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1.
Cereb Cortex ; 29(5): 2010-2033, 2019 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-29912316

RESUMEN

Mutations in PRoline-Rich Transmembrane protein 2 (PRRT2) underlie a group of paroxysmal disorders including epilepsy, kinesigenic dyskinesia and migraine. Most of the mutations lead to impaired PRRT2 expression and/or function, emphasizing the pathogenic role of the PRRT2 deficiency. In this work, we investigated the phenotype of primary hippocampal neurons obtained from mouse embryos in which the PRRT2 gene was constitutively inactivated. Although PRRT2 is expressed by both excitatory and inhibitory neurons, its deletion decreases the number of excitatory synapses without significantly affecting the number of inhibitory synapses or the nerve terminal ultrastructure. Analysis of synaptic function in primary PRRT2 knockout excitatory neurons by live imaging and electrophysiology showed slowdown of the kinetics of exocytosis, weakened spontaneous and evoked synaptic transmission and markedly increased facilitation. Inhibitory neurons showed strengthening of basal synaptic transmission, accompanied by faster depression. At the network level these complex synaptic effects resulted in a state of heightened spontaneous and evoked activity that was associated with increased excitability of excitatory neurons in both PRRT2 knockout primary cultures and acute hippocampal slices. The data indicate the existence of network instability/hyperexcitability as the possible basis of the paroxysmal phenotypes associated with PRRT2 mutations.


Asunto(s)
Hipocampo/fisiología , Proteínas de la Membrana/fisiología , Plasticidad Neuronal , Neuronas/fisiología , Transmisión Sináptica , Animales , Células Cultivadas , Exocitosis , Masculino , Potenciales de la Membrana , Proteínas de la Membrana/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Vías Nerviosas/fisiología , Sinapsis/fisiología , Sinapsis/ultraestructura
2.
Brain ; 141(4): 1000-1016, 2018 04 01.
Artículo en Inglés | MEDLINE | ID: mdl-29554219

RESUMEN

See Lerche (doi:10.1093/brain/awy073) for a scientific commentary on this article.Proline-rich transmembrane protein 2 (PRRT2) is the causative gene for a heterogeneous group of familial paroxysmal neurological disorders that include seizures with onset in the first year of life (benign familial infantile seizures), paroxysmal kinesigenic dyskinesia or a combination of both. Most of the PRRT2 mutations are loss-of-function leading to haploinsufficiency and 80% of the patients carry the same frameshift mutation (c.649dupC; p.Arg217Profs*8), which leads to a premature stop codon. To model the disease and dissect the physiological role of PRRT2, we studied the phenotype of neurons differentiated from induced pluripotent stem cells from previously described heterozygous and homozygous siblings carrying the c.649dupC mutation. Single-cell patch-clamp experiments on induced pluripotent stem cell-derived neurons from homozygous patients showed increased Na+ currents that were fully rescued by expression of wild-type PRRT2. Closely similar electrophysiological features were observed in primary neurons obtained from the recently characterized PRRT2 knockout mouse. This phenotype was associated with an increased length of the axon initial segment and with markedly augmented spontaneous and evoked firing and bursting activities evaluated, at the network level, by multi-electrode array electrophysiology. Using HEK-293 cells stably expressing Nav channel subtypes, we demonstrated that the expression of PRRT2 decreases the membrane exposure and Na+ current of Nav1.2/Nav1.6, but not Nav1.1, channels. Moreover, PRRT2 directly interacted with Nav1.2/Nav1.6 channels and induced a negative shift in the voltage-dependence of inactivation and a slow-down in the recovery from inactivation. In addition, by co-immunoprecipitation assays, we showed that the PRRT2-Nav interaction also occurs in brain tissue. The study demonstrates that the lack of PRRT2 leads to a hyperactivity of voltage-dependent Na+ channels in homozygous PRRT2 knockout human and mouse neurons and that, in addition to the reported synaptic functions, PRRT2 is an important negative modulator of Nav1.2 and Nav1.6 channels. Given the predominant paroxysmal character of PRRT2-linked diseases, the disturbance in cellular excitability by lack of negative modulation of Na+ channels appears as the key pathogenetic mechanism.


Asunto(s)
Regulación de la Expresión Génica/genética , Proteínas de la Membrana/metabolismo , Mutación/genética , Canal de Sodio Activado por Voltaje NAV1.2/metabolismo , Canal de Sodio Activado por Voltaje NAV1.6/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neuronas/fisiología , Animales , Segmento Inicial del Axón/fisiología , Diferenciación Celular , Corteza Cerebral/citología , Consanguinidad , Fibroblastos/patología , Células HEK293 , Humanos , Células Madre Pluripotentes Inducidas , Potenciales de la Membrana/genética , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Canal de Sodio Activado por Voltaje NAV1.6/genética , Proteína Homeótica Nanog/genética , Proteína Homeótica Nanog/metabolismo , Proteínas del Tejido Nervioso/genética , Enfermedades del Sistema Nervioso/genética , Enfermedades del Sistema Nervioso/patología , Neuronas/citología , Factor de Transcripción PAX6/genética , Factor de Transcripción PAX6/metabolismo , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXB1/metabolismo , Hermanos
3.
J Neurosci ; 37(7): 1747-1756, 2017 02 15.
Artículo en Inglés | MEDLINE | ID: mdl-28073939

RESUMEN

Gephyrin is a key scaffold protein mediating the anchoring of GABAA receptors at inhibitory synapses. Here, we exploited superresolution techniques combined with proximity-based clustering analysis and model simulations to investigate the single-molecule gephyrin reorganization during plasticity of inhibitory synapses in mouse hippocampal cultured neurons. This approach revealed that, during the expression of inhibitory LTP, the increase of gephyrin density at postsynaptic sites is associated with the promoted formation of gephyrin nanodomains. We demonstrate that the gephyrin rearrangement in nanodomains stabilizes the amplitude of postsynaptic currents, indicating that, in addition to the number of synaptic GABAA receptors, the nanoscale distribution of GABAA receptors in the postsynaptic area is a crucial determinant for the expression of inhibitory synaptic plasticity. In addition, the methodology implemented here clears the way to the application of the graph-based theory to single-molecule data for the description and quantification of the spatial organization of the synapse at the single-molecule level.SIGNIFICANCE STATEMENT The mechanisms of inhibitory synaptic plasticity are poorly understood, mainly because the size of the synapse is below the diffraction limit, thus reducing the effectiveness of conventional optical and imaging techniques. Here, we exploited superresolution approaches combined with clustering analysis to study at unprecedented resolution the distribution of the inhibitory scaffold protein gephyrin in response to protocols inducing LTP of inhibitory synaptic responses (iLTP). We found that, during the expression of iLTP, the increase of synaptic gephyrin is associated with the fragmentation of gephyrin in subsynaptic nanodomains. We demonstrate that such synaptic gephyrin nanodomains stabilize the amplitude of inhibitory postsynaptic responses, thus identifying the nanoscale gephyrin rearrangement as a key determinant for inhibitory synaptic plasticity.


Asunto(s)
Proteínas Portadoras/metabolismo , Neuronas GABAérgicas/citología , Depresión Sináptica a Largo Plazo/fisiología , Proteínas de la Membrana/metabolismo , Densidad Postsináptica/metabolismo , 6-Ciano 7-nitroquinoxalina 2,3-diona/farmacología , Algoritmos , Animales , Células Cultivadas , Simulación por Computador , Agonistas de Aminoácidos Excitadores/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Femenino , Neuronas GABAérgicas/efectos de los fármacos , Hipocampo/citología , Depresión Sináptica a Largo Plazo/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Neurológicos , N-Metilaspartato/farmacología , Péptidos/metabolismo , Polímeros , Densidad Postsináptica/efectos de los fármacos , Receptores de GABA-A/metabolismo , Valina/análogos & derivados , Valina/farmacología
4.
PLoS Comput Biol ; 13(7): e1005672, 2017 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-28749937

RESUMEN

Developing neuronal systems intrinsically generate coordinated spontaneous activity that propagates by involving a large number of synchronously firing neurons. In vivo, waves of spikes transiently characterize the activity of developing brain circuits and are fundamental for activity-dependent circuit formation. In vitro, coordinated spontaneous spiking activity, or network bursts (NBs), interleaved within periods of asynchronous spikes emerge during the development of 2D and 3D neuronal cultures. Several studies have investigated this type of activity and its dynamics, but how a neuronal system generates these coordinated events remains unclear. Here, we investigate at a cellular level the generation of network bursts in spontaneously active neuronal cultures by exploiting high-resolution multielectrode array recordings and computational network modelling. Our analysis reveals that NBs are generated in specialized regions of the network (functional neuronal communities) that feature neuronal links with high cross-correlation peak values, sub-millisecond lags and that share very similar structural connectivity motifs providing recurrent interactions. We show that the particular properties of these local structures enable locally amplifying spontaneous asynchronous spikes and that this mechanism can lead to the initiation of NBs. Through the analysis of simulated and experimental data, we also show that AMPA currents drive the coordinated activity, while NMDA and GABA currents are only involved in shaping the dynamics of NBs. Overall, our results suggest that the presence of functional neuronal communities with recurrent local connections allows a neuronal system to generate spontaneous coordinated spiking activity events. As suggested by the rules used for implementing our computational model, such functional communities might naturally emerge during network development by following simple constraints on distance-based connectivity.


Asunto(s)
Potenciales de Acción/fisiología , Modelos Neurológicos , Red Nerviosa/citología , Neuronas/citología , Animales , Células Cultivadas , Biología Computacional , Hipocampo/citología , Red Nerviosa/fisiología , Neuronas/fisiología , Ratas
5.
J Biol Chem ; 290(29): 18045-18055, 2015 Jul 17.
Artículo en Inglés | MEDLINE | ID: mdl-26037926

RESUMEN

Kidins220 (kinase D-interacting substrate of 220 kDa)/ankyrin repeat-rich membrane spanning (ARMS) acts as a signaling platform at the plasma membrane and is implicated in a multitude of neuronal functions, including the control of neuronal activity. Here, we used the Kidins220(-/-) mouse model to study the effects of Kidins220 ablation on neuronal excitability. Multielectrode array recordings showed reduced evoked spiking activity in Kidins220(-/-) hippocampal networks, which was compatible with the increased excitability of GABAergic neurons determined by current-clamp recordings. Spike waveform analysis further indicated an increased sodium conductance in this neuronal subpopulation. Kidins220 association with brain voltage-gated sodium channels was shown by co-immunoprecipitation experiments and Na(+) current recordings in transfected HEK293 cells, which revealed dramatic alterations of kinetics and voltage dependence. Finally, an in silico interneuronal model incorporating the Kidins220-induced Na(+) current alterations reproduced the firing phenotype observed in Kidins220(-/-) neurons. These results identify Kidins220 as a novel modulator of Nav channel activity, broadening our understanding of the molecular mechanisms regulating network excitability.


Asunto(s)
Hipocampo/citología , Proteínas de la Membrana/metabolismo , Neuronas/metabolismo , Canales de Sodio Activados por Voltaje/metabolismo , Animales , Células COS , Células Cultivadas , Chlorocebus aethiops , Potenciales Evocados , Eliminación de Gen , Células HEK293 , Hipocampo/metabolismo , Hipocampo/fisiología , Humanos , Proteínas de la Membrana/genética , Ratones , Ratones Endogámicos C57BL , Red Nerviosa , Neuronas/citología
6.
PLoS Genet ; 8(5): e1002706, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22589743

RESUMEN

It has been suggested that imprinted genes are important in the regulation of sleep. However, the fundamental question of whether genomic imprinting has a role in sleep has remained elusive up to now. In this work we show that REM and NREM sleep states are differentially modulated by the maternally expressed imprinted gene Gnas. In particular, in mice with loss of imprinting of Gnas, NREM and complex cognitive processes are enhanced while REM and REM-linked behaviors are inhibited. This is the first demonstration that a specific overexpression of an imprinted gene affects sleep states and related complex behavioral traits. Furthermore, in parallel to the Gnas overexpression, we have observed an overexpression of Ucp1 in interscapular brown adipose tissue (BAT) and a significant increase in thermoregulation that may account for the REM/NREM sleep phenotypes. We conclude that there must be significant evolutionary advantages in the monoallelic expression of Gnas for REM sleep and for the consolidation of REM-dependent memories. Conversely, biallelic expression of Gnas reinforces slow wave activity in NREM sleep, and this results in a reduction of uncertainty in temporal decision-making processes.


Asunto(s)
Cognición/fisiología , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Impresión Genómica , Sueño REM/genética , Sueño REM/fisiología , Tejido Adiposo Pardo , Alelos , Animales , Temperatura Corporal , Regulación de la Temperatura Corporal/genética , Regulación de la Temperatura Corporal/fisiología , Cromograninas , Metilación de ADN , Electroencefalografía , Exones , Subunidades alfa de la Proteína de Unión al GTP Gs/fisiología , Regulación de la Expresión Génica , Canales Iónicos , Ratones , Proteínas Mitocondriales , Eliminación de Secuencia , Proteína Desacopladora 1 , Vigilia
7.
Cereb Cortex ; 23(3): 581-93, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22368083

RESUMEN

Synapsins (Syn I, Syn II, and Syn III) are a family of synaptic vesicle phosphoproteins regulating synaptic transmission and plasticity. SYN1/2 genes have been identified as major epilepsy susceptibility genes in humans and synapsin I/II/III triple knockout (TKO) mice are epileptic. However, excitatory and inhibitory synaptic transmission and short-term plasticity have never been analyzed in intact neuronal circuits of TKO mice. To clarify the generation and expression of the epileptic phenotype, we performed patch-clamp recordings in the CA1 region of acute hippocampal slices from 1-month-old presymptomatic and 6-month-old epileptic TKO mice and age-matched controls. We found a strong imbalance between basal glutamatergic and γ-aminobutyric acid (GABA)ergic transmission with increased evoked excitatory postsynaptic current and impaired evoked inhibitory postsynaptic current amplitude. This imbalance was accompanied by a parallel derangement of short-term plasticity paradigms, with enhanced facilitation of glutamatergic transmission in the presymptomatic phase and milder depression of inhibitory synapses in the symptomatic phase. Interestingly, a lower tonic GABA(A) current due to the impaired GABA release is responsible for the more depolarized resting potential found in TKO CA1 neurons, which makes them more susceptible to fire. All these changes preceded the appearance of epilepsy, indicating that the distinct changes in excitatory and inhibitory transmission due to the absence of Syns initiate the epileptogenic process.


Asunto(s)
Región CA1 Hipocampal/fisiología , Epilepsia/fisiopatología , Plasticidad Neuronal/fisiología , Transmisión Sináptica/fisiología , Animales , Epilepsia/genética , Potenciales Postsinápticos Excitadores/fisiología , Femenino , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Modelos Teóricos , Técnicas de Placa-Clamp , Sinapsis/fisiología , Sinapsinas/deficiencia , Sinapsinas/genética
8.
J Neurosci ; 32(17): 5868-79, 2012 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-22539848

RESUMEN

A large amount of experimental evidence has highlighted the rapid changes in synaptic efficacy induced by high-frequency stimulation and BDNF at central excitatory synapses. We clarified the quantal mechanisms and the involvement of Synapsin I (SynI) phosphorylation in the expression of post-tetanic potentiation (PTP) and in its modulation by BDNF in mouse glutamatergic autapses. We found that PTP is associated with an elevation in the probability of release and a concomitant increase in the size of the readily releasable pool (RRP). The latter component was virtually absent in SynI knock-out (KO) neurons, which indeed displayed impaired PTP. PTP was fully rescued by the expression of wild-type SynI, but not of its dephosphomimetic mutants in the phosphorylation sites for cAMP-dependent protein kinase and Ca²âº/calmodulin-dependent protein kinases I/II. BDNF potently enhanced PTP through a further increase in the RRP size, which was missing in SynI KO neurons. In these neurons, the BDNF-induced PTP enhancement was rescued by the expression of wild-type SynI, but not of its dephosphomimetic mutant at the mitogen-dependent protein kinase sites. The results indicate that the increase in RRP size necessary for the full expression of PTP, and its sensitivity to BDNF, involve phosphorylation of SynI at distinct sites, thus implicating SynI as an essential downstream effector for the expression of PTP and for its enhancement by BDNF.


Asunto(s)
Fenómenos Biofísicos/genética , Factor Neurotrófico Derivado del Encéfalo/farmacología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Regulación de la Expresión Génica/fisiología , Neuronas/efectos de los fármacos , Mutación Puntual/fisiología , Sinapsinas/deficiencia , 6-Ciano 7-nitroquinoxalina 2,3-diona/farmacología , Análisis de Varianza , Animales , Fenómenos Biofísicos/efectos de los fármacos , Calcio/metabolismo , Carbazoles/farmacología , Células Cultivadas , Estimulación Eléctrica , Embrión de Mamíferos , Inhibidores Enzimáticos/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Potenciales Postsinápticos Excitadores/genética , Potenciales Postsinápticos Excitadores/fisiología , Regulación de la Expresión Génica/efectos de los fármacos , Hipocampo/citología , Alcaloides Indólicos/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Modelos Biológicos , Mutagénesis Sitio-Dirigida/métodos , Neuronas/fisiología , Técnicas de Placa-Clamp , Fosforilación/fisiología , Sinapsinas/genética , Transfección
9.
J Neurosci ; 31(5): 1752-61, 2011 Feb 02.
Artículo en Inglés | MEDLINE | ID: mdl-21289185

RESUMEN

To reach the open state, the GABA(A) receptor (GABA(A)R) is assumed to bind two agonist molecules. Although it is currently believed that GABA(A)R could also operate in the monoliganded state, the gating properties of singly bound GABA(A)R are poorly understood and their physiological role is still obscure. In the present study, we characterize for the first time the gating properties of singly bound GABA(A)Rs by using a mutagenesis approach and we propose that monoliganded GABA(A)R contribute in shaping synaptic responses. At saturating GABA concentrations, currents mediated by recombinant GABA(A)Rs with a single functional binding site display slow onset, fast deactivation kinetics, and slow rate of desensitization-resensitization. GABA(A)Rs with two binding sites activated by brief pulses of subsaturating GABA concentrations (in the range of the GABA concentration profile in the synaptic cleft) could also mediate fast deactivating currents, displaying deactivation kinetics similar to those mediated by GABA(A)Rs with a single functional binding site. Model simulations of receptors activated by realistic synaptic GABA waves revealed that a considerable proportion of GABA(A) receptors open in the monoliganded state during synaptic transmission, therefore contributing in shaping IPSCs.


Asunto(s)
Inhibición Neural/fisiología , Receptores de GABA-A/metabolismo , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/metabolismo , Electrofisiología , Células HEK293 , Humanos , Inmunohistoquímica , Microscopía Confocal , Plásmidos , Reacción en Cadena de la Polimerasa , Receptores de GABA-A/genética , Transfección/métodos
10.
Brain Sci ; 11(11)2021 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-34827452

RESUMEN

Dissociated cortical neurons in vitro display spontaneously synchronized, low-frequency firing patterns, which can resemble the slow wave oscillations characterizing sleep in vivo. Experiments in humans, rodents, and cortical slices have shown that awakening or the administration of activating neuromodulators decrease slow waves, while increasing the spatio-temporal complexity of responses to perturbations. In this study, we attempted to replicate those findings using in vitro cortical cultures coupled with micro-electrode arrays and chemically treated with carbachol (CCh), to modulate sleep-like activity and suppress slow oscillations. We adapted metrics such as neural complexity (NC) and the perturbational complexity index (PCI), typically employed in animal and human brain studies, to quantify complexity in simplified, unstructured networks, both during resting state and in response to electrical stimulation. After CCh administration, we found a decrease in the amplitude of the initial response and a marked enhancement of the complexity during spontaneous activity. Crucially, unlike in cortical slices and intact brains, PCI in cortical cultures displayed only a moderate increase. This dissociation suggests that PCI, a measure of the complexity of causal interactions, requires more than activating neuromodulation and that additional factors, such as an appropriate circuit architecture, may be necessary. Exploring more structured in vitro networks, characterized by the presence of strong lateral connections, recurrent excitation, and feedback loops, may thus help to identify the features that are more relevant to support causal complexity.

11.
Neural Comput ; 22(8): 2031-58, 2010 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-20438336

RESUMEN

A nerve cell receives multiple inputs from upstream neurons by way of its synapses. Neuron processing functions are thus influenced by changes in the biophysical properties of the synapse, such as long-term potentiation (LTP) or depression (LTD). This observation has opened new perspectives on the biophysical basis of learning and memory, but its quantitative impact on the information transmission of a neuron remains partially elucidated. One major obstacle is the high dimensionality of the neuronal input-output space, which makes it unfeasible to perform a thorough computational analysis of a neuron with multiple synaptic inputs. In this work, information theory was employed to characterize the information transmission of a cerebellar granule cell over a region of its excitatory input space following synaptic changes. Granule cells have a small dendritic tree (on average, they receive only four mossy fiber afferents), which greatly bounds the input combinatorial space, reducing the complexity of information-theoretic calculations. Numerical simulations and LTP experiments quantified how changes in neurotransmitter release probability (p) modulated information transmission of a cerebellar granule cell. Numerical simulations showed that p shaped the neurotransmission landscape in unexpected ways. As p increased, the optimality of the information transmission of most stimuli did not increase strictly monotonically; instead it reached a plateau at intermediate p levels. Furthermore, our results showed that the spatiotemporal characteristics of the inputs determine the effect of p on neurotransmission, thus permitting the selection of distinctive preferred stimuli for different p values. These selective mechanisms may have important consequences on the encoding of cerebellar mossy fiber inputs and the plasticity and computation at the next circuit stage, including the parallel fiber-Purkinje cell synapses.


Asunto(s)
Cerebelo/fisiología , Modelos Neurológicos , Neuronas/fisiología , Transmisión Sináptica/fisiología , Animales , Plasticidad Neuronal/fisiología , Técnicas de Placa-Clamp , Ratas , Ratas Wistar
12.
Cell Rep ; 31(10): 107735, 2020 06 09.
Artículo en Inglés | MEDLINE | ID: mdl-32521260

RESUMEN

Kainate receptors (KARs) mediate postsynaptic currents with a key impact on neuronal excitability. However, the molecular determinants controlling KAR postsynaptic localization and stabilization are poorly understood. Here, we exploit optogenetic and single-particle tracking approaches to study the role of KAR conformational states induced by glutamate binding on KAR lateral mobility at synapses. We report that following glutamate binding, KARs are readily and reversibly trapped at glutamatergic synapses through increased interaction with the ß-catenin/N-cadherin complex. We demonstrate that such activation-dependent synaptic immobilization of KARs is crucial for the modulation of short-term plasticity of glutamatergic synapses. Thus, the present study unveils the crosstalk between conformational states and lateral mobility of KARs, a mechanism regulating glutamatergic signaling, particularly in conditions of sustained synaptic activity.


Asunto(s)
Ácido Glutámico/metabolismo , Ácido Kaínico/metabolismo , Plasticidad Neuronal/genética , Transmisión Sináptica/genética , Humanos
13.
Sci Rep ; 8(1): 5578, 2018 04 03.
Artículo en Inglés | MEDLINE | ID: mdl-29615719

RESUMEN

Neuronal responses to external stimuli vary from trial to trial partly because they depend on continuous spontaneous variations of the state of neural circuits, reflected in variations of ongoing activity prior to stimulus presentation. Understanding how post-stimulus responses relate to the pre-stimulus spontaneous activity is thus important to understand how state dependence affects information processing and neural coding, and how state variations can be discounted to better decode single-trial neural responses. Here we exploited high-resolution CMOS electrode arrays to record simultaneously from thousands of electrodes in in-vitro cultures stimulated at specific sites. We used information-theoretic analyses to study how ongoing activity affects the information that neuronal responses carry about the location of the stimuli. We found that responses exhibited state dependence on the time between the last spontaneous burst and the stimulus presentation and that the dependence could be described with a linear model. Importantly, we found that a small number of selected neurons carry most of the stimulus information and contribute to the state-dependent information gain. This suggests that a major value of large-scale recording is that it individuates the small subset of neurons that carry most information and that benefit the most from knowledge of its state dependence.


Asunto(s)
Estimulación Eléctrica , Electrofisiología/instrumentación , Metales/química , Neuronas/citología , Óxidos , Semiconductores , Animales , Células Cultivadas , Electrodos , Hipocampo/citología , Modelos Lineales , Neuronas/metabolismo , Norepinefrina/metabolismo , Ratas
14.
Sci Rep ; 7(1): 2460, 2017 05 26.
Artículo en Inglés | MEDLINE | ID: mdl-28550283

RESUMEN

Neurotoxicity and the accumulation of extracellular amyloid-beta1-42 (Aß) peptides are associated with the development of Alzheimer's disease (AD) and correlate with neuronal activity and network dysfunctions, ultimately leading to cellular death. However, research on neurodegenerative diseases is hampered by the paucity of reliable readouts and experimental models to study such functional decline from an early onset and to test rescue strategies within networks at cellular resolution. To overcome this important obstacle, we demonstrate a simple yet powerful in vitro AD model based on a rat hippocampal cell culture system that exploits large-scale neuronal recordings from 4096-electrodes on CMOS-chips for electrophysiological quantifications. This model allows us to monitor network activity changes at the cellular level and to uniquely uncover the early activity-dependent deterioration induced by Aß-neurotoxicity. We also demonstrate the potential of this in vitro model to test a plausible hypothesis underlying the Aß-neurotoxicity and to assay potential therapeutic approaches. Specifically, by quantifying N-methyl D-aspartate (NMDA) concentration-dependent effects in comparison with low-concentration allogenic-Aß, we confirm the role of extrasynaptic-NMDA receptors activation that may contribute to Aß-neurotoxicity. Finally, we assess the potential rescue of neural stem cells (NSCs) and of two pharmacotherapies, memantine and saffron, for reversing Aß-neurotoxicity and rescuing network-wide firing.


Asunto(s)
Péptidos beta-Amiloides/antagonistas & inhibidores , Antiparkinsonianos/farmacología , Hipocampo/efectos de los fármacos , Memantina/farmacología , Neuronas/efectos de los fármacos , Fragmentos de Péptidos/antagonistas & inhibidores , Receptores de N-Metil-D-Aspartato/genética , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Células Madre Adultas/citología , Células Madre Adultas/efectos de los fármacos , Células Madre Adultas/metabolismo , Péptidos beta-Amiloides/toxicidad , Animales , Crocus/química , Embrión de Mamíferos , Femenino , Expresión Génica , Hipocampo/metabolismo , Dispositivos Laboratorio en un Chip , Microelectrodos , N-Metilaspartato/farmacología , Células-Madre Neurales/citología , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/metabolismo , Neuronas/citología , Neuronas/metabolismo , Fragmentos de Péptidos/toxicidad , Extractos Vegetales/química , Ratas , Ratas Endogámicas F344 , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/agonistas , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Técnicas de Cultivo de Tejidos
15.
Neuron ; 95(1): 63-69.e5, 2017 Jul 05.
Artículo en Inglés | MEDLINE | ID: mdl-28683270

RESUMEN

The lateral mobility of neurotransmitter receptors has been shown to tune synaptic signals. Here we report that GABAA receptors (GABAARs) can diffuse between adjacent dendritic GABAergic synapses in long-living desensitized states, thus laterally spreading "activation memories" between inhibitory synapses. Glutamatergic activity limits this inter-synaptic diffusion by trapping GABAARs at excitatory synapses. This novel form of activity-dependent hetero-synaptic interplay is likely to modulate dendritic synaptic signaling.


Asunto(s)
Dendritas/metabolismo , Potenciales Postsinápticos Inhibidores , Plasticidad Neuronal/fisiología , Receptores de GABA-A/metabolismo , Sinapsis/metabolismo , Animales , Calcio/metabolismo , Difusión , Hipocampo/citología , Hipocampo/metabolismo , Inmunohistoquímica , Ratones , Neuronas/metabolismo , Imagen Óptica , Técnicas de Placa-Clamp , Puntos Cuánticos , Receptores de Ácido Kaínico/metabolismo , Receptor de Ácido Kaínico GluK2
16.
Sci Rep ; 7(1): 17765, 2017 12 19.
Artículo en Inglés | MEDLINE | ID: mdl-29259298

RESUMEN

Circadian clock is known to adapt to environmental changes and can significantly influence cognitive and physiological functions. In this work, we report specific behavioral, cognitive, and sleep homeostatic defects in the after hours (Afh) circadian mouse mutant, which is characterized by lengthened circadian period. We found that the circadian timing irregularities in Afh mice resulted in higher interval timing uncertainty and suboptimal decisions due to incapability of processing probabilities. Our phenotypic observations further suggested that Afh mutants failed to exhibit the necessary phenotypic plasticity for adapting to temporal changes at multiple time scales (seconds-to-minutes to circadian). These behavioral effects of Afh mutation were complemented by the specific disruption of the Per/Cry circadian regulatory complex in brain regions that govern food anticipatory behaviors, sleep, and timing. We derive statistical predictions, which indicate that circadian clock and sleep are complementary processes in controlling behavioral/cognitive performance during 24 hrs. The results of this study have pivotal implications for understanding how the circadian clock modulates sleep and behavior.


Asunto(s)
Adaptación Fisiológica/fisiología , Conducta Animal/fisiología , Relojes Circadianos/fisiología , Ritmo Circadiano/fisiología , Homeostasis/fisiología , Sueño/fisiología , Adaptación Fisiológica/genética , Animales , Encéfalo/fisiología , Relojes Circadianos/genética , Ritmo Circadiano/genética , Femenino , Homeostasis/genética , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Mutación/genética , Sueño/genética
17.
Front Cell Neurosci ; 10: 30, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26909023

RESUMEN

[This corrects the article on p. 246 in vol. 8, PMID: 25202237.].

18.
Front Neurosci ; 10: 121, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27065786

RESUMEN

The recent availability of human induced pluripotent stem cells (hiPSCs) holds great promise as a novel source of human-derived neurons for cell and tissue therapies as well as for in vitro drug screenings that might replace the use of animal models. However, there is still a considerable lack of knowledge on the functional properties of hiPSC-derived neuronal networks, thus limiting their application. Here, upon optimization of cell culture protocols, we demonstrate that both spontaneous and evoked electrical spiking activities of these networks can be characterized on-chip by taking advantage of the resolution provided by CMOS multielectrode arrays (CMOS-MEAs). These devices feature a large and closely-spaced array of 4096 simultaneously recording electrodes and multi-site on-chip electrical stimulation. Our results show that networks of human-derived neurons can respond to electrical stimulation with a physiological repertoire of spike waveforms after 3 months of cell culture, a period of time during which the network undergoes the expression of developing patterns of spontaneous spiking activity. To achieve this, we have investigated the impact on the network formation and on the emerging network-wide functional properties induced by different biochemical substrates, i.e., poly-dl-ornithine (PDLO), poly-l-ornithine (PLO), and polyethylenimine (PEI), that were used as adhesion promoters for the cell culture. Interestingly, we found that neuronal networks grown on PDLO coated substrates show significantly higher spontaneous firing activity, reliable responses to low-frequency electrical stimuli, and an appropriate level of PSD-95 that may denote a physiological neuronal maturation profile and synapse stabilization. However, our results also suggest that even 3-month culture might not be sufficient for human-derived neuronal network maturation. Taken together, our results highlight the tight relationship existing between substrate coatings and emerging network properties, i.e., spontaneous activity, responsiveness, synapse formation and maturation. Additionally, our results provide a baseline on the functional properties expressed over 3 months of network development for a commercially available line of hiPSC-derived neurons. This is a first step toward the development of functional pre-clinical assays to test pharmaceutical compounds on human-derived neuronal networks with CMOS-MEAs.

19.
Prog Brain Res ; 148: 69-80, 2005.
Artículo en Inglés | MEDLINE | ID: mdl-15661182

RESUMEN

In the last decade, the physiology of cerebellar neurons and synapses has been extended to a considerable extent. We have found that the mossy fiber-granule cell relay can generate a complex form of long-term potentiation (mf-GrC LTP) following high-frequency mf discharge. Induction. Mf-GrC LTP depends on NMDA and mGlu receptor activation, intracellular Ca(2+) increase, PKC activation, and NO production. The preventative action of intracellular agents (BAPTA, PKC-inhibitors) and of membrane hyperpolarization, and the correlated increase in intracellular Ca(2+) observed using fluorescent dyes, indicate that induction occurs postsynaptically. Expression. Expression includes three components: (a) an increase of synaptic currents, (b) an increase of intrinsic excitability in GrC, and (c) an increase of intrinsic excitability in mf terminals. Based on quantal analysis, the EPSC increase is mostly explained by enhanced neurotransmitter release. NO is a candidate retrograde neurotransmitter which could determine both presynaptic current changes and LTP. NO cascade blockers inhibit both presynaptic current changes and LTP. The increase in intrinsic excitability involves a raise in apparent input resistance in the subthreshold region and a spike threshold reduction. Together with other forms of cerebellar plasticity, mf-GrC LTP opens new hypothesis on how the cerebellum processes incoming information.


Asunto(s)
Cerebelo/citología , Cerebelo/fisiología , Potenciación a Largo Plazo/fisiología , Fibras Nerviosas/fisiología , Transmisión Sináptica/fisiología , Animales , Humanos
20.
Annu Int Conf IEEE Eng Med Biol Soc ; 2015: 3759-62, 2015 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-26737111

RESUMEN

High density multielectrode array recordings with CMOS-MEAs allow to monitor cell culture activity with unprecedent details respect to previous recording techniques. This is clarifying how network activity develops and is motivating the development of novel data analysis tools. Here, in order to advance in the exploitation of the richness of these large-scale array recordings, we introduce a principal component analysis approach that aims at improving on existing methodologies to describe neural activity events within large networks.


Asunto(s)
Neuronas/fisiología , Potenciales de Acción , Agonistas alfa-Adrenérgicos/farmacología , Animales , Técnicas de Cultivo de Célula/instrumentación , Técnicas de Cultivo de Célula/métodos , Células Cultivadas , Estimulación Eléctrica , Electrodos , Hipocampo/citología , Red Nerviosa/citología , Norepinefrina/farmacología , Análisis de Componente Principal
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