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In this work, we highlight an electrophysiological feature often observed in recordings from mouse CA1 pyramidal cells that has so far been ignored by experimentalists and modelers. It consists of a large and dynamic increase in the depolarization baseline (i.e., the minimum value of the membrane potential between successive action potentials during a sustained input) in response to strong somatic current injections. Such an increase can directly affect neurotransmitter release properties and, more generally, the efficacy of synaptic transmission. However, it cannot be explained by any currently available conductance-based computational model. Here we present a model addressing this issue, demonstrating that experimental recordings can be reproduced by assuming that an input current modifies, in a time-dependent manner, the electrical and permeability properties of the neuron membrane by shifting the ionic reversal potentials and channel kinetics. For this reason, we propose that any detailed model of ion channel kinetics for neurons exhibiting this characteristic should be adapted to correctly represent the response and the synaptic integration process during strong and sustained inputs.
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Hipocampo , Células Piramidales , Potenciales de Acción/fisiología , Animales , Hipocampo/fisiología , Ratones , Neuronas , Transmisión SinápticaRESUMEN
Synaptic dysfunction plays a central role in Alzheimer's disease (AD), since it drives the cognitive decline. An association between a polymorphism of the adenosine A2A receptor (A2AR) encoding gene-ADORA2A, and hippocampal volume in AD patients was recently described. In this study, we explore the synaptic function of A2AR in age-related conditions. We report, for the first time, a significant overexpression of A2AR in hippocampal neurons of aged humans, which is aggravated in AD patients. A similar profile of A2AR overexpression in rats was sufficient to drive age-like memory impairments in young animals and to uncover a hippocampal LTD-to-LTP shift. This was accompanied by increased NMDA receptor gating, dependent on mGluR5 and linked to enhanced Ca2+ influx. We confirmed the same plasticity shift in memory-impaired aged rats and APP/PS1 mice modeling AD, which was rescued upon A2AR blockade. This A2AR/mGluR5/NMDAR interaction might prove a suitable alternative for regulating aberrant mGluR5/NMDAR signaling in AD without disrupting their constitutive activity.
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Envejecimiento/metabolismo , Depresión Sináptica a Largo Plazo , Neuronas/metabolismo , Receptor de Adenosina A2A/metabolismo , Receptor del Glutamato Metabotropico 5/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Adenosina/metabolismo , Enfermedad de Alzheimer/metabolismo , Animales , Células Cultivadas , Hipocampo/metabolismo , Humanos , Ratones , Ratas , Ratas Sprague-Dawley , Memoria EspacialRESUMEN
SCN1A (NaV1.1 sodium channel) mutations cause Dravet syndrome (DS) and GEFS+ (which is in general milder), and are risk factors in other epilepsies. Phenotypic variability limits precision medicine in epilepsy, and it is important to identify factors that set phenotype severity and their mechanisms. It is not yet clear whether SCN1A mutations are necessary for the development of severe phenotypes or just for promoting seizures. A relevant example is the pleiotropic R1648H mutation that can cause either mild GEFS+ or severe DS. We used a R1648H knock-in mouse model (Scn1aRH/+) with mild/asymptomatic phenotype to dissociate the effects of seizures and of the mutation per se. The induction of short repeated seizures, at the age of disease onset for Scn1a mouse models (P21), had no effect in WT mice, but transformed the mild/asymptomatic phenotype of Scn1aRH/+ mice into a severe DS-like phenotype, including frequent spontaneous seizures and cognitive/behavioral deficits. In these mice, we found no major modifications in cytoarchitecture or neuronal death, but increased excitability of hippocampal granule cells, consistent with a pathological remodeling. Therefore, we demonstrate for our model that an SCN1A mutation is a prerequisite for a long term deleterious effect of seizures on the brain, indicating a clear interaction between seizures and the mutation for the development of a severe phenotype generated by pathological remodeling. Applied to humans, this result suggests that genetic alterations, even if mild per se, may increase the risk of second hits to develop severe phenotypes.
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Epilepsia/genética , Epilepsia/patología , Canal de Sodio Activado por Voltaje NAV1.1/genética , Convulsiones/genética , Convulsiones/patología , Animales , Técnicas de Sustitución del Gen , Hipocampo/patología , Ratones , Mutación , FenotipoRESUMEN
NMDA receptors (NMDARs) are ionotropic receptors crucial for brain information processing. Yet, evidence also supports an ion-flux-independent signaling mode mediating synaptic long-term depression (LTD) and spine shrinkage. Here, we identify AETA (Aη), an amyloid-ß precursor protein (APP) cleavage product, as an NMDAR modulator with the unique dual regulatory capacity to impact both signaling modes. AETA inhibits ionotropic NMDAR activity by competing with the co-agonist and induces an intracellular conformational modification of GluN1 subunits. This favors non-ionotropic NMDAR signaling leading to enhanced LTD and favors spine shrinkage. Endogenously, AETA production is increased by in vivo chemogenetically induced neuronal activity. Genetic deletion of AETA production alters NMDAR transmission and prevents LTD, phenotypes rescued by acute exogenous AETA application. This genetic deletion also impairs contextual fear memory. Our findings demonstrate AETA-dependent NMDAR activation (ADNA), characterizing AETA as a unique type of endogenous NMDAR modulator that exerts bidirectional control over NMDAR signaling and associated information processing.
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Precursor de Proteína beta-Amiloide , Receptores de N-Metil-D-Aspartato , Transducción de Señal , Animales , Humanos , Ratones , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Espinas Dendríticas/metabolismo , Miedo/fisiología , Hipocampo/metabolismo , Depresión Sináptica a Largo Plazo/fisiología , Depresión Sináptica a Largo Plazo/efectos de los fármacos , Memoria/fisiología , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Transducción de Señal/fisiología , RatasRESUMEN
The amyloid-ß precursor protein (APP) is a ubiquitous protein with a strong genetic link to Alzheimer's disease. Although the protein was identified more than forty years ago, its physiological function is still unclear. In recent years, advances in technology have allowed researchers to tackle APP functions in greater depth. In this review, we discuss the latest research pertaining to APP functions from development to aging. We also address the different roles that APP could play in specific types of cells of the central and peripheral nervous system and in other organs of the body. We argue that, until we fully identify the functions of APP in space and time, we will be missing important pieces of the puzzle to solve its pathological implication in Alzheimer's disease and beyond.
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Enfermedad de Alzheimer , Precursor de Proteína beta-Amiloide , Humanos , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/patología , Sinapsis/metabolismo , Sistema Nervioso Periférico/patología , Péptidos beta-Amiloides/metabolismoRESUMEN
N-methyl-D-aspartate receptors (NMDARs) are critical for the maturation and plasticity of glutamatergic synapses. In the hippocampus, NMDARs mainly contain GluN2A and/or GluN2B regulatory subunits. The amyloid precursor protein (APP) has emerged as a putative regulator of NMDARs, but the impact of this interaction to their function is largely unknown. By combining patch-clamp electrophysiology and molecular approaches, we unravel a dual mechanism by which APP controls GluN2B-NMDARs, depending on the life stage. We show that APP is highly abundant specifically at the postnatal postsynapse. It interacts with GluN2B-NMDARs, controlling its synaptic content and mediated currents, both in infant mice and primary neuronal cultures. Upon aging, the APP amyloidogenic-derived C-terminal fragments, rather than APP full-length, contribute to aberrant GluN2B-NMDAR currents. Accordingly, we found that the APP processing is increased upon aging, both in mice and human brain. Interfering with stability or production of the APP intracellular domain normalized the GluN2B-NMDARs currents. While the first mechanism might be essential for synaptic maturation during development, the latter could contribute to age-related synaptic impairments.
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Precursor de Proteína beta-Amiloide , Receptores de N-Metil-D-Aspartato , Ratones , Humanos , Animales , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Transducción de Señal , Hipocampo/metabolismo , Sinapsis/metabolismoRESUMEN
BACKGROUND: Amyloid precursor protein (APP) processing is central to Alzheimer's disease (AD) etiology. As early cognitive alterations in AD are strongly correlated to abnormal information processing due to increasing synaptic impairment, it is crucial to characterize how peptides generated through APP cleavage modulate synapse function. We previously described a novel APP processing pathway producing η-secretase-derived peptides (Aη) and revealed that Aη-α, the longest form of Aη produced by η-secretase and α-secretase cleavage, impaired hippocampal long-term potentiation (LTP) ex vivo and neuronal activity in vivo. METHODS: With the intention of going beyond this initial observation, we performed a comprehensive analysis to further characterize the effects of both Aη-α and the shorter Aη-ß peptide on hippocampus function using ex vivo field electrophysiology, in vivo multiphoton calcium imaging, and in vivo electrophysiology. RESULTS: We demonstrate that both synthetic peptides acutely impair LTP at low nanomolar concentrations ex vivo and reveal the N-terminus to be a primary site of activity. We further show that Aη-ß, like Aη-α, inhibits neuronal activity in vivo and provide confirmation of LTP impairment by Aη-α in vivo. CONCLUSIONS: These results provide novel insights into the functional role of the recently discovered η-secretase-derived products and suggest that Aη peptides represent important, pathophysiologically relevant, modulators of hippocampal network activity, with profound implications for APP-targeting therapeutic strategies in AD.
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Enfermedad de Alzheimer , Potenciación a Largo Plazo , Secretasas de la Proteína Precursora del Amiloide , Péptidos beta-Amiloides , Precursor de Proteína beta-Amiloide/genética , Humanos , NeuronasRESUMEN
Neuroinflammation in patients with Alzheimer's disease (AD) and related mouse models has been recognized for decades, but the contribution of the recently described meningeal immune population to AD pathogenesis remains to be addressed. Here, using the 3xTg-AD model, we report an accumulation of interleukin-17 (IL-17)-producing cells, mostly γδ T cells, in the brain and the meninges of female, but not male, mice, concomitant with the onset of cognitive decline. Critically, IL-17 neutralization into the ventricles is sufficient to prevent short-term memory and synaptic plasticity deficits at early stages of disease. These effects precede blood-brain barrier disruption and amyloid-beta or tau pathology, implying an early involvement of IL-17 in AD pathology. When IL-17 is neutralized at later stages of disease, the onset of short-memory deficits and amyloidosis-related splenomegaly is delayed. Altogether, our data support the idea that cognition relies on a finely regulated balance of "inflammatory" cytokines derived from the meningeal immune system.
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Enfermedad de Alzheimer/metabolismo , Conducta Animal , Encéfalo/metabolismo , Cognición , Mediadores de Inflamación/metabolismo , Interleucina-17/metabolismo , Linfocitos Intraepiteliales/metabolismo , Enfermedades Neuroinflamatorias/metabolismo , Sinapsis/metabolismo , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/prevención & control , Enfermedad de Alzheimer/psicología , Animales , Antiinflamatorios/farmacología , Anticuerpos Monoclonales/farmacología , Anticuerpos Neutralizantes/farmacología , Conducta Animal/efectos de los fármacos , Encéfalo/efectos de los fármacos , Encéfalo/patología , Cognición/efectos de los fármacos , Modelos Animales de Enfermedad , Femenino , Mediadores de Inflamación/antagonistas & inhibidores , Interleucina-17/antagonistas & inhibidores , Linfocitos Intraepiteliales/efectos de los fármacos , Masculino , Memoria a Corto Plazo , Ratones de la Cepa 129 , Ratones Transgénicos , Enfermedades Neuroinflamatorias/patología , Enfermedades Neuroinflamatorias/prevención & control , Enfermedades Neuroinflamatorias/psicología , Plasticidad Neuronal , Sinapsis/efectos de los fármacos , Sinapsis/patologíaRESUMEN
Adenosine-induced modulation of neuromuscular transmission in young (3-4-week-old) rats was evaluated. Inhibition of adenosine kinase with iodotubercidin (ITU; 10 microM), which is known to induce adenosine release, enhanced the amplitude of evoked end-plate potentials (EPPs) recorded from innervated diaphragm muscle fibers. This facilitatory effect was transformed into an inhibitory one upon blockade of adenosine A(2A) receptors with 4-(2-[7-amino-2-(2-furly)[1,2,4]triazolo[2,3-a][1,3,5]triazin5ylamino] ethyl) phenol (ZM 241385) (50 nM); further blockade of adenosine A(1) receptors with the selective antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX; 10 nM) abolished that inhibition. Adenosine or 2-chloroadenosine (CADO), at submicromolar concentrations, increased the amplitude and the quantal content of EPPs, whereas at low micromolar concentrations they decreased EPP amplitude. Blockade of A(1) receptors with DPCPX (10 nM) prevented both excitatory and inhibitory effects, whereas blockade of A(2A) receptors with ZM241385 (50 nM) prevented only the excitatory effects. DPCPX and ZM241385 also prevented the excitatory effect of the selective A(2A) receptor agonist 2-[p-(2-carboxyethyl) phenethylamino]-5'-N-ethylcarboxamido adenosine hydrochloride (CGS 21680; 10 nM). CADO (30 nM) also increased neuromuscular transmission in adult (12-16-week-old) rats. It is suggested that at the motor nerve endings, low extracellular concentrations of adenosine activate both A(2A) and A(1) receptors, but activation of A(2A) receptors predominates over A(1) receptors; the activity of A(2A) receptors might, however, require coactivation of A(1) receptors. This facilitatory action of low concentrations of extracellular adenosine upon acetylcholine release may be particularly relevant at developing neuromuscular junctions, where subtle changes in synaptic levels of acetylcholine might influence synaptic stabilization.
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Adenosina/fisiología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Potenciales Postsinápticos Miniatura/efectos de los fármacos , Unión Neuromuscular/efectos de los fármacos , 2-Cloroadenosina/farmacología , Adenosina/farmacología , Antagonistas del Receptor de Adenosina A1 , Antagonistas del Receptor de Adenosina A2 , Animales , Diafragma/efectos de los fármacos , Diafragma/inervación , Diafragma/fisiología , Potenciales Postsinápticos Excitadores/fisiología , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/fisiología , Ligandos , Masculino , Potenciales Postsinápticos Miniatura/fisiología , Unión Neuromuscular/fisiología , Ratas , Ratas Wistar , Receptor de Adenosina A1/fisiología , Receptor de Adenosina A2A/fisiología , Tubercidina/análogos & derivados , Tubercidina/farmacologíaRESUMEN
While neuronal loss has long been considered as the main contributor to age-related cognitive decline, these alterations are currently attributed to gradual synaptic dysfunction driven by calcium dyshomeostasis and alterations in ionotropic/metabotropic receptors. Given the key role of the hippocampus in encoding, storage, and retrieval of memory, the morpho- and electrophysiological alterations that occur in the major synapse of this network-the glutamatergic-deserve special attention. We guide you through the hippocampal anatomy, circuitry, and function in physiological context and focus on alterations in neuronal morphology, calcium dynamics, and plasticity induced by aging and Alzheimer's disease (AD). We provide state-of-the art knowledge on glutamatergic transmission and discuss implications of these novel players for intervention. A link between regular consumption of caffeine-an adenosine receptor blocker-to decreased risk of AD in humans is well established, while the mechanisms responsible have only now been uncovered. We review compelling evidence from humans and animal models that implicate adenosine A2A receptors (A2AR) upsurge as a crucial mediator of age-related synaptic dysfunction. The relevance of this mechanism in patients was very recently demonstrated in the form of a significant association of the A2AR-encoding gene with hippocampal volume (synaptic loss) in mild cognitive impairment and AD. Novel pathways implicate A2AR in the control of mGluR5-dependent NMDAR activation and subsequent Ca2+ dysfunction upon aging. The nature of this receptor makes it particularly suited for long-term therapies, as an alternative for regulating aberrant mGluR5/NMDAR signaling in aging and disease, without disrupting their crucial constitutive activity.
RESUMEN
The notion of "immune privilege" of the brain has been revised to accommodate its infiltration, at steady state, by immune cells that participate in normal neurophysiology. However, the immune mechanisms that regulate learning and memory remain poorly understood. Here, we show that noninflammatory interleukin-17 (IL-17) derived from a previously unknown fetal-derived meningeal-resident γδ T cell subset promotes cognition. When tested in classical spatial learning paradigms, mice lacking γδ T cells or IL-17 displayed deficient short-term memory while retaining long-term memory. The plasticity of glutamatergic synapses was reduced in the absence of IL-17, resulting in impaired long-term potentiation in the hippocampus. Conversely, IL-17 enhanced glial cell production of brain-derived neurotropic factor, whose exogenous provision rescued the synaptic and behavioral phenotypes of IL-17-deficient animals. Together, our work provides previously unknown clues on the mechanisms that regulate short-term versus long-term memory and on the evolutionary and functional link between the immune and nervous systems.
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Interleucina-17/inmunología , Memoria a Corto Plazo , Meninges/inmunología , Plasticidad Neuronal/inmunología , Linfocitos T/inmunología , Animales , Interleucina-17/deficiencia , Ratones , Ratones Endogámicos C57BL , Ratones NoqueadosRESUMEN
There is a growing consensus that Alzheimer's disease (AD) involves failure of the homeostatic machinery, which underlies the firing stability of neural circuits. What are the culprits leading to neuron firing instability? The amyloid precursor protein (APP) is central to AD pathogenesis, and we recently showed that its intracellular domain (AICD) could modify synaptic signal integration. We now hypothesize that AICD modifies neuron firing activity, thus contributing to the disruption of memory processes. Using cellular, electrophysiological, and behavioral techniques, we show that pathological AICD levels weaken CA1 neuron firing activity through a gene-transcription-dependent mechanism. Furthermore, increased AICD production in hippocampal neurons modifies oscillatory activity, specifically in the γ-frequency range, and disrupts spatial memory task. Collectively, our data suggest that AICD pathological levels, observed in AD mouse models and in human patients, might contribute to progressive neuron homeostatic failure, driving the shift from normal aging to AD.
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Potenciales de Acción/fisiología , Precursor de Proteína beta-Amiloide/química , Precursor de Proteína beta-Amiloide/metabolismo , Región CA1 Hipocampal/fisiología , Neuronas/fisiología , Memoria Espacial/fisiología , Animales , Canales de Calcio/metabolismo , Ritmo Gamma/fisiología , Humanos , Masculino , Ratones Endogámicos C57BL , Modelos Biológicos , Canales de Potasio/metabolismo , Dominios Proteicos , Ratas Sprague-Dawley , Relación Estructura-Actividad , Transcripción GenéticaRESUMEN
The amyloid precursor protein (APP) harbors physiological roles at synapses and is central to Alzheimer's disease (AD) pathogenesis. Evidence suggests that APP intracellular domain (AICD) could regulate synapse function, but the underlying molecular mechanisms remain unknown. We addressed AICD actions at synapses, per se, combining in vivo AICD expression, ex vivo AICD delivery or APP knock-down by in utero electroporation of shRNAs with whole-cell electrophysiology. We report a critical physiological role of AICD in controlling GluN2B-containing NMDA receptors (NMDARs) at immature excitatory synapses, via a transcription-dependent mechanism. We further show that AICD increase in mature neurons, as reported in AD, alters synaptic NMDAR composition to an immature-like GluN2B-rich profile. This disrupts synaptic signal integration, via over-activation of SK channels, and synapse plasticity, phenotypes rescued by GluN2B antagonism. We provide a new physiological role for AICD, which becomes pathological upon AICD increase in mature neurons. Thus, AICD could contribute to AD synaptic failure.
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Enfermedad de Alzheimer/fisiopatología , Péptidos beta-Amiloides/farmacología , Precursor de Proteína beta-Amiloide/metabolismo , Hipocampo/patología , Neurogénesis/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapsis/patología , Precursor de Proteína beta-Amiloide/genética , Animales , Modelos Animales de Enfermedad , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Potenciación a Largo Plazo/efectos de los fármacos , Ratones , Dominios Proteicos , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/genética , Sinapsis/efectos de los fármacos , Sinapsis/metabolismoRESUMEN
Motor nerve terminals possess adenosine A(2A) receptors and brain derived neurotrophic factor (BDNF) TrkB receptors. In the present work we evaluated how BDNF actions on neuromuscular transmission would be influenced by adenosine A(2A) receptors activation. BDNF (20-100 ng/ml) on its own was devoid of effect on evoked endplate potentials (EPPs) recorded intracellularly from rat innervated diaphragms paralysed with tubocurarine. However, when BDNF was applied 45 min after a brief (2 min) depolarizing KCl (10 mM) pulse or when the adenosine A(2A) receptors were activated with CGS 21680 (10 nM), BDNF (20 ng/ml) increased EPPs amplitude without influencing the resting membrane potential of the muscle fibre. The action of BDNF was prevented by the adenosine A(2A) receptor antagonist, ZM 241385 (50 nM) as well as by the TrkB receptor phosphorylation inhibitor, K252a (200 nM). The PKA inhibitor, H-89 (1 microM), prevented the excitatory effect of CGS 21680 (10 nM) on EPPs as well as prevented its ability to trigger a BDNF effect. The PLCgamma inhibitor, U73122 (5 microM), did not prevent the excitatory action of CGS 21680 (10 nM) on neuromuscular transmission, but abolished the action of BDNF in the presence of the A(2A) receptor agonist. The results suggest the following sequence of events in what concerns cooperativity between A(2A) receptors and TrkB receptors at the neuromuscular junction: A(2A) receptor activates the PKA pathway, which promotes the action of BDNF through TrkB receptors coupled to PLCgamma, leading to enhancement of neuromuscular transmission.
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Factor Neurotrófico Derivado del Encéfalo/farmacología , Unión Neuromuscular/fisiología , Receptor de Adenosina A2A/fisiología , Transmisión Sináptica/fisiología , Adenosina/análogos & derivados , Adenosina/farmacología , Antagonistas del Receptor de Adenosina A2 , Animales , Diafragma/efectos de los fármacos , Diafragma/fisiología , Potenciales Evocados/efectos de los fármacos , Potenciales Evocados/fisiología , Unión Neuromuscular/efectos de los fármacos , Fenetilaminas/farmacología , Ratas , Transmisión Sináptica/efectos de los fármacosRESUMEN
At the neuromuscular junction, spontaneous giant events (GMEPPs) are enhanced in different conditions when degenerative and/or remodeling processes take place, but no one investigated their incidence upon aging. In the present work, we evaluated evoked and spontaneous neuromuscular transmission events recorded from single muscle fibers. Phrenic-diaphragm preparations of 3-4, 12-16, 36-40 and 70-80 weeks old rat males were used. We found that the occurrence of GMEPPs significantly increases in aged rats. Moreover, in old rats the neuromuscular transmission was significantly impaired due to a significant decrease in the amplitude and quantal content of evoked endplate potentials. Interestingly, the number of observed EPPs failures were â¼ 3 times lower than the predicted value based on the quantal content. This discrepancy was not observed in infant or adult rats. The coincidence of a high GMEPPs frequency with a lower than expected EPPs failure rate suggests that GMEPPs events are needed to preserve effective neuromuscular transmission in aged animals.
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Envejecimiento/fisiología , Potenciales Postsinápticos Miniatura , Acetilcolina/metabolismo , Animales , Diafragma/inervación , Diafragma/fisiología , Masculino , Fibras Musculares Esqueléticas/fisiología , Unión Neuromuscular/fisiología , Nervio Frénico/fisiología , Ratas WistarRESUMEN
Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease leading to motor neuron dysfunction resulting in impairment of neuromuscular transmission. A2A adenosine receptors have already been considered as a potential therapeutical target for ALS but their neuromodulatory role at the neuromuscular junction in ALS remains to be clarified. In the present work, we evaluated the effects of A2A receptors on neuromuscular transmission of an animal model of ALS: SOD1(G93A) mice either in the pre-symptomatic (4-6 weeks old) or in the symptomatic (12-14 weeks old) stage. Electrophysiological experiments were performed obtaining intracellular recordings in Mg2+ paralyzed phrenic nerve-hemidiaphragm preparations. Endplate potentials (EPPs), quantal content (q. c.) of EPPs, miniature endplate potentials (MEPPs) and giant miniature endplate potential (GMEPPs) were recorded. In the pre-symptomatic phase of the disease (4-6 weeks old mice), the selective A2A receptor agonist, CGS 21680, significantly enhanced (p<0.05 Unpaired t-test) the mean amplitude and q.c. of EPPs, and the frequency of MEPPs and GMEPPs at SOD1(G93A) neuromuscular junctions, the effect being of higher magnitude (p<0.05, Unpaired t-test) than age-matched control littermates. On the contrary, in symptomatic mice (12-14 weeks old), CGS 21680 was devoid of effect on both the amplitude and q.c. of EPPs and the frequency of MEPPs and GMEPPs (p<0.05 Paired t-test). The results herein reported clearly document that at the neuromuscular junction of SOD1(G93A) mice there is an exacerbation of A2A receptor-mediated excitatory effects at the pre-symptomatic phase, whereas in the symptomatic phase A2A receptor activation is absent. The results thus suggest that A2A receptors function changes with ALS progression.
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Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/fisiopatología , Mutación/genética , Unión Neuromuscular/fisiopatología , Receptor de Adenosina A2A/metabolismo , Superóxido Dismutasa/genética , Transmisión Sináptica , Adenosina/análogos & derivados , Esclerosis Amiotrófica Lateral/patología , Animales , Progresión de la Enfermedad , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Humanos , Masculino , Ratones Transgénicos , Placa Motora/efectos de los fármacos , Placa Motora/patología , Placa Motora/fisiopatología , Unión Neuromuscular/patología , Fenetilaminas , Transmisión Sináptica/efectos de los fármacosRESUMEN
Amyotrophic lateral sclerosis is characterized by a progressive degeneration of the corticospinal tract motor neurons. Growing evidence suggests that degeneration may begin at the distal axon proceeding in a dying-back pattern. It seemed therefore of interest to investigate synaptic transmission at the neuromuscular junction (NMJ) in pre- and symptomatic phases of the disease. Endplate potentials (EPPs), miniatures endplate potentials (MEPPs) and giant MEPPs (GMEPPs) were recorded from innervated diaphragm muscle fibers from 4-6 and 12-15 weeks-old SOD1(G93A) mice and non-transgenic aged-matched littermates (WT). In the pre-symptomatic phase, SOD1(G93A) mice exhibited a significant increase in the mean amplitude of EPPs together with an increase in the mean quantal content of EPPs, suggesting that more acetylcholine is being released into the synaptic cleft. SOD1(G93A) mice presented a higher frequency of GMEPPs, suggestive of intracellular Ca(2+) deregulation in nerve terminals. The increase in the mean amplitude of MEPPs and the decreased mean rise-time of MEPPs in SOD1(G93A) mice point to post-synaptic related changes. In the symptomatic phase, electrophysiological data showed evidence for two NMJ groups in SOD1(G93A) mice: SOD1a and SOD1b. SOD1a group presented reduced mean amplitude of both EPPs and MEPPs. The mean rise-time of MEPPs was increased, when compared to WT and to SOD1b group, indicating impairments in the neuromuscular transmission. In contrast, the neuromuscular transmission of SOD1b group was not different from age-matched WT nor pre-symptomatic SOD1(G93A) mice, being somehow in between both groups. Altogether these results show that the neuromuscular transmission of SOD1(G93A) mice is enhanced in the pre-symptomatic phase. In the symptomatic phase our results are consistent with the hypothesis that the diaphragm of SOD1(G93A) mice is undergoing cycles of denervation/re-innervation supported by mixed neuromuscular junction populations. These early changes in the neuromuscular transmission of SOD1(G93A) mice suggest that the ALS associated events start long before symptoms onset.
Asunto(s)
Esclerosis Amiotrófica Lateral/fisiopatología , Modelos Animales de Enfermedad , Unión Neuromuscular/fisiología , Superóxido Dismutasa/metabolismo , Transmisión Sináptica/fisiología , Esclerosis Amiotrófica Lateral/genética , Esclerosis Amiotrófica Lateral/patología , Animales , Diafragma/inervación , Diafragma/metabolismo , Femenino , Humanos , Masculino , Ratones , Ratones Transgénicos , Actividad Motora/genética , Actividad Motora/fisiología , Músculo Esquelético/inervación , Músculo Esquelético/metabolismo , Unión Neuromuscular/metabolismo , Superóxido Dismutasa/genética , Potenciales Sinápticos/genética , Potenciales Sinápticos/fisiología , Transmisión Sináptica/genética , Factores de TiempoRESUMEN
In infant rats adenosine A(2A) receptor-mediated modulation of neuromuscular transmission predominates over A1 receptor-mediated neuromodulation. We investigated whether aging affects this A(2A)/A(1) receptor balance. Evoked (EPPs) and miniature end plate potentials (MEPPs) were recorded from single fibers of (weeks-old) infant (3-4), young adult (12-16), older (36-38), and aged (80-90) male rat-diaphragm. The non A1/A(2A) selective agonist, 2-chloroadenosine (CADO; 30 nM) and the adenosine kinase inhibitor, iodotubericidin (ITU; 10 µM) increased mean amplitude and quantal content of EPPs in infant, young adult, and older adult rats, but not in aged rats. The facilitatory effects were prevented by the A(2A) receptor antagonist, ZM241385 (50 nM) and mimicked by the A(2A) receptor agonist, CGS21680 (10 nM). The A1 receptor agonist, 6-cyclopentyladenosine (CPA; 100 nM), decreased EPPs amplitude in all age groups. It is concluded that aging differently influences adenosine A1 receptor and A(2A) receptor-mediated presynaptic modulation of neuromuscular transmission, so that the facilitatory influence decreases upon aging, whereas the inhibitory influence remains unchanged in aged animals. The reduction of adenosine A(2A) receptors upon aging may contribute to the age-related changes in neuromuscular function.