RESUMO
BACKGROUND: In advanced stages of Parkinson's disease (PD), dyskinesia and motor fluctuations become seriously debilitating and therapeutic options become scarce. Aberrant activity of striatal cholinergic interneurons (SCIN) has been shown to be critical to PD and dyskinesia, but the systemic administration of cholinergic medications can exacerbate extrastriatal-related symptoms. Thus, targeting the mechanisms causing pathological SCIN activity in severe PD with motor fluctuations and dyskinesia is a promising therapeutic alternative. METHODS: We used ex vivo electrophysiological recordings combined with pharmacology to study the alterations in intracellular signaling that contribute to the altered SCIN physiology observed in the 6-hydroxydopamine mouse model of PD treated with levodopa. RESULTS: The altered phenotypes of SCIN of parkinsonian mice during the "off levodopa" state resulting from aberrant Kir/leak and Kv1.3 currents can be rapidly reverted by acute inhibition of cAMP-ERK1/2 signaling. Inverse agonists that inhibit the ligand-independent activity of D5 receptors, like clozapine, restore Kv1.3 and Kir/leak currents and SCIN normal physiology in dyskinetic mice. CONCLUSION: Our work unravels a signaling pathway involved in the dysregulation of membrane currents causing SCIN hyperexcitability and burst-pause activity in parkinsonian mice treated with levodopa (l-dopa). These changes persist during off-medication periods due to tonic mechanisms that can be acutely reversed by pharmacological interventions. Thus, targeting the D5-cAMP-ERK1/2 signaling pathway selectively in SCIN may have therapeutic effects in PD and dyskinesia by restoring the normal SCIN function. © 2022 International Parkinson and Movement Disorder Society.
Assuntos
Discinesia Induzida por Medicamentos , Doença de Parkinson , Animais , Antiparkinsonianos/farmacologia , Antiparkinsonianos/uso terapêutico , Colinérgicos/metabolismo , Colinérgicos/farmacologia , Colinérgicos/uso terapêutico , Corpo Estriado/metabolismo , Modelos Animais de Doenças , Discinesia Induzida por Medicamentos/patologia , Interneurônios/metabolismo , Levodopa/farmacologia , Levodopa/uso terapêutico , Camundongos , Oxidopamina/toxicidadeRESUMO
BACKGROUND: Self-limited Childhood Epilepsies are the most prevalent epileptic syndrome in children. Its pathogenesis is unknown. In this disease, symptoms resolve spontaneously in approximately 50% of patients when maturity is reached, prompting to a maturation problem. The purpose of this study was to understand the molecular bases of this disease by generating and analyzing induced pluripotent stem cell-derived neurons from a family with 7 siblings, among whom 4 suffer from this disease. METHODS: Two affected siblings and, as controls, a healthy sister and the unaffected mother of the family were studied. Using exome sequencing, a homozygous variant in the FYVE, RhoGEF and PH Domain Containing 6 gene was identified in the patients as a putative genetic factor that could contribute to the development of this familial disorder. After informed consent was signed, skin biopsies from the 4 individuals were collected, fibroblasts were derived and reprogrammed and neurons were generated and characterized by markers and electrophysiology. Morphological, electrophysiological and gene expression analyses were performed on these neurons. RESULTS: Bona fide induced pluripotent stem cells and derived neurons could be generated in all cases. Overall, there were no major shifts in neuronal marker expression among patient and control-derived neurons. Compared to two familial controls, neurons from patients showed shorter axonal length, a dramatic reduction in synapsin-1 levels and cytoskeleton disorganization. In addition, neurons from patients developed a lower action potential threshold with time of in vitro differentiation and the amount of current needed to elicit an action potential (rheobase) was smaller in cells recorded from NE derived from patients at 12 weeks of differentiation when compared with shorter times in culture. These results indicate an increased excitability in patient cells that emerges with the time in culture. Finally, functional genomic analysis showed a biased towards immaturity in patient-derived neurons. CONCLUSIONS: We are reporting the first in vitro model of self-limited childhood epilepsy, providing the cellular bases for future in-depth studies to understand its pathogenesis. Our results show patient-specific neuronal features reflecting immaturity, in resonance with the course of the disease and previous imaging studies.
Assuntos
Epilepsia , Células-Tronco Pluripotentes Induzidas , Potenciais de Ação/fisiologia , Diferenciação Celular/genética , Criança , Epilepsia/genética , Epilepsia/metabolismo , Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Neurônios/metabolismoRESUMO
BACKGROUND: Enhanced striatal cholinergic interneuron activity contributes to the striatal hypercholinergic state in Parkinson's disease (PD) and to levodopa-induced dyskinesia. In severe PD, dyskinesia and motor fluctuations become seriously debilitating, and the therapeutic strategies become scarce. Given that the systemic administration of anticholinergics can exacerbate extrastriatal-related symptoms, targeting cholinergic interneurons is a promising therapeutic alternative. Therefore, unraveling the mechanisms causing pathological cholinergic interneuron activity in severe PD with motor fluctuations and dyskinesia may provide new molecular therapeutic targets. METHODS: We used ex vivo electrophysiological recordings combined with pharmacological and morphological studies to investigate the intrinsic alterations of cholinergic interneurons in the 6-hydroxydopamine mouse model of PD treated with levodopa. RESULTS: Cholinergic interneurons exhibit pathological burst-pause activity in the parkinsonian "off levodopa" state. This is mediated by a persistent ligand-independent activity of dopamine D1/D5 receptor signaling, involving a cyclic adenosine monophosphate (cAMP) pathway. Dysregulation of membrane ion channels that results in increased inward-rectifier potassium type 2 (Kir2) and decreased leak currents causes the burst pause activity, which can be dampened by pharmacological inhibition of intracellular cAMP. A single challenge with a dyskinetogenic dose of levodopa is sufficient to induce persistent cholinergic interneuron burst-pause firing. CONCLUSION: Our data unravel a mechanism causing aberrant cholinergic interneuron burst-pause activity in parkinsonian mice treated with levodopa. Targeting D5-cAMP signaling and the regulation of Kir2 and leak channels may alleviate parkinsonism and dyskinesia by restoring normal cholinergic interneuron function. © 2021 International Parkinson and Movement Disorder Society.
Assuntos
Corpo Estriado , Levodopa , Animais , Colinérgicos/farmacologia , Interneurônios , Levodopa/farmacologia , Camundongos , Oxidopamina/toxicidadeRESUMO
A variety of glial cell functions are supported by connexin and pannexin proteins. These functions include the modulation of synaptic gain, the control of excitability through regulation of the ion and neurotransmitter composition of the extracellular milieu and the promotion of neuronal survival. Connexins and pannexins support these functions through diverse molecular mechanisms, including channel and non-channel functions. The former comprise the formation of gap junction-mediated networks supported by connexin intercellular channels and the formation of pore-like membrane structures or hemichannels formed by both connexins and pannexins. Non-channel functions involve adhesion properties and the participation in signaling intracellular cascades. Pathological conditions of the nervous system such as ischemia, neurodegeneration, pathogen infection, trauma and tumors are characterized by distinctive remodeling of connexin expression and function. However, whether these changes can be interpreted as part of the pathogenesis, or as beneficial compensatory effects, remains under debate. Here we review the available evidence addressing this matter with a special emphasis in mouse models with selective manipulation of glial connexin and pannexin proteins in vivo. We postulate that the beneficial vs. detrimental effects of glial connexin remodeling in pathological conditions depend on the impact of remodeling on the different connexin and pannexin channel and non-channel functions, on the characteristics of the inflammatory environment and on the type of interaction among glial cells types.
RESUMO
Many functions of glial cells depend on the formation of selective glial networks mediated by gap junctions formed by members of the connexin family. Olfactory ensheathing cells (OECs) are specialized glia associated with olfactory sensory neuron axons. Like other glia, they form selective networks, however, the connexins that support OEC connectivity in vivo have not been identified. We used an in vivo mouse model to selectively delete candidate connexin genes with temporal control from OECs and address the physiological consequences. Using this model, we effectively abolished the expression of connexin 43 (Cx43) in OECs in both juvenile and adult mice. Cx43-deleted OECs exhibited features consistent with the loss of gap junctions including reduced membrane conductance, largely reduced sensitivity to the gap junction blocker meclofenamic acid and loss of dye coupling. This indicates that Cx43, a typically astrocytic connexin, is the main connexin forming functional channels in OECs. Despite these changes in functional properties, the deletion of Cx43 deletion did not alter the density of OECs. The strategy used here may prove useful to delete other candidate genes to better understand the functional roles of OECs in vivo.
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Conexina 43/fisiologia , Junções Comunicantes/fisiologia , Técnicas de Inativação de Genes , Neuroglia/fisiologia , Bulbo Olfatório/citologia , Envelhecimento/metabolismo , Animais , Conexina 43/deficiência , Conexina 43/genética , Cruzamentos Genéticos , Feminino , Junções Comunicantes/efeitos dos fármacos , Genes Reporter , Genes Sintéticos , Integrases/genética , Masculino , Ácido Meclofenâmico/farmacologia , Camundongos , Camundongos Knockout , Proteína Proteolipídica de Mielina/genética , Bulbo Olfatório/metabolismo , Técnicas de Patch-Clamp , Tamoxifeno/farmacologiaRESUMO
Striatal cholinergic interneurons provide modulation to striatal circuits involved in voluntary motor control and goal-directed behaviors through their autonomous tonic discharge and their firing "pause" responses to novel and rewarding environmental events. Striatal cholinergic interneuron hyperactivity was linked to the motor deficits associated with Parkinson's disease and the adverse effects of chronic antiparkinsonian therapy like l-DOPA-induced dyskinesia. Here we addressed whether Kv7 channels, which provide negative feedback to excitation in other neuron types, are involved in the control of striatal cholinergic interneuron tonic activity and response to excitatory inputs. We found that autonomous firing of striatal cholinergic interneurons is not regulated by Kv7 channels. In contrast, Kv7 channels limit the summation of excitatory postsynaptic potentials in cholinergic interneurons through a postsynaptic mechanism. Striatal cholinergic interneurons have a high reserve of Kv7 channels, as their opening using pharmacological tools completely silenced the tonic firing and markedly reduced their intrinsic excitability. A strong inhibition of striatal cholinergic interneurons was also observed in response to the anti-inflammatory drugs diclofenac and meclofenamic acid, however, this effect was independent of Kv7 channels. These data bring attention to new potential molecular targets and pharmacological tools to control striatal cholinergic interneuron activity in pathological conditions where they are believed to be hyperactive, including Parkinson's disease.
Assuntos
Carbamatos/farmacologia , Corpo Estriado/efeitos dos fármacos , Diclofenaco/farmacologia , Interneurônios/efeitos dos fármacos , Moduladores de Transporte de Membrana/farmacologia , Fenilenodiaminas/farmacologia , Canais de Potássio/metabolismo , Acetilcolina/metabolismo , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Corpo Estriado/fisiologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Interneurônios/fisiologia , Masculino , Camundongos Transgênicos , Bloqueadores dos Canais de Potássio/farmacologia , Técnicas de Cultura de TecidosRESUMO
Enormous advances have been made in the recent years in regard to the mechanisms and neural circuits by which odors are sensed and perceived. Part of this understanding has been gained from parallel studies in insects and rodents that show striking similarity in the mechanisms they use to sense, encode, and perceive odors. In this review, we provide a short introduction to the functioning of olfactory systems from transduction of odorant stimuli into electrical signals in sensory neurons to the anatomical and functional organization of the networks involved in neural representation of odors in the central nervous system. We make emphasis on the functional and anatomical architecture of the first synaptic relay of the olfactory circuit, the olfactory bulb in vertebrates and the antennal lobe in insects. We discuss how the exquisite and conserved architecture of this structure is established and how different odors are encoded in mosaic activity patterns. Finally, we discuss the validity of methods used to compare activation patterns in relation to perceptual similarity.
Assuntos
Rede Nervosa/fisiologia , Odorantes , Condutos Olfatórios/citologia , Percepção Olfatória/fisiologia , Neurônios Receptores Olfatórios/fisiologia , Olfato/fisiologia , Animais , Humanos , Transdução de Sinais/fisiologiaRESUMO
Signal processing in neuritic trees is ruled by the concerted action of passive and active membrane properties that, together, determine the degree of electrical compartmentalization of these trees. We analyzed how active properties modulate spatial propagation of graded signals in a pair of nonspiking (NS) neurons of the leech. NS neurons present a very extensive neuritic tree that mediates the interaction with all the excitatory motoneurons in leech ganglia. NS cells express voltage-activated Ca(2+) conductances (VACCs) that, under certain experimental conditions, evoke low-threshold spikes. We studied the distribution of calcium transients in NS neurons loaded with fluorescent calcium probes in response to low-threshold spikes, electrical depolarizing pulses, and synaptic inputs. The three types of stimuli evoked calcium transients of similar characteristics in the four main branches of the neuron. The magnitude of the calcium transients evoked by electrical pulses was a graded function of the change in NS membrane potential and depended on the baseline potential level. The underlying VACCs were partially inactivated at rest and strongly inactivated at -20 mV. Stimulation of mechanosensory pressure cells evoked calcium transients in NS neurons whose amplitude was a linear function of the amplitude of the postsynaptic response. The results evidenced that VACCs aid an efficient propagation of graded signals, turning the vast neuritic tree of NS cells into an electrically compact structure.
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Potenciais de Ação , Neurônios Motores/fisiologia , Potenciais Sinápticos , Animais , Cálcio/metabolismo , Canais de Cálcio/fisiologia , Gânglios dos Invertebrados/citologia , Gânglios dos Invertebrados/fisiologia , Sanguessugas , Mecanorreceptores/fisiologia , Mecanotransdução CelularRESUMO
Striatal cholinergic interneurons show tonic spiking activity in the intact and sliced brain, which stems from intrinsic mechanisms. Because of it, they are also known as "tonically active neurons" (TANs). Another hallmark of TAN electrophysiology is a pause response to appetitive and aversive events and to environmental cues that have predicted these events during learning. Notably, the pause response is lost after the degeneration of dopaminergic neurons in animal models of Parkinson's disease. Moreover, Parkinson's disease patients are in a hypercholinergic state and find some clinical benefit in anticholinergic drugs. Current theories propose that excitatory thalamic inputs conveying information about salient sensory stimuli trigger an intrinsic hyperpolarizing response in the striatal cholinergic interneurons. Moreover, it has been postulated that the loss of the pause response in Parkinson's disease is related to a diminution of I(sAHP), a slow outward current that mediates an afterhyperpolarization following a train of action potentials. Here we report that I(sAHP) induces a marked spike-frequency adaptation in adult rat striatal cholinergic interneurons, inducing an abrupt end of firing during sustained excitation. Chronic loss of dopaminergic neurons markedly reduces I(sAHP) and spike-frequency adaptation in cholinergic interneurons, allowing them to fire continuously and at higher rates during sustained excitation. These findings provide a plausible explanation for the hypercholinergic state in Parkinson's disease. Moreover, a reduction of I(sAHP) may alter synchronization of cholinergic interneurons with afferent inputs, thus contributing to the loss of the pause response in Parkinson's disease.
Assuntos
Acetilcolina/metabolismo , Potenciais de Ação/fisiologia , Corpo Estriado/patologia , Interneurônios/fisiologia , Transtornos Parkinsonianos/patologia , Análise de Variância , Animais , Antracenos/farmacologia , Apamina/farmacologia , Ácido Ascórbico/efeitos adversos , Bário/farmacologia , Simulação por Computador , Corpo Estriado/lesões , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Estimulação Elétrica/métodos , Técnicas In Vitro , Indóis/farmacologia , Masculino , Modelos Neurológicos , Fármacos Neuroprotetores/farmacologia , Oxidopamina/efeitos adversos , Transtornos Parkinsonianos/induzido quimicamente , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Substância Negra/lesões , Substância Negra/fisiologiaRESUMO
The NS neurons are nonspiking cells, present as pairs in each midbody ganglion of the leech nervous system, which display a very extensive arborization. They were shown to regulate the coactivation of motoneurons. Here we have investigated the electrophysiological properties of these neurons under the hypothesis that transmission along the extensive neurites requires the aid of voltage-dependent conductances. The results indicate that NS neurons respond to electrical stimulation with a spike-like event, which was not an all-or-none but rather a graded phenomenon that depended on the intensity and duration of the electrical stimulus. The spike-like response was activated at a membrane potential of approximately -50 mV; its amplitude was a logarithmic function of the extracellular Ca2+ concentration and was unaffected by a broad range of changes in the extracellular Na+ concentration; intracellular application of tetraethylammonium (TEA) caused a large increase in its amplitude and duration. These data indicate that NS neurons bear voltage-dependent low-threshold Ca2+ and TEA-sensitive K+ conductances that could contribute to shaping synaptic signals, or transmission along the extensive neuritic tree.
Assuntos
Cálcio/metabolismo , Sanguessugas/fisiologia , Potenciais da Membrana/fisiologia , Condução Nervosa/fisiologia , Neurônios/fisiologia , Animais , Técnicas de Patch-Clamp , Potássio/metabolismoRESUMO
Electrical coupling through gap junctions constitutes a mode of signal transmission between neurons (electrical synaptic transmission). Originally discovered in invertebrates and in lower vertebrates, electrical synapses have recently been reported in immature and adult mammalian nervous systems. This has renewed the interest in understanding the role of electrical synapses in neural circuit function and signal processing. The present review focuses on the role of gap junctions in shaping the dynamics of neural networks by forming electrical synapses between neurons. Electrical synapses have been shown to be important elements in coincidence detection mechanisms and they can produce complex input-output functions when arranged in combination with chemical synapses. We postulate that these synapses may also be important in redefining neuronal compartments, associating anatomically distinct cellular structures into functional units. The original view of electrical synapses as static connecting elements in neural circuits has been revised and a considerable amount of evidence suggests that electrical synapses substantially affect the dynamics of neural circuits.
Assuntos
Junções Comunicantes/fisiologia , Rede Nervosa/fisiologia , Animais , Humanos , Transmissão Sináptica/fisiologiaRESUMO
Electrical transmission among neurons has been considered a mechanism to synchronize neuronal activity, and rectification provides a mechanism to confine the flow of signals among the connected neurons. The question is how this type of transmission operates within complex neuronal networks. In the leech, the neurons located in position 151 of the midbody ganglion map are connected to virtually every motoneuron via rectifying electrical synapses that pass negative current to the motoneurons. These are nonspiking neurons, and here we have labeled them NS neurons. The goal of this investigation has been to assess their role in regulating motor activity and how rectifying electrical synapses contribute to the function of motor networks. The coupling between NS neurons and motoneurons was voltage sensitive: it increased as motoneurons were depolarized. In addition, excitation of motoneurons evoked hyperpolarizing synaptic responses in NS neurons, the amplitude of which depended on the membrane potential of the latter and on the motoneuron firing frequency. This hyperpolarization was mediated by chemical transmission through an interneuronal layer that spanned the nerve cord. These interactions established a feedback loop between NS and motoneurons that was regulated by the membrane potential of NS. This mechanism was responsible for the uncoupling between otherwise electrically coupled motoneurons. In this way, the NS neurons can act as "electrical neuromodulators," modifying the interaction of other neurons, depending on the activity of the system as a whole.
Assuntos
Neurônios Motores/fisiologia , Rede Nervosa/fisiologia , Sinapses/fisiologia , Potenciais de Ação/fisiologia , Animais , Estimulação Elétrica , Retroalimentação/fisiologia , Gânglios dos Invertebrados/citologia , Gânglios dos Invertebrados/fisiologia , Junções Comunicantes/fisiologia , Técnicas In Vitro , Sanguessugas , Potenciais da Membrana/fisiologia , Rede Nervosa/citologia , Inibição Neural/fisiologia , Vias Neurais/fisiologia , Transmissão Sináptica/fisiologiaRESUMO
Se analizó el efecto que la estimulación aversiva predecible e impredecible provoca sobre el rango social en ratas. Se formaron parejas de ratas Sprague Dawley macho adultas y se procedió a evaluar y caracterizar el rango jerárquico de cada miembro del par. Para ello, se registró la prioridad y desplazamientos de los animales para acceder a una fuente limitada de agua. Una vez verificado el rango de cada individuo, se procedió a someter a los animales dominantes de cada pareja a una sesión de 100 shocks eléctricos no escapables, en donde mitad de los animales lo recibieron a intervalos constantes y la otra a intervalos variables. Posteriormente, se volvió a analizar la jerarquía. Los resultados mostraron que la aplicación de descargas a intervalos variables (descargas no predecibles) provocaron una caída del status de los dominantes, mientras que la misma cantidad de descargas presentadas a intervalos fijos no produjo cambios en la jerarquía de los dominantes.