1,720,988 research outputs found
Reduced inhibitory gate in the barrel cortex of Neuroligin3R451C knock-in mice, an animal model of Autism Spectrum Disorders
Full text linkNeuroligins are postsynaptic adhesion molecules that interacting with presynaptic neurexins ensure the cross-talk between pre-and postsynaptic specializations. Rare mutations in neurexin-neuroligin genes have been linked to autism spectrum disorders (ASDs). One of these, the R451C mutation of the gene encoding for Neuroligin3 (Nlgn3), has been found in patients with familial forms of ASDs. Animals carrying this mutation (NL3R451C knock-in mice) exhibit impaired social behaviors, reminiscent of those observed in ASD patients, associated with major alterations in both GABAergic and glutamatergic transmission, which vary among different brain regions and at different developmental stages. Here, pair recordings from parvalbumin-(PV) expressing basket cells and spiny neurons were used to study GABAergic synaptic signaling in layer IV barrel cortex of NL3R451C mutant mice. We found that the R451C mutation severely affects the probability of GABA release from PV-expressing basket cells, responsible for controlling via thalamo-cortical inputs the feed-forward inhibition. No changes in excitatory inputs to parvalbumin-positive basket cells or spiny neurons were detected. These data clearly show that primary targets of the NL3 mutation are PV-expressing basket cells, independently of the brain region where they are localized. Changes in the inhibitory gate of layer IV somatosensory cortex may alter sensory processing in ASD patients leading to misleading sensory representations with difficulties to combine pieces of information into a unified perceptual whole. © 2014 The Authors
GABAergic signaling as therapeutic target for Autism Spectrum Disorders
Full text linkGABA, the main inhibitory neurotransmitter in the adult brain, early in postnatal life exerts a depolarizing and excitatory action. This depends on accumulation of chloride inside the cell via the cation-chloride importer NKCC1, being the expression of the chloride exporter KCC2 very low at birth. The developmentally regulated expression of KCC2 results in extrusion of chloride with age and a shift of GABA from the depolarizing to the hyperpolarizing direction. The depolarizing action of GABA leads to intracellular calcium rise through voltage-dependent calcium channels and/or NMDA receptors. GABA-mediated calcium signals regulate a variety of developmental processes from cell proliferation migration, differentiation, synapse maturation and neuronal wiring. Therefore, it is not surprising that some forms of neuro-developmental disorders such as Autism Spectrum Disorders (ASDs) are associated with alterations of GABAergic signaling and impairment of the excitatory/inhibitory balance in selective neuronal circuits. In this review we will discuss how changes of GABAA-mediated neurotransmission affect several forms of ASDs including the Fragile X, the Angelman and Rett syndromes. Then, we will describe various animal models of ASDs with GABAergic dysfunctions, highlighting their behavioral deficits and the possibility to rescue them by targeting selective components of the GABAergic synapse. In particular, we will discuss how in some cases, reverting the polarity of GABA responses from the depolarizing to the hyperpolarizing direction with the diuretic bumetanide, a selective blocker of NKCC1, may have beneficial effects on ASDs, thus opening new therapeutic perspectives for the treatment of these devastating disorders
Graphene Oxide Nanosheets Reduce Astrocyte Reactivity to Inflammation and Ameliorate Experimental Autoimmune Encephalomyelitis
Full text linkIn neuroinflammation, astrocytes play multifaceted roles
that regulate
the neuronal environment. Astrocytes sense and respond to pro-inflammatory
cytokines (CKs) and, by a repertoire of intracellular Ca2+ signaling, contribute to disease progression. Therapeutic approaches
wish to reduce the overactivation in Ca2+ signaling in
inflammatory-reactive astrocytes to restore dysregulated cellular
changes. Cell-targeting therapeutics might take advantage by the use
of nanomaterial-multifunctional platforms such as graphene oxide (GO).
GO biomedical applications in the nervous system involve therapeutic
delivery and sensing, and GO flakes were shown to enable interfacing
of neuronal and glial membrane dynamics. We exploit organotypic spinal
cord cultures and optical imaging to explore Ca2+ changes
in astrocytes, and we report, when spinal tissue is exposed to CKs,
neuroinflammatory-associated modulation of resident glia. We show
the efficacy of GO to revert these dynamic changes in astrocytic reactivity
to CKs, and we translate this potential in an animal model of immune-mediated
neuroinflammatory disease
Thin graphene oxide nanoflakes modulate glutamatergic synapses in the amygdala cultured circuits: exploiting synaptic approaches to anxiety disorders
Full text linkAnxiety disorders (ADs) are nervous system maladies involving changes in the amygdala synaptic circuitry, such as an upregulation of excitatory neurotransmission at glutamatergic synapses. In the field of nanotechnology, thin graphene oxide flakes with nanoscale lateral size (s-GO) have shown outstanding promise for the manipulation of excitatory neuronal transmission with high temporal and spatial precision, thus they were considered as ideal candidates for modulating amygdalar glutamatergic transmission. Here, we validated an in vitro model of amygdala circuitry as a screening tool to target synapses, towards development of future ADs treatments. After one week in vitro, dissociated amygdalar neurons reconnected forming functional networks, whose development recapitulated that of the tissue of origin. When acutely applied to these cultures, s-GO flakes induced a selective modification of excitatory activity. This type of interaction between s-GO and amygdalar neurons may form the basis for the exploitation of alternative approaches in the treatment of ADs
Bilirubin Triggers Calcium Elevations and Dysregulates Giant Depolarizing Potentials During Rat Hippocampus Maturation
Full text linkNeonatal hyperbilirubinemia may result in long-lasting motor, auditory and learning impairments. The mechanisms responsible for the localization of unconjugated bilirubin (UCB) to specific brain areas as well as those involved in potentially permanent central nervous system (CNS) dysfunctions are far from being clear. One area of investigation includes exploring how hyperbilirubinemia determines neuronal alterations predisposing to neurodevelopmental disorders. We focused on the hippocampus and pyramidal cell dysregulation of calcium homeostasis and synaptic activity, with a particular focus on early forms of correlated network activity, i.e., giant depolarizing potentials (GDPs), crucially involved in shaping mature synaptic networks. We performed live calcium imaging and patch clamp recordings from acute hippocampal slices isolated from wild-type rats exposed to exogenous high bilirubin concentration. We then explored the impact of endogenous bilirubin accumulation in hippocampal slices isolated from a genetic model of hyperbilirubinemia, i.e., Gunn rats. Our data show in both models an age-dependent dysregulation of calcium dynamics accompanied by severe alterations in GDPs, which were strongly reduced in hippocampal slices of hyperbilirubinemic rats, where the expression of GABAergic neurotransmission markers was also altered. We propose that hyperbilirubinemia damages neurons and affects the refinement of GABAergic synaptic circuitry during a critical period of hippocampal development
Reprogramming fibroblasts to neural-precursor-like cells by structured overexpression of pallial patterning genes
No full textIn this study, we assayed the capability of four genes implicated in embryonic specification of the cortico-cerebral field, Foxg1, Pax6, Emx2 and Lhx2, to reprogramme mouse embryonic fibroblasts towards neural identities. Lentivirus-mediated, TetON-dependent overexpression of Pax6 and Foxg1 transgenes specifically activated the neural stem cell (NSC) reporter Sox1-EGFP in a substantial fraction of engineered cells. The efficiency of this process was enhanced up to ten times by simultaneous inactivation of Trp53 and co-administration of a specific drug mix inhibiting HDACs, H3K27-HMTase and H3K4m2-demethylase. Remarkably, a fraction of the reprogrammed population expressed other NSC markers and retained its new identity, even after switching off the reprogramming transgenes. When transferred into a pro-differentiative environment, Pax6/Foxg1-overexpressing cells activated the neuronal marker Tau-EGFP. Frequency of Tau-EGFP positive cells was almost doubled upon delayed delivery of Emx2 and Lhx2 transgenes. A further improvement of the neuron-like cell output was achieved by inhibition of the BMP and TGFβ pathways. Tau-EGFP positive cells were able to generate action potentials upon injection of depolarizing current pulses, further indicating their neuron-like phenotype
Towards new generation of neuro-implantable devices : engineering neuron/carbon nanotubes integrated functional units
No full text2008/2009Le nanotecnologie sono un campo delle scienze che utilizza materiali e dispositivi ingegnerizzati aventi la più piccola organizzazione funzionale a livello di dimensioni nanometriche. Questo implica che nanodispositivi e nanomateriali possano interagire con i sistemi biologici a livello molecolare con un elevato grado di specificità. É largamente accettato che l’applicazione delle nanotecnologie nell’ambito delle neuroscienze abbia un forte potenziale (Silva, 2006). In questo contesto, i nanotubi di carbonio (CNT), un’innovativa forma di carbonio composta da strutture tubulari di grafite dalle dimensioni nanometriche dotate di buone proprietà di conduzione elettrica, si sono dimostrati promettenti candidati per sviluppare la tecnologia di dispositivi impiantabili in ambito biomedico.
Diversi studi hanno dimostrato la biocompatibilità dei substrati di CNT per i neuroni in termini di adesione, crescita e differenziamento cellulare (riassunti in Sucapane et al., 2009).
Al fine di aumentare la nostra conoscenza riguardo alle interazioni presenti in sistemi ibridi formati da CNT e neuroni, abbiamo caratterizzato l’attività di reti neuronali cresciuti su supporti di CNT attraverso la tecnica del patch clamp.
Il nostro gruppo ha riportato che circuti neuronali cresciuti in vitro su substrati di CNT presentano un’aumentata attività sinaptica spontanea rispetto al controllo a fronte di comparabili proprietà base (proprietà passive di membrana, morfologia e densità dei neuroni) delle colture nelle due condizioni di crescita (Lovat et al., 2005).
Si è quindi ipotizzato che tale aumentata attività spontanea potesse originare da una modificazione nel modo in cui i singoli neuroni generano il segnale elettrico.
A tal fine, si sono monitorate variazioni nelle proprietà elettrogeniche di singoli neuroni, utilizzando un protocollo standard per caratterizzare l’integrazione di potenziali d’azione retropropaganti nei dendriti (Larkum et al., 1999). In configurazione current clamp, attraverso brevi iniezioni di corrente nel soma della cellula, abbiamo indotto una serie di regolari potenziali d’azione (PA) a varie frequenze nel neurone sotto registrazione, quindi abbiamo studiato la presenza di un’addizionale depolarizzazione somatica dopo l’ultimo PA del treno.
Abbiamo osservato che neuroni di controllo mostrano nella maggioranza dei casi una iperpolarizzazione (AHP) del potenziale di membrana dopo l’ultimo PA del treno, mentre una depolarizzazione (ADP) è presente solo in una piccola quota di casi. In presenza di CNT, invece, l’ADP risulta essere l’evento predominante.
L’ADP è inoltre abolita dall’applicazione di CoCl2, un bloccante non specifico dei canali calcio voltaggio dipendenti. Per di più, l’area dell’ADP può essere diminuita dall’applicazione di nifedipina (10 μM) e l’ulteriore coapplicazione di NiCl2 (50 μM) elimina totalmente l’ADP, suggerendo che sia i canali calcio voltaggio dipendenti ad alta soglia di attivazione, sia quelli a bassa soglia, siano coinvolti in questo processo (Cellot et al., 2009).
Attraverso la microscopia elettronica a trasmissione (TEM) e, più recentemente, mediante quella a scansione (SEM) è stata messa in evidenza la presenza di discontinui punti di stretto contatto tra CNT e membrane neuronali: la nostra ipotesi è che tali strutture ibride siano in grado di favorire la retropropagazione dei PA nei dendriti distali.
La maggiore eccitabilità a livello del singolo neurone, inoltre, potrebbe essere responsabile dell’incremento di attività spontanea della rete neuronale.
Abbiamo quindi ulteriormente caratterizzato l’attività della rete neuronale attraverso registrazioni da coppie di neuroni, dove il neurone presinaptico veniva stimolato ad avere treni di potenziali d’azione a 20 Hz in configurazione current clamp e simultaneamente il neurone postsinaptico era monitorato in configurazione voltage clamp per vedere la presenza o l’assenza di una risposta sinaptica.
I nostri esperimenti indicano che la probabilità di trovare connessioni monosinaptiche gabaergiche tra neuroni è aumentata in presenza di CNT (56% vs 40% in controllo). Inoltre, è stato rilevato un ulteriore effetto dei CNT sulla plasticità a breve termine delle sinapsi: nelle condizioni di controllo, treni di potenziali d’azione nella cellula presinaptica evocano nella cellula postsinaptica nel 90% dei casi una chiara depressione nell’ampiezza di consecutivi ePSCs, mentre solo in meno del 10% è possibile rilevare una facilitazione. Al contrario, in presenza di CNT, nel 39% delle coppie, il neurone postsinaptico risponde in modo chiaramente facilitativo.
Nelle più recenti serie di esperimenti, abbiamo voluto indagare più approfonditamente l’origine di questa modificazione in termini di plasticità sinaptica; a tal fine, abbiamo trattato neuroni in controllo e su CNT con tetrodotossina 1 µM per 5 giorni, al fine di bloccare completamente l’attività elettrica della rete neuronale, e abbiamo compiuto delle registrazioni da coppie di neuroni.
Mentre la risposta prevalentemente di depressione dei controlli non è modificata da tale trattamento, neuroni cresciuti su substrati di cnt in condizioni di blocco dell’attività elettrica non presentano più sinapsi con caratteristiche di facilitazione, ma hanno un comportamento simile ai contolli. Questi risultati indicano che la facilitazione è una proprietà tipica di sinapsi attive sviluppatesi in presenza di CNT.XXII Ciclo198
Interfacing neurons with carbon nanotubes:(re)engineering neuronal signaling
No full textAbstract: Carbon nanotubes (CNTs) are cylindrically shaped nanostructures made by sheets of
graphene rolled up to form hollow tubes. Owing to their unique range of thermal, electronic, and
structural properties, CNTs have been rapidly developing as a technology platform for biological and
medical applications, including those designed to develop novel neuro-implantable devices. Depending
on their structure, CNTs combine an incredible strength with an extreme flexibility. Further, these
materials exhibit physical and chemical properties which allow them to efficiently conduit electrical
current in electrochemical interfaces. CNTs can be organized in scaffolds made up of small fibers or
tubes with diameters similar to those of neural processes such as axons and dendrites. Recently, CNT
scaffolds have been found to promote growth, differentiation, and survival of neurons and to modify
their electrophysiological properties. These features make CNTs an attractive material for the design
of nano–bio hybrid systems able to govern cell-specific behaviors in cultured neuronal networks.
The leading scope of this short review is to highlight how nanotube scaffolds can impact on neuronal
signaling ability. In particular, we will focus on the direct and specific interactions between this synthetic
nanomaterial and biological cell membranes, and on the ability of CNTs to improve interfaces developed
to record or to stimulate neuronal activity.
CNTs hold the potential for the development of innovative nanomaterial-based neurological implants.
Therefore, it is particularly relevant to improve our knowledge on the impact on neuronal performance
of interfacing nerve cells with CNTs
Interplay Among Synaptic Glutamate Release and Excitotoxicity: Neuronal Damage and Graphene-Based Materials Related Protection
Full text linkGlutamate-related excitotoxicity represents a fundamental pathological process underlying both acute and chronic disorders of the central nervous system. Excessive stimulation of ionotropic and metabotropic glutamate receptors induces ionic dysregulation, mitochondrial dysfunction, and oxidative stress, which can activate necrotic and apoptotic pathways, processes further amplified by defective glutamate clearance and astrocytic impairment. These mechanisms are recognized as key contributors to neuronal damage in ischemic stroke, Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease, identifying excitotoxicity as a convergent hallmark of neurodegeneration. Despite considerable progress in elucidating its molecular mechanisms, clinical translation of excitotoxicity-targeted interventions remains limited, largely due to the difficulty of selectively attenuating pathological glutamatergic activity while preserving physiological neurotransmission. Recent advances in nanotechnology, particularly the development of graphene-based materials (GBMs), have offered innovative approaches for neuroprotection. Owing to their unique physicochemical properties and compatibility with neural tissue, GBMs have been investigated as platforms for neural interfacing, regenerative scaffolds, drug delivery platforms, and direct modulators of glutamatergic transmission. In particular, small graphene oxide nanosheets exhibit the capacity to downregulate glutamate release and confer anti-inflammatory and neuroprotective effects. These findings suggest that GBMs may represent a promising class of neuromodulatory tools for mitigating excitotoxic injury, warranting further preclinical and translational investigations
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