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1.
Hum Mol Genet ; 29(12): 2051-2064, 2020 07 29.
Artículo en Inglés | MEDLINE | ID: mdl-32426821

RESUMEN

Prader-Willi syndrome (PWS) is a neurodevelopmental disorder that is characterized by metabolic alteration and sleep abnormalities mostly related to rapid eye movement (REM) sleep disturbances. The disease is caused by genomic imprinting defects that are inherited through the paternal line. Among the genes located in the PWS region on chromosome 15 (15q11-q13), small nucleolar RNA 116 (Snord116) has been previously associated with intrusions of REM sleep into wakefulness in humans and mice. Here, we further explore sleep regulation of PWS by reporting a study with PWScrm+/p- mouse line, which carries a paternal deletion of Snord116. We focused our study on both macrostructural electrophysiological components of sleep, distributed among REMs and nonrapid eye movements. Of note, here, we study a novel electroencephalography (EEG) graphoelements of sleep for mouse studies, the well-known spindles. EEG biomarkers are often linked to the functional properties of cortical neurons and can be instrumental in translational studies. Thus, to better understand specific properties, we isolated and characterized the intrinsic activity of cortical neurons using in vitro microelectrode array. Our results confirm that the loss of Snord116 gene in mice influences specific properties of REM sleep, such as theta rhythms and, for the first time, the organization of REM episodes throughout sleep-wake cycles. Moreover, the analysis of sleep spindles present novel specific phenotype in PWS mice, indicating that a new catalog of sleep biomarkers can be informative in preclinical studies of PWS.


Asunto(s)
Impresión Genómica/genética , Síndrome de Prader-Willi/genética , ARN Nucleolar Pequeño/genética , Sueño/genética , Animales , Modelos Animales de Enfermedad , Electroencefalografía , Humanos , Ratones , Neuronas/metabolismo , Neuronas/patología , Fenotipo , Síndrome de Prader-Willi/fisiopatología , Sueño/fisiología , Sueño REM/genética
2.
Mol Ther ; 29(10): 3072-3092, 2021 10 06.
Artículo en Inglés | MEDLINE | ID: mdl-34058387

RESUMEN

A common feature of diverse brain disorders is the alteration of GABA-mediated inhibition because of aberrant, intracellular chloride homeostasis induced by changes in the expression and/or function of chloride transporters. Notably, pharmacological inhibition of the chloride importer NKCC1 is able to rescue brain-related core deficits in animal models of these pathologies and in some human clinical studies. Here, we show that reducing NKCC1 expression by RNA interference in the Ts65Dn mouse model of Down syndrome (DS) restores intracellular chloride concentration, efficacy of gamma-aminobutyric acid (GABA)-mediated inhibition, and neuronal network dynamics in vitro and ex vivo. Importantly, adeno-associated virus (AAV)-mediated, neuron-specific NKCC1 knockdown in vivo rescues cognitive deficits in diverse behavioral tasks in Ts65Dn animals. Our results highlight a mechanistic link between NKCC1 expression and behavioral abnormalities in DS mice and establish a molecular target for new therapeutic approaches, including gene therapy, to treat brain disorders characterized by neuronal chloride imbalance.


Asunto(s)
Síndrome de Down/terapia , Terapia Genética/métodos , Miembro 2 de la Familia de Transportadores de Soluto 12/genética , Animales , Cloruros/metabolismo , Modelos Animales de Enfermedad , Síndrome de Down/genética , Síndrome de Down/psicología , Técnicas de Silenciamiento del Gen , Homeostasis , Masculino , Ratones , Neuronas/metabolismo , Interferencia de ARN
3.
iScience ; 27(4): 109438, 2024 Apr 19.
Artículo en Inglés | MEDLINE | ID: mdl-38544574

RESUMEN

Gamma-aminobutyric acid (GABA) is the main inhibitory neurotransmitter in adults. Depolarizing GABA responses have been well characterized at neuronal-population average level during typical neurodevelopment and partially in brain disorders. However, no investigation has specifically assessed whether a mosaicism of cells with either depolarizing or hyperpolarizing/inhibitory GABAergic responses exists in animals in health/disease at diverse developmental stages, including adulthood. Here, we showed that such mosaicism is present in wild-type (WT) and down syndrome (DS) neuronal networks, as assessed at increasing scales of complexity (cultures, brain slices, behaving mice). Nevertheless, WT mice presented a much lower percentage of cells with depolarizing GABA than DS mice. Restoring the mosaicism of hyperpolarizing and depolarizing GABA-responding neurons to WT levels rescued anxiety behavior in DS mice. Moreover, we found heterogeneous GABAergic responses in developed control and trisomic human induced-pluripotent-stem-cells-derived neurons. Thus, a heterogeneous subpopulation of GABA-responding cells exists in physiological/pathological conditions in mouse and human neurons, possibly contributing to disease-associated behaviors.

4.
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.

5.
Epigenetics Chromatin ; 14(1): 1, 2021 01 06.
Artículo en Inglés | MEDLINE | ID: mdl-33407878

RESUMEN

BACKGROUND: DNA methylation has emerged as an important epigenetic regulator of brain processes, including circadian rhythms. However, how DNA methylation intervenes between environmental signals, such as light entrainment, and the transcriptional and translational molecular mechanisms of the cellular clock is currently unknown. Here, we studied the after-hours mice, which have a point mutation in the Fbxl3 gene and a lengthened circadian period. METHODS: In this study, we used a combination of in vivo, ex vivo and in vitro approaches. We measured retinal responses in Afh animals and we have run reduced representation bisulphite sequencing (RRBS), pyrosequencing and gene expression analysis in a variety of brain tissues ex vivo. In vitro, we used primary neuronal cultures combined to micro electrode array (MEA) technology and gene expression. RESULTS: We observed functional impairments in mutant neuronal networks, and a reduction in the retinal responses to light-dependent stimuli. We detected abnormalities in the expression of photoreceptive melanopsin (OPN4). Furthermore, we identified alterations in the DNA methylation pathways throughout the retinohypothalamic tract terminals and links between the transcription factor Rev-Erbα and Fbxl3. CONCLUSIONS: The results of this study, primarily represent a contribution towards an understanding of electrophysiological and molecular phenotypic responses to external stimuli in the Afh model. Moreover, as DNA methylation has recently emerged as a new regulator of neuronal networks with important consequences for circadian behaviour, we discuss the impact of the Afh mutation on the epigenetic landscape of circadian biology.


Asunto(s)
Ritmo Circadiano , Metilación de ADN , Animales , Ritmo Circadiano/genética , Ratones , Mutación , Fenotipo , Factores de Transcripción/genética
6.
Adv Neurobiol ; 22: 351-387, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31073944

RESUMEN

One of the main limitations preventing the realization of a successful dialogue between the brain and a putative enabling device is the intricacy of brain signals. In this perspective, closed-loop in vitro systems can be used to investigate the interactions between a network of neurons and an external system, such as an interacting environment or an artificial device. In this chapter, we provide an overview of closed-loop in vitro systems, which have been developed for investigating potential neuroprosthetic applications. In particular, we first explore how to modify or set a target dynamical behavior in a network of neurons. We then analyze the behavior of in vitro systems connected to artificial devices, such as robots. Finally, we provide an overview of biological neuronal networks interacting with artificial neuronal networks, a configuration currently offering a promising solution for clinical applications.


Asunto(s)
Técnicas de Cultivo de Célula/métodos , Técnicas In Vitro/métodos , Red Nerviosa/citología , Redes Neurales de la Computación , Neuronas/citología , Robótica/métodos , Encéfalo/citología , Humanos
7.
iScience ; 19: 402-414, 2019 Sep 27.
Artículo en Inglés | MEDLINE | ID: mdl-31421595

RESUMEN

Recent advances in bioelectronics and neural engineering allowed the development of brain machine interfaces and neuroprostheses, capable of facilitating or recovering functionality in people with neurological disability. To realize energy-efficient and real-time capable devices, neuromorphic computing systems are envisaged as the core of next-generation systems for brain repair. We demonstrate here a real-time hardware neuromorphic prosthesis to restore bidirectional interactions between two neuronal populations, even when one is damaged or missing. We used in vitro modular cell cultures to mimic the mutual interaction between neuronal assemblies and created a focal lesion to functionally disconnect the two populations. Then, we employed our neuromorphic prosthesis for bidirectional bridging to artificially reconnect two disconnected neuronal modules and for hybrid bidirectional bridging to replace the activity of one module with a real-time hardware neuromorphic Spiking Neural Network. Our neuroprosthetic system opens avenues for the exploitation of neuromorphic-based devices in bioelectrical therapeutics for health care.

8.
J Vis Exp ; (135)2018 05 15.
Artículo en Inglés | MEDLINE | ID: mdl-29863681

RESUMEN

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) is a promising approach when pharmacological treatment fails or neurosurgery is not recommended. Acute brain slices coupled to microelectrode arrays (MEAs) represent a valuable tool to study neuronal network interactions and their modulation by electrical stimulation. As compared to conventional extracellular recording techniques, they provide the added advantages of a greater number of observation points and a known inter-electrode distance, which allow studying the propagation path and speed of electrophysiological signals. However, tissue oxygenation may be greatly impaired during MEA recording, requiring a high perfusion rate, which comes at the cost of decreased signal-to-noise ratio and higher oscillations in the experimental temperature. Electrical stimulation further stresses the brain tissue, making it difficult to pursue prolonged recording/stimulation epochs. Moreover, electrical modulation of brain slice activity needs to target specific structures/pathways within the brain slice, requiring that electrode mapping be easily and quickly performed live during the experiment. Here, we illustrate how to perform the recording and electrical modulation of 4-aminopyridine (4AP)-induced epileptiform activity in rodent brain slices using planar MEAs. We show that the brain tissue obtained from mice outperforms rat brain tissue and is thus better suited for MEA experiments. This protocol guarantees the generation and maintenance of a stable epileptiform pattern that faithfully reproduces the electrophysiological features observed with conventional field potential recording, persists for several hours, and outlasts sustained electrical stimulation for prolonged epochs. Tissue viability throughout the experiment is achieved thanks to the use of a small-volume custom recording chamber allowing for laminar flow and quick solution exchange even at low (1 mL/min) perfusion rates. Quick MEA mapping for real-time monitoring and selection of stimulating electrodes is performed by a custom graphic user interface (GUI).


Asunto(s)
Encéfalo/cirugía , Estimulación Eléctrica/métodos , Fenómenos Electrofisiológicos/fisiología , Microelectrodos/estadística & datos numéricos , Neuronas/metabolismo , Animales , Encéfalo/patología , Ratas , Roedores
9.
Brain Neurosci Adv ; 2: 2398212818776475, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-32166141

RESUMEN

BACKGROUND: In recent years, biomedical devices have proven to be able to target also different neurological disorders. Given the rapid ageing of the population and the increase of invalidating diseases affecting the central nervous system, there is a growing demand for biomedical devices of immediate clinical use. However, to reach useful therapeutic results, these tools need a multidisciplinary approach and a continuous dialogue between neuroscience and engineering, a field that is named neuroengineering. This is because it is fundamental to understand how to read and perturb the neural code in order to produce a significant clinical outcome. RESULTS: In this review, we first highlight the importance of developing novel neurotechnological devices for brain repair and the major challenges expected in the next years. We describe the different types of brain repair strategies being developed in basic and clinical research and provide a brief overview of recent advances in artificial intelligence that have the potential to improve the devices themselves. We conclude by providing our perspective on their implementation to humans and the ethical issues that can arise. CONCLUSIONS: Neuroengineering approaches promise to be at the core of future developments for clinical applications in brain repair, where the boundary between biology and artificial intelligence will become increasingly less pronounced.

10.
Front Neurosci ; 10: 315, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27458335

RESUMEN

In this paper, we show that neuronal assemblies plated on Micro Electrode Arrays present synchronized, low frequency firing patterns similar to in vivo slow wave oscillations, which are a key parameter of sleep-like state. Although neuronal cultures lack the characteristic high-frequency waves of wakefulness, it is possible to modulate their spontaneous firing pattern through the administration of specific neurotransmitters such as acetylcholine. We thus stimulated the cortical cultures with an agonist of acetylcholine receptor, Carbachol, which caused a desynchronization of the spontaneous firing of the cultures. We recorded and monitored the cultures for a period of over 31 h. We analyzed the electrophysiological signals by exploiting novel methodological approaches, taking into account the different temporal scales of the recorded signals, and considering both spikes and local field potentials. Supporting the electrophysiological analysis results, gene expressions of targeted genes showed the activation of specific markers involved in sleep-wake rhythms. Our results demonstrate that the Carbachol treatment induces desynchronization of neuronal activity, altering sleep-like properties in an in vitro model.

11.
Front Neurosci ; 10: 409, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27610075

RESUMEN

[This corrects the article on p. 315 in vol. 10, PMID: 27458335.].

12.
Front Neuroeng ; 6: 10, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-24312049

RESUMEN

Hippocampal networks exhibit spontaneous electrophysiological activity that can be modulated by pharmacological manipulation and can be monitored over time using Micro-Electrode Arrays (MEAs), devices composed by a glass substrate and metal electrodes. The typical mode of activity of these dissociated cultures is the network-wide bursting pattern, which, if properly chemically modulated, can recall the ictal events of the epileptic phenotypes and is well-suited to study the effects of antiepileptic compounds. In this paper, we analyzed the changes induced by Carbamazepine (CBZ) and Valproate (VPA) on mature networks of hippocampal neurons in "control" condition (i.e., in the culturing medium) and upon treatment with the pro-convulsant bicuculline (BIC). We found that, in both control and BIC-treated networks, high doses (100 µM-1 mM) of CBZ almost completely suppressed the spiking and bursting activity of hippocampal neurons. On the contrary, VPA never completely abolish the electrophysiological activity in both experimental designs. Interestingly, VPA cultures pre-treated with BIC showed dual effects. In fact, in some cultures, at low VPA concentrations (100 nM-1 µM), we observed decreased firing/bursting levels, which returned to values comparable to BIC-evoked activity at high VPA concentrations (100 µM-1 mM). In other cultures, VPA reduced BIC-evoked activity in a concentration-independent manner. In conclusion, our study demonstrates that MEA-coupled hippocampal networks are responsive to chemical manipulations and, upon proper pharmacological modulation, might provide model systems to detect acute pharmacological effects of antiepileptic drugs.

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