RESUMEN
SYNGAP1 haploinsufficiency in humans causes intellectual disability (ID). SYNGAP1 is highly expressed in cortical excitatory neurons and, reducing its expression in mice accelerates the maturation of excitatory synapses during sensitive developmental periods, restricts the critical period window for plasticity, and impairs cognition. However, its specific role in interneurons remains largely undetermined. In this study, we investigated the effects of conditional Syngap1 disruption in medial ganglionic eminence (MGE)-derived interneurons on hippocampal interneuron firing properties and excitatory synaptic inputs, as well as on pyramidal cell synaptic inhibition and synaptic integration. We show that conditional Syngap1 disruption in MGE-derived interneurons results in cell-specific impairment of firing properties of hippocampal Nkx2.1 fast-spiking interneurons, with enhancement of their AMPA receptor (AMPAR)-mediated excitatory synaptic inputs but compromised short-term plasticity. In contrast, regular-spiking Nkx2.1 interneurons are largely unaffected. These changes are associated with impaired pyramidal cell synaptic inhibition and enhanced summation of excitatory responses. Unexpectedly, we found that the Syngap1flox allele used in this study contains inverted loxP sites and that its targeted recombination in MGE-derived interneurons induces some cell loss during embryonic development and the reversible inversion of the sequence flanked by the loxP sites in postmitotic cells. Together, these results suggest that Syngap1 plays a role in cell-specific regulation of hippocampal interneuron function and inhibition of pyramidal cells in mice. However, because of our finding that the Syngap1flox allele used in this study contains inverted loxP sites, it will be important to further investigate interneuron function using a different Syngap1 conditional allele.
Asunto(s)
Interneuronas , Células Piramidales , Humanos , Ratones , Animales , Ratones Transgénicos , Interneuronas/fisiología , Células Piramidales/fisiología , Hipocampo/metabolismo , Recombinación Genética , Proteínas Activadoras de ras GTPasa/genética , Proteínas Activadoras de ras GTPasa/metabolismoRESUMEN
General DNA hypomethylation is associated with Alzheimer's disease (AD), but it is unclear when DNA hypomethylation starts or plays a role in AD pathology or whether DNA re-methylation would rescue early amyloid-related cognitive impairments. In an APP transgenic mouse model of AD-like amyloid pathology we found that early intraneuronal amyloid beta build-up is sufficient to unleash a global and beta-site amyloid precursor protein cleaving enzyme 1 (bace-1) DNA demethylation in AD-vulnerable brain regions. S-adenosylmethionine administration at these early stages abolished this hypomethylation, diminished the amyloid pathology and restored cognitive capabilities. To assess a possible human significance of findings, we examined the methylation at 12 CpGs sites in the bace-1 promoter, using genome-wide DNA methylation data from 740 postmortem human brains. Thus, we found significant associations of bace-1 promoter methylation with ß-amyloid load among persons with AD dementia, and PHFtau tangle density. Our results support a plausible causal role for the earliest amyloid beta accumulation to provoke DNA hypomethylation, influencing AD pathological outcomes.
RESUMEN
Current therapies for Alzheimer's disease (AD) offer partial symptomatic relief and do not modify disease progression. There is substantial evidence indicating a disease onset years before clinical diagnosis, at which point no effective therapy has been found. In this study, we investigated the efficacy of a new multi-target drug, M30, at relatively early stages of the AD-like amyloid pathology in a robust rat transgenic model. McGill-R-Thy1-APP transgenic rats develop the full AD-like amyloid pathology in a progressive fashion, and have a minimal genetic burden. McGill rats were given 5âmg/kg M30 or vehicle per os, every 2 days for 4 months, starting at a stage where the transgenic animals suffer detectable cognitive impairments. At the completion of the treatment, cognitive functions were assessed with Novel Object Location and Novel Object Recognition tests. The brains were then analyzed to assess amyloid-ß (Aß) burden and the levels of key inflammatory markers. Long-term treatment with M30 was associated with both the prevention and the reversal of transgene-related cognitive decline. The effects on cognition were accompanied by a shift of the Aß-immunoreactive material toward an amyloid plaque aggregated molecular form, diminished molecular signs of CNS inflammation and a change in microglia morphology toward a surveying phenotype. This study is the first to demonstrate the therapeutic potential of M30 in a rat model of the AD amyloid pathology. It provides a rationale for further investigations with M30 and with potential multi-target approaches to delay, prevent or reverse the progression the AD pathology at early disease-stages.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Antiinflamatorios/farmacología , Encéfalo/efectos de los fármacos , Cognición/efectos de los fármacos , Hidroxiquinolinas/farmacología , Nootrópicos/farmacología , Enfermedad de Alzheimer/inmunología , Péptidos beta-Amiloides/metabolismo , Animales , Encéfalo/inmunología , Cognición/fisiología , Modelos Animales de Enfermedad , Femenino , Peroxidación de Lípido/efectos de los fármacos , Peroxidación de Lípido/fisiología , Masculino , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Fármacos Neuroprotectores/farmacología , Ratas Transgénicas , Reconocimiento en Psicología/efectos de los fármacosRESUMEN
Mesencephalic dopamine (DA) neurons have been suggested to use glutamate as a cotransmitter. Here, we suggest a mechanism for this form of cotransmission by showing that a subset of DA neurons both in vitro and in vivo expresses vesicular glutamate transporter 2 (VGluT2). Expression of VGluT2 decreases with age. Moreover, when DA neurons are grown in isolation using a microculture system, there is a marked upregulation of VGluT2 expression. We provide evidence that expression of this transporter is normally repressed through a contact-dependent interaction with GABA and other DA neurons, thus providing a partial explanation for the highly restricted expression of VGluT2 in DA neurons in vivo. Our results demonstrate that the neurotransmitter phenotype of DA neurons is both developmentally and dynamically regulated. These findings may have implications for a better understanding of the fast synaptic action of DA neurons as well as basal ganglia circuitry.
Asunto(s)
Dopamina/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Neuronas/fisiología , Proteína 2 de Transporte Vesicular de Glutamato/biosíntesis , Animales , Células Cultivadas , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neuronas/metabolismo , Proteína 2 de Transporte Vesicular de Glutamato/genéticaRESUMEN
The medial septum diagonal band area (MS/DB) projects to the hippocampus through the fornix/fimbria pathway and is implicated in generating hippocampal theta oscillations. The hippocampus also projects back to the MS/DB, but very little is known functionally about this input. Here, we investigated the physiological role of hippocamposeptal feedback to the MS/DB in a complete in vitro septohippocampal preparation containing the intact interconnecting fornix/fimbria pathway. We demonstrated that carbachol-induced rhythmic theta-like hippocampal oscillations recorded extracellularly were synchronized with powerful rhythmic IPSPs in whole-cell recorded MS/DB neurons. Interestingly, we found that these IPSPs evoked rebound spiking in GABAergic MS/DB neurons. In contrast, putative cholinergic and glutamatergic MS/DB neurons responded only weakly with rebound spiking and, as a result, were mostly silent during theta-like oscillations. We next determined the mechanism underlying the rebound spiking that followed the IPSPs in MS/DB GABAergic neurons using phasic electrical stimulation of the fornix/fimbria pathway. We demonstrate that the increased rebound spiking was attributable to the activation of I(h) current, because it was significantly reduced by low concentrations of the I(h) antagonist ZD7288 [4-(N-ethyl-N-phenylamino)-1,2-dimethyl-6-(methylamino) pyridinium chloride]. Together, these results suggest that rhythmical activity in hippocampus is transferred to the MS/DB and can preferentially phase the spiking of GABAergic MS/DB neurons because of their significant expression of I(h) currents. Our data demonstrate that hippocamposeptal inhibition facilitates theta rhythmic discharges in MS/DB GABAergic neurons while favoring the inhibition of most ACh and glutamate neurons.
Asunto(s)
Banda Diagonal de Broca/fisiología , Hipocampo/fisiología , Neuronas/fisiología , Núcleos Septales/fisiología , Ácido gamma-Aminobutírico/metabolismo , Potenciales de Acción , Animales , Animales Recién Nacidos , Carbacol/farmacología , Agonistas Colinérgicos/farmacología , Banda Diagonal de Broca/citología , Estimulación Eléctrica , Electrofisiología , Hipocampo/efectos de los fármacos , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores , Inhibición Neural/fisiología , Vías Nerviosas/efectos de los fármacos , Vías Nerviosas/fisiología , Neuronas/metabolismo , Oscilometría , Periodicidad , Ratas , Ratas Sprague-Dawley , Tiempo de Reacción , Núcleos Septales/citología , Transmisión Sináptica/fisiologíaRESUMEN
The medial septum and diagonal band complex (MS-DB) is believed to play a key role in generating theta oscillations in the hippocampus, a phenomenon critical for learning and memory. Although the importance of the MS-DB in hippocampal theta rhythm generation is generally accepted, it remains to be determined whether the MS-DB alone can generate hippocampal oscillations or is only a transducer of rhythmic activity from other brain areas. Secondly, it is known that hippocampal theta rhythm can be separated into an atropine-sensitive and insensitive component. However, it remains to be established if the MS-DB can generate both types of rhythm. To answer these questions, we used a new in vitro rat septohippocampal preparation placed in a hermetically separated two side recording chamber. We showed that carbachol activation of the MS-DB generated large theta oscillations in the CA1 and CA3 regions of the hippocampus. These oscillations were blocked by applying either the GABA(A) receptor antagonist bicuculline or the AMPA/kainate antagonist DNQX to the hippocampus. Interestingly, the application of the muscarinic receptor antagonist atropine produced only a partial decrease in the amplitude, without modification of the frequency, of theta. These results show for the first time, that upon optimal excitation, the MS-DB alone is able to generate hippocampal oscillations in the theta frequency band. Moreover, these MS-DB generated theta oscillations are mediated by muscarinic and nonmuscarinic receptors and have a pharmacological profile similar to theta rhythm observed in awake animals.
Asunto(s)
Atropina/farmacología , Hipocampo/efectos de los fármacos , Antagonistas Muscarínicos/farmacología , Tabique Pelúcido/efectos de los fármacos , Ritmo Teta/efectos de los fármacos , Animales , Bicuculina/farmacología , Carbacol/farmacología , Antagonistas de Receptores de GABA-A , Ácido Glutámico/fisiología , Hipocampo/fisiología , Técnicas In Vitro , Quinoxalinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores de GABA-A/fisiología , Receptores de Ácido Kaínico/antagonistas & inhibidores , Receptores Muscarínicos/efectos de los fármacos , Receptores Muscarínicos/fisiología , Tabique Pelúcido/fisiología , Tetrodotoxina/farmacología , Ácido gamma-Aminobutírico/fisiologíaRESUMEN
Basal forebrain neurons play an important role in memory and attention. In addition to cholinergic and GABAergic neurons, glutamatergic neurons and neurons that can corelease acetylcholine and glutamate have recently been described in the basal forebrain. Although it is well known that nerve growth factor (NGF) promotes synaptic function of cholinergic basal forebrain neurons, how NGF affects the newly identified basal forebrain neurons remains undetermined. Here, we examined the effects of NGF on synaptic transmission of medial septum and diagonal band of Broca (MS-DBB) neurons expressing different neurotransmitter phenotypes. We used MS-DBB neurons from 10- to 13-d-old rats, cultured on astrocytic microislands to promote the development of autaptic connections. Evoked and spontaneous postsynaptic currents were recorded, and neurotransmitters released were characterized pharmacologically. We found that chronic exposure to NGF significantly increased acetylcholine and glutamate release from cholinergic MS-DBB neurons, whereas glutamate and GABA transmission from noncholinergic MS-DBB neurons were not affected by NGF. Interestingly, the NGF-induced increase in neurotransmission was mediated by p75(NTR). These results demonstrate a previously unidentified role of NGF and its receptor p75(NTR); their interactions are crucial for cholinergic and glutamatergic transmission in the septohippocampal pathway.
Asunto(s)
Acetilcolina/metabolismo , Banda Diagonal de Broca/metabolismo , Ácido Glutámico/metabolismo , Factor de Crecimiento Nervioso/administración & dosificación , Receptor de Factor de Crecimiento Nervioso/metabolismo , Núcleos Septales/metabolismo , Animales , Células Cultivadas , Fibras Colinérgicas/metabolismo , Factor de Crecimiento Nervioso/fisiología , Vías Nerviosas/fisiología , Ratas , Ratas Sprague-Dawley , Receptor de Factor de Crecimiento Nervioso/fisiología , Transmisión Sináptica/fisiologíaRESUMEN
BACKGROUND: Several clinical studies suggested that antipsychotic-based medications could ameliorate cognitive functions impaired in certain schizophrenic patients. Accordingly, we investigated the effects of various dopaminergic receptor antagonists--including atypical antipsychotics that are prescribed for the treatment of schizophrenia--in a model of toxicity using cultured hippocampal neurons, the hippocampus being a region of particular relevance to cognition. RESULTS: Hippocampal cell death induced by deprivation of growth medium constituents was strongly blocked by drugs including antipsychotics (10(-10)-10(-6) M) that display nM affinities for D2 and/or D4 receptors (clozapine, haloperidol, (+/-)-sulpiride, domperidone, clozapine, risperidone, chlorpromazine, (+)-butaclamol and L-741,742). These effects were shared by some caspases inhibitors and were not accompanied by inhibition of reactive oxygen species. In contrast, (-)-raclopride and remoxipride, two drugs that preferentially bind D2 over D4 receptors were ineffective, as well as the selective D3 receptor antagonist U 99194. Interestingly, (-)-raclopride (10(-6) M) was able to block the neuroprotective effect of the atypical antipsychotic clozapine (10(-6) M). CONCLUSION: Taken together, these data suggest that D2-like receptors, particularly the D4 subtype, mediate the neuroprotective effects of antipsychotic drugs possibly through a ROS-independent, caspase-dependent mechanism.
Asunto(s)
Antipsicóticos/farmacología , Antagonistas de Dopamina/farmacología , Hipocampo/citología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Animales , Anisoles/farmacología , Benzazepinas/farmacología , Butaclamol/farmacología , Inhibidores de Caspasas , Muerte Celular , Células Cultivadas/efectos de los fármacos , Células Cultivadas/patología , Clorpromazina/farmacología , Clozapina/farmacología , Domperidona/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Ginkgo biloba , Haloperidol/farmacología , Hipocampo/embriología , Indanos/farmacología , Isoxazoles/farmacología , Neuronas/patología , Péptidos/farmacología , Piperidinas/farmacología , Extractos Vegetales/farmacología , Propilaminas/farmacología , Racloprida/farmacología , Ratas , Ratas Sprague-Dawley , Especies Reactivas de Oxígeno/metabolismo , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Remoxiprida/farmacología , Risperidona/farmacología , Sulpirida/farmacologíaRESUMEN
It is widely believed that expression of the vesicular glutamate transporter genes VGLUT1 and VGLUT2 is restricted to glutamatergic neurons and that the two transporters segregate in different sets of neurons. Using single-cell multiplex RT-PCR (sc-RT-mPCR), we show that VGLUT1 and VGLUT2 mRNAs were coexpressed in most of the sampled neurons from the rat hippocampus, cortex, and cerebellum at postnatal Day (P)14 but not P60. In accordance, changes in VGLUT1 and VGLUT2 mRNA concentrations were found to occur in these and other brain areas between P14 and P60, as revealed by semiquantitative RT-PCR and quantitated by ribonuclease protection assay. VGLUT1 and -2 coexpression in the hippocampal formation is supported further by in situ hybridization data showing that virtually all cells in the CA1-CA3 pyramidal and granule cell layers were highly positive for both transcripts until P14. It was revealed using sc-RT-mPCR that transcripts for VGLUT1 and VGLUT2 were also present in neurons of the cerebellum, striatum, and septum that expressed markers for gamma-aminobutyric acid (GABA)ergic or cholinergic phenotypes, as well as in hippocampal cells containing transcripts for the glial fibrillary acidic protein. Our study suggests that VGLUT1 and VGLUT2 proteins may often transport glutamate into vesicles within the same neuron, especially during early postnatal development, and that they are expressed widely in presumed glutamatergic, GABAergic, and cholinergic neurons, as well as in astrocytes. Furthermore, our study shows that such coexpressing neurons remain in the adult brain and identifies several areas that contain them in both young and adult rats.
Asunto(s)
Colina O-Acetiltransferasa/genética , Glutamato Descarboxilasa/genética , Hipocampo/fisiología , Proteínas de Transporte de Membrana/genética , Neuronas/fisiología , Animales , Astrocitos/fisiología , Electrofisiología , Regulación del Desarrollo de la Expresión Génica , Regulación Enzimológica de la Expresión Génica , Hipocampo/citología , Hipocampo/crecimiento & desarrollo , ARN Mensajero/análisis , Ratas , Ratas Sprague-Dawley , Proteína 1 de Transporte Vesicular de Glutamato , Proteína 2 de Transporte Vesicular de GlutamatoRESUMEN
Chronic inflammation has been reported to be a significant factor in the induction and progression of a number of chronic neurological disorders including Alzheimer's disease and Down syndrome. It is believed that inflammation may promote synaptic dysfunction, an effect that is mediated in part by pro-inflammatory cytokines such as interleukin-1beta (IL-1beta). However, the role of IL-1beta and other cytokines in synaptic transmission is still poorly understood. In this study, we have investigated how synaptic transmission and neuronal excitability in hippocampal pyramidal neurons are affected by chronic inflammation induced by exposing organotypic slices to the bacterial cell-wall product lipopolysaccharide (LPS). We report that CA1 pyramidal neurons recorded in whole cell from slices previously exposed to LPS for 7 days had resting membrane potential and action potential properties similar to those of the controls. However, they had significantly lower membrane resistance and a more elevated action potential threshold, and displayed a slower frequency of action potential discharge. Moreover, the amplitude of pharmacologically isolated postsynaptic gamma-aminobutyric acid (GABA)ergic potentials, but not excitatory glutamatergic postsynaptic potentials, was significantly larger following chronic LPS exposure. Interestingly, co-incubation of the IL-1 receptor antagonist (IL-1Ra) concurrently with LPS prevented the increase in GABAergic transmission, but not the reduction in intrinsic neuronal excitability. Finally, we confirmed that LPS dramatically increased IL-1beta, and IL-1beta-dependent IL-6 levels in the culture medium for 2 days before returning to baseline. We conclude that CA1 pyramidal neurons in slices chronically exposed to LPS show a persistent decrease in excitability due to a combined decrease in intrinsic membrane excitability and an enhancement in synaptic GABAergic input, the latter being dependent on IL-1beta. Therefore, chronic inflammation in hippocampus produces IL-1beta-dependent and -independent effects in neuronal and synaptic function that could contribute significantly to cognitive disturbances.
Asunto(s)
Potenciales de Acción/efectos de los fármacos , Hipocampo/efectos de los fármacos , Interleucina-1/metabolismo , Lipopolisacáridos/farmacología , Inhibición Neural/efectos de los fármacos , Potenciales de Acción/fisiología , Animales , Encefalitis/inmunología , Encefalitis/fisiopatología , Hipocampo/citología , Hipocampo/fisiología , Interleucina-6/metabolismo , Inhibición Neural/fisiología , Enfermedades Neurodegenerativas/inmunología , Enfermedades Neurodegenerativas/fisiopatología , Neuroinmunomodulación/efectos de los fármacos , Neuroinmunomodulación/fisiología , Técnicas de Cultivo de Órganos , Técnicas de Placa-Clamp , Células Piramidales/efectos de los fármacos , Células Piramidales/metabolismo , Células Piramidales/fisiología , Ratas , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Ácido gamma-Aminobutírico/fisiologíaRESUMEN
Dopamine neurons have been suggested to use glutamate as a cotransmitter. To identify the basis of such a phenotype, we have examined the expression of the three recently identified vesicular glutamate transporters (VGLUT1-3) in postnatal rat dopamine neurons in culture. We found that the majority of isolated dopamine neurons express VGLUT2, but not VGLUT1 or 3. In comparison, serotonin neurons express only VGLUT3. Single-cell RT-PCR experiments confirmed the presence of VGLUT2 mRNA in dopamine neurons. Arguing for phenotypic heterogeneity among axon terminals, we find that only a proportion of terminals established by dopamine neurons are VGLUT2-positive. Taken together, our results provide a basis for the ability of dopamine neurons to release glutamate as a cotransmitter. A detailed analysis of the conditions under which DA neurons gain or loose a glutamatergic phenotype may provide novel insight into pathophysiological processes that underlie diseases such as schizophrenia, Parkinson's disease and drug dependence.
Asunto(s)
Proteínas Portadoras/biosíntesis , Dopamina/metabolismo , Ácido Glutámico/metabolismo , Proteínas de Transporte de Membrana , Neuronas/metabolismo , Sinapsis/metabolismo , Proteínas de Transporte Vesicular , Sistemas de Transporte de Aminoácidos Acídicos/biosíntesis , Sistemas de Transporte de Aminoácidos Acídicos/genética , Animales , Proteínas Portadoras/genética , Células Cultivadas , Inmunohistoquímica , Mesencéfalo/citología , Mesencéfalo/metabolismo , Neuronas/citología , Técnicas de Placa-Clamp , Terminales Presinápticos/metabolismo , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Serotonina/biosíntesis , Factores de Tiempo , Tirosina 3-Monooxigenasa/genética , Proteína 1 de Transporte Vesicular de Glutamato , Proteína 2 de Transporte Vesicular de Glutamato , Proteínas de Transporte Vesicular de Glutamato , Ácido gamma-Aminobutírico/biosíntesisRESUMEN
Transforming growth factors betas (TGFbetas) are known to have important roles in neuronal survival and can be upregulated in disease. However, unlike many other trophic factors, nothing is known about the rapid neurotransmitter-like actions of TGFbeta in the CNS. We explored this by examining the effects of TGFbeta on calcium influx of large enzymatically dissociated basal forebrain neurons. We show that brief application of TGFbeta2, but not TGFbeta1, to fura-2AM-loaded neurons reversibly and acutely (within seconds) inhibited K(+)-evoked calcium influx. Moreover, using single-cell RT-PCR, we confirmed that the large TGFbeta2-responsive neurons presented a cholinergic phenotype. Investigation of the signaling mechanism underlying TGFbeta2 actions using whole-cell recordings of calcium currents revealed that TGFbeta2-mediated responses were insensitive to the nonhydrolyzable GTP analogue GTPgammaS. However, TGFbeta2-mediated calcium current reductions were prevented by intracellular perfusion of a Smad2/3 peptide antagonist. Together, these results suggest that TGFbeta2 can acutely regulate the excitability of basal forebrain cholinergic neurons through an atypical signaling mechanism.