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1.
Adv Funct Mater ; 32(8)2022 Feb 16.
Artículo en Inglés | MEDLINE | ID: mdl-35603230

RESUMEN

We report innovative scalable, vertical, ultra-sharp nanowire arrays that are individually addressable to enable long-term, native recordings of intracellular potentials. Stable amplitudes of intracellular potentials from 3D tissue-like networks of neurons and cardiomyocytes are obtained. Individual electrical addressability is necessary for high-fidelity intracellular electrophysiological recordings. This study paves the way toward predictive, high-throughput, and low-cost electrophysiological drug screening platforms.

2.
Hum Mol Genet ; 26(14): 2649-2666, 2017 07 15.
Artículo en Inglés | MEDLINE | ID: mdl-28444183

RESUMEN

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurological disorder that affects premutation carriers with 55-200 CGG-expansion repeats (preCGG) in FMR1, presenting with early alterations in neuronal network formation and function that precede neurodegeneration. Whether intranuclear inclusions containing DNA damage response (DDR) proteins are causally linked to abnormal synaptic function, neuronal growth and survival are unknown. In a mouse that harbors a premutation CGG expansion (preCGG), cortical and hippocampal FMRP expression is moderately reduced from birth through adulthood, with greater FMRP reductions in the soma than in the neurite, despite several-fold elevation of Fmr1 mRNA levels. Resting cytoplasmic calcium concentration ([Ca2+]i) in cultured preCGG hippocampal neurons is chronically elevated, 3-fold compared to Wt; elevated ROS and abnormal glutamatergic responses are detected at 14 DIV. Elevated µ-calpain activity and a higher p25/p35 ratio in the cortex of preCGG young adult mice indicate abnormal Cdk5 regulation. In support, the Cdk5 substrate, ATM, is upregulated by 1.5- to 2-fold at P0 and 6 months in preCGG brain, as is p-Ser1981-ATM. Bax:Bcl-2 is 30% higher in preCGG brain, indicating a greater vulnerability to apoptotic activation. Elevated [Ca2+]i, ROS, and DDR signals are normalized with dantrolene. Chronic [Ca2+]i dysregulation amplifies Cdk5-ATM signaling, possibly linking impaired glutamatergic signaling and DDR to neurodegeneration in preCGG brain.


Asunto(s)
Proteínas de la Ataxia Telangiectasia Mutada/metabolismo , Ataxia/genética , Ataxia/metabolismo , Calcio/metabolismo , Quinasa 5 Dependiente de la Ciclina/metabolismo , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/metabolismo , Temblor/genética , Temblor/metabolismo , Animales , Proteínas de la Ataxia Telangiectasia Mutada/genética , Quinasa 5 Dependiente de la Ciclina/genética , Modelos Animales de Enfermedad , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Neuronas/patología , ARN Mensajero/metabolismo , Expansión de Repetición de Trinucleótido
3.
Cell Tissue Res ; 371(2): 309-323, 2018 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-29018970

RESUMEN

Andersen's syndrome (AS) is a rare autosomal disorder that has been defined by the triad of periodic paralysis, cardiac arrhythmia, and developmental anomalies. AS has been directly linked to over 40 different autosomal dominant negative loss-of-function mutations in the KCNJ2 gene, encoding for the tetrameric strong inward rectifying K+ channel KIR2.1. While KIR2.1 channels have been suggested to contribute to setting the resting membrane potential (RMP) and to control the duration of the action potential (AP) in skeletal and cardiac muscle, the mechanism by which AS mutations produce such complex pathophysiological symptoms is poorly understood. Thus, we use an adenoviral transduction strategy to study in vivo subcellular distribution of wild-type (WT) and AS-associated mutant KIR2.1 channels in mouse skeletal muscle. We determined that WT and D71V AS mutant KIR2.1 channels are localized to the sarcolemma and the transverse tubules (T-tubules) of skeletal muscle fibers, while the ∆314-315 AS KIR2.1 mutation prevents proper trafficking of the homo- or hetero-meric channel complexes. Whole-cell voltage-clamp recordings in individual skeletal muscle fibers confirmed the reduction of inwardly rectifying K+ current (IK1) after transduction with ∆314-315 KIR2.1 as compared to WT channels. Analysis of skeletal muscle function revealed reduced force generation during isometric contraction as well as reduced resistance to muscle fatigue in extensor digitorum longus muscles transduced with AS mutant KIR2.1. Together, these results suggest that KIR2.1 channels may be involved in the excitation-contraction coupling process required for proper skeletal muscle function. Our findings provide clues to mechanisms associated with periodic paralysis in AS.


Asunto(s)
Síndrome de Andersen/genética , Técnicas de Silenciamiento del Gen , Músculo Esquelético/patología , Mutación/genética , Canales de Potasio de Rectificación Interna/genética , Adenoviridae/metabolismo , Síndrome de Andersen/patología , Síndrome de Andersen/fisiopatología , Animales , Células COS , Chlorocebus aethiops , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Activación del Canal Iónico , Contracción Isométrica , Ratones , Fatiga Muscular , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/fisiopatología
4.
Environ Sci Technol ; 51(24): 14406-14416, 2017 Dec 19.
Artículo en Inglés | MEDLINE | ID: mdl-29131945

RESUMEN

Nineteen ortho-substituted PCBs are chiral and found enantioselectively enriched in ecosystems. Their differential actions on biological targets are not understood. PCB 95 (2,2',3,5',6-pentachlorobiphenyl), a chiral PCB of current environmental relevance, is among the most potent toward modifying ryanodine receptors (RyR) function and Ca2+ signaling. PCB 95 enantiomers are separated and assigned aR- and aS-PCB 95 using three chiral-column HPLC and circular dichroism spectroscopy. Studies of RyR1-enriched microsomes show aR-PCB 95 with >4× greater potency (EC50 = 0.20 ± 0.05 µM), ∼ 1.3× higher efficacy (Bmax = 3.74 ± 0.07 µM) in [3H]Ryanodine-binding and >3× greater rates (R = 7.72 ± 0.31 nmol/sec/mg) of Ca2+ efflux compared with aS-PCB 95, whereas racemate has intermediate activity. aR-PCB 95 has modest selectivity for RyR2, and lower potency than racemate toward the RyR isoform mixture in brain membranes. Chronic exposure of hippocampal neuronal networks to nanomolar PCB 95 during a critical developmental period shows divergent influences on synchronous Ca2+ oscillation (SCO): rac-PCB 95 increasing and aR-PCB 95 decreasing SCO frequency at 50 nM, although the latter's effects are nonmonotonic at higher concentration. aS-PCB95 shows the greatest influence on inhibiting responses to 20 Hz electrical pulse trains. Considering persistence of PCB 95 in the environment, stereoselectivity toward RyRs and developing neuronal networks may clarify health risks associated with enantioisomeric enrichment of PCBs.


Asunto(s)
Hipocampo/efectos de los fármacos , Bifenilos Policlorados , Canal Liberador de Calcio Receptor de Rianodina/efectos de los fármacos , Animales , Ratones , Neuronas
5.
J Physiol ; 593(21): 4781-97, 2015 Nov 01.
Artículo en Inglés | MEDLINE | ID: mdl-26383921

RESUMEN

Muscle contraction is triggered by Ca(2+) ions released from the sarcoplasmic reticulum (SR) in response to depolarization of skeletal muscle fibres. Muscle activation is also associated with a voltage-activated trans-sarcolemmal Ca(2+) influx early identified as a current flowing through L-type Ca(2+) channels. Because removal of external Ca(2+) does not impede fibres from contracting, a negligible role was given to this voltage-activated Ca(2+) entry, although the decline of Ca(2+) release is more pronounced in the absence of Ca(2+) during long-lasting activation. Furthermore, it is not clearly established whether Ca(2+) exclusively flows through L-type channels or in addition through a parallel voltage-activated pathway distinct from L-type channels. Here, by monitoring the quenching of fura-2 fluorescence resulting from Mn(2+) influx in voltage-controlled mouse and zebrafish isolated muscle fibres, we show that the L-type current is the only contributor to Ca(2+) influx during long-lasting depolarizations in skeletal muscle. Calibration of the Mn(2+) quenching signal allowed us to estimate a mean Mn(2+) current of 0.31 ± 0.06 A F(-1) flowing through L-type channels during a train of action potentials. Measurements of SR Ca(2+) changes with fluo-5N in response to depolarization revealed that an elevated voltage-activated Ca(2+) current potentiated SR Ca(2+) loading and addition of external Mn(2+) produced quenching of fluo-5N in the SR, indicating that voltage-activated Ca(2+) /Mn(2+) influx contributes to SR Ca(2+) /Mn(2+) loading.


Asunto(s)
Canales de Calcio Tipo L/metabolismo , Señalización del Calcio , Músculo Esquelético/metabolismo , Retículo Sarcoplasmático/metabolismo , Potenciales de Acción , Animales , Calcio/metabolismo , Células Cultivadas , Manganeso/metabolismo , Contracción Muscular , Músculo Esquelético/fisiología , Pez Cebra
6.
J Biol Chem ; 289(27): 19180-90, 2014 Jul 04.
Artículo en Inglés | MEDLINE | ID: mdl-24847052

RESUMEN

Malignant hyperthermia (MH) is potentially fatal pharmacogenetic disorder of skeletal muscle caused by intracellular Ca(2+) dysregulation. NCX is a bidirectional transporter that effluxes (forward mode) or influxes (reverse mode) Ca(2+) depending on cellular activity. Resting intracellular calcium ([Ca(2+)]r) and sodium ([Na(+)]r) concentrations are elevated in MH susceptible (MHS) swine and murine muscles compared with their normal (MHN) counterparts, although the contribution of NCX is unclear. Lowering [Na(+)]e elevates [Ca(2+)]r in both MHN and MHS swine muscle fibers and it is prevented by removal of extracellular Ca(2+) or reduced by t-tubule disruption, in both genotypes. KB-R7943, a nonselective NCX3 blocker, reduced [Ca(2+)]r in both swine and murine MHN and MHS muscle fibers at rest and decreased the magnitude of the elevation of [Ca(2+)]r observed in MHS fibers after exposure to halothane. YM-244769, a high affinity reverse mode NCX3 blocker, reduces [Ca(2+)]r in MHS muscle fibers and decreases the amplitude of [Ca(2+)]r rise triggered by halothane, but had no effect on [Ca(2+)]r in MHN muscle. In addition, YM-244769 reduced the peak and area under the curve of the Ca(2+) transient elicited by high [K(+)]e and increased its rate of decay in MHS muscle fibers. siRNA knockdown of NCX3 in MHS myotubes reduced [Ca(2+)]r and the Ca(2+) transient area induced by high [K(+)]e. These results demonstrate a functional NCX3 in skeletal muscle whose activity is enhanced in MHS. Moreover reverse mode NCX3 contributes to the Ca(2+) transients associated with K(+)-induced depolarization and the halothane-triggered MH episode in MHS muscle fibers.


Asunto(s)
Calcio/metabolismo , Hipertermia Maligna/metabolismo , Músculo Esquelético/metabolismo , Intercambiador de Sodio-Calcio/metabolismo , Animales , Espacio Extracelular/efectos de los fármacos , Espacio Extracelular/metabolismo , Técnicas de Silenciamiento del Gen , Halotano/farmacología , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Hipertermia Maligna/patología , Potenciales de la Membrana/efectos de los fármacos , Ratones , Fibras Musculares Esqueléticas/efectos de los fármacos , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/patología , Niacinamida/análogos & derivados , Niacinamida/farmacología , Potasio/metabolismo , Sodio/metabolismo , Intercambiador de Sodio-Calcio/genética , Porcinos , Tiourea/análogos & derivados , Tiourea/farmacología
7.
Soins Psychiatr ; (291): 16-20, 2014.
Artículo en Francés | MEDLINE | ID: mdl-24741824

RESUMEN

Although the mode of onset of schizophrenia can be acute, it is important to remember that the disorder rarely starts as a "clap of thunder in a quiet sky", and that it is more often gradual and insidious, with negative and affective symptoms. Acute and transient psychotic disorder, on the other hand, is a short delusional episode forming suddenly and lasting a few days, sometimes a few hours. Schizophrenic evolution forms only part of the possible evolutions. It is therefore necessary to disassociate acute and transient psychotic disorder from schizophrenic disorders, which gives a wrong representation of the onset of schizophrenia.


Asunto(s)
Trastornos Psicóticos/enfermería , Trastornos Psicóticos/psicología , Esquizofrenia/diagnóstico , Esquizofrenia/enfermería , Psicología del Esquizofrénico , Enfermedad Aguda , Diagnóstico Diferencial , Humanos , Síntomas Prodrómicos , Psicopatología , Trastornos Psicóticos/diagnóstico , Trastorno de la Personalidad Esquizotípica/diagnóstico , Trastorno de la Personalidad Esquizotípica/enfermería , Trastorno de la Personalidad Esquizotípica/psicología
8.
Acta Paediatr ; 102(6): e288-90, 2013 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-23647528

RESUMEN

UNLABELLED: We report that daily administration of the diuretic NKCC1 chloride co-transporter, bumetanide, reduces the severity of autism in a 10-year-old Fragile X boy using CARS, ADOS, ABC, RDEG and RRB before and after treatment. In keeping with extensive clinical use of this diuretic, the only side effect was a small hypokalaemia. A double-blind clinical trial is warranted to test the efficacy of bumetanide in FRX. CONCLUSION: This single case report showed an improvement of the scores of each test used after 3 months of treatment. Double-blind clinical trials are warranted to test the efficacy of bumetanide in FRX.


Asunto(s)
Bumetanida/uso terapéutico , Trastornos Generalizados del Desarrollo Infantil/complicaciones , Diuréticos/uso terapéutico , Síndrome del Cromosoma X Frágil/complicaciones , Síndrome del Cromosoma X Frágil/tratamiento farmacológico , Bumetanida/efectos adversos , Niño , Diuréticos/efectos adversos , Humanos , Hipopotasemia/inducido químicamente , Masculino
9.
J Physiol ; 590(23): 6027-36, 2012 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-23006480

RESUMEN

Contraction of skeletal muscle is triggered by the release of Ca(2+) from the sarcoplasmic reticulum (SR) in response to depolarization of the muscle membrane. Depolarization is known to elicit a conformational change of the dihydropyridine receptor (DHPR) in the tubular membrane that controls in a time- and voltage-dependent manner the opening of the ryanodine receptor (RyR), the SR Ca(2+) release channel. At rest, it is assumed that RyRs are kept in a closed state imposed by the repressive action of DHPRs; however, a direct control of the RyR gating by the DHPR has up to now never been demonstrated in resting adult muscle. In this study, we monitored slow changes in SR Ca(2+) content using the Ca(2+) indicator fluo-5N loaded in the SR of voltage-clamped mouse muscle fibres. We first show that external Ca(2+) removal induced a reversible SR Ca(2+) efflux at -80 mV and prevented SR Ca(2+) refilling following depolarization-evoked SR Ca(2+) depletion. The dihydropyridine compound nifedipine induced similar effects. The rate of SR Ca(2+) efflux was also shown to be controlled in a time- and voltage-dependent manner within a membrane potential range more negative than -50 mV. Finally, intracellular addition of ryanodine produced an irreversible SR Ca(2+) efflux and kept the SR in a highly depleted state following depolarization-evoked SR Ca(2+) depletion. The fact that resting SR Ca(2+) efflux is modulated by conformational changes of DHPRs induced by external Ca(2+), nifedipine and voltage demonstrates that DHPRs exert an active control on gating of RyRs in resting skeletal muscle.


Asunto(s)
Canales de Calcio Tipo L/fisiología , Fibras Musculares Esqueléticas/fisiología , Canal Liberador de Calcio Receptor de Rianodina/fisiología , Animales , Calcio/fisiología , Masculino , Potenciales de la Membrana , Ratones , Ratones Endogámicos C57BL , Retículo Sarcoplasmático/fisiología
10.
Soins Psychiatr ; (275): 38-41, 2011.
Artículo en Francés | MEDLINE | ID: mdl-21793375

RESUMEN

Harassment may be either moral, physical or sexual. It is defined as a phenomenon that happens repeatedly. It is underestimated in professional environments and probably even more so in private life. Without referring to a pathological personality, harassers have common pathological traits.


Asunto(s)
Determinación de la Personalidad , Acoso Sexual/legislación & jurisprudencia , Acoso Sexual/psicología , Conducta Social , Lugar de Trabajo , Víctimas de Crimen/psicología , Femenino , Francia , Humanos , Masculino , Factores de Riesgo , Medio Social
11.
Front Aging Neurosci ; 12: 576678, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33281596

RESUMEN

The disease biology of frontotemporal lobe dementia (FTD) is complex and not fully understood, with limited translational value appreciated from animal models to date. Human cellular systems that can recapitulate phenotypic features of disease offer promise as translational tools to not only increase our understanding of disease processes but also increase the probability of success of translating novel treatment options to patients. However not all researchers may necessarily have access to well-characterized induced pluripotent stem cell (iPSC)-derived human neurons. As an example, we therefore comprehensively profiled phenotypic features over time in one commercially-available IPSC-derived human neuron cell line. This included systems-level assessments of neurite outgrowth dynamics, neuronal network function, and genome-wide gene expression. By investigating progranulin biology as an example we then demonstrated the utility of these cells as a tool to investigate human disease biology. For example, by using the siRNA-mediated knockdown of the progranulin (GRN) gene, we demonstrated the establishment of an isogenic human cellular model to facilitate translational FTD research. We reproduced findings from rodent neurons by demonstrating that recombinant progranulin (rPGRN) mediated neuroprotection. Contrary to previous rodent data, in our human cellular models, growth factor treatment showed no consistent sensitivity to modulate neurite outgrowth dynamics. Our study further provides the first evidence that rRPGRN modulated neuronal firing and synchrony in human neurons. Taken together, our datasets are a valuable systems-level resource demonstrating the utility of the tested commercially-available human iPSC neurons for investigating basic human neurobiology, translational neuroscience, and drug discovery applications in neurodegenerative and other CNS diseases.

12.
J Gen Physiol ; 141(5): 557-65, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23630339

RESUMEN

Depolarization of skeletal muscle fibers induces sarcoplasmic reticulum (SR) Ca(2+) release and contraction that progressively decline while depolarization is maintained. Voltage-dependent inactivation of SR Ca(2+) release channels and SR Ca(2+) depletion are the two processes proposed to explain the decline of SR Ca(2+) release during long-lasting depolarizations. However, the relative contribution of these processes, especially under physiological conditions of activation, is not clearly established. Using Fura-2 and Fluo-5N to monitor cytosolic and SR Ca(2+) changes, respectively, in voltage-controlled mouse muscle fibers, we show that 2-min conditioning depolarizations reduce voltage-activated cytosolic Ca(2+) signals with a V1/2 of -53 mV but also induce SR Ca(2+) depletion that decreased the releasable pool of Ca(2+) with the same voltage sensitivity. In contrast, measurement of SR Ca(2+) changes indicated that SR Ca(2+) release channels were inactivated after SR had been depleted and in response to much higher depolarizations with a V1/2 of -13 mV. In response to trains of action potentials, cytosolic Ca(2+) signals decayed with time, whereas SR Ca(2+) changes remained stable over 1-min stimulation, demonstrating that SR Ca(2+) depletion is exclusively responsible for the decline of SR Ca(2+) release under physiological conditions of excitation. These results suggest that previous studies using steady-state inactivation protocols to investigate the voltage dependence of Ca(2+) release inactivation in fact probed the voltage dependence of SR Ca(2+) depletion, and that SR Ca(2+) depletion is the only process that leads to Ca(2+) release decline during continuous stimulation of skeletal muscle.


Asunto(s)
Calcio/metabolismo , Retículo Sarcoplasmático/metabolismo , Potenciales de Acción/fisiología , Animales , Citosol/metabolismo , Masculino , Potenciales de la Membrana/fisiología , Ratones
13.
J Gen Physiol ; 139(3): 209-18, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22371362

RESUMEN

Under resting conditions, external Ca(2+) is known to enter skeletal muscle cells, whereas Ca(2+) stored in the sarcoplasmic reticulum (SR) leaks into the cytosol. The nature of the pathways involved in the sarcolemmal Ca(2+) entry and in the SR Ca(2+) leak is still a matter of debate, but several lines of evidence suggest that these Ca(2+) fluxes are up-regulated in Duchenne muscular dystrophy. We investigated here SR calcium permeation at resting potential and in response to depolarization in voltage-controlled skeletal muscle fibers from control and mdx mice, the mouse model of Duchenne muscular dystrophy. Using the cytosolic Ca(2+) dye Fura2, we first demonstrated that the rate of Ca(2+) increase in response to cyclopiazonic acid (CPA)-induced inhibition of SR Ca(2+)-ATPases at resting potential was significantly higher in mdx fibers, which suggests an elevated SR Ca(2+) leak. However, removal of external Ca(2+) reduced the rate of CPA-induced Ca(2+) increase in mdx and increased it in control fibers, which indicates an up-regulation of sarcolemmal Ca(2+) influx in mdx fibers. Fibers were then loaded with the low-affinity Ca(2+) dye Fluo5N-AM to measure intraluminal SR Ca(2+) changes. Trains of action potentials, chloro-m-cresol, and depolarization pulses evoked transient Fluo5N fluorescence decreases, and recovery of voltage-induced Fluo5N fluorescence changes were inhibited by CPA, demonstrating that Fluo5N actually reports intraluminal SR Ca(2+) changes. Voltage dependence and magnitude of depolarization-induced SR Ca(2+) depletion were found to be unchanged in mdx fibers, but the rate of the recovery phase that followed depletion was found to be faster, indicating a higher SR Ca(2+) reuptake activity in mdx fibers. Overall, CPA-induced SR Ca(2+) leak at -80 mV was found to be significantly higher in mdx fibers and was potentiated by removal of external Ca(2+) in control fibers. The elevated passive SR Ca(2+) leak may contribute to alteration of Ca(2+) homeostasis in mdx muscle.


Asunto(s)
Señalización del Calcio/fisiología , Fibras Musculares Esqueléticas/metabolismo , Retículo Sarcoplasmático/metabolismo , Animales , ATPasas Transportadoras de Calcio/antagonistas & inhibidores , ATPasas Transportadoras de Calcio/metabolismo , Permeabilidad de la Membrana Celular , Citosol/metabolismo , Modelos Animales de Enfermedad , Fenómenos Electrofisiológicos , Colorantes Fluorescentes , Técnicas In Vitro , Masculino , Potenciales de la Membrana , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos mdx , Microscopía Confocal , Modelos Biológicos , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo
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