive potency than its regio-isomer Z-BOX B. Importantly, Z-BOX A had the same vasoconstrictive effect, independent of its origin from oxidative degradation or chemical synthesis. Finally, in slices of Slo1-deficient knockout mice, paxilline and Z-BOX A remained ineffective in changing arteriole diameter. CONCLUSIONS: We identified individual components of the oxidative bilirubin degradation that led to vasoconstriction of cerebral arterioles. The vasoconstrictive effect of Z-BOX A and Z-BOX B was mediated by BKC a channel activity that might represent a signaling pathway in the occurrence of delayed cerebral vasospasm in subarachnoid hemorrhage patients.


Assuntos
Arteríolas/efeitos dos fármacos , Hemina/farmacologia , Indóis/farmacologia , Peptídeos/farmacologia , Vasoconstrição/efeitos dos fármacos , Córtex Visual/irrigação sanguínea , Animais , Arteríolas/patologia , Bilirrubina/metabolismo , Heme/metabolismo , Canais de Potássio Ativados por Cálcio de Condutância Alta/antagonistas & inibidores , Camundongos , Microscopia de Interferência , Bloqueadores dos Canais de Potássio , Pirróis , Hemorragia Subaracnóidea/complicações , Vasoespasmo Intracraniano/etiologia , Córtex Visual/efeitos dos fármacos
19.
Cell Calcium ; 52(2): 182-9, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22658827

RESUMO

The development of genetically modified mice in which subpopulations of cortical neurons are labelled by fluorescent proteins has greatly facilitated single-cellular imaging and electrophysiology studies in vitro and in vivo. However, the parallel visualization of both inhibitory and excitatory neocortical neurons remains problematic. We here provide an alternative approach to identify GABAergic neurons in the context of in vivo calcium imaging. The method relies on the Emx1(IREScre) recombinase driven expression of a red fluorescent protein in excitatory neurons and glia. We quantitatively examined the upper layers of the visual neocortex in vivo and found that due to pronounced neuropil staining Emx1(IREScre)-negative and Emx1(IREScre)-positive neurons can be reliably differentiated based on negative and positive contrast, respectively. Immunohistochemical analyses confirmed that the entire population of GABAergic interneurons is represented by Emx1(IREScre)-negative cells. The potential usefulness of the method is exemplified by calcium imaging of sensory-evoked responses in the primary visual cortex. We conclude that the proposed method extends the repertoire of strategies aimed at discriminating two major neocortical neuron populations in situ.


Assuntos
Neurônios GABAérgicos/metabolismo , Regulação da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Integrases/metabolismo , Proteínas Luminescentes/metabolismo , Fatores de Transcrição/metabolismo , Animais , Cálcio/metabolismo , Neurônios GABAérgicos/patologia , Genes Reporter/genética , Proteínas de Homeodomínio/genética , Integrases/genética , Proteínas Luminescentes/genética , Camundongos , Microscopia de Fluorescência , Neocórtex/citologia , Neocórtex/metabolismo , Neocórtex/patologia , Fatores de Transcrição/genética , Proteína Vermelha Fluorescente
20.
Epilepsia ; 52(9): 1532-43, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21668443

RESUMO

Epileptic seizures rank among the most frequent neurologic symptoms during the neonatal period. Accumulating data from experimental animal studies and clinical trials in humans suggest that neonatal seizures could adversely affect normal brain development and result in long-term neurologic sequelae. Unfortunately, currently used anticonvulsive drugs are often ineffective in the neonatal period. One particularity of the immature neuronal network during neonatal development is that the neurotransmitter γ-aminobutyric acid (GABA) is mainly depolarizing, rather than hyperpolarizing as commonly observed in adults. This might, in part, explain not only the higher seizure propensity of the immature neuronal network, but also the limited anticonvulsive efficacy of GABA-enhancing drugs during early postnatal life. Accordingly, pharmacologic attenuation of GABAergic depolarization has been proposed as a strategy for neonatal seizure control. However, the underlying conjecture of a depolarizing mode of GABA action has been seriously challenged recently. In the present review, we will summarize the state of knowledge regarding GABAergic depolarization in early life and discuss how these data might impact a currently tested anticonvulsive strategy.


Assuntos
Anticonvulsivantes/farmacologia , Córtex Cerebral/efeitos dos fármacos , Epilepsia/patologia , Potenciais da Membrana/efeitos dos fármacos , Ácido gama-Aminobutírico/metabolismo , Animais , Animais Recém-Nascidos , Anticonvulsivantes/uso terapêutico , Córtex Cerebral/crescimento & desenvolvimento , Modelos Animais de Doenças , Epilepsia/tratamento farmacológico , Humanos , Recém-Nascido , Modelos Biológicos
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