RESUMEN
Growing evidence points to a disruption of cortico-thalamo-cortical circuits in schizophrenia (SZ) and bipolar disorder (BD). Clues for a specific involvement of the thalamic reticular nucleus (TRN) come from its unique neuronal characteristics and neural connectivity, allowing it to shape the thalamo-cortical information flow. A direct involvement of the TRN in SZ and BD has not been tested thus far. We used a combination of human postmortem and rodent studies to test the hypothesis that neurons expressing parvalbumin (PV neurons), a main TRN neuronal population, and associated Wisteria floribunda agglutinin-labeled perineuronal nets (WFA/PNNs) are altered in SZ and BD, and that these changes may occur early in the course of the disease as a consequence of oxidative stress. In both disease groups, marked decreases of PV neurons (immunoreactive for PV) and WFA/PNNs were observed in the TRN, with no effects of duration of illness or age at onset. Similarly, in transgenic mice with redox dysregulation, numbers of PV neurons and WFA/PNN+PV neurons were decreased in transgenic compared with wild-type mice; these changes were present at postnatal day (P) 20 for PV neurons and P40 for WFA/PNN+PV neurons, accompanied by alterations of their firing properties. These results show profound abnormalities of PV neurons in the TRN of subjects with SZ and BD, and offer support for the hypothesis that oxidative stress may play a key role in impacting TRN PV neurons at early stages of these disorders. We put forth that these TRN abnormalities may contribute to disruptions of sleep spindles, focused attention and emotion processing in these disorders.
Asunto(s)
Trastorno Bipolar/fisiopatología , Esquizofrenia/fisiopatología , Núcleos Talámicos/fisiopatología , Animales , Trastorno Bipolar/metabolismo , Encéfalo/fisiopatología , Femenino , Neuronas GABAérgicas/metabolismo , Hipocampo/metabolismo , Humanos , Masculino , Ratones , Ratones Noqueados , Red Nerviosa/metabolismo , Estrés Oxidativo/fisiología , Parvalbúminas/metabolismo , Parvalbúminas/fisiología , Esquizofrenia/metabolismo , Tálamo/fisiopatologíaRESUMEN
Xanthurenic acid (XA), formed from 3-hydroxykynurenine (3-HK) in the kynurenine pathway of tryptophan degradation, may modulate glutamatergic neurotransmission by inhibiting the vesicular glutamate transporter and/or activating Group II metabotropic glutamate receptors. Here we examined the molecular and cellular mechanisms by which 3-HK controls the neosynthesis of XA in rat, mouse and human brain, and compared the physiological actions of 3-HK and XA in the rat brain. In tissue homogenates, XA formation from 3-HK was observed in all three species and traced to a major role of kynurenine aminotransferase II (KAT II). Transamination of 3-HK to XA was also demonstrated using human recombinant KAT II. Neosynthesis of XA was significantly increased in the quinolinate-lesioned rat striatum, indicating a non-neuronal localization of the process. Studies using rat cortical slices revealed that newly produced XA is rapidly released into the extracellular compartment, and that XA biosynthesis can be manipulated experimentally in the same way as the production of kynurenic acid from kynurenine (omission of Na+ or glucose, depolarizing conditions, or addition of 2-oxoacids). The synthesis of XA from 3-HK was confirmed in vivo by striatal microdialysis. In slices from the rat hippocampus, both 3-HK and XA reduced the slopes of dentate gyrus field EPSPs. The effect of 3-HK was reduced in the presence of the KAT inhibitor aminooxyacetic acid. Finally, both 3-HK and XA reduced the power of gamma-oscillatory activity recorded from the hippocampal CA3 region. Endogenous XA, newly formed from 3-HK, may therefore play a physiological role in attentional and cognitive processes.
Asunto(s)
Encéfalo/citología , Encéfalo/metabolismo , Quinurenina/análogos & derivados , Xanturenatos/química , Xanturenatos/metabolismo , Anciano , Animales , Encéfalo/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Femenino , Fluoroquinolonas/farmacología , Glucosa/metabolismo , Glutamina/farmacología , Humanos , Técnicas In Vitro , Quinurenina/metabolismo , Quinurenina/farmacología , Masculino , Ratones , Persona de Mediana Edad , Piperazinas/farmacología , Cambios Post Mortem , Ácido Pirúvico/metabolismo , Ácido Quinolínico/farmacología , Ratas , Ratas Sprague-Dawley , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Tacrolimus/análogos & derivados , Tacrolimus/metabolismo , Temperatura , Distribución Tisular/efectos de los fármacos , Distribución Tisular/fisiología , Transaminasas/metabolismo , Veratridina/metabolismoRESUMEN
Astrocytes possess many of the same signalling molecules as neurons. However, the role of astrocytes in information processing, if any, is unknown. Using electrophysiological and imaging methods, we report the first evidence that astrocytes modulate neuronal sensory inhibition in the rodent thalamus. We found that mGlu2 receptor activity reduces inhibitory transmission from the thalamic reticular nucleus to the somatosensory ventrobasal thalamus (VB): mIPSC frequencies in VB slices were reduced by the Group II mGlu receptor agonist LY354740, an effect potentiated by mGlu2 positive allosteric modulator (PAM) LY487379 co-application (30 nM LY354740: 10.0 ± 1.6% reduction; 30 nM LY354740 & 30 µM LY487379: 34.6 ± 5.2% reduction). We then showed activation of mGlu2 receptors on astrocytes: astrocytic intracellular calcium levels were elevated by the Group II agonist, which were further potentiated upon mGlu2 PAM co-application (300 nM LY354740: ratio amplitude 0.016 ± 0.002; 300 nM LY354740 & 30 µM LY487379: ratio amplitude 0.035 ± 0.003). We then demonstrated mGlu2-dependent astrocytic disinhibition of VB neurons in vivo: VB neuronal responses to vibrissae stimulation trains were disinhibited by the Group II agonist and the mGlu2 PAM (LY354740: 156 ± 12% of control; LY487379: 144 ± 10% of control). Presence of the glial inhibitor fluorocitrate abolished the mGlu2 PAM effect (91 ± 5% of control), suggesting the mGlu2 component to the Group II effect can be attributed to activation of mGlu2 receptors localised on astrocytic processes within the VB. Gating of thalamocortical function via astrocyte activation represents a novel sensory processing mechanism. As this thalamocortical circuitry is important in discriminative processes, this demonstrates the importance of astrocytes in synaptic processes underlying attention and cognition.
Asunto(s)
Astrocitos/fisiología , Células Receptoras Sensoriales/fisiología , Tálamo/citología , Vibrisas/fisiología , Animales , Animales Recién Nacidos , Astrocitos/efectos de los fármacos , Calcio/metabolismo , Citratos/farmacología , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Fármacos actuantes sobre Aminoácidos Excitadores/farmacología , Técnicas In Vitro , Iontoforesis , Masculino , N-Metilaspartato/farmacología , Ratas , Ratas Sprague-Dawley , Bloqueadores de los Canales de Sodio/farmacología , Tetrodotoxina/farmacología , Valina/análogos & derivados , Valina/farmacologíaRESUMEN
The mediodorsal thalamus (MD) likely plays an important role in cognition as it receives abundant afferent connections from the amygdala and prefrontal cortex (PFC). Indeed, disturbed activity within the MD is thought to precipitate cognitive deficits associated with schizophrenia. As compounds acting at the Group II metabotropic glutamate (mGlu) receptors (subtypes mGlu2/mGlu3) have efficacy in animal models of schizophrenia, we investigated whether a Group II agonist and an mGlu2 positive allosteric modulator (PAM) could modulate MD activity. Extracellular single-unit recordings were made in vivo from MD neurones in anaesthetised rats. Responses were elicited by electrical stimulation of the PFC and/or amygdala, with Group II compounds locally applied as required. The Group II agonist reduced inhibition evoked in the MD: an effect manifested as an increase in short-latency responses, and a decrease in long-latency burst-firing. This disinhibitory action of the Group II receptors in the MD represents a mechanism of potential therapeutic importance as increased inhibition in the MD has been associated with cognitive deficit-onset. Furthermore, as co-application of the mGlu2 PAM did not potentiate the Group II agonist effects in the MD, we suggest that the Group II disinhibitory effect is majority-mediated via mGlu3. This heterogeneity in Group II receptor thalamic physiology bears consequence, as compounds active exclusively at the mGlu2 subtype are unlikely to perturb maladapted MD firing patterns associated with cognitive deficits, with activity at mGlu3 receptors possibly more appropriate. Indeed, polymorphisms in the mGlu3, but not the mGlu2, gene have been detected in patients with schizophrenia.
Asunto(s)
Potenciales de Acción/fisiología , Cognición/fisiología , Núcleo Talámico Mediodorsal/citología , Red Nerviosa/fisiología , Neuronas/fisiología , Receptores de Glutamato Metabotrópico/metabolismo , Potenciales de Acción/efectos de los fármacos , Animales , Biofisica , Cognición/efectos de los fármacos , Estimulación Eléctrica , Fármacos actuantes sobre Aminoácidos Excitadores/farmacología , Iontoforesis , Masculino , Red Nerviosa/efectos de los fármacos , Inhibición Neural/efectos de los fármacos , Vías Nerviosas/efectos de los fármacos , Vías Nerviosas/fisiología , Neuronas/efectos de los fármacos , Estimulación Física , Ratas , Ratas Wistar , Tiempo de Reacción/efectos de los fármacos , Vibrisas/inervación , Ácido gamma-Aminobutírico/farmacologíaRESUMEN
Several lines of evidence suggest that there are similarities in the pathomechanisms of glaucoma and Alzheimer's disease, and that amyloid-beta (Aß) could be a new, promising target for neuroprotective therapy of glaucoma. In the present study, we evaluated the effect of the Aß aggregation modulator MRZ-99030 in the Morrison model of glaucoma based on increased intraocular pressure (IOP) in rats. MRZ-99030 provided dose-dependent neuroprotection and at the highest dose (240 mg/kg) reduced the degree of RGC apoptosis to 33 % of that seen after vehicle (P < 0.05; one-way ANOVA). No significant effect on IOP was observed. Pharmacokinetic experiments showed that following systemic injection of MRZ-99030, concentrations above affinity for Aß were reached. Hence the present results are consistent with the notion that Aß is a promising target for neuroprotective intervention in glaucoma and that MRZ-99030 may be a good drug candidate for further development.
Asunto(s)
Dipéptidos/farmacología , Glaucoma/tratamiento farmacológico , Fármacos Neuroprotectores/farmacología , Animales , Apoptosis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Dipéptidos/farmacocinética , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Glaucoma/patología , Glaucoma/fisiopatología , Presión Intraocular/efectos de los fármacos , Masculino , Fármacos Neuroprotectores/farmacocinética , Ratas , Retina/efectos de los fármacos , Retina/patología , Retina/fisiopatologíaRESUMEN
As postsynaptic metabotropic subtype 1 (mGlu1) receptors are present in the thalamus, we have investigated the effect of potentiating and antagonising mGlu1 receptors on responses of thalamic neurones to noxious sensory stimulation. Extracellular recordings were made in vivo with multi-barrel iontophoretic electrodes from single neurones in the thalamus of urethane-anaesthetised rats. Responses to iontophoretic applications of the Group I mGlu agonist 3,5-dihydroxy-phenylglycine (DHPG) were selectively potentiated by co-application of the mGlu1 positive allosteric modulator Ro67-4853, whereas they were selectively reduced upon co-application of the mGlu1 receptor orthosteric antagonist LY367385. This indicates that thalamic DHPG responses are mediated primarily via mGlu1 receptors, consistent with the high postsynaptic levels of this receptor in the thalamus. Furthermore, potentiation of DHPG responses by Ro67-4853 were greater when the initial DHPG response was of a low magnitude. Ro67-4853 also potentiated responses of thalamic neurones to noxious thermal stimulation, whilst having little effect on the baseline activity of nociceptive neurones. By contrast, nociceptive responses were reduced by LY367385. In a further series of experiments we found that inactivation of somatosensory cortex by cooling resulted in a reduction of thalamic nociceptive responses. These results underline the importance of mGlu1 receptors in the processing of sensory information in the thalamus, particularly with respect to nociceptive responses. Furthermore, the involvement of mGlu1 receptors may reflect the activity of descending cortico-thalamic afferents.
Asunto(s)
Neuronas/fisiología , Nocicepción/fisiología , Receptores de Glutamato Metabotrópico/metabolismo , Tálamo/fisiología , Potenciales de Acción/efectos de los fármacos , Animales , Benzoatos/farmacología , Carbamatos/farmacología , Frío , Fármacos actuantes sobre Aminoácidos Excitadores/farmacología , Agonistas de Aminoácidos Excitadores/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Glicina/análogos & derivados , Glicina/farmacología , Calor , Masculino , Vías Nerviosas/fisiopatología , Neuronas/efectos de los fármacos , Nocicepción/efectos de los fármacos , Ratas , Ratas Wistar , Receptores de Glutamato Metabotrópico/agonistas , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Resorcinoles/farmacología , Corteza Somatosensorial/fisiopatología , Tálamo/efectos de los fármacos , Xantenos/farmacologíaRESUMEN
Vesicular glutamate transporters (VGLUTs) are known to be important in the uptake of glutamate into vesicles in the presynaptic terminal; thereby playing a role in synaptic function. VGLUT dysfunction has also been suggested in neurological and psychiatric disorders such as epilepsy and schizophrenia. A number of compounds have been identified as VGLUT inhibitors; however, little is known as to how these compounds affect synaptic transmission. We therefore investigated the effects of structurally unrelated VGLUT inhibitors on synaptic transmission in the rodent hippocampus and prefrontal cortex. In the CA1 and dentate gyrus regions of the in vitro slice preparation of mouse hippocampus, AMPA receptor-mediated field excitatory postsynaptic potentials (fEPSPs) were evoked in response to Schaffer collateral/commissural pathway stimulation. Application of the VGLUT inhibitors Rose Bengal (RB), Congo Red (CR) or Chicago Sky Blue 6B (CB) resulted in a concentration-related reduction of fEPSP amplitudes. RB (30µM) or CB (300µM) also depressed NMDA receptor-mediated responses in the CA1 region. The naturally occurring kynurenine Xanthurenic Acid (XA) is reported to be a VGLUT inhibitor. We found XA attenuated both AMPA and NMDA receptor-mediated synaptic transmission. The potency order of the VGLUT inhibitors was consistent with literature Ki values for VGLUT inhibition. Impaired glutamatergic neurotransmission is believed to contribute to schizophrenia, and VGLUTs have also been implicated in this disease. We therefore investigated the effect of VGLUT inhibition in the prefrontal cortex. Application of the VGLUT inhibitors RB or CB resulted in a concentration-dependent reduction in the amplitude of glutamate receptor-mediated fEPSPs recorded in layer V/VI in response to stimulation in the forceps minor. We conclude that VGLUT inhibitors can modulate glutamatergic synaptic transmission in the PFC and hippocampus. This could be important in the pathophysiology of nervous system disorders and might represent a target for developing novel pharmacological therapies.
Asunto(s)
Sistema de Transporte de Aminoácidos X-AG/antagonistas & inhibidores , Ácido Glutámico/fisiología , Hipocampo/fisiología , Corteza Prefrontal/fisiología , Transmisión Sináptica/efectos de los fármacos , 2-Amino-5-fosfonovalerato/farmacología , Animales , Región CA1 Hipocampal/efectos de los fármacos , Giro Dentado/efectos de los fármacos , Antagonistas de Aminoácidos Excitadores/farmacología , Femenino , Hipocampo/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Corteza Prefrontal/efectos de los fármacos , Quinoxalinas/farmacologíaRESUMEN
The feedback connections from the cortical middle temporal (MT) motion area, to layer 6 of the primary visual cortex (V1), have the capacity to drive a cascaded feedback influence from the layer 6 cortico-geniculate cells back to the lateral geniculate nucleus (LGN) relay cells. This introduces the possibility of a re-entrant motion signal affecting the relay of the retinal input through the LGN to the visual cortex. The question is whether the response of LGN cells to moving stimuli involves a component derived from this feedback. By producing a reversible focal pharmacological block of the activity of an MT direction column we show the presence of such an influence from MT on the responses of magno, parvo and koniocellular cells in the macaque LGN. The pattern of effect in the LGN reflects the direction bias of the MT location inactivated. This suggests a moving stimulus is captured by iterative interactions in the circuit formed by visual cortical areas and visual thalamus.
Asunto(s)
Retroalimentación Fisiológica/fisiología , Cuerpos Geniculados/fisiología , Percepción de Movimiento/fisiología , Estimulación Luminosa/métodos , Corteza Visual/fisiología , Animales , Femenino , Macaca mulattaRESUMEN
Xanthurenic acid (XA), an endogenous kynurenine, is a known vesicular glutamate transport (VGLUT) inhibitor and has also been proposed as an mGlu2/3 receptor agonist. Changes in these systems have been implicated in the pathophysiology of schizophrenia and other psychiatric disorders; however, little is known of how XA affects synaptic transmission. We therefore investigated the effects of XA on synaptic transmission at two hippocampal glutamatergic pathways and evaluated the ability of XA to bind to mGlu2/3 receptors. Field excitatory postsynaptic potentials (fEPSPs) were recorded from either the dentate gyrus (DG) or CA1 region of mouse hippocampal slices in vitro. Addition of XA to the bathing medium (1-10 mM) resulted in a dose-related reduction of fEPSP amplitudes (up to 52% reduction) in both hippocampal regions. In the DG, the VGLUT inhibitors Congo Red and Rose Bengal, and the mGlu2/3 agonist LY354740, also reduced fEPSPs (up to 80% reduction). The mGlu2/3 antagonist LY341495 reversed the LY354740 effect, but not the XA effect. LY354740, but not XA, also reduced DG paired-pulse depression. XA had no effect on specific binding of 1 nM [(3)H]LY341495 to membranes with human mGlu2 receptors. We conclude that XA can modulate synaptic transmission via a mechanism that may involve VGLUT inhibition rather than activation of mGlu2/3 receptors. This could be important in the pathophysiology of nervous system disorders including schizophrenia and might represent a target for developing novel pharmacological therapies.
Asunto(s)
Hipocampo/metabolismo , Quinurenina/fisiología , Transmisión Sináptica/fisiología , Proteínas de Transporte Vesicular de Glutamato/antagonistas & inhibidores , Proteínas de Transporte Vesicular de Glutamato/fisiología , Xanturenatos/farmacología , Animales , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Hipocampo/efectos de los fármacos , Humanos , Ratones , Ratones Endogámicos C57BL , Técnicas de Cultivo de Órganos , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Transmisión Sináptica/efectos de los fármacosRESUMEN
Xanthurenic acid (XA), a molecule arising from tryptophan metabolism by transamination of 3-hydroxykynurenine, has recently been identified as an endogenous Group II (mGlu2 and mGlu3) metabotropic glutamate (mGlu) receptor ligand in vitro. Impairments in Group II mGlu receptor expression and function have been implicated in the pathophysiology of schizophrenia, as have multiple steps in the kynurenine metabolism pathway. Therefore, we examined XA in vivo to further investigate its potential as a Group II mGlu receptor ligand using a preparation that has been previously demonstrated to efficiently reveal the action of other Group II mGlu receptor ligands in vivo. Extracellular single-neurone recordings were made in the rat ventrobasal thalamus (VB) in conjunction with iontophoresis of agonists, an antagonist and a positive allosteric modulator and/or intravenous (i.v.) injection of XA. We found the XA effect on sensory inhibition, when applied iontophoretically and i.v., was similar to that of other Group II mGlu receptor agonists in reducing inhibition evoked in the VB from the thalamic reticular nucleus upon physiological sensory stimulation. Furthermore, we postulate that XA may be the first potential endogenous allosteric agonist (termed 'endocoid') for the mGlu receptors. As the Group II receptors and kynurenine metabolism pathway have both been heavily implicated in the pathophysiology of schizophrenia, XA could play a pivotal role in antipsychotic research as this potential endocoid represents both a convergence within these two biological parameters and a novel class of Group II mGlu receptor ligand. This article is part of a Special Issue entitled 'Metabotropic Glutamate Receptors'.
Asunto(s)
Vías Aferentes/efectos de los fármacos , Vías Aferentes/fisiología , Agonistas de Aminoácidos Excitadores/farmacología , Núcleos Talámicos Ventrales/efectos de los fármacos , Núcleos Talámicos Ventrales/fisiología , Vibrisas/fisiología , Xanturenatos/farmacología , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Administración Intravenosa , Regulación Alostérica/fisiología , Aminoácidos/farmacología , Animales , Compuestos Bicíclicos con Puentes/farmacología , Interacciones Farmacológicas , Antagonistas de Aminoácidos Excitadores/farmacología , Iontoforesis , Masculino , Inhibición Neural/efectos de los fármacos , Inhibición Neural/fisiología , Neuronas/fisiología , Ratas , Ratas Wistar , Xantenos/farmacología , Xanturenatos/administración & dosificación , Xanturenatos/agonistas , Xanturenatos/antagonistas & inhibidoresRESUMEN
Group II metabotropic glutamate receptor (mGlu) modulation of sensory processing in the rat ventrobasal thalamic nucleus (VB) has been extensively studied in vivo. However, it is not yet known what the relative contributions are of the Group II mGlu receptor subtypes (mGlu2 and mGlu3) to this modulation, nor to what extent these receptors may be activated under physiological conditions during this process. Using single-neurone recording in the rat VB in vivo with local application of the selective Group II agonist LY354740 and the subtype selective mGlu2 positive allosteric modulator (PAM) LY487379, our findings were twofold. Firstly, we found that there is an mGlu2 component to the effects of LY354740 on sensory responses in the VB. Secondly, we have demonstrated that application of the PAM alone can modulate sensory responses of single neurones in vivo. This indicates that mGlu2 receptors can be activated by endogenous agonist following physiological sensory stimulation. We speculate that the mGlu2 subtype could be activated under physiological stimulus-evoked conditions by 'glutamate spillover' from synapses between excitatory sensory afferents and VB neurones that can lead to a reduction in sensory-evoked inhibition arising from the thalamic reticular nucleus (TRN). We propose that this potential mGlu2 receptor modulation of inhibition could play an important role in discerning relevant information from background activity upon physiological sensory stimulation. Furthermore, this could be a site of action for mGlu2 PAMs to modulate cognitive processes.
Asunto(s)
Receptores de Glutamato Metabotrópico/fisiología , Sensación/fisiología , Tálamo/fisiología , Animales , Compuestos Bicíclicos con Puentes/farmacología , Agonistas de Aminoácidos Excitadores/farmacología , Masculino , Estimulación Física , Piridinas/farmacología , Ratas , Ratas Wistar , Células Receptoras Sensoriales/fisiología , Sulfonamidas/farmacología , Tálamo/efectos de los fármacos , Vibrisas/fisiologíaRESUMEN
Metabotropic glutamate subtype 1 (mGlu1) receptor is thought to play a role in synaptic responses in thalamic relay nuclei. The aim of this study was to evaluate the positive allosteric modulator (PAM) Ro67-4853 as a tool to modulate thalamic mGlu1 receptors on single thalamic neurones in vivo. Ro67-4853, applied by iontophoresis onto ventrobasal thalamus neurones of urethane-anaesthetised rats, selectively enhanced responses to the agonist (S)-3,5-dihydroxy-phenylglycine (DHPG), an effect consistent with mGlu1 potentiation. The PAM was also able to enhance maintained responses to 10 Hz trains of sensory stimulation of the vibrissae, but had little effect on responses to single sensory stimuli. Thus Ro67-4853 appears to be a highly selective tool that can be useful in investigating how mGlu1 receptor potentiation can alter neural processing in vivo. Our results show the importance of mGlu1 in sensory processing and attention mechanisms at the thalamic level and suggest that positive modulation of mGlu1 receptors might be a useful mechanism for enhancing cognitive and attentional processes.
Asunto(s)
Carbamatos/farmacología , Neuronas Aferentes/efectos de los fármacos , Neuronas/efectos de los fármacos , Receptores de Glutamato Metabotrópico/metabolismo , Tálamo/efectos de los fármacos , Percepción del Tacto/efectos de los fármacos , Xantenos/farmacología , Animales , Masculino , Neuronas/metabolismo , Neuronas Aferentes/fisiología , Ratas , Ratas Wistar , Tálamo/metabolismo , Percepción del Tacto/fisiología , Vibrisas/fisiologíaRESUMEN
In the age-related, blinding disease glaucoma, retinal ganglion cells (RGCs) degenerate, possibly affecting glutamatergic retinofugal transmission to the brain. The superior colliculus (SC) is a major central target of retinofugal axons in the rodent, a much used disease model. We investigated the contribution of NMDA-type glutamate receptors to retinocollicular transmission in a rat glaucoma model, using a SC brain slice preparation to determine the sensitivity of synaptic responses to the NMDAR antagonist D-AP5. At 32weeks after induction of experimental glaucoma, but not earlier, there was an increase in NMDAR contribution to SC synaptic responses in slices receiving input from glaucomatous eyes. This suggests that there are changes in NMDAR function after RGC degeneration in experimental glaucoma, which may represent functional SC compensation through plasticity via NMDARs. This has implications for studies carried out using rodent glaucoma models, especially those evaluating potential treatment strategies, as it suggests that functional changes in the central visual system need to be considered in addition to those in the eye. Furthermore, the data underline the need for early therapeutic intervention in order to pre-empt subsequent central functional changes.
Asunto(s)
Glaucoma/fisiopatología , Receptores de N-Metil-D-Aspartato/metabolismo , Células Ganglionares de la Retina/fisiología , Colículos Superiores/fisiopatología , Transmisión Sináptica/fisiología , Vías Visuales/fisiopatología , Análisis de Varianza , Animales , Electrofisiología , Potenciales Postsinápticos Excitadores/fisiología , Glaucoma/metabolismo , Masculino , Ratas , Colículos Superiores/metabolismo , Sinapsis/metabolismo , Vías Visuales/metabolismoRESUMEN
Retinal ganglion cell (RGC) death in glaucoma models is associated with accumulation of activated microglia, a sign of neural degeneration which has been shown to constitute a barrier for transplant cell survival and migration. This study investigated the use of triamcinolone (TA) to control macrophage/microglia accumulation in a model of RGC depletion to create a permissive environment for stem cell grafting. Intravitreal NMDA alone or in combination with TA was used to induce rapid onset of RGC death in 3-4 week old Lister hooded (LH) rat eyes prior to Müller stem cell transplantation into the vitreoretinal space. The effect of NMDA on RGC death and microglial accumulation was assessed immuno-histochemically, whilst electroretinography (ERG) was used to assess RGC function. Post transplantation, survival of grafted cells and their association with microglia were also examined by immunohistochemical methods. Intravitreal injection of NMDA alone resulted in severe macrophage/microglia accumulation associated with extensive RGC death 4-7 days post-treatment. Combination of NMDA with TA significantly reduced microglial numbers in the RGC when compared to NMDA only treated eyes while still depleting the retina of RGC. At the same time, NMDA/TA treatment also caused functional RGC loss as demonstrated by reduction of the scotopic threshold response. Upon transplantation with Müller stem cells, NMDA/TA treatment caused significant reduction in the number of transplant associated macrophage/microglia compared to eyes treated with NMDA alone. On this basis it is proposed that intravitreal injection of TA may be useful as an effective anti-inflammatory agent to control macrophage/microglia accumulation induced by RGC death, thereby creating a retinal environment permissive to cell transplantation studies.
Asunto(s)
Antiinflamatorios/farmacología , Supervivencia de Injerto/fisiología , Macrófagos/metabolismo , Microglía/metabolismo , N-Metilaspartato/toxicidad , Células Ganglionares de la Retina/patología , Trasplante de Células Madre , Triamcinolona Acetonida/farmacología , Animales , Apoptosis/efectos de los fármacos , Biomarcadores/metabolismo , Supervivencia Celular , Electrorretinografía , Técnica del Anticuerpo Fluorescente Indirecta , Hibridación Fluorescente in Situ , Etiquetado Corte-Fin in Situ , Inyecciones , Microglía/citología , Ratas , Células Ganglionares de la Retina/metabolismo , Cuerpo VítreoRESUMEN
Nerve cell death is the key event in all neurodegenerative disorders, with apoptosis and necrosis being central to both acute and chronic degenerative processes. However, until now, it has not been possible to study these dynamically and in real time. In this study, we use spectrally distinct, well-recognised fluorescent cell death markers to enable the temporal resolution and quantification of the early and late phases of apoptosis and necrosis of single nerve cells in different disease models. The tracking of single-cell death profiles in the same living eye over hours, days, weeks and months is a significant advancement on currently available techniques. We identified a numerical preponderance of late-phase versus early-phase apoptotic cells in chronic models, reinforcing the commonalities between cellular mechanisms in different disease models. We showed that MK801 effectively inhibited both apoptosis and necrosis, but our findings support the use of our technique to investigate more specific anti-apoptotic and anti-necrotic strategies with well-defined targets, with potentially greater clinical application. The optical properties of the eye provide compelling opportunities for the quantitative monitoring of disease mechanisms and dynamics in experimental neurodegeneration. Our findings also help to directly observe retinal nerve cell death in patients as an adjunct to refining diagnosis, tracking disease status and assessing therapeutic intervention.
Asunto(s)
Apoptosis , Enfermedades Neurodegenerativas/diagnóstico , Enfermedad de Alzheimer/patología , Péptidos beta-Amiloides/metabolismo , Animales , Modelos Animales de Enfermedad , Maleato de Dizocilpina/farmacología , Ratones , Necrosis , Células Ganglionares de la Retina/metabolismo , Células Ganglionares de la Retina/patologíaRESUMEN
Photoreceptor loss causes irreversible blindness in many retinal diseases. Repair of such damage by cell transplantation is one of the most feasible types of central nervous system repair; photoreceptor degeneration initially leaves the inner retinal circuitry intact and new photoreceptors need only make single, short synaptic connections to contribute to the retinotopic map. So far, brain- and retina-derived stem cells transplanted into adult retina have shown little evidence of being able to integrate into the outer nuclear layer and differentiate into new photoreceptors. Furthermore, there has been no demonstration that transplanted cells form functional synaptic connections with other neurons in the recipient retina or restore visual function. This might be because the mature mammalian retina lacks the ability to accept and incorporate stem cells or to promote photoreceptor differentiation. We hypothesized that committed progenitor or precursor cells at later ontogenetic stages might have a higher probability of success upon transplantation. Here we show that donor cells can integrate into the adult or degenerating retina if they are taken from the developing retina at a time coincident with the peak of rod genesis. These transplanted cells integrate, differentiate into rod photoreceptors, form synaptic connections and improve visual function. Furthermore, we use genetically tagged post-mitotic rod precursors expressing the transcription factor Nrl (ref. 6) (neural retina leucine zipper) to show that successfully integrated rod photoreceptors are derived only from immature post-mitotic rod precursors and not from proliferating progenitor or stem cells. These findings define the ontogenetic stage of donor cells for successful rod photoreceptor transplantation.
Asunto(s)
Tratamiento Basado en Trasplante de Células y Tejidos , Células Fotorreceptoras de Vertebrados/citología , Células Fotorreceptoras de Vertebrados/trasplante , Retina/citología , Retina/patología , Trasplante de Células Madre , Animales , Diferenciación Celular , Proliferación Celular , Supervivencia Celular , Pollos/genética , Luz , Ratones , Células Fotorreceptoras de Vertebrados/efectos de la radiación , Retina/embriología , Retina/efectos de la radiación , Degeneración Retiniana/patología , Degeneración Retiniana/terapia , Sinapsis/metabolismo , Factores de TiempoRESUMEN
Previous work has indicated that metabotropic glutamate receptors (mGluRs) modulate visual responses of superior colliculus (SC) neurones in vivo in a variety of ways, in a manner that can be dependent upon visual stimulus properties. How this occurs remains unclear. In this study we aimed to determine how activation of mGluR2 and mGluR3 receptors (Group II) might modulate visual responses, by using field potential and whole-cell patch clamp recording techniques in rat SC slice. Stimulation within the superficial layers of the SC, in the presence of ionotropic glutamate receptor antagonists, evoked IPSCs that were blocked by bicuculline indicating that they are mediated via GABAA receptors. It is likely that these IPSCs were of heterogeneous origin as they showed substantial variation in paired-pulse behaviour. Nevertheless, activation of Group II mGluRs with the group-selective agonist LY354740 (300 nM, bath application) resulted in a reduction of these IPSCs (to 56% of control amplitude), and this was associated with a decrease in paired-pulse depression. At the same concentration, LY354740 did not reduce the EPSC or field-EPSP evoked by stimulation of the retinal input to the SC. The effects of LY354740 on IPSCs were not mimicked by the mGluR3-selective agonist N-acetyl-aspartyl-glutamate (NAAG, 200-500 microM). Stimulation of IPSCs with trains of impulses (10 at 20 Hz) in order to mimic natural activation patterns resulted in sequences of IPSCs that were reduced in amplitude towards the end of the stimulus train. Application of the Group II antagonist LY341495 (100 nM) under these conditions resulted in an increase in later IPSCs in a third of neurones tested. These findings indicate that mGluR2 (but not mGluR3) can selectively modulate GABAergic inhibition in SC, probably via a presynaptic mechanism. Furthermore, these receptors may be activated by synaptically released transmitter during patterns of activation similar to those seen during visual processing. Thus mGluR2 receptors could have a function in activity-dependent modulation of inhibitory processing during visual responses.
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Inhibición Neural , Receptores de Glutamato Metabotrópico/metabolismo , Receptores Presinapticos/metabolismo , Colículos Superiores/metabolismo , Transmisión Sináptica , Vías Visuales/metabolismo , Ácido gamma-Aminobutírico/metabolismo , Animales , Bicuculina/farmacología , Compuestos Bicíclicos con Puentes/farmacología , Dipéptidos/farmacología , Relación Dosis-Respuesta a Droga , Estimulación Eléctrica , Agonistas de Aminoácidos Excitadores/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Antagonistas del GABA/farmacología , Antagonistas de Receptores de GABA-A , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Inhibición Neural/efectos de los fármacos , Técnicas de Placa-Clamp , Ratas , Ratas Endogámicas , Receptores de GABA-A/metabolismo , Receptores de Glutamato Metabotrópico/efectos de los fármacos , Receptores Presinapticos/efectos de los fármacos , Colículos Superiores/citología , Colículos Superiores/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Factores de Tiempo , Vías Visuales/citología , Vías Visuales/efectos de los fármacosRESUMEN
The local synaptic connectivity in the superficial gray layer of the superior colliculus (SC) was assessed following retinal ganglion cell axonal regeneration through a peripheral nerve graft into the SC of Lister Hooded rats, using in vitro brain slice techniques. Repair was effected between the ipsilateral eye and SC, following bilateral lesion of optic nerves and ablation of ipsilateral occipital cortex. Deafferentation surgery alone resulted in a complete loss of synaptic potentials of extrinsic origin, once both retinal and cortical inputs were removed. Stimulation of graft insertion sites elicited synaptic responses comprising monosynaptic and network-mediated depolarising events. This activity, together with similar spontaneous bursts of depolarising events and action potential firing, was generated by the activation of non-N-methyl-D-aspartate glutamate receptors. This behaviour may reflect the development of a local recurrent synaptic connectivity following the repair surgery, as both evoked and spontaneous responses developed into large long-lasting bursts of excitatory activity when inhibition mediated by GABA receptors was blocked. These results suggest that the ultrastructural changes in the superficial layers of the SC resulting from deafferentation are reflected functionally at the synaptic level in the target structure even after repair. Such changes are likely to compromise the ability of the target structure to function normally during information processing. Therefore, although axons regenerating along peripheral nerve grafts can make functional synaptic connections, their efficacy in activating the target structure will probably be compromised by local changes in synaptic connectivity.
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Red Nerviosa/fisiología , Neuronas/fisiología , Regeneración/fisiología , Colículos Superiores/citología , Transmisión Sináptica/fisiología , Vías Visuales/fisiología , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Potenciales de Acción/efectos de la radiación , Animales , Animales Recién Nacidos , Axones/fisiología , Axotomía/métodos , Estado de Descerebración , Interacciones Farmacológicas , Estimulación Eléctrica/métodos , Antagonistas de Aminoácidos Excitadores/farmacología , Lateralidad Funcional/fisiología , Antagonistas del GABA/farmacología , Técnicas In Vitro , Neuronas/efectos de los fármacos , Neuronas/efectos de la radiación , Nervios Periféricos/anatomía & histología , Nervios Periféricos/trasplante , Ácidos Fosfínicos/farmacología , Picrotoxina/farmacología , Propanolaminas/farmacología , Quinoxalinas/farmacología , Ratas , Células Ganglionares de la Retina/patología , Células Ganglionares de la Retina/fisiología , Trasplantes , Corteza Visual/fisiología , Vías Visuales/lesiones , Vías Visuales/fisiopatologíaRESUMEN
Group III metabotropic glutamate receptors (especially mGlu4, mGlu7, mGlu8) are thought to be involved in modulating visual processing in the adult superior colliculus, a major termination site of retinal input in the rodent brain. We have investigated this role by making field EPSP recordings in response to optic tract stimulation in superior colliculus slices taken from rats aged from P14 to P180. Application of the Group III agonist L-AP4 at a concentration (10 microM) effective to activate mGlu4 and mGlu8 receptors, but not mGlu7 receptors, resulted in reductions of the field EPSP in all ages, although the effect was greatest in slices taken from P14 rats. Increasing the L-AP4 concentration to 100 microM so as to also activate mGlu7 receptors resulted in further field EPSP reductions. Similar reductions were seen in the combined presence of the GABA antagonists picrotoxin and CGP55845A, indicating a lack of involvement of GABAergic mechanisms in the action of L-AP4. Pairing of optic tract stimuli (20 ms separation) resulted in paired-pulse depression at all ages. L-AP4 was found to reduce paired-pulse depression at both 10 microM and 100 microM in slices from all ages of rat. The results of this study suggest that mGlu4/mGlu8 and mGlu7 receptors modulate retino-tectal transmission via a presynaptic mechanism, and that these effects are greatest in young animals. This is the first demonstration of a functional change in Group III receptor effect with aging, and this would be consistent with developmental regulation of these receptors.