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1.
Nat Commun ; 14(1): 3103, 2023 05 29.
Artículo en Inglés | MEDLINE | ID: mdl-37248289

RESUMEN

The mechanisms of communication between the brain and the immune cells are still largely unclear. Here, we characterize the populations of resident natural killer (NK) cells and innate lymphoid cells (ILC) 1 in the meningeal dura layer of adult mice. We describe that ILC1/NK cell-derived interferon-γ and acetylcholine can contribute to the modulation of brain homeostatic functions, shaping synaptic neuronal transmission and neurotransmitter levels with effects on mice behavior. In detail, the interferon-γ plays a role in the formation of non-spatial memory, tuning the frequency of GABAergic neurotransmission on cortical pyramidal neurons, while the acetylcholine is a mediator involved in the modulation of brain circuitries that regulate anxiety-like behavior. These findings disclose mechanisms of immune-to-brain communication that modulate brain functions under physiological conditions.


Asunto(s)
Acetilcolina , Interferón gamma , Animales , Ratones , Linfocitos , Inmunidad Innata , Células Asesinas Naturales , Ansiedad
2.
Molecules ; 28(3)2023 Jan 27.
Artículo en Inglés | MEDLINE | ID: mdl-36770914

RESUMEN

α3ß4 nicotinic acetylcholine receptors (nARs) are pentameric ligand-gated cation channels that function in peripheral tissue and in the peripheral and central nervous systems, where they are critical mediators of ganglionic synaptic transmission and modulators of reward-related behaviours. In the pentamer, two α3ß4 subunit couples provide ligand-binding sites, and the fifth single (accessory) subunit (α3 or ß4) regulates receptor trafficking from the endoplasmic reticulum to the cell surface. A number of rare missense variants of the human ß4 subunit have recently been linked to nicotine dependence and/or sporadic amyotrophic lateral sclerosis, and altered responses to nicotine have been reported for these variants; however, it is unknown whether the effects of mutations depend on the subunit within the ligand-binding couples and/or on the fifth subunit. Here, by expressing single populations of pentameric receptors with fixed stoichiometry in cultured cells, we investigated the effect of ß4 variants in the fifth position on the assembly and surface exposure of α3ß4 nAChRs. The results demonstrate that the missense mutations in the accessory subunit alone, despite not affecting the assembly of α3ß4 receptors, alter their trafficking and surface localisation. Thus, altered trafficking of an otherwise functional nAChR may underlie the pathogenic effects of these mutations.


Asunto(s)
Mutación Missense , Receptores Nicotínicos , Humanos , Ligandos , Receptores Nicotínicos/metabolismo , Nicotina/metabolismo , Membrana Celular/metabolismo
3.
Bioengineering (Basel) ; 9(11)2022 Oct 27.
Artículo en Inglés | MEDLINE | ID: mdl-36354532

RESUMEN

Nano- or microdevices, enabling simultaneous, long-term, multisite, cellular recording and stimulation from many excitable cells, are expected to make a strategic turn in basic and applied cardiology (particularly tissue engineering) and neuroscience. We propose an innovative approach aiming to elicit bioelectrical information from the cell membrane using an integrated circuit (IC) bearing a coating of nanowires on the chip surface. Nanowires grow directly on the backend of the ICs, thus allowing on-site amplification of bioelectric signals with uniform and controlled morphology and growth of the NWs on templates. To implement this technology, we evaluated the biocompatibility of silicon and zinc oxide nanowires (NWs), used as a seeding substrate for cells in culture, on two different primary cell lines. Human cardiac stromal cells were used to evaluate the effects of ZnO NWs of different lengths on cell behavior, morphology and growth, while BV-2 microglial-like cells and GH4-C1 neuroendocrine-like cell lines were used to evaluate cell membrane-NW interaction and contact when cultured on Si NWs. As the optimization of the contact between integrated microelectronics circuits and cellular membranes represents a long-standing issue, our technological approach may lay the basis for a new era of devices exploiting the microelectronics' sensitivity and "smartness" to both improve investigation of biological systems and to develop suitable NW-based systems available for tissue engineering and regenerative medicine.

4.
Exp Neurol ; 350: 113969, 2022 04.
Artículo en Inglés | MEDLINE | ID: mdl-34973962

RESUMEN

Gradual decline in cognitive and non-cognitive functions are considered clinical hallmarks of Alzheimer's Disease (AD). Post-mortem autoptic analysis shows the presence of amyloid ß deposits, neuroinflammation and severe brain atrophy. However, brain circuit alterations and cellular derailments, assessed in very early stages of AD, still remain elusive. The understanding of these early alterations is crucial to tackle defective mechanisms. In a previous study we proved that the Tg2576 mouse model of AD displays functional deficits in the dorsal hippocampus and relevant behavioural AD-related alterations. We had shown that these deficits in Tg2576 mice correlate with the precocious degeneration of dopamine (DA) neurons in the Ventral Tegmental Area (VTA) and can be restored by L-DOPA treatment. Due to the distinct functionality and connectivity of dorsal versus ventral hippocampus, here we investigated neuronal excitability and synaptic functionality in the ventral CA1 hippocampal sub-region of Tg2576 mice. We found an age-dependent alteration of cell excitability and firing in pyramidal neurons starting at 3 months of age, that correlates with reduced levels in the ventral CA1 of tyrosine hydroxylase - the rate-limiting enzyme of DA synthesis. Additionally, at odds with the dorsal hippocampus, we found no alterations in basal glutamatergic transmission and long-term plasticity of ventral neurons in 8-month old Tg2576 mice compared to age-matched controls. Last, we used computational analysis to model the early derailments of firing properties observed and hypothesize that the neuronal alterations found could depend on dysfunctional sodium and potassium conductances, leading to anticipated depolarization-block of action potential firing. The present study depicts that impairment of cell excitability and homeostatic control of firing in ventral CA1 pyramidal neurons is a prodromal feature in Tg2576 AD mice.


Asunto(s)
Enfermedad de Alzheimer/fisiopatología , Región CA1 Hipocampal/fisiopatología , Fenómenos Electrofisiológicos , Células Piramidales , Potenciales de Acción , Envejecimiento , Animales , Dopaminérgicos/farmacología , Neuronas Dopaminérgicas , Femenino , Levodopa/farmacología , Masculino , Ratones , Ratones Transgénicos , Canales de Potasio , Canales de Sodio , Tirosina 3-Monooxigenasa/metabolismo , Área Tegmental Ventral/fisiopatología
5.
Glia ; 70(1): 173-195, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34661306

RESUMEN

Microglia cells are active players in regulating synaptic development and plasticity in the brain. However, how they influence the normal functioning of synapses is largely unknown. In this study, we characterized the effects of pharmacological microglia depletion, achieved by administration of PLX5622, on hippocampal CA3-CA1 synapses of adult wild type mice. Following microglial depletion, we observed a reduction of spontaneous and evoked glutamatergic activity associated with a decrease of dendritic spine density. We also observed the appearance of immature synaptic features and higher levels of plasticity. Microglia depleted mice showed a deficit in the acquisition of the Novel Object Recognition task. These events were accompanied by hippocampal astrogliosis, although in the absence ofneuroinflammatory condition. PLX-induced synaptic changes were absent in Cx3cr1-/- mice, highlighting the role of CX3CL1/CX3CR1 axis in microglia control of synaptic functioning. Remarkably, microglia repopulation after PLX5622 withdrawal was associated with the recovery of hippocampal synapses and learning functions. Altogether, these data demonstrate that microglia contribute to normal synaptic functioning in the adult brain and that their removal induces reversible changes in organization and activity of glutamatergic synapses.


Asunto(s)
Microglía , Neuronas , Animales , Encéfalo , Fármacos actuantes sobre Aminoácidos Excitadores/farmacología , Hipocampo , Ratones , Compuestos Orgánicos/farmacología , Sinapsis/fisiología
6.
Pharmacol Res ; 175: 105959, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34756924

RESUMEN

Glioblastomas (GBMs), the most frequent brain tumours, are highly invasive and their prognosis is still poor despite the use of combination treatment. MG624 is a 4-oxystilbene derivative that is active on α7- and α9-containing neuronal nicotinic acetylcholine receptor (nAChR) subtypes. Hybridisation of MG624 with a non-nicotinic resveratrol-derived pro-oxidant mitocan has led to two novel compounds (StN-4 and StN-8) that are more potent than MG624 in reducing the viability of GBM cells, but less potent in reducing the viability of mouse astrocytes. Functional analysis of their activity on α7 receptors showed that StN-4 is a silent agonist, whereas StN-8 is a full antagonist, and neither alters intracellular [Ca2+] levels when acutely applied to U87MG cells. After 72 h of exposure, both compounds decreased U87MG cell proliferation, and pAKT and oxphos ATP levels, but only StN-4 led to a significant accumulation of cells in phase G1/G0 and increased apoptosis. One hour of exposure to either compound also decreased the mitochondrial and cytoplasmic ATP production of U87MG cells, and this was not paralleled by any increase in the production of reactive oxygen species. Knocking down the α9 subunit (which is expressed at relatively high levels in U87MG cells) decreased the potency of the effects of both compounds on cell viability, but cell proliferation, ATP production, pAKT levels were unaffected by the presence of the noncell-permeable α7/α9-selective antagonist αBungarotoxin. These last findings suggest that the anti-tumoral effects of StN-4 and StN-8 on GBM cells are not only due to their action on nAChRs, but also to other non-nicotinic mechanisms.


Asunto(s)
Compuestos de Amonio/farmacología , Antineoplásicos/farmacología , Neoplasias Encefálicas/tratamiento farmacológico , Glioblastoma/tratamiento farmacológico , Estilbenos/farmacología , Adenosina Trifosfato/metabolismo , Animales , Astrocitos/efectos de los fármacos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Línea Celular Tumoral , Fenómenos Fisiológicos Celulares/efectos de los fármacos , Glioblastoma/genética , Glioblastoma/metabolismo , Humanos , Ligandos , Ratones , Proteínas Proto-Oncogénicas c-akt/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Receptores Nicotínicos/genética , Receptor Nicotínico de Acetilcolina alfa 7/genética
7.
Neurobiol Stress ; 14: 100324, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33937445

RESUMEN

Exposure to aversive events during sensitive developmental periods can affect the preferential coping strategy adopted by individuals later in life, leading to either stress-related psychiatric disorders, including depression, or to well-adaptation to future adversity and sources of stress, a behavior phenotype termed "resilience". We have previously shown that interfering with the development of mother-pups bond with the Repeated Cross Fostering (RCF) stress protocol can induce resilience to depression-like phenotype in adult C57BL/6J female mice. Here, we used patch-clamp recording in midbrain slice combined with both in vivo and ex vivo pharmacology to test our hypothesis of a link between electrophysiological modifications of dopaminergic neurons in the intermediate Ventral Tegmental Area (VTA) of RCF animals and behavioral resilience. We found reduced hyperpolarization-activated (Ih) cation current amplitude and evoked firing in VTA dopaminergic neurons from both young and adult RCF female mice. In vivo, VTA-specific pharmacological manipulation of the Ih current reverted the pro-resilient phenotype in adult early-stressed mice or mimicked behavioral resilience in adult control animals. This is the first evidence showing how pro-resilience behavior induced by early events is linked to a long-lasting reduction of Ih current and excitability in VTA dopaminergic neurons.

8.
J Med Chem ; 63(24): 15668-15692, 2020 12 24.
Artículo en Inglés | MEDLINE | ID: mdl-33325696

RESUMEN

A series of diastereomeric 2-(2-pyrrolidinyl)-1,4-benzodioxanes bearing a small, hydrogen-bonding substituent at the 7-, 6-, or 5-position of benzodioxane have been studied for α4ß2 and α3ß4 nicotinic acetylcholine receptor affinity and activity. Analogous to C(5)H replacement with N and to a much greater extent than decoration at C(7), substitution at benzodioxane C(5) confers very high α4ß2/α3ß4 selectivity to the α4ß2 partial agonism. Docking into the two receptor structures recently determined by cryo-electron microscopy and site-directed mutagenesis at the minus ß2 side converge in indicating that the limited accommodation capacity of the ß2 pocket, compared to that of the ß4 pocket, makes substitution at C(5) rather than at more projecting C(7) position determinant for this pursued subtype selectivity.


Asunto(s)
Dioxanos/química , Agonistas Nicotínicos/química , Receptores Nicotínicos/química , Sitios de Unión , Microscopía por Crioelectrón , Dioxanos/síntesis química , Dioxanos/metabolismo , Humanos , Enlace de Hidrógeno , Simulación del Acoplamiento Molecular , Mutagénesis Sitio-Dirigida , Agonistas Nicotínicos/síntesis química , Agonistas Nicotínicos/metabolismo , Antagonistas Nicotínicos/química , Antagonistas Nicotínicos/metabolismo , Pirrolidinas/química , Receptores Nicotínicos/genética , Receptores Nicotínicos/metabolismo , Estereoisomerismo , Relación Estructura-Actividad
9.
Environ Sci Pollut Res Int ; 25(36): 35866-35873, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-29532382

RESUMEN

There is a growing market demand for small-scale biomass gasifiers that is driven by the economic incentives and the legislative framework. Small-scale gasifiers produce a gaseous fuel, commonly referred to as producer gas, with relatively low heating value. Thus, the most common energy conversion systems that are coupled with small-scale gasifiers are internal combustion engines. In order to increase the electrical efficiency, the operators choose dual fuel engines and mix the producer gas with diesel. The Wiebe function has been a valuable tool for assessing the efficiency of dual fuel internal combustion engines. This study introduces a thermodynamic model that works in parallel with the Wiebe function and calculates the emissions of the engines. This "vis-à-vis" approach takes into consideration the actual conditions inside the cylinders-as they are returned by the Wiebe function-and calculates the final thermodynamic equilibrium of the flue gases mixture. This approach aims to enhance the operation of the dual fuel internal combustion engines by identifying the optimal operating conditions and-at the same time-advance pollution control and minimize the environmental impact.


Asunto(s)
Biocombustibles/análisis , Fuentes Generadoras de Energía , Gases/análisis , Modelos Teóricos , Madera/química , Biomasa , Termodinámica
10.
PLoS One ; 11(8): e0160950, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27525414

RESUMEN

Noise can enhance perception of tactile and proprioceptive stimuli by stochastic resonance processes. However, the mechanisms underlying this general phenomenon remain to be characterized. Here we studied how externally applied noise influences action potential firing in mouse primary sensory neurons of dorsal root ganglia, modelling a basic process in sensory perception. Since noisy mechanical stimuli may cause stochastic fluctuations in receptor potential, we examined the effects of sub-threshold depolarizing current steps with superimposed random fluctuations. We performed whole cell patch clamp recordings in cultured neurons of mouse dorsal root ganglia. Noise was added either before and during the step, or during the depolarizing step only, to focus onto the specific effects of external noise on action potential generation. In both cases, step + noise stimuli triggered significantly more action potentials than steps alone. The normalized power norm had a clear peak at intermediate noise levels, demonstrating that the phenomenon is driven by stochastic resonance. Spikes evoked in step + noise trials occur earlier and show faster rise time as compared to the occasional ones elicited by steps alone. These data suggest that external noise enhances, via stochastic resonance, the recruitment of transient voltage-gated Na channels, responsible for action potential firing in response to rapid step-wise depolarizing currents.


Asunto(s)
Potenciales de Acción , Ruido , Células Receptoras Sensoriales/citología , Animales , Femenino , Humanos , Masculino , Ratones , Procesos Estocásticos
11.
J Neurosci ; 32(29): 9796-804, 2012 Jul 18.
Artículo en Inglés | MEDLINE | ID: mdl-22815494

RESUMEN

Ionotropic glutamate receptors, which underlie a majority of excitatory synaptic transmission in the CNS, associate with transmembrane proteins that modify their intracellular trafficking and channel gating. Significant advances have been made in our understanding of AMPA-type glutamate receptor (AMPAR) regulation by transmembrane AMPAR regulatory proteins. Less is known about the functional influence of cornichons-unrelated AMPAR-interacting proteins, identified by proteomic analysis. Here we confirm that cornichon homologs 2 and 3 (CNIH-2 and CNIH-3), but not CNIH-1, slow the deactivation and desensitization of both GluA2-containing calcium-impermeable and GluA2-lacking calcium-permeable (CP) AMPARs expressed in tsA201 cells. CNIH-2 and -3 also enhanced the glutamate sensitivity, single-channel conductance, and calcium permeability of CP-AMPARs while decreasing their block by intracellular polyamines. We examined the potential effects of CNIHs on native AMPARs by recording from rat optic nerve oligodendrocyte precursor cells (OPCs), known to express a significant population of CP-AMPARs. These glial cells exhibited surface labeling with an anti-CNIH-2/3 antibody. Two features of their AMPAR-mediated currents-the relative efficacy of the partial agonist kainate (I(KA)/I(Glu) ratio 0.4) and a greater than fivefold potentiation of kainate responses by cyclothiazide-suggest AMPAR association with CNIHs. Additionally, overexpression of CNIH-3 in OPCs markedly slowed AMPAR desensitization. Together, our experiments support the view that CNIHs are capable of altering key properties of AMPARs and suggest that they may do so in glia.


Asunto(s)
Proteínas del Huevo/metabolismo , Proteínas de la Membrana/metabolismo , Neuroglía/metabolismo , Neuronas/metabolismo , Receptores AMPA/metabolismo , Animales , Calcio/metabolismo , Línea Celular , Células Cultivadas , Proteínas del Huevo/genética , Agonistas de Aminoácidos Excitadores/farmacología , Femenino , Ácido Glutámico/farmacología , Humanos , Activación del Canal Iónico/efectos de los fármacos , Activación del Canal Iónico/genética , Ácido Kaínico/farmacología , Masculino , Proteínas de la Membrana/genética , Neuroglía/citología , Neuroglía/efectos de los fármacos , Neuronas/citología , Neuronas/efectos de los fármacos , Nervio Óptico/citología , Nervio Óptico/efectos de los fármacos , Nervio Óptico/metabolismo , Ratas , Receptores AMPA/genética , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/genética , Transfección
12.
J Neurosci ; 32(17): 5853-67, 2012 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-22539847

RESUMEN

The kinetics of IPSCs influence many neuronal processes, such as the frequencies of oscillations and the duration of shunting inhibition. The subunit composition of recombinant GABA(A) receptors (GABA(A)Rs) strongly affects the deactivation kinetics of GABA-evoked currents. However, for GABAergic synapses, the relationship between subunit composition and IPSC decay is less clear. Here we addressed this by combining whole-cell recordings of miniature IPSCs (mIPSCs) and quantitative immunolocalization of synaptic GABA(A)R subunits. In cerebellar stellate, thalamic relay, and main olfactory bulb (MOB) deep short-axon cells of Wistar rats, the only synaptic α subunit was α1, and zolpidem-sensitive mIPSCs had weighted decay time constants (τ(w)) of 4-6 ms. Nucleus reticularis thalami neurons expressed only α3 as the synaptic α subunit and exhibited slow (τ(w) = 28 ms), zolpidem-insensitive mIPSCs. By contrast, MOB external tufted cells contained two α subunit types (α1 and α3) at their synapses. Quantitative analysis of multiple immunolabeled images revealed small within-cell, but large between-cell, variability in synaptic α1/α3 ratios. This corresponded to large cell-to-cell variability in the decay (τ(w) = 3-30 ms) and zolpidem sensitivity of mIPSCs. Currents evoked by rapid application of GABA to patches excised from HEK cells expressing different mixtures of α1 and α3 subunits displayed highly variable deactivation times that correlated with the α1/α3 cDNA ratio. Our results demonstrate that diversity in the decay of IPSCs can be generated by varying the expression of different GABA(A)R subunits that alone confer different decay kinetics, allowing the time course of inhibition to be tuned to individual cellular requirements.


Asunto(s)
Potenciales Postsinápticos Inhibidores/fisiología , Inhibición Neural/fisiología , Neuronas/fisiología , Receptores de GABA-A/metabolismo , Sinapsis/fisiología , Animales , Animales Recién Nacidos , Proteínas Portadoras/metabolismo , Cerebelo/citología , Cerebelo/metabolismo , Relación Dosis-Respuesta a Droga , Estimulación Eléctrica , GABAérgicos/farmacología , Expresión Génica/fisiología , Humanos , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Masculino , Proteínas de la Membrana/metabolismo , Inhibición Neural/efectos de los fármacos , Vías Nerviosas/fisiología , Bulbo Olfatorio/citología , Bulbo Olfatorio/metabolismo , Técnicas de Placa-Clamp , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratas , Ratas Wistar , Receptores de GABA-A/genética , Sinapsis/efectos de los fármacos , Tálamo/citología , Tálamo/metabolismo , Factores de Tiempo , Transfección , Ácido gamma-Aminobutírico/farmacología
13.
Nat Neurosci ; 14(11): 1430-8, 2011 Oct 09.
Artículo en Inglés | MEDLINE | ID: mdl-21983683

RESUMEN

Oligodendrocyte precursor cells (OPCs), a major glial cell type that gives rise to myelinating oligodendrocytes in the CNS, express calcium-permeable AMPA receptors (CP-AMPARs). Although CP-AMPARs are important for OPC proliferation and neuron-glia signaling, they render OPCs susceptible to ischemic damage in early development. We identified factors controlling the dynamic regulation of AMPAR subtypes in OPCs from rat optic nerve and mouse cerebellar cortex. We found that activation of group 1 mGluRs drove an increase in the proportion of CP-AMPARs, reflected by an increase in single-channel conductance and inward rectification. This plasticity required the elevation of intracellular calcium and used PI3K, PICK-1 and the JNK pathway. In white matter, neurons and astrocytes release both ATP and glutamate. Unexpectedly, activation of purinergic receptors in OPCs decreased CP-AMPAR expression, suggesting a capacity for homeostatic regulation. Finally, we found that stargazin-related transmembrane AMPAR regulatory proteins, which are critical for AMPAR surface expression in neurons, regulate CP-AMPAR plasticity in OPCs.


Asunto(s)
Calcio/metabolismo , Activación del Canal Iónico/fisiología , Plasticidad Neuronal/fisiología , Oligodendroglía/fisiología , Receptores AMPA/metabolismo , Animales , Animales Recién Nacidos , Antígenos/metabolismo , Fenómenos Biofísicos/efectos de los fármacos , Fenómenos Biofísicos/genética , Fenómenos Biofísicos/fisiología , Canales de Calcio/metabolismo , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Linaje de la Célula , Cerebelo/citología , Inhibidores Enzimáticos/farmacología , Fármacos actuantes sobre Aminoácidos Excitadores/farmacología , Femenino , Galactosilceramidasa/metabolismo , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/fisiología , Ácido Glutámico/farmacología , Glicinérgicos/farmacología , Técnicas In Vitro , Activación del Canal Iónico/efectos de los fármacos , Proteínas Luminiscentes/genética , Masculino , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/genética , Metoxihidroxifenilglicol/análogos & derivados , Metoxihidroxifenilglicol/farmacología , Ratones , Ratones Transgénicos , Mutación/genética , Plasticidad Neuronal/genética , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Oligodendroglía/citología , Nervio Óptico/citología , Proteoglicanos/metabolismo , Ratas , Receptores AMPA/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Células Madre/metabolismo , Estricnina/farmacología , Tetrodotoxina/farmacología
14.
J Neurosci ; 31(2): 753-63, 2011 Jan 12.
Artículo en Inglés | MEDLINE | ID: mdl-21228184

RESUMEN

High-affinity extrasynaptic GABA(A) receptors (GABA(A)Rs) are a prominent feature of cerebellar granule neurons and thalamic relay neurons. In both cell types, the presence of synaptic glomeruli would be expected to promote activation of these GABA(A)Rs, contributing to phasic spillover-mediated currents and tonic inhibition. However, the precise role of different receptor subtypes in these two phenomena is unclear. To address this question, we made recordings from neurons in acute brain slices from mice, and from tsA201 cells expressing recombinant GABA(A)Rs. We found that δ subunit-containing GABA(A)Rs of both cerebellar granule neurons and thalamic relay neurons of the lateral geniculate nucleus contributed to tonic conductance caused by ambient GABA but not to spillover-mediated currents. In the presence of a low "ambient" GABA concentration, recombinant "extrasynaptic" δ subunit-containing GABA(A)Rs exhibited profound desensitization, rendering them insensitive to brief synaptic- or spillover-like GABA transients. Together, our results demonstrate that phasic spillover and tonic inhibition reflect the activation of distinct receptor populations.


Asunto(s)
Receptores de GABA-A/fisiología , Ácido gamma-Aminobutírico/fisiología , Animales , Línea Celular , Cerebelo/citología , Cerebelo/fisiología , Humanos , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/fisiología , Técnicas de Placa-Clamp , Subunidades de Proteína/fisiología , Ratas , Receptores de GABA-A/genética , Sinapsis/fisiología , Tálamo/citología , Tálamo/fisiología , Transfección
15.
Nat Neurosci ; 12(8): 1042-9, 2009 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-19578381

RESUMEN

Although feedforward inhibition onto Purkinje cells was first documented 40 years ago, we understand little of how inhibitory interneurons contribute to cerebellar function in behaving animals. Using a mouse line (PC-Deltagamma2) in which GABA(A) receptor-mediated synaptic inhibition is selectively removed from Purkinje cells, we examined how feedforward inhibition from molecular layer interneurons regulates adaptation of the vestibulo-ocular reflex. Although impairment of baseline motor performance was relatively mild, the ability to adapt the phase of the vestibulo-ocular reflex and to consolidate gain adaptations was strongly compromised. Purkinje cells showed abnormal patterns of simple spikes, both during and in the absence of evoked compensatory eye movements. On the basis of modeling our experimental data, we propose that feedforward inhibition, by controlling the fine-scale patterns of Purkinje cell activity, enables the induction of plasticity in neurons of the cerebellar and vestibular nuclei.


Asunto(s)
Corteza Cerebelosa/fisiología , Aprendizaje/fisiología , Inhibición Neural/fisiología , Plasticidad Neuronal/fisiología , Células de Purkinje/fisiología , Núcleos Vestibulares/fisiología , Potenciales de Acción/fisiología , Adaptación Fisiológica/fisiología , Animales , Corteza Cerebelosa/citología , Interneuronas/citología , Interneuronas/fisiología , Ratones , Ratones Noqueados , Técnicas de Cultivo de Órganos , Células de Purkinje/citología , Receptores de GABA-A/genética , Reflejo Vestibuloocular/fisiología , Ácido gamma-Aminobutírico/metabolismo
16.
Nat Neurosci ; 12(3): 277-85, 2009 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-19234459

RESUMEN

Although the properties and trafficking of AMPA-type glutamate receptors (AMPARs) depend critically on associated transmembrane AMPAR regulatory proteins (TARPs) such as stargazin (gamma-2), no TARP has been described that can specifically regulate the important class of calcium-permeable (CP-) AMPARs. We examined the stargazin-related protein gamma-5, which is highly expressed in Bergmann glia, a cell type possessing only CP-AMPARs. gamma-5 was previously thought not to be a TARP, and it has been widely used as a negative control. Here we find that, contrary to expectation, gamma-5 acts as a TARP and serves this role in Bergmann glia. Whereas gamma-5 interacts with all AMPAR subunits, and modifies their behavior to varying extents, its main effect is to regulate the function of AMPAR subunit combinations that lack short-form subunits, which constitute predominantly CP-AMPARs. Our results suggest an important role for gamma-5 in regulating the functional contribution of CP-AMPARs.


Asunto(s)
Señalización del Calcio/fisiología , Calcio/metabolismo , Permeabilidad de la Membrana Celular/fisiología , Receptores AMPA/clasificación , Receptores AMPA/fisiología , Animales , Canales de Calcio/genética , Canales de Calcio/fisiología , Señalización del Calcio/genética , Línea Celular , Permeabilidad de la Membrana Celular/genética , Humanos , Neuroglía/química , Neuroglía/metabolismo , Neuroglía/fisiología , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiología , Transporte de Proteínas/genética , Ratas , Receptores AMPA/genética , Receptores AMPA/metabolismo
17.
J Physiol ; 585(Pt 1): 91-101, 2007 Nov 15.
Artículo en Inglés | MEDLINE | ID: mdl-17901118

RESUMEN

Among principal neurons, adult Purkinje cells have long been considered unusual in lacking functional NMDA receptors. This view has emerged largely from studies on rats, where NMDA receptors are expressed in Purkinje cells of newborn animals, but are lost after 2 weeks. By contrast, immunolabelling data have shown that Purkinje cells from adult mice express multiple NMDA receptor subunits, suggesting a possible species difference. To investigate the presence of functional NMDA receptors in Purkinje cells of mice, and to explore the contribution of different receptor subunits, we made whole-cell and single-channel patch-clamp recordings from Purkinje cells of wild-type and NR2D-/- mice of different ages. Here we report that multiple NMDA receptor subtypes are indeed expressed in Purkinje cells of young and adult mice; in the adult, both NR2A- and NR2B-containing subtypes are present. Furthermore, we show that NMDA receptor-mediated EPSCs can be evoked by climbing fibre stimulation, and appear to be mediated mainly by NR2A-containing receptors.


Asunto(s)
Fibras Nerviosas/fisiología , Células de Purkinje/fisiología , Receptores de N-Metil-D-Aspartato/fisiología , Envejecimiento/fisiología , Animales , Potenciales Evocados/fisiología , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Técnicas de Placa-Clamp , Receptores de N-Metil-D-Aspartato/genética , Transmisión Sináptica/fisiología
18.
Nat Neurosci ; 10(7): 923-9, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17572671

RESUMEN

In mammals, identifying the contribution of specific neurons or networks to behavior is a key challenge. Here we describe an approach that facilitates this process by enabling the rapid modulation of synaptic inhibition in defined cell populations. Binding of zolpidem, a systemically active allosteric modulator that enhances the function of the GABAA receptor, requires a phenylalanine residue (Phe77) in the gamma2 subunit. Mice in which this residue is changed to isoleucine are insensitive to zolpidem. By Cre recombinase-induced swapping of the gamma2 subunit (that is, exchanging Ile77 for Phe77), zolpidem sensitivity can be restored to GABAA receptors in chosen cell types. We demonstrate the power of this method in the cerebellum, where zolpidem rapidly induces significant motor deficits when Purkinje cells are made uniquely sensitive to its action. This combined molecular and pharmacological technique has demonstrable advantages over targeted cell ablation and will be invaluable for investigating many neuronal circuits.


Asunto(s)
Conducta Animal/efectos de los fármacos , Neuronas/fisiología , Receptores de GABA-A/fisiología , Sinapsis/fisiología , Animales , Autorradiografía , Electrofisiología , Femenino , Agonistas del GABA/farmacología , Genotipo , Proteínas Fluorescentes Verdes/genética , Inmunohistoquímica , Hibridación in Situ , Masculino , Ratones , Ratones Noqueados , Neuronas/efectos de los fármacos , Técnicas de Placa-Clamp , Equilibrio Postural/efectos de los fármacos , Equilibrio Postural/fisiología , Ingeniería de Proteínas , Células de Purkinje/efectos de los fármacos , Células de Purkinje/fisiología , Piridinas/farmacología , Receptores de GABA-A/genética , Receptores de GABA-B/genética , Receptores de GABA-B/fisiología , Sinapsis/efectos de los fármacos , Zolpidem
19.
Ig Sanita Pubbl ; 63(2): 113-25, 2007.
Artículo en Italiano | MEDLINE | ID: mdl-18223698

RESUMEN

INTRODUCTION: The Italian Heat Health Watch Warning System (HHWWS) was set up following the over 3000 excess deaths which occurred in Italy during the heat wave of 2003, In June 2005 the warning system issued a heat warning in various Italian cities. METHODS: A case control study was performed in one of these cities (Bari) in order to identify individual and environmental risk factors as well as preventive strategies for reducing mortality during future heat waves. Cases were defined as subjects aged <65 years who had died during the heat wave and whose death certificate listed one of the following as the cause of death: heat, cardiovascular or cerebrovascular disorders, neurocognitive disorders, dehydration or fever/infection not otherwise specified. For each case, three age-matched controls were randomly selected among individuals followed by the same general practitioner as the case. All variables significantly associated with mortality (<0.1) in the univariate analysis were entered into a conditional logistic regression model and the population attributable fraction (PAF) was calculated for significant variables (at p<0.05). RESULTS: Twenty cases and sixty controls were included in the study. In 17 cases (89%) death had occurred at home and 11(55%) of these were cardiovascular- related deaths. At the multivariate analysis, the factors significantly associated with mortality risk during the heat wave were: having a functioning air conditioner at home [OR:0.09(95% CI 0.01-1.00)], having an Activities of Daily Living score <2 [OR:21.0(95%CI 1.81-242.47)] and having been hospitalized the year preceding death [OR:18.1(95%CI 2.04-160.51)]. CONCLUSIONS: Public health interventions during heat waves should include the provision of access to an air conditioned environment. Subjects with impaired health (especially if recently hospitalized) and with significant limitations in their activities of daily living are probably at higher risk during heat waves.


Asunto(s)
Actividades Cotidianas , Calor , Estudios de Casos y Controles , Humanos , Italia/epidemiología , Factores de Riesgo
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