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1.
Proc Natl Acad Sci U S A ; 115(19): 5004-5009, 2018 05 08.
Artículo en Inglés | MEDLINE | ID: mdl-29691318

RESUMEN

Tonic inhibition in the brain is mediated through an activation of extrasynaptic GABAA receptors by the tonically released GABA, resulting in a persistent GABAergic inhibitory action. It is one of the key regulators for neuronal excitability, exerting a powerful action on excitation/inhibition balance. We have previously reported that astrocytic GABA, synthesized by monoamine oxidase B (MAOB), mediates tonic inhibition via GABA-permeable bestrophin 1 (Best1) channel in the cerebellum. However, the role of astrocytic GABA in regulating neuronal excitability, synaptic transmission, and cerebellar brain function has remained elusive. Here, we report that a reduction of tonic GABA release by genetic removal or pharmacological inhibition of Best1 or MAOB caused an enhanced neuronal excitability in cerebellar granule cells (GCs), synaptic transmission at the parallel fiber-Purkinje cell (PF-PC) synapses, and motor performance on the rotarod test, whereas an augmentation of tonic GABA release by astrocyte-specific overexpression of MAOB resulted in a reduced neuronal excitability, synaptic transmission, and motor performance. The bidirectional modulation of astrocytic GABA by genetic alteration of Best1 or MAOB was confirmed by immunostaining and in vivo microdialysis. These findings indicate that astrocytes are the key player in motor coordination through tonic GABA release by modulating neuronal excitability and could be a good therapeutic target for various movement and psychiatric disorders, which show a disturbed excitation/inhibition balance.


Asunto(s)
Astrocitos/metabolismo , Cerebelo/metabolismo , Desempeño Psicomotor/fisiología , Células de Purkinje/metabolismo , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/metabolismo , Animales , Astrocitos/citología , Bestrofinas/genética , Bestrofinas/metabolismo , Cerebelo/citología , Ratones Endogámicos BALB C , Ratones Noqueados , Monoaminooxidasa/genética , Monoaminooxidasa/metabolismo , Células de Purkinje/citología , Ácido gamma-Aminobutírico/genética
2.
J Nanobiotechnology ; 13: 92, 2015 Dec 18.
Artículo en Inglés | MEDLINE | ID: mdl-26683698

RESUMEN

BACKGROUND: Multi-walled carbon nanotubes (MW-CNTs) have been extensively explored for their possible beneficial use in the nervous system. CNTs have shown to modulate neuronal growth and electrical properties, but its effect that varying length of MW-CNTs on primary astrocyte roles have not been clearly demonstrated yet. RESULTS: We investigate here the effect of MW-CNTs on astrocytic morphology, cell-cell interaction and the distribution of intracellular GABA (gamma-amino butyric acid). Primary cultured cortical astrocytes on MW-CNT-coated glass coverslips grow rounder and make more cell-cell interactions, with many cell processes, compared to astrocytes on poly-D-lysine (PDL) coverslips. In addition, intracellular GABA spreads into the cell processes of astrocytes on MW-CNT coverslips. When this GABA spreads into cell processes from the cell body GABA can be released more easily and in larger quantities compared to astrocytes on PDL coverslips. CONCLUSIONS: Our result confirm that MW-CNTs modulate astrocytic morphology, the distribution of astrocytic GABA, cell-cell interactions and the extension of cell processes. CNTs look to be a promising material for use neuroprosthetics such as brain-machine interface technologies.


Asunto(s)
Astrocitos/metabolismo , Uniones Intercelulares/metabolismo , Nanotubos de Carbono/química , Ácido gamma-Aminobutírico/metabolismo , Animales , Animales Recién Nacidos , Astrocitos/efectos de los fármacos , Astrocitos/ultraestructura , Interfaces Cerebro-Computador/estadística & datos numéricos , Comunicación Celular , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Corteza Cerebral/ultraestructura , Uniones Intercelulares/efectos de los fármacos , Uniones Intercelulares/ultraestructura , Ratones , Ratones Endogámicos BALB C , Polilisina/química , Polilisina/farmacología , Cultivo Primario de Células
3.
J Physiol ; 592(22): 4951-68, 2014 Nov 15.
Artículo en Inglés | MEDLINE | ID: mdl-25239459

RESUMEN

GABA is the major inhibitory transmitter in the brain and is released not only from a subset of neurons but also from glia. Although neuronal GABA is well known to be synthesized by glutamic acid decarboxylase (GAD), the source of glial GABA is unknown. After estimating the concentration of GABA in Bergmann glia to be around 5-10 mM by immunogold electron microscopy, we demonstrate that GABA production in glia requires MAOB, a key enzyme in the putrescine degradation pathway. In cultured cerebellar glia, both Ca(2+)-induced and tonic GABA release are significantly reduced by both gene silencing of MAOB and the MAOB inhibitor selegiline. In the cerebellum and striatum of adult mice, general gene silencing, knock out of MAOB or selegiline treatment resulted in elimination of tonic GABA currents recorded from granule neurons and medium spiny neurons. Glial-specific rescue of MAOB resulted in complete rescue of tonic GABA currents. Our results identify MAOB as a key synthesizing enzyme of glial GABA, which is released via bestrophin 1 (Best1) channel to mediate tonic inhibition in the brain.


Asunto(s)
Monoaminooxidasa/metabolismo , Neuroglía/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Animales , Calcio/metabolismo , Células Cultivadas , Cerebelo/citología , Cerebelo/metabolismo , Cuerpo Estriado/citología , Cuerpo Estriado/metabolismo , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Monoaminooxidasa/genética , Inhibición Neural , Neuroglía/fisiología
4.
Cell Death Dis ; 14(7): 474, 2023 07 27.
Artículo en Inglés | MEDLINE | ID: mdl-37500624

RESUMEN

Cell-to-cell propagation of protein aggregates has been implicated in the progression of neurodegenerative diseases. However, the underlying mechanism and modulators of this process are not fully understood. Here, we screened a small-molecule library in a search for agents that suppress the propagation of α-synuclein and mutant huntingtin (mHtt). These screens yielded several molecules, some of which were effective against both α-synuclein and mHtt. Among these molecules, we focused on simvastatin and pravastatin. Simvastatin administration in a transgenic model of synucleinopathy effectively ameliorated behavioral deficits and α-synuclein accumulation, whereas pravastatin had no effect. Because only simvastatin enters the brain effectively, these results suggest that inhibition of brain cholesterol synthesis is important in simvastatin effects. In cultured cells, accumulation of intracellular cholesterol, induced by genetic ablation of the NPC1 gene or by pharmacological treatment with U18666A, increased α-synuclein aggregation and secretion. In contrast, lowering cholesterol using methyl-ß-cyclodextrin or statins reversed α-synuclein aggregation and secretion in NPC1-knockout cells. Consistent with these observations, feeding a high-fat diet aggravated α-synuclein pathology and behavioral deficits in the preformed fibril-injected mouse model, an effect that was also reversed by simvastatin administration. These results suggest that statins suppress propagation of protein aggregates by lowering cholesterol in the brain.


Asunto(s)
Inhibidores de Hidroximetilglutaril-CoA Reductasas , Animales , Ratones , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismo , Colesterol/metabolismo , Inhibidores de Hidroximetilglutaril-CoA Reductasas/farmacología , Pravastatina/farmacología , Agregado de Proteínas , Simvastatina/farmacología
5.
Mol Neurodegener ; 18(1): 41, 2023 Jun 24.
Artículo en Inglés | MEDLINE | ID: mdl-37355598

RESUMEN

BACKGROUND: There are currently no disease-modifying therapeutics for Parkinson's disease (PD). Although extensive efforts were undertaken to develop therapeutic approaches to delay the symptoms of PD, untreated α-synuclein (α-syn) aggregates cause cellular toxicity and stimulate further disease progression. PROTAC (Proteolysis-Targeting Chimera) has drawn attention as a therapeutic modality to target α-syn. However, no PROTACs have yet shown to selectively degrade α-syn aggregates mainly owing to the limited capacity of the proteasome to degrade aggregates, necessitating the development of novel approaches to fundamentally eliminate α-syn aggregates. METHODS: We employed AUTOTAC (Autophagy-Targeting Chimera), a macroautophagy-based targeted protein degradation (TPD) platform developed in our earlier studies. A series of AUTOTAC chemicals was synthesized as chimeras that bind both α-syn aggregates and p62/SQSTM1/Sequestosome-1, an autophagic receptor. The efficacy of Autotacs was evaluated to target α-syn aggregates to phagophores and subsequently lysosomes for hydrolysis via p62-dependent macroautophagy. The target engagement was monitored by oligomerization and localization of p62 and autophagic markers. The therapeutic efficacy to rescue PD symptoms was characterized in cultured cells and mice. The PK/PD (pharmacokinetics/pharmacodynamics) profiles were investigated to develop an oral drug for PD. RESULTS: ATC161 induced selective degradation of α-syn aggregates at DC50 of ~ 100 nM. No apparent degradation was observed with monomeric α-syn. ATC161 mediated the targeting of α-syn aggregates to p62 by binding the ZZ domain and accelerating p62 self-polymerization. These p62-cargo complexes were delivered to autophagic membranes for lysosomal degradation. In PD cellular models, ATC161 exhibited therapeutic efficacy to reduce cell-to-cell transmission of α-syn and to rescue cells from the damages in DNA and mitochondria. In PD mice established by injecting α-syn preformed fibrils (PFFs) into brain striata via stereotaxic surgery, oral administration of ATC161 at 10 mg/kg induced the degradation of α-syn aggregates and reduced their propagation. ATC161 also mitigated the associated glial inflammatory response and improved muscle strength and locomotive activity. CONCLUSION: AUTOTAC provides a platform to develop drugs for PD. ATC161, an oral drug with excellent PK/PD profiles, induces selective degradation of α-syn aggregates in vitro and in vivo. We suggest that ATC161 is a disease-modifying drug that degrades the pathogenic cause of PD.


Asunto(s)
Enfermedad de Parkinson , Ratones , Animales , Enfermedad de Parkinson/metabolismo , alfa-Sinucleína/metabolismo , Autofagia , Proteolisis , Células Cultivadas , Encéfalo/metabolismo
6.
Mol Brain ; 15(1): 27, 2022 03 28.
Artículo en Inglés | MEDLINE | ID: mdl-35346306

RESUMEN

Abnormal deposition of α-synuclein aggregates in Lewy bodies and Lewy neurites is the hallmark lesion in Parkinson's disease (PD). These aggregates, thought to be the culprit of disease pathogenesis, spread throughout the brain as the disease progresses. Agents that inhibit α-synuclein aggregation and/or spread of aggregates would thus be candidate disease-modifying drugs. Here, we found that Chicago sky blue 6B (CSB) may be such a drug, showing that it inhibits α-synuclein aggregation and cell-to-cell propagation in both in vitro and in vivo models of synucleinopathy. CSB inhibited the fibrillation of α-synuclein in a concentration-dependent manner through direct binding to the N-terminus of α-synuclein. Furthermore, both seeded polymerization and cell-to-cell propagation of α-synuclein were inhibited by CSB treatment. Notably, CSB alleviated behavioral deficits and neuropathological features, such as phospho-α-synuclein and astrogliosis, in A53T α-synuclein transgenic mice. These results indicate that CSB directly binds α-synuclein and inhibits its aggregation, thereby blocking α-synuclein cell-to-cell propagation.


Asunto(s)
Enfermedad de Parkinson , alfa-Sinucleína , Animales , Cuerpos de Lewy/patología , Ratones , Enfermedad de Parkinson/patología , Azul de Tripano/metabolismo , alfa-Sinucleína/metabolismo
7.
Sci Rep ; 8(1): 11589, 2018 08 02.
Artículo en Inglés | MEDLINE | ID: mdl-30072733

RESUMEN

For decades, the glial function has been highlighted not only as the 'structural glue', but also as an 'active participant' in neural circuits. Here, we suggest that tumor necrosis factor α (TNF-α), a key inflammatory cytokine, alters the neural activity of the cerebellar Purkinje cells (PCs) by facilitating gliotransmission in the juvenile male rat cerebellum. A bath application of TNF-α (100 ng/ml) in acute cerebellar slices elevates spiking activity of PCs with no alterations in the regularity of PC firings. Interestingly, the effect of TNF-α on the intrinsic excitability of PCs was abolished under a condition in which the type1 TNF receptor (TNFR1) in Bergmann glia (BG) was genetically suppressed by viral delivery of an adeno-associated virus (AAV) containing TNFR1-shRNA. In addition, we measured the concentration of glutamate derived from dissociated cerebellar cortical astrocyte cultures treated with TNF-α and observed a progressive increase of glutamate in a time-dependent manner. We hypothesised that TNF-α-induced elevation of glutamate from BGs enveloping the synaptic cleft may directly activate metabotropic glutamate receptor1 (mGluR1). Pharmacological inhibition of mGluR1, indeed, prevented the TNF-α-mediated elevation of the intrinsic excitability in PCs. Taken together, our study reveals that TNF-α triggers glutamate release in BG, thereby increasing the intrinsic excitability of cerebellar PCs in a mGluR1-dependent manner.


Asunto(s)
Astrocitos/metabolismo , Ácido Glutámico/metabolismo , Células de Purkinje/metabolismo , Transmisión Sináptica , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Astrocitos/citología , Masculino , Ratones , Células de Purkinje/citología , Ratas , Receptores de Glutamato Metabotrópico/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo
9.
Nano Rev Exp ; 8(1): 1323853, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-30410703

RESUMEN

Introduction: Functionalised carbon nanotubes (CNTs) have been shown to be promising biomaterials in neural systems, such as CNT -based nerve scaffolds to drive nerve regeneration. CNTs have been shown to modulate neuronal growth and improve electrical conductivity of neurons. Methods: Cultured astrocytes on the functionalized CNTs (PEG, caroboxyl group) were assessed for distribution of GABA, glutamate uptake assay using isotope and change of conductance of CNTs by ATP. Immunostaining of GABA using anti-GABA (red), anti-GFAP (green) antibody in primary cortical astrocytes on MW-CNT and PDL coverslips. Results: The functionalization of CNTs has improved their solubility and biocompatibility and alters their cellular interaction pathways. Recently, CNTs have been shown to modulate morphofunctional characteristics of glia as well as neurons. Among the various types of glia, astrocytes express diverse receptors for corresponding neurotransmitters and release gliotransmitters, including glutamate, adenosine triphosphate, and γ-amino butyric acid. Gliotransmitters are primarily released from astrocytes and play important roles in glia-neuron crosstalk. Conclusion: This review focuses on the effects of CNTs on glial cells and discusses how functionalized CNTs can modulate morphology and gliotransmitters of glial cells. Based on exciting new findings, they look to be a promising material for use in brain disease therapy or neuroprosthetics.

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