RESUMEN
BACKGROUND: The effects of Alzheimer's disease (AD) pathology on the experience of pain are poorly understood. OBJECTIVE: To understand the pathophysiological mechanisms underlying pain sensory transmission in the transgenic mouse model of AD, CRND8. METHODS: We explored AD-related pathology in the spinal cord and dorsal root ganglia of 18-week-old female CRND8 mice. We assessed nociceptive responses to both acute heat stimuli and persistent inflammatory pain in CRND8 mice and non-transgenic (non-Tg) littermates. In addition, we searched for differences in biochemical correlates of inflammatory pain between CRND8 and non-Tg mice. Finally, we investigated the excitability of dorsal horn noc iceptive neurons in spinal cord slices from CRND8 and non-Tg mice. RESULTS: We demonstrated the presence of intracellular AD-like pathology in the spinal cord and in the dorsal root ganglia nociceptive sensory neurons of CRND8 mice. We found that CRND8 mice had a reduced susceptibility to acute noxious heat stimuli and an increased sensitivity to tonic inflammatory pain. Tonic inflammatory pain correlated with a lack of induction of pro-opiomelanocortin in the spinal cord of CRND8 mice as compared to non-Tg mice. Electrophysiological recording in acute spinal cord slice preparations indicated an increased probability of glutamate release at the membrane of dorsal horn nociceptive neurons in CRND8 mice. CONCLUSION: This study suggests that an increased thermal tolerance and a facilitation of nociception by peripheral inflammation can coexist in AD.
Asunto(s)
Enfermedad de Alzheimer , Hiperalgesia , Ratones , Femenino , Animales , Enfermedad de Alzheimer/complicaciones , Calor , Dolor/etiología , Ratones TransgénicosRESUMEN
Microglia represent the resident immune system in the brain. They mediate neuroinflammatory processes and have been described as important regulators of homeostasis in the central nervous system (CNS). Among several players and mechanisms contributing to microglial function in inflammation, ATP and glutamate have been shown to be involved in microgliosis. In this study, we focused on receptor subtypes that respond to these neurotransmitters, purinergic ionotropic P2X7 receptor and metabotropic glutamate mGlu5 receptor. We found that both receptors are functionally expressed in a murine microglia cell line, BV2 cells, and we performed patch-clamp experiments to measure purinergic ionotropic P2X7 receptor ion flux in control condition and after metabotropic glutamate mGlu5 receptor activation. The selective purinergic ionotropic P2X7 receptor agonist, 2'(3')-O-(4-benzoylbenzoyl)adenosine-5'-triphosphate (BzATP, 100 µM), elicited a robust current that was prevented by the selective purinergic ionotropic P2X7 receptor antagonist A438079 (10 µM). When BV2 cells were acutely stimulated with the selective metabotropic glutamate mGlu5 agonist, (RS)-2-chloro-5-hydroxyphenylglycine (CHPG, 200 µM), purinergic ionotropic P2X7 receptor current was increased. This positive modulation was prevented by the selective metabotropic glutamate mGlu5 receptor antagonist 3-((2-Methyl-4-thiazolyl)ethynyl)pyridine (MTEP, 1 µM). Moreover, nitric oxide synthesis elicited by purinergic ionotropic P2X7 receptor activation was enhanced by metabotropic glutamate mGlu5 receptor co-stimulation. Taken together, our results suggest an important crosstalk between ATP and glutamate in inflammation. Pro-inflammatory effects mediated by purinergic ionotropic P2X7 receptor might be exacerbated by simultaneous exposure of microglia to ATP and glutamate, suggesting new pharmacological targets to modulate neuroinflammation.
Asunto(s)
Microglía , Receptor del Glutamato Metabotropico 5 , Receptores Purinérgicos P2X7 , Animales , Ratones , Adenosina Trifosfato/farmacología , Células Cultivadas , Agonistas de Aminoácidos Excitadores , Ácido Glutámico/metabolismo , Inflamación/metabolismo , Receptor del Glutamato Metabotropico 5/metabolismo , Receptores Purinérgicos P2X7/metabolismoRESUMEN
Fragile X Syndrome is a genetic form of intellectual disability associated with autism, epilepsy and mood disorders. Electrophysiology studies in Fmr1 knockout (KO) mice, a murine model of Fragile X Syndrome, have demonstrated alterations of synaptic plasticity, with exaggerated long-term depression induced by activation of metabotropic glutamate receptors (mGluR-LTD) in Fmr1 KO hippocampus. We have previously demonstrated that activation of serotonin 5-HT7 receptors reverses mGluR-LTD in the hippocampus of wild-type and Fmr1 KO mice, thus correcting a synaptic dysfunction typically observed in this disease model. Here we show that pharmacological inhibition of cyclin-dependent kinase 5 (Cdk5, a signaling molecule recently shown to be a modulator of brain synaptic plasticity) enhanced mGluR-LTD in wild-type hippocampal neurons, which became comparable to exaggerated mGluR-LTD observed in Fmr1 KO neurons. Furthermore, Cdk5 inhibition prevented 5-HT7 receptor-mediated reversal of mGluR-LTD both in wild-type and in Fmr1 KO neurons. Our results show that Cdk5 modulates hippocampal synaptic plasticity. 5-HT7 receptors require Cdk5 to modulate synaptic plasticity in wild-type and rescue abnormal plasticity in Fmr1 KO neurons, pointing out Cdk5 as a possible novel target in Fragile X Syndrome.
RESUMEN
Mitochondria in neurons contribute to energy supply, the regulation of synaptic transmission, Ca2+ homeostasis, neuronal excitability, and stress adaptation. In recent years, several studies have highlighted that the neurotransmitter serotonin (5-HT) plays an important role in mitochondrial biogenesis in cortical neurons, and regulates mitochondrial activity and cellular function in cardiomyocytes. 5-HT exerts its diverse actions by binding to cell surface receptors that are classified into seven distinct families (5-HT1 to 5-HT7). Recently, it was shown that 5-HT3 and 5-HT4 receptors are located on the mitochondrial membrane and participate in the regulation of mitochondrial function. Furthermore, it was observed that activation of brain 5-HT7 receptors rescued mitochondrial dysfunction in female mice from two models of Rett syndrome, a rare neurodevelopmental disorder characterized by severe behavioral and physiological symptoms. Our Western blot analyses performed on cell-lysate and purified mitochondria isolated from neuronal cell line SH-SY5Y showed that 5-HT7 receptors are also expressed into mitochondria. Maximal binding capacity (Bmax) obtained by Scatchard analysis on purified mitochondrial membranes was 0.081 pmol/mg of 5-HT7 receptor protein. Lastly, we evaluated the effect of selective 5-HT7 receptor agonist LP-211 and antagonist (inverse agonist) SB-269970 on mitochondrial respiratory chain (MRC) cytochrome c oxidase activity on mitochondria from SH-SY5Y cells. Our findings provide the first evidence that 5-HT7 receptor is also expressed in mitochondria.
Asunto(s)
Membranas Mitocondriales/metabolismo , Neuroblastoma/metabolismo , Receptores de Serotonina/metabolismo , Serotonina/metabolismo , Humanos , Membranas Mitocondriales/efectos de los fármacos , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/patología , Receptores de Serotonina/química , Antagonistas de la Serotonina/farmacología , Agonistas de Receptores de Serotonina/farmacología , Células Tumorales CultivadasRESUMEN
Recent preclinical studies have shown that activation of the serotonin 5-HT7 receptor has the potential to treat neurodevelopmental disorders such as Fragile X syndrome, a rare disease characterized by autistic features. With the aim to provide the scientific community with diversified drug-like 5-HT7 receptor-preferring agonists, we designed a set of new long-chain arylpiperazines by exploiting structural fragments present in clinically approved drugs or in preclinical candidates (privileged scaffolds). The new compounds were synthesized, tested for their affinity at 5-HT7 and 5-HT1A receptors, and screened for their in vitro stability to microsomal degradation and toxicity. Selected compounds were characterized as 5-HT7 receptor-preferring ligands, endowed with high metabolic stability and low toxicity. Compound 7g emerged as a drug-like 5-HT7 receptor-preferring agonist capable to rescue synaptic plasticity and attenuate stereotyped behavior in a mouse model of Fragile X syndrome.
Asunto(s)
Diseño de Fármacos , Síndrome del Cromosoma X Frágil/tratamiento farmacológico , Piperazinas/farmacología , Receptores de Serotonina/metabolismo , Agonistas de Receptores de Serotonina/farmacología , Relación Dosis-Respuesta a Droga , Síndrome del Cromosoma X Frágil/metabolismo , Células HEK293 , Humanos , Estructura Molecular , Piperazinas/síntesis química , Piperazinas/química , Agonistas de Receptores de Serotonina/síntesis química , Agonistas de Receptores de Serotonina/química , Relación Estructura-ActividadRESUMEN
Pituitary adenylate cyclase-activating polypeptide (PACAP) modulates glutamatergic synaptic transmission and plasticity in the hippocampus, a brain area with a key role in learning and memory. In agreement, several studies have demonstrated that PACAP modulates learning in physiological conditions. Recent publications show reduced PACAP levels and/or alterations in PACAP receptor expression in different conditions associated with cognitive disability. It is noteworthy that PACAP administration rescued impaired synaptic plasticity and learning in animal models of aging, Alzheimer's disease, Parkinson's disease, and Huntington's chorea. In this context, results from our laboratory demonstrate that PACAP rescued metabotropic glutamate receptor-mediated synaptic plasticity in the hippocampus of a mouse model of fragile X syndrome (FXS), a genetic form of intellectual disability. PACAP is actively transported through the blood-brain barrier and reaches the brain following intranasal or intravenous administration. Besides, new studies have identified synthetic PACAP analog peptides with improved selectivity and pharmacokinetic properties with respect to the native peptide. Our review supports the shared idea that pharmacological activation of PACAP receptors might be beneficial for brain pathologies with cognitive disability. In addition, we suggest that the effects of PACAP treatment might be further studied as a possible therapy in FXS.
RESUMEN
The fragile X mental retardation protein (FMRP) is an RNA-binding protein involved in translational regulation of mRNAs that play key roles in synaptic morphology and plasticity. The functional absence of FMRP causes the fragile X syndrome (FXS), the most common form of inherited intellectual disability and the most common monogenic cause of autism. No effective treatment is available for FXS. We recently identified the Phosphodiesterase 2A (Pde2a) mRNA as a prominent target of FMRP. PDE2A enzymatic activity is increased in the brain of Fmr1-KO mice, a recognized model of FXS, leading to decreased levels of cAMP and cGMP. Here, we pharmacologically inhibited PDE2A in Fmr1-KO mice and observed a rescue both of the maturity of dendritic spines and of the exaggerated hippocampal mGluR-dependent long-term depression. Remarkably, PDE2A blockade rescued the social and communicative deficits of both mouse and rat Fmr1-KO animals. Importantly, chronic inhibition of PDE2A in newborn Fmr1-KO mice followed by a washout interval, resulted in the rescue of the altered social behavior observed in adolescent mice. Altogether, these results reveal the key role of PDE2A in the physiopathology of FXS and suggest that its pharmacological inhibition represents a novel therapeutic approach for FXS.
Asunto(s)
Comunicación Animal , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 2/metabolismo , Espinas Dendríticas/efectos de los fármacos , Síndrome del Cromosoma X Frágil/enzimología , Hipocampo/efectos de los fármacos , Imidazoles/farmacología , Depresión Sináptica a Largo Plazo/efectos de los fármacos , Neuronas/efectos de los fármacos , Inhibidores de Fosfodiesterasa/farmacología , Conducta Social , Triazinas/farmacología , Animales , Animales Recién Nacidos , AMP Cíclico/metabolismo , GMP Cíclico/metabolismo , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 2/antagonistas & inhibidores , Espinas Dendríticas/patología , Embrión de Mamíferos , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/patología , Síndrome del Cromosoma X Frágil/fisiopatología , Técnicas de Inactivación de Genes , Hipocampo/metabolismo , Ratones , Ratones Noqueados , Neuronas/metabolismo , Neuronas/patología , Cultivo Primario de Células , Ratas , Receptores de Glutamato Metabotrópico/efectos de los fármacos , Receptores de Glutamato Metabotrópico/metabolismoRESUMEN
We have previously demonstrated that activation of serotonin 5-HT7 receptors (5-HT7R) reverses metabotropic glutamate receptor-mediated long term depression (mGluR-LTD) in the hippocampus of wild-type (WT) and Fmr1 Knockout (KO) mice, a model of Fragile X Syndrome (FXS) in which mGluR-LTD is abnormally enhanced. Here, we have investigated intracellular mechanisms underlying the effect of 5-HT7R activation using patch clamp on hippocampal slices. Furthermore, we have tested whether in vivo administration of LP-211, a selective 5-HT7R agonist, can rescue learning and behavior in Fmr1 KO mice. In the presence of an adenylate cyclase blocker, mGluR-LTD was slightly enhanced in WT and therefore the difference between mGluR-LTD in WT and Fmr1 KO slices was no longer present. Conversely, activation of adenylate cyclase by either forskolin or Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) completely reversed mGluR-LTD in WT and Fmr1 KO. 5-HT7R activation reversed mGluR-LTD in WT and corrected exaggerated mGluR-LTD in Fmr1 KO; this effect was abolished by blockade of either adenylate cyclase or protein kinase A (PKA). Exposure of hippocampal slices to LP-211 caused an increased phosphorylation of extracellular signal regulated kinase (ERK), an intracellular effector involved in mGluR-LTD, in WT mice. Conversely, this effect was barely detectable in Fmr1 KO mice, suggesting that 5-HT7R-mediated reversal of mGluR-LTD does not require ERK stimulation. Finally, an acute in vivo administration of LP-211 improved novel object recognition (NOR) performance in WT and Fmr1 KO mice and reduced stereotyped behavior in Fmr1 KO mice. Our results indicate that mGluR-LTD in WT and Fmr1 KO slices is bidirectionally modulated in conditions of either reduced or enhanced cAMP formation. Activation of 5-HT7 receptors reverses mGluR-LTD by activation of the cAMP/PKA intracellular pathway. Importantly, a systemic administration of a 5-HT7R agonist to Fmr1 KO mice corrected learning deficits and repetitive behavior. We suggest that selective 5-HT7R agonists might become novel pharmacological tools for FXS therapy.
RESUMEN
Metabotropic glutamate receptor subtype 5 (mGluR5) is crucially implicated in the pathophysiology of Fragile X Syndrome (FXS); however, its dysfunction at the sub-cellular level, and related synaptic and cognitive phenotypes are unexplored. Here, we probed the consequences of mGluR5/Homer scaffold disruption for mGluR5 cell-surface mobility, synaptic N-methyl-D-aspartate receptor (NMDAR) function, and behavioral phenotypes in the second-generation Fmr1 knockout (KO) mouse. Using single-molecule tracking, we found that mGluR5 was significantly more mobile at synapses in hippocampal Fmr1 KO neurons, causing an increased synaptic surface co-clustering of mGluR5 and NMDAR. This correlated with a reduced amplitude of synaptic NMDAR currents, a lack of their mGluR5-activated long-term depression, and NMDAR/hippocampus dependent cognitive deficits. These synaptic and behavioral phenomena were reversed by knocking down Homer1a in Fmr1 KO mice. Our study provides a mechanistic link between changes of mGluR5 dynamics and pathological phenotypes of FXS, unveiling novel targets for mGluR5-based therapeutics.
Asunto(s)
Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/psicología , Receptor del Glutamato Metabotropico 5/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapsis/metabolismo , Animales , Cognición , Modelos Animales de Enfermedad , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/genética , Hipocampo/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Plasticidad Neuronal , Receptor del Glutamato Metabotropico 5/genética , Receptores de N-Metil-D-Aspartato/genética , Sinapsis/genéticaRESUMEN
Astrocytes regulate neuronal activity and blood brain barrier through tiny plasma membrane branches or astrocytic processes (APs) making contact with synapses and brain vessels. Several transmitters released by astrocytes and exerting their action on several receptor classes expressed by astrocytes themselves influence their physiology. Here we found that APs are dynamically modulated by purines. In live imaging experiments carried out in rat hippocampal astrocytes, Gq-coupled P2Y1 receptor blockade with the selective antagonist MRS2179 (1 µM) or inhibition of its effector phospholipase C using U73122 (3 µM) produced APs retraction, while stimulation of the same receptor with the selective agonist 2MeSADP (100 µM) increased their number. Since astrocytes, among other transmitters, release ATP by several mechanisms including connexin hemichannels, we used the connexin hemichannel inhibitor carbenoxolone (100 µM) and APs retraction was observed. In our system we then measured expression or function of channels important for modulation of volume transmission and K+ buffering, aquaporin-4, and K+ inward rectifying (Kir) channels, respectively. Aquaporin-4 expression level did not change whereas, in whole-cell patch-clamp recordings performed to measure Kir current, we observed an increase in K+ current in all conditions where APs number was reduced. These data are supporting the idea of a dynamic modulation of astrocytic processes by purinergic signal, strengthening the role of purines in brain homeostasis.
Asunto(s)
Astrocitos/metabolismo , Membrana Celular/metabolismo , Hipocampo/metabolismo , Receptores Purinérgicos P2Y1/metabolismo , Animales , Acuaporina 4/metabolismo , Señalización del Calcio , Células Cultivadas , Conexinas/metabolismo , Canales de Potasio Rectificados Internamente Asociados a la Proteína G/metabolismo , Ratas Sprague-Dawley , Fosfolipasas de Tipo C/antagonistas & inhibidores , Fosfolipasas de Tipo C/metabolismoRESUMEN
The neuropeptide PACAP modulates synaptic transmission in the hippocampus exerting multiple effects through different receptor subtypes: the underlying mechanisms have not yet been completely elucidated. The neurotransmitter acetylcholine (ACh) also exerts a well-documented modulation of hippocampal synaptic transmission and plasticity. Since PACAP was shown to stimulate ACh release in the hippocampus, we tested whether PACAP acting through ACh might indirectly modulate glutamate-mediated synaptic transmission at a pre- and/or at a post-synaptic level. Using patch clamp on rat hippocampal slices, we tested PACAP effects on stimulation-evoked AMPA receptor-mediated excitatory post-synaptic currents (EPSCsAMPA) in the CA3-CA1 synapse and on spontaneous miniature EPSCs (mEPSCs) in CA1 pyramidal neurons. A subnanomolar dose of PACAP (0.5nM) decreased EPSCsAMPA amplitude, enhanced EPSC paired-pulse facilitation (PPF) and reduced mEPSC frequency, indicating a pre-synaptic decrease of glutamate release probability: these effects were abolished by simultaneous blockade of muscarinic and nicotinic ACh receptors, indicating the involvement of endogenous ACh. The effect of subnanomolar PACAP was abolished by a PAC1 receptor antagonist but not by a VPAC receptor blocker. At a higher concentration (10nM), PACAP inhibited EPSCsAMPA: this effect persisted in the presence of ACh receptor antagonists and did not involve any change in PPF or in mEPSC frequency, thus was not mediated by ACh and was exerted post- synaptically on CA1 pyramidal neurons. We suggest that a high-affinity PAC1 receptor pre-synaptically modulates hippocampal glutamatergic transmission acting through ACh. Therefore, administration of PACAP at very low doses might be envisaged in cognitive diseases with reduced cholinergic transmission.
Asunto(s)
Acetilcolina/metabolismo , Ácido Glutámico/metabolismo , Hipocampo/metabolismo , Polipéptido Hipofisario Activador de la Adenilato-Ciclasa/metabolismo , Transmisión Sináptica/fisiología , Animales , Polipéptido Hipofisario Activador de la Adenilato-Ciclasa/administración & dosificación , Terminales Presinápticos/efectos de los fármacos , Terminales Presinápticos/metabolismo , Células Piramidales/efectos de los fármacos , Células Piramidales/metabolismo , Ratas Sprague-Dawley , Ratas Wistar , Receptores AMPA/antagonistas & inhibidores , Receptores AMPA/metabolismo , Receptores del Polipéptido Activador de la Adenilato-Ciclasa Hipofisaria/metabolismo , Receptores de Tipo II del Péptido Intestinal Vasoactivo/metabolismo , Receptores de Tipo I del Polipéptido Intestinal Vasoactivo/metabolismo , Técnicas de Cultivo de TejidosRESUMEN
Serotonin 5-HT7 receptors are expressed in the hippocampus and modulate the excitability of hippocampal neurons. We have previously shown that 5-HT7 receptors modulate glutamate-mediated hippocampal synaptic transmission and long-term synaptic plasticity. In particular, we have shown that activation of 5-HT7 receptors reversed metabotropic glutamate receptor-mediated long-term depression (mGluR-LTD) in wild-type (wt) and in Fmr1 KO mice, a mouse model of Fragile X Syndrome in which mGluR-LTD is abnormally enhanced, suggesting that 5-HT7 receptor agonists might be envisaged as a novel therapeutic strategy for Fragile X Syndrome. In this perspective, we have characterized the basic in vitro pharmacokinetic properties of novel molecules with high binding affinity and selectivity for 5-HT7 receptors and we have tested their effects on synaptic plasticity using patch clamp on acute hippocampal slices. Here we show that LP-211, a high affinity selective agonist of 5-HT7 receptors, reverses mGluR-LTD in wt and Fmr1 KO mice, correcting a synaptic malfunction in the mouse model of Fragile X Syndrome. Among novel putative agonists of 5-HT7 receptors, the compound BA-10 displayed improved affinity and selectivity for 5-HT7 receptors and improved in vitro pharmacokinetic properties with respect to LP-211. BA-10 significantly reversed mGluR-LTD in the CA3-CA1 synapse in wt and Fmr1KO mice, indicating that BA-10 behaved as a highly effective agonist of 5-HT7 receptors and reduced exaggerated mGluR-LTD in a mouse model of Fragile X Syndrome. On the other side, the compounds RA-7 and PM-20, respectively arising from in vivo metabolism of LP-211 and BA-10, had no effect on mGluR-LTD thus did not behave as agonists of 5-HT7 receptors in our conditions. The present results provide information about the structure-activity relationship of novel 5-HT7 receptor agonists and indicate that LP-211 and BA-10 might be used as novel pharmacological tools for the therapy of Fragile X Syndrome.
RESUMEN
Serotonin type 7 receptors (5-HT7) are expressed in several brain areas, regulate brain development, synaptic transmission and plasticity, and therefore are involved in various brain functions such as learning and memory. A number of studies suggest that 5-HT7 receptors could be potential pharmacotherapeutic target for cognitive disorders. Several abnormalities of serotonergic system have been described in patients with autism spectrum disorder (ASD), including abnormal activity of 5-HT transporter, altered blood and brain 5-HT levels, reduced 5-HT synthesis and altered expression of 5-HT receptors in the brain. A specific role for 5-HT7 receptors in ASD has not yet been demonstrated but some evidence implicates their possible involvement. We have recently shown that 5-HT7 receptor activation rescues hippocampal synaptic plasticity in a mouse model of Fragile X Syndrome, a monogenic cause of autism. Several other studies have shown that 5-HT7 receptors modulate behavioral flexibility, exploratory behavior, mood disorders and epilepsy, which include core and co-morbid symptoms of ASD. These findings further suggest an involvement of 5-HT7 receptors in ASD. Here, we review the physiological roles of 5-HT7 receptors and their implications in Fragile X Syndrome and other ASD.
RESUMEN
Activation of group-I metabotropic glutamate receptors, mGlu1 and mGlu5, triggers a variety of signalling pathways in neurons and glial cells, which are differently implicated in synaptic plasticity. The earliest and much of key studies discovered abnormal mGlu5 receptor function in Fragile X syndrome (FXS) mouse models which then motivated more recent work that finds mGlu5 receptor dysfunction in related disorders such as intellectual disability (ID), obsessive-compulsive disorder (OCD) and autism. Therefore, mGlu1/5 receptor dysfunction may represent a common aetiology of these complex diseases. Furthermore, many studies have focused on dysregulation of mGlu5 signalling to synaptic protein synthesis. However, emerging evidence finds abnormal mGlu5 receptor interactions with its scaffolding proteins in FXS which results in mGlu5 receptor dysfunction and phenotypes independent of signalling to protein synthesis. Finally, both an increased and reduced mGlu5 functioning seem to be associated with ID and autism spectrum disorders, with important consequences for potential treatment of these developmental disorders.
Asunto(s)
Trastorno Autístico/metabolismo , Síndrome del Cromosoma X Frágil/metabolismo , Discapacidad Intelectual/metabolismo , Receptor del Glutamato Metabotropico 5/metabolismo , Receptores de Glutamato Metabotrópico/metabolismo , Transducción de Señal , Animales , Humanos , Plasticidad NeuronalRESUMEN
A three-step synthetic pathway has been employed to synthesize a small library of 2-(4-arylpiperidin-1-yl)-1-(1H-indol-3-yl)ethanone and 2-(4-arylpiperidin-1-yl)-1-(1H-indol-3-yl)ethane-1,2-dione derivatives that have been screened in [(3)H]ifenprodil competition binding assay. Some compounds exhibited significant binding affinity at nanomolar concentration, the most active being ligand 35 (IC50=5.5nM). Docking experiments suggested the main interactions between 35 and GluN2B-containing NMDA receptors. Notably, the compound 35 reduced NMDA-mediated excitatory post-synaptic currents recorded in mouse hippocampal slices indicating antagonistic effects (50nM). Moreover, the compound 35 has shown antioxidant effects in a preliminary screening, thus suggesting that it might be considered prototype for future drug development of novel 'dual target' neuroprotective agents.
Asunto(s)
Anticonvulsivantes/farmacología , Indoles/química , Receptores de N-Metil-D-Aspartato/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Anticonvulsivantes/química , Antioxidantes/química , Antioxidantes/farmacología , Unión Competitiva , Técnicas de Química Sintética , Hipocampo/efectos de los fármacos , Técnicas In Vitro , Ligandos , Ratones , Ratones Endogámicos DBA , Ratones Endogámicos , Simulación del Acoplamiento Molecular , Estructura Molecular , Piperidinas/metabolismo , Bibliotecas de Moléculas Pequeñas/síntesis químicaRESUMEN
In the course of the identification of new indole derivatives targeting GluN2B-subunit-containing N-methyl-D-aspartate (NMDA) receptor, the (N-1H-indol-6-methanesulfonamide-3-yl)-2-(4-benzylpiperidin-1-yl)ethanone (10b) was identified as a potent ligand for this NMDA receptor subunit. It displays very high binding affinity (IC50 of 8.9 nmol) for displacement of [3H]ifenprodil, thus showing improved potency with respect to the previously reported analogues as confirmed by functional assay. This finding was consistent with the docking pose of compound 10b within the binding pocket localized in the GluN1-GluN2B subunit interface of NMDA receptor tetraheteromeric complex.
Asunto(s)
Indoles/síntesis química , Indoles/farmacología , Receptores de N-Metil-D-Aspartato/efectos de los fármacos , Animales , Ligandos , Espectroscopía de Resonancia Magnética , Masculino , Modelos Moleculares , Ratas , Ratas Wistar , Receptores de N-Metil-D-Aspartato/química , Difracción de Rayos XRESUMEN
BACKGROUND: Fragile X syndrome (FXS) is a genetic cause of intellectual disability and autism. Fmr1 knockout (Fmr1 KO) mice, an animal model of FXS, exhibit spatial memory impairment and synapse malfunctioning in the hippocampus, with abnormal enhancement of long-term depression mediated by metabotropic glutamate receptors (mGluR-LTD). The neurotransmitter serotonin (5-HT) modulates hippocampal-dependent learning through serotonin 1A (5-HT1A) and serotonin 7 (5-HT7) receptors; the underlying mechanisms are unknown. METHODS: We used electrophysiology to test the effects of 5-HT on mGluR-LTD in wild-type and Fmr1 KO mice and immunocytochemistry and biotinylation assay to study related changes of 2-amino-3-(5-methyl-3-oxo-1,2-oxazol-4-yl)propanoic acid (AMPA) glutamate receptor surface expression. RESULTS: Application of 5-HT or 8-OH-DPAT (a mixed 5-HT1A/5-HT7 agonist) reversed mGluR-LTD in hippocampal slices. Reversal of mGluR-LTD by 8-OH-DPAT persisted in the presence of the 5-HT1A receptor antagonist WAY-100635, was abolished by SB-269970 (5-HT7 receptor antagonist), and was mimicked by LP-211, a novel selective 5-HT7 receptor agonist. Consistently, 8-OH-DPAT decreased mGluR-mediated reduction of AMPA glutamate receptor 2 (GluR2) subunit surface expression in hippocampal slices and cultured hippocampal neurons, an effect mimicked by LP-211 and blocked by SB-269970. In Fmr1 KO mice, mGluR-LTD was abnormally enhanced; similarly to wild-type, 8-OH-DPAT reversed mGluR-LTD and decreased mGluR-induced reduction of surface AMPA receptors, an effect antagonized by SB-269970. CONCLUSIONS: Serotonin 7 receptor activation reverses metabotropic glutamate receptor-induced AMPA receptor internalization and LTD both in wild-type and in Fmr1 KO mice, correcting excessive mGluR-LTD. Therefore, selective activation of 5-HT7 receptors may represent a novel strategy in the therapy of FXS.
Asunto(s)
Síndrome del Cromosoma X Frágil/metabolismo , Hipocampo/efectos de los fármacos , Depresión Sináptica a Largo Plazo/efectos de los fármacos , Receptores de Serotonina/metabolismo , Agonistas de Receptores de Serotonina/farmacología , 8-Hidroxi-2-(di-n-propilamino)tetralin/farmacología , Animales , Modelos Animales de Enfermedad , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Potenciales Postsinápticos Excitadores/fisiología , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Síndrome del Cromosoma X Frágil/fisiopatología , Hipocampo/metabolismo , Hipocampo/fisiopatología , Depresión Sináptica a Largo Plazo/fisiología , Ratones , Ratones Noqueados , Potenciales Postsinápticos Miniatura/efectos de los fármacos , Potenciales Postsinápticos Miniatura/fisiología , Serotonina/farmacologíaRESUMEN
As an extension of our studies, novel indole derivatives were rationally designed and synthesized as ligands targeted to GluN2B/NMDA receptors. The 2-(4-benzylpiperidin-1-yl)-1-(6-hydroxy-1H-indol-3-yl)ethanone (4i) and 1-(4-benzylpiperidin-1-yl)-2-(6-hydroxy-1H-indol-3-yl)ethane-1,2-dione (6i) showed high binding affinity in [3H]ifenprodil displacement assay. By computational studies, we suggested the hypothetical interactions playing a significant role during the binding process. However, in functional and in vivo studies the most potent compound 4i did not show any activity whereas it displayed relevant affinity toward the σ2 receptor.