RESUMO
Pediatric Acute-onset Neuropsychiatric Syndrome (PANS) is characterized by the abrupt onset of significant obsessive-compulsive symptoms (OCS) and/or severe food restriction, together with other neuropsychiatric manifestations. An autoimmune pathogenesis triggered by infection has been proposed for at least a subset of PANS. The older diagnosis of Pediatric Autoimmune Neuropsychiatric Disorder Associated with Streptococcus (PANDAS) describes rapid onset of OCD and/or tics associated with infection with Group A Streptococcus. The pathophysiology of PANS and PANDAS remains incompletely understood. We recently found serum antibodies from children with rigorously defined PANDAS to selectively bind to cholinergic interneurons (CINs) in the striatum. Here we examine this binding in children with relapsing and remitting PANS, a more heterogeneous condition, collected in a distinct clinical context from those examined in our previous work, from children with a clinical history of Streptococcus infection. IgG from PANS cases showed elevated binding to striatal CINs in both mouse and human brain. Patient plasma collected during symptom flare decreased a molecular marker of CIN activity, phospho-riboprotein S6, in ex vivo brain slices; control plasma did not. Neither elevated antibody binding to CINs nor diminished CIN activity was seen with plasma collected from the same children during remission. These findings replicate what we have seen previously in PANDAS and support the hypothesis that at least a subset of PANS cases have a neuroimmune pathogenesis. Given the critical role of CINs in modulating basal ganglia function, these findings confirm striatal CINs as a locus of interest in the pathophysiology of both PANS and PANDAS.
RESUMO
Both the NMDA receptor (NMDAR) positive allosteric modulator (PAM), and antagonist, can exert rapid antidepressant effects as shown in several animal and human studies. However, how this bidirectional modulation of NMDARs causes similar antidepressant effects remains unknown. Notably, the initial cellular trigger, specific cell-type(s), and subunit(s) of NMDARs mediating the antidepressant-like effects of a PAM or an antagonist have not been identified. Here, we used electrophysiology, microdialysis, and NMR spectroscopy to evaluate the effect of a NMDAR PAM (rapastinel) or NMDAR antagonist, ketamine on NMDAR function and disinhibition-mediated glutamate release. Further, we used cell-type specific knockdown (KD), pharmacological, and behavioral approaches to dissect the cell-type specific role of GluN2B, GluN2A, and dopamine receptor subunits in the actions of NMDAR PAM vs. antagonists. We demonstrate that rapastinel directly enhances NMDAR activity on principal glutamatergic neurons in medial prefrontal cortex (mPFC) without any effect on glutamate efflux, while ketamine blocks NMDAR on GABA interneurons to cause glutamate efflux and indirect activation of excitatory synapses. Behavioral studies using cell-type-specific KD in mPFC demonstrate that NMDAR-GluN2B KD on Camk2a- but not Gad1-expressing neurons blocks the antidepressant effects of rapastinel. In contrast, GluN2B KD on Gad1- but not Camk2a-expressing neurons blocks the actions of ketamine. The results also demonstrate that Drd1-expressing pyramidal neurons in mPFC mediate the rapid antidepressant actions of ketamine and rapastinel. Together, these results demonstrate unique initial cellular triggers as well as converging effects on Drd1-pyramidal cell signaling that underlie the antidepressant actions of NMDAR-positive modulation vs. NMDAR blockade.
Assuntos
Ketamina , Receptores de N-Metil-D-Aspartato , Animais , Antidepressivos/farmacologia , Humanos , Interneurônios/metabolismo , Ketamina/farmacologia , Córtex Pré-Frontal/metabolismo , Células Piramidais/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismoRESUMO
Ketamine, a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist, produces rapid and long-lasting antidepressant effects in major depressive disorder (MDD) patients. (2R,6R)-Hydroxynorketamine [(2R,6R)-HNK], a metabolite of ketamine, is reported to produce rapid antidepressant effects in rodent models without the side effects of ketamine. Importantly, (2R,6R)-HNK does not block NMDA receptors like ketamine, and the molecular signaling mechanisms for (2R,6R)-HNK remain unknown. Here, we examined the involvement of BDNF/TrkB/mechanistic target of rapamycin complex 1 (mTORC1) signaling in the antidepressant actions of (2R,6R)-HNK. Intramedial prefrontal cortex (intra-mPFC) infusion or systemic (2R,6R)-HNK administration induces rapid and long-lasting antidepressant effects in behavioral tests, identifying the mPFC as a key region for the actions of (2R,6R)-HNK. The antidepressant actions of (2R,6R)-HNK are blocked in mice with a knockin of the BDNF Val66Met allele (which blocks the processing and activity-dependent release of BDNF) or by intra-mPFC microinjection of an anti-BDNF neutralizing antibody. Blockade of L-type voltage-dependent Ca2+ channels (VDCCs), required for activity-dependent BDNF release, also blocks the actions of (2R,6R)-HNK. Intra-mPFC infusion of pharmacological inhibitors of TrkB or mTORC1 signaling, which are downstream of BDNF, also block the actions of (2R,6R)-HNK. Moreover, (2R,6R)-HNK increases synaptic function in the mPFC. These findings indicate that activity-dependent BDNF release and downstream TrkB and mTORC1 signaling, which increase synaptic function in the mPFC, are required for the rapid and long-lasting antidepressant effects of (2R,6R)-HNK, supporting the potential use of this metabolite for the treatment of MDD.
Assuntos
Antidepressivos/farmacologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Ketamina/análogos & derivados , Animais , Células Cultivadas , Depressão/tratamento farmacológico , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Ketamina/farmacologia , Alvo Mecanístico do Complexo 1 de Rapamicina/antagonistas & inibidores , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacosRESUMO
Dysfunction of medial prefrontal cortex (mPFC) in association with imbalance of inhibitory and excitatory neurotransmission has been implicated in depression. However, the precise cellular mechanisms underlying this imbalance, particularly for GABAergic transmission in the mPFC, and the link with the rapid acting antidepressant ketamine remains poorly understood. Here we determined the influence of chronic unpredictable stress (CUS), an ethologically validated model of depression, on synaptic markers of GABA neurotransmission, and the influence of a single dose of ketamine on CUS-induced synaptic deficits in mPFC of male rodents. The results demonstrate that CUS decreases GABAergic proteins and the frequency of inhibitory post synaptic currents (IPSCs) of layer V mPFC pyramidal neurons, concomitant with depression-like behaviors. In contrast, a single dose of ketamine can reverse CUS-induced deficits of GABA markers, in conjunction with reversal of CUS-induced depressive-like behaviors. These findings provide further evidence of impairments of GABAergic synapses as key determinants of depressive behavior and highlight ketamine-induced synaptic responses that restore GABA inhibitory, as well as glutamate neurotransmission.
Assuntos
Antidepressivos/administração & dosagem , Depressão/fisiopatologia , Ketamina/administração & dosagem , Neurônios/efeitos dos fármacos , Córtex Pré-Frontal/efeitos dos fármacos , Estresse Psicológico/fisiopatologia , Transmissão Sináptica/efeitos dos fármacos , Ácido gama-Aminobutírico/fisiologia , Animais , Modelos Animais de Doenças , Potenciais Pós-Sinápticos Inibidores/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Neurônios/fisiologia , Córtex Pré-Frontal/fisiopatologiaRESUMO
The striatum contains several types of neurons including medium spiny projection neurons (MSNs), cholinergic interneurons (ChIs), and fast-spiking interneurons (FSIs). Modulating the activity of these neurons by the dopamine D2 receptor (D2R) can greatly impact motor control and movement disorders. D2R exists in two isoforms: D2L and D2S. Here, we assessed whether alterations in the D2L and D2S expression levels affect neuronal excitability and synaptic function in striatal neurons. We observed that quinpirole inhibited the firing rate of all three types of striatal neurons in wild-type (WT) mice. However, in D2L knockout (KO) mice, quinpirole enhanced the excitability of ChIs, lost influence on spike firing of MSNs, and remained inhibitory effect on spike firing of FSIs. Additionally, we showed mIPSC frequency (but not mIPSC amplitude) was reduced in ChIs from D2L KO mice compared with WT mice, suggesting spontaneous GABA release is reduced at GABAergic terminals onto ChIs in D2L KO mice. Furthermore, we found D2L deficiency resulted in reduced dendritic spine density in ChIs, suggesting D2L activation plays a role in the formation/maintenance of dendritic spines of ChIs. These findings suggest new molecular and cellular mechanisms for causing ChIs abnormality seen in Parkinson's disease or drug-induced dyskinesias.
RESUMO
The lateral hypothalamus and the nucleus accumbens shell (AcbSh) are brain regions important for food intake. The AcbSh contains high levels of receptor for melanin-concentrating hormone (MCH), a lateral hypothalamic peptide critical for feeding and metabolism. MCH receptor (MCHR1) activation in the AcbSh increases food intake, while AcbSh MCHR1 blockade reduces feeding. Here biochemical and cellular mechanisms of MCH action in the rodent AcbSh are described. A reduction of phosphorylation of GluR1 at serine 845 (pSer(845)) is shown to occur after both pharmacological and genetic manipulations of MCHR1 activity. These changes depend upon signaling through G(i/o), and result in decreased surface expression of GluR1-containing AMPA receptors (AMPARs). Electrophysiological analysis of medium spiny neurons (MSNs) in the AcbSh revealed decreased amplitude of AMPAR-mediated synaptic events (mEPSCs) with MCH treatment. In addition, MCH suppressed action potential firing MSNs through K(+) channel activation. Finally, in vivo recordings confirmed that MCH reduces neuronal cell firing in the AcbSh in freely moving animals. The ability of MCH to reduce cell firing in the AcbSh is consistent with a general model from other pharmacological and electrophysiological studies whereby reduced AcbSh neuronal firing leads to food intake. The current work integrates the hypothalamus into this model, providing biochemical and cellular mechanisms whereby metabolic and limbic signals converge to regulate food intake.
Assuntos
Hormônios Hipotalâmicos/metabolismo , Hipotálamo/metabolismo , Melaninas/metabolismo , Núcleo Accumbens/fisiologia , Hormônios Hipofisários/metabolismo , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/genética , Animais , Compostos de Bário/farmacologia , Biotina/análogos & derivados , Biotina/metabolismo , Cloretos/farmacologia , Fosfoproteína 32 Regulada por cAMP e Dopamina/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Potenciais Pós-Sinápticos Excitadores/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônios Hipotalâmicos/genética , Hormônios Hipotalâmicos/farmacologia , Hipotálamo/citologia , Técnicas In Vitro , Masculino , Melaninas/genética , Melaninas/farmacologia , Camundongos , Camundongos Transgênicos , Vias Neurais/fisiologia , Neurônios/classificação , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Núcleo Accumbens/citologia , Técnicas de Patch-Clamp/métodos , Hormônios Hipofisários/genética , Hormônios Hipofisários/farmacologia , Bloqueadores dos Canais de Potássio/farmacologia , Ratos , Ratos Long-Evans , Ratos Wistar , Receptores de AMPA/genética , Receptores de AMPA/metabolismo , Serina/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Morphological studies show that repeated restraint stress leads to selective atrophy in the apical dendritic field of pyramidal cells in the medial prefrontal cortex (mPFC). However, the functional consequence of this selectivity remains unclear. The apical dendrite of layer V pyramidal neurons in the mPFC is a selective locus for the generation of increased excitatory postsynaptic currents (EPSCs) by serotonin (5-HT) and hypocretin (orexin). On that basis, we hypothesized that apical dendritic atrophy might result in a blunting of 5-HT- and hypocretin-induced excitatory responses. Using a combination of whole-cell recording and two-photon imaging in rat mPFC slices, we were able to correlate electrophysiological and morphological changes in the same layer V pyramidal neurons. Repeated mild restraint stress produced a decrement in both 5-HT- and hypocretin-induced EPSCs, an effect that was correlated with a decrease in apical tuft dendritic branch length and spine density in the distal tuft branches. Chronic treatment with the stress hormone corticosterone, while reducing 5-HT responses and generally mimicking the morphological effects of stress, failed to produce a significant decrease in hypocretin-induced EPSCs. Accentuating this difference, pretreatment of stressed animals with the glucocorticoid receptor antagonist RU486 blocked reductions in 5-HT-induced EPSCs but not hypocretin-induced EPSCs. We conclude: (i) stress-induced apical dendritic atrophy results in diminished responses to apically targeted excitatory inputs and (ii) corticosterone plays a greater role in stress-induced reductions in EPSCs evoked by 5-HT as compared with hypocretin, possibly reflecting the different pathways activated by the two transmitters.
Assuntos
Corticosterona/farmacologia , Dendritos/patologia , Potenciais Pós-Sinápticos Excitadores , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neuropeptídeos/metabolismo , Córtex Pré-Frontal/metabolismo , Serotonina/metabolismo , Corticosteroides/metabolismo , Animais , Atrofia , Eletrofisiologia/métodos , Glucocorticoides/metabolismo , Masculino , Modelos Biológicos , Neurônios/metabolismo , Orexinas , Fótons , Ratos , Receptores de Glucocorticoides/metabolismo , Serotonina/farmacologiaRESUMO
OBJECTIVE: Pediatric obsessive-compulsive disorder (OCD) sometimes appears rapidly, even overnight, often after an infection. Pediatric autoimmune neuropsychiatric disorders associated with streptococcal infections, or PANDAS, describes such a situation after infection with Streptococcus pyogenes. PANDAS may result from induced autoimmunity against brain antigens, although this remains unproven. Pilot work suggests that IgG antibodies from children with PANDAS bind to cholinergic interneurons (CINs) in the striatum. CIN deficiency has been independently associated with tics in humans and with repetitive behavioral pathology in mice, making it a plausible locus of pathology. The authors sought to replicate and extend earlier work and to investigate the cellular effects of PANDAS antibodies on cholinergic interneurons. METHODS: Binding of IgG to specific neurons in human and mouse brain slices was evaluated ex vivo after incubation with serum from 27 children with rigorously characterized PANDAS, both at baseline and after intravenous immunoglobulin (IVIG) treatment, and 23 matched control subjects. Binding was correlated with symptom measures. Neural activity after serum incubation was assessed in mouse slices using molecular markers and electrophysiological recording. RESULTS: IgG from children with PANDAS bound to CINs, but not to several other neuron types, more than IgG from control subjects, in three independent cohorts of patients. Post-IVIG serum had reduced IgG binding to CINs, and this reduction correlated with symptom improvement. Baseline PANDAS sera decreased activity of striatal CINs, but not of parvalbumin-expressing GABAergic interneurons, and altered their electrophysiological responses, in acute mouse brain slices. Post-IVIG PANDAS sera and IgG-depleted baseline sera did not alter the activity of striatal CINs. CONCLUSIONS: These findings provide strong evidence for striatal CINs as a critical cellular target that may contribute to pathophysiology in children with rapid-onset OCD symptoms, and perhaps in other conditions.
Assuntos
Autoanticorpos/imunologia , Doenças Autoimunes/imunologia , Neurônios Colinérgicos/imunologia , Corpo Estriado/imunologia , Transtorno Obsessivo-Compulsivo/imunologia , Infecções Estreptocócicas/imunologia , Animais , Doenças Autoimunes/complicações , Estudos de Casos e Controles , Criança , Pré-Escolar , Neurônios Colinérgicos/fisiologia , Corpo Estriado/fisiopatologia , Feminino , Humanos , Imunoglobulina G/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Transtorno Obsessivo-Compulsivo/complicações , Transtorno Obsessivo-Compulsivo/etiologia , Infecções Estreptocócicas/complicaçõesRESUMO
Dysregulation of the glutamatergic system and its receptors in medial prefrontal cortex (mPFC) has been implicated in major depressive disorder. Recent preclinical studies have shown that enhancing NMDA receptor (NMDAR) activity can exert rapid antidepressant-like effects. AGN-241751, an NMDAR positive allosteric modulator (PAM), is currently being tested as an antidepressant in clinical trials, but the mechanism and NMDAR subunit(s) mediating its antidepressant-like effects are unknown. We therefore used molecular, biochemical, and electrophysiological approaches to examine the cell-type-specific role of GluN2B-containing NMDAR in mediating antidepressant-like behavioral effects of AGN-241751. We demonstrate that AGN-241751 exerts antidepressant-like effects and reverses behavioral deficits induced by chronic unpredictable stress in mice. AGN-241751 treatment enhances NMDAR activity of excitatory and parvalbumin-inhibitory neurons in mPFC, activates Akt/mTOR signaling, and increases levels of synaptic proteins crucial for synaptic plasticity in the prefrontal cortex. Furthermore, cell-type-specific knockdown of GluN2B-containing NMDARs in mPFC demonstrates that GluN2B subunits on excitatory, but not inhibitory, neurons are necessary for antidepressant-like effects of AGN-241751. Together, these results demonstrate antidepressant-like actions of the NMDAR PAM AGN-241751 and identify GluN2B on excitatory neurons of mPFC as initial cellular trigger underlying these behavioral effects.
Assuntos
Transtorno Depressivo Maior , Receptores de N-Metil-D-Aspartato , Animais , Antidepressivos/farmacologia , Antidepressivos/uso terapêutico , Transtorno Depressivo Maior/tratamento farmacológico , Camundongos , Neurônios/metabolismo , Córtex Pré-Frontal/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismoRESUMO
We previously reported that the serotonergic system is important for the antidepressant-like effects of ketamine, a non-competitive N-methyl-D-aspartate receptor antagonist, which produces rapid and long-lasting antidepressant effects in patients with major depressive disorder (MDD). In particular, selective stimulation of the 5-HT1A receptor in the medial prefrontal cortex (mPFC), as opposed to the somatic 5-HT1A autoreceptor, has been shown to play a critical role in the antidepressant-like actions of ketamine. However, the detailed mechanisms underlying mPFC 5-HT1A receptor-mediated antidepressant-like effects are not fully understood. Here we examined the involvement of the glutamate AMPA receptor and brain-derived neurotrophic factor (BDNF) in the antidepressant-like effects of 5-HT1A receptor activation in the mPFC. The results show that intra-mPFC infusion of the 5-HT1A receptor agonist 8-OH-DPAT induces rapid and long-lasting antidepressant-like effects in the forced swim, novelty-suppressed feeding, female urine sniffing, and chronic unpredictable stress tests. In addition, the results demonstrate that the antidepressant-like effects of intra-mPFC infusion of 8-OH-DPAT are blocked by co-infusion of an AMPA receptor antagonist or an anti-BDNF neutralizing antibody. In addition, mPFC infusion of 8-OH-DPAT increased the phosphorylation of signaling proteins downstream of BDNF, including mTOR, ERK, 4EBP1, and p70S6K. Finally, selective stimulation of the 5-HT1A receptor increased levels of synaptic proteins and synaptic function in the mPFC. Collectively, these results indicate that selective stimulation of 5-HT1A receptor in the mPFC exerts rapid and sustained antidepressant-like effects via activation of AMPA receptor/BDNF/mTOR signaling in mice, which subsequently increase synaptic function in the mPFC, and provide evidence for the 5-HT1A receptor as a target for the treatment of MDD.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Transtorno Depressivo Maior , Animais , Antidepressivos/farmacologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Depressão/tratamento farmacológico , Transtorno Depressivo Maior/tratamento farmacológico , Feminino , Humanos , Camundongos , Córtex Pré-Frontal/metabolismo , Receptor 5-HT1A de Serotonina , Receptores de AMPARESUMO
A single subanesthetic dose of ketamine, an NMDA receptor (NMDAR) antagonist, produces rapid and sustained antidepressant actions in depressed patients, addressing a major unmet need for the treatment of mood disorders. Ketamine produces a rapid increase in extracellular glutamate and synaptic formation in the prefrontal cortex, but the initial cellular trigger that initiates this increase and ketamine's behavioral actions has not been identified. To address this question, we used a combination of viral shRNA and conditional mutation to produce cell-specific knockdown or deletion of a key NMDAR subunit, GluN2B, implicated in the actions of ketamine. The results demonstrated that the antidepressant actions of ketamine were blocked by GluN2B-NMDAR knockdown on GABA (Gad1) interneurons, as well as subtypes expressing somatostatin (Sst) or parvalbumin (Pvalb), but not glutamate principle neurons in the medial prefrontal cortex (mPFC). Further analysis of GABA subtypes showed that cell-specific knockdown or deletion of GluN2B in Sst interneurons blocked or occluded the antidepressant actions of ketamine and revealed sex-specific differences that are associated with excitatory postsynaptic currents on mPFC principle neurons. These findings demonstrate that GluN2B-NMDARs on GABA interneurons are the initial cellular trigger for the rapid antidepressant actions of ketamine and show sex-specific adaptive mechanisms to GluN2B modulation.
Assuntos
Antidepressivos/farmacologia , Neurônios GABAérgicos/metabolismo , Interneurônios/metabolismo , Ketamina/farmacologia , Caracteres Sexuais , Animais , Feminino , Neurônios GABAérgicos/patologia , Técnicas de Inativação de Genes , Glutamato Descarboxilase/genética , Glutamato Descarboxilase/metabolismo , Interneurônios/patologia , Masculino , Camundongos , Camundongos Transgênicos , Parvalbuminas/genética , Parvalbuminas/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Somatostatina/genética , Somatostatina/metabolismoRESUMO
Impaired function in the medial prefrontal cortex (mPFC) contributes to depression, and the therapeutic response produced by novel rapid-acting antidepressants such as ketamine are mediated by mPFC activity. The mPFC contains multiple types of pyramidal cells, but it is unclear whether a particular subtype mediates the rapid antidepressant actions of ketamine. Here we tested two major subtypes, Drd1 and Drd2 dopamine receptor expressing pyramidal neurons and found that activating Drd1 expressing pyramidal cells in the mPFC produces rapid and long-lasting antidepressant and anxiolytic responses. In contrast, photostimulation of Drd2 expressing pyramidal cells was ineffective across anxiety-like and depression-like measures. Disruption of Drd1 activity also blocked the rapid antidepressant effects of ketamine. Finally, we demonstrate that stimulation of mPFC Drd1 terminals in the BLA recapitulates the antidepressant effects of somatic stimulation. These findings aid in understanding the cellular target neurons in the mPFC and the downstream circuitry involved in rapid antidepressant responses.
Assuntos
Antidepressivos/farmacologia , Ketamina/farmacologia , Optogenética , Córtex Pré-Frontal/efeitos dos fármacos , Receptores de Dopamina D1/metabolismo , Animais , Complexo Nuclear Basolateral da Amígdala/efeitos da radiação , Agonistas de Dopamina , Feminino , Masculino , Camundongos Endogâmicos C57BL , Córtex Pré-Frontal/metabolismo , Receptores de Dopamina D2/metabolismoRESUMO
Currently available antidepressants have a delayed onset and limited efficacy, highlighting the need for new, rapid and more efficacious agents. Ketamine, an NMDA receptor antagonist, has emerged as a new rapid-acting antidepressant, effective even in treatment resistant patients. However, ketamine induces undesired psychotomimetic and dissociative side effects that limit its clinical use. The d-stereoisomer of methadone (dextromethadone; REL-1017) is a noncompetitive NMDA receptor antagonist with an apparently favorable safety and tolerability profile. The current study examined the rapid and sustained antidepressant actions of d-methadone in several behavioral paradigms, as well as on mTORC1 signaling and synaptic changes in the medial prefrontal cortex (mPFC). A single dose of d-methadone promoted rapid and sustained antidepressant responses in the novelty-suppressed feeding test (NSFT), a measure of anxiety, and in the female urine sniffing test (FUST), a measure of motivation and reward. D-methadone also produced a rapid reversal of the sucrose preference deficit, a measure of anhedonia, in rats exposed to chronic unpredictable stress. D-methadone increased phospho-p70S6 kinase, a downstream target of mTORC1 in the mPFC, and intra-mPFC infusion of the selective mTORC1 inhibitor rapamycin blocked the antidepressant actions of d-methadone in the FUST and NSFT. D-methadone administration also increased levels of the synaptic proteins, PSD95, GluA1, and Synapsin 1 and enhanced synaptic function in the mPFC. Studies in primary cortical cultures show that d-methadone also increases BDNF release, as well as phospho-p70S6 kinase. These findings indicate that d-methadone induces rapid antidepressant actions through mTORC1-mediated synaptic plasticity in the mPFC similar to ketamine.
Assuntos
Antidepressivos/administração & dosagem , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metadona/administração & dosagem , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/metabolismo , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Animais , Espinhas Dendríticas/efeitos dos fármacos , Espinhas Dendríticas/fisiologia , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Ketamina/administração & dosagem , Masculino , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
Preclinical studies demonstrate that rapid acting antidepressants, including ketamine require stimulation of mTORC1 signaling. This pathway is regulated by neuronal activity, endocrine and metabolic signals, notably the amino acid leucine, which activates mTORC1 signaling via binding to the upstream regulator sestrin. Here, we examined the antidepressant actions of NV-5138, a novel highly selective small molecule modulator of sestrin that penetrates the blood brain barrier. The results demonstrate that a single dose of NV-5138 produced rapid and long-lasting antidepressant effects, and rapidly reversed anhedonia caused by chronic stress exposure. The antidepressant actions of NV-5138 required BDNF release as the behavioral responses are blocked by infusion of a BDNF neutralizing antibody into the medial prefrontal cortex (mPFC) or in mice with a knock-in of a BDNF polymorphism that blocks activity dependent BDNF release. NV-5138 administration also rapidly increased synapse number and function in the mPFC, and reversed the synaptic deficits caused by chronic stress. Together, the results demonstrate that NV-5138 produced rapid synaptic and antidepressant behavioral responses via activation of the mTORC1 pathway and BDNF signaling, indicating that pharmacological modulation of sestrin is a novel approach for development of rapid acting antidepressants.
Assuntos
Antidepressivos , Comportamento Animal/efeitos dos fármacos , Proteínas de Choque Térmico/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Transmissão Sináptica/efeitos dos fármacos , Animais , Antidepressivos/química , Antidepressivos/farmacocinética , Antidepressivos/farmacologia , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Proteínas de Choque Térmico/genética , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Camundongos , Camundongos Knockout , Ratos , Ratos Sprague-Dawley , Transmissão Sináptica/genéticaRESUMO
The transcription factor cAMP response element-binding protein (CREB) is implicated in mediating the actions of chronic morphine in the locus ceruleus (LC), but direct evidence to support such a role is limited. Here, we investigated the influence of CREB on LC neuronal activity and opiate withdrawal behaviors by selectively manipulating CREB activity in the LC using viral vectors encoding genes for CREBGFP (wild-type CREB tagged with green fluorescent protein), caCREBGFP (a constitutively active CREB mutant), dnCREBGFP (a dominant-negative CREB mutant), or GFP alone as a control. Our results show that in vivo overexpression of CREBGFP in the LC significantly aggravated particular morphine withdrawal behaviors, whereas dnCREBGFP expression attenuated these behaviors. At the cellular level, CREBGFP expression in the LC in vivo and in vitro had no significant effect on neuronal firing at baseline but enhanced the excitatory effect of forskolin (an activator of adenylyl cyclase) on these neurons, which suggests that the cAMP signaling pathway in these neurons was sensitized after CREB expression. Moreover, in vitro studies showed that caCREBGFP-expressing LC neurons fired significantly faster and had a more depolarized resting membrane potential compared with GFP-expressing control cells. Conversely, LC neuronal activity was decreased by dnCREBGFP, and the neurons were hyperpolarized by this treatment. Together, these data provide direct evidence that CREB plays an important role in controlling the electrical excitability of LC neurons and that morphine-induced increases in CREB activity contribute to the behavioral and neural adaptations associated with opiate dependence and withdrawal.
Assuntos
Comportamento Animal/efeitos dos fármacos , Proteína de Ligação a CREB/metabolismo , Locus Cerúleo/fisiopatologia , Transtornos Mentais/induzido quimicamente , Transtornos Mentais/fisiopatologia , Neurônios , Ópio/efeitos adversos , Potenciais de Ação/efeitos dos fármacos , Adaptação Fisiológica/efeitos dos fármacos , Animais , Locus Cerúleo/efeitos dos fármacos , Masculino , Plasticidade Neuronal/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Síndrome de Abstinência a Substâncias , Transmissão Sináptica/efeitos dos fármacosRESUMO
Diminished connectivity between midline-intralaminar thalamic nuclei and prefrontal cortex has been suggested to contribute to cognitive deficits that are detectable even in early stages of schizophrenia. The midline-intralaminar relay cells comprise the final link in the ascending arousal pathway and are selectively excited by the wake-promoting peptides hypocretin 1 and 2 (orexin A and B). This excitation occurs both at the level of the relay cell bodies and their axon terminals within prefrontal cortex. In rat brain slices, the release of glutamate from midline-intralaminar thalamocortical terminals induces excitatory postsynaptic currents (EPSCs) in layer V pyramidal cells in prefrontal cortex. When hypocretin is infused into medial prefrontal cortex of behaving animals, it improves performance in a complex cognitive task requiring divided attention. Chronic restraint stress causes atrophy of the apical dendritic arbors in layer V prefrontal pyramidal cells and leads to a reduction in hypocretin-induced EPSCs, indicating impairment in excitatory thalamocortical transmission. Thus, taken together with evidence for an underlying loss of excitatory thalamocortical connectivity in schizophrenia, stress in this illness could further exacerbate a breakdown in cortical processing of incoming information from the ascending arousal system.
Assuntos
Córtex Cerebral/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neuropeptídeos/metabolismo , Córtex Pré-Frontal/metabolismo , Esquizofrenia/metabolismo , Tálamo/metabolismo , Dendritos/fisiologia , Humanos , Orexinas , Esquizofrenia/fisiopatologiaRESUMO
GLYX-13 is a putative NMDA receptor modulator with glycine-site partial agonist properties that produces rapid antidepressant effects, but without the psychotomimetic side effects of ketamine. Studies were conducted to examine the molecular, cellular, and behavioral actions of GLYX-13 to further characterize the mechanisms underlying the antidepressant actions of this agent. The results demonstrate that a single dose of GLYX-13 rapidly activates the mTORC1 pathway in the prefrontal cortex (PFC), and that infusion of the selective mTORC1 inhibitor rapamycin into the medial PFC (mPFC) blocks the antidepressant behavioral actions of GLYX-13, indicating a requirement for mTORC1 similar to ketamine. The results also demonstrate that GLYX-13 rapidly increases the number and function of spine synapses in the apical dendritic tuft of layer V pyramidal neurons in the mPFC. Notably, GLYX-13 significantly increased the synaptic responses to hypocretin, a measure of thalamocortical synapses, compared with its effects on 5-HT responses, a measure of cortical-cortical responses mediated by the 5-HT2A receptor. Behavioral studies further demonstrate that GLYX-13 does not influence 5-HT2 receptor induced head twitch response or impulsivity in a serial reaction time task (SRTT), whereas ketamine increased responses in both tests. In contrast, both GLYX-13 and ketamine increased attention in the SRTT task, which is linked to hypocretin-thalamocortical responses. The differences in the 5-HT2 receptor synaptic and behavioral responses may be related to the lack of psychotomimetic side effects of GLYX-13 compared with ketamine, whereas regulation of the hypocretin responses may contribute to the therapeutic benefits of both rapid acting antidepressants.
Assuntos
Antidepressivos/farmacologia , Comportamento Animal/efeitos dos fármacos , Ketamina/farmacologia , Oligopeptídeos/farmacologia , Córtex Pré-Frontal/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Sinapses/efeitos dos fármacos , Animais , Antidepressivos/administração & dosagem , Ketamina/administração & dosagem , Masculino , Camundongos Endogâmicos C57BL , Oligopeptídeos/administração & dosagem , Ratos , Ratos Sprague-DawleyRESUMO
The hypocretins (hcrt1 and hcrt2) are expressed by a discrete population of hypothalamic neurons projecting to many regions of the CNS, including the dorsal raphe nucleus (DRN), where serotonin (5-HT) neurons are concentrated. In this study, we investigated responses to hcrts in 216 physiologically identified 5-HT and non-5-HT neurons of the DRN using intracellular and whole-cell recording in rat brain slices. Hcrt1 and hcrt2 induced similar amplitude and dose-dependent inward currents in most 5-HT neurons tested (EC50, approximately 250 nm). This inward current was not blocked by the fast Na+ channel blocker TTX or in a Ca2+-free solution, indicating a direct postsynaptic action. The hcrt-induced inward current reversed near -18 mV and was primarily dependent on external Na+ but not on external or internal Ca2+, features typical of Na+/K+ nonselective cation channels. At higher concentrations, hcrts also increased spontaneous postsynaptic currents in 5-HT neurons (EC50, approximately 450-600 nm), which were TTX-sensitive and mostly blocked by the GABA(A) antagonist bicuculline, indicating increased impulse flow in local GABA interneurons. Accordingly, hcrts were found to increase the basal firing of presumptive GABA interneurons. Immunolabeling showed that hcrt fibers projected to both 5-HT and GABA neurons in the DRN. We conclude that hcrts act directly to excite 5-HT neurons primarily via a TTX-insensitive, Na+/K+ nonselective cation current, and indirectly to activate local inhibitory GABA inputs to 5-HT cells. The greater potency of hcrts in direct excitation compared with indirect inhibition suggests a negative feedback function for the latter at higher levels of hcrt activity.
Assuntos
Proteínas de Transporte/farmacologia , Peptídeos e Proteínas de Sinalização Intracelular , Condução Nervosa/fisiologia , Inibição Neural/fisiologia , Neurônios/metabolismo , Neuropeptídeos/farmacologia , Núcleos da Rafe/metabolismo , Serotonina/metabolismo , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Axônios/metabolismo , Cálcio/metabolismo , Relação Dose-Resposta a Droga , Potenciais Pós-Sinápticos Excitadores/efeitos dos fármacos , Corantes Fluorescentes , Antagonistas GABAérgicos/farmacologia , Antagonistas de Receptores de GABA-A , Imuno-Histoquímica , Técnicas In Vitro , Interneurônios/efeitos dos fármacos , Interneurônios/fisiologia , Masculino , Microeletrodos , Condução Nervosa/efeitos dos fármacos , Inibição Neural/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Receptores de Orexina , Orexinas , Técnicas de Patch-Clamp , Núcleos da Rafe/citologia , Núcleos da Rafe/efeitos dos fármacos , Ratos , Receptores Acoplados a Proteínas G , Receptores de Neuropeptídeos , Serotonina/farmacologia , Tetrodotoxina/farmacologia , Ácido gama-Aminobutírico/metabolismoRESUMO
Chronic opiate exposure induces numerous neurochemical adaptations in the noradrenergic system, including upregulation of the cAMP-signaling pathway and increased expression of tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis. These adaptations are thought to compensate for opiate-mediated neuronal inhibition but also contribute to physical dependence, including withdrawal after abrupt cessation of drug exposure. Little is known about molecules that regulate the noradrenergic response to opiates. Here we report that noradrenergic locus ceruleus (LC) neurons of mice with a conditional deletion of BDNF in postnatal brain respond to chronic morphine treatment with a paradoxical downregulation of cAMP-mediated excitation and lack of dynamic regulation of TH expression. This was accompanied by a threefold reduction in opiate withdrawal symptoms despite normal antinociceptive tolerance in the BDNF-deficient mice. Although expression of TrkB, the receptor for BDNF, was high in the LC, endogenous BDNF expression was absent there and in the large majority of other noradrenergic neurons. Therefore, a BDNF-signaling pathway originating from non-noradrenergic sources is essential for opiate-induced molecular adaptations of the noradrenergic system.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Entorpecentes/farmacologia , Plasticidade Neuronal/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Animais , Comportamento Animal/fisiologia , Fator Neurotrófico Derivado do Encéfalo/deficiência , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Contagem de Células , Colforsina/farmacologia , AMP Cíclico/metabolismo , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Deleção de Genes , Genes Reporter , Genótipo , Hibridização In Situ , Técnicas In Vitro , Integrases/genética , Locus Cerúleo/citologia , Locus Cerúleo/efeitos dos fármacos , Locus Cerúleo/fisiologia , Camundongos , Camundongos Mutantes , Camundongos Transgênicos , Entorpecentes/efeitos adversos , Plasticidade Neuronal/fisiologia , Neurônios/metabolismo , Norepinefrina/metabolismo , Receptores Opioides mu/agonistas , Recombinação Genética , Índice de Gravidade de Doença , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Síndrome de Abstinência a Substâncias , Transgenes , Tirosina 3-Mono-Oxigenase/metabolismo , Proteínas Virais/genéticaRESUMO
A single sub-anesthetic dose of ketamine, a short-acting NMDA receptor blocker, induces a rapid and prolonged antidepressant effect in treatment-resistant major depression. In animal models, ketamine (24 h) reverses depression-like behaviors and associated deficits in excitatory postsynaptic currents (EPSCs) generated in apical dendritic spines of layer V pyramidal cells of medial prefrontal cortex (mPFC). However, little is known about the effects of ketamine on basal dendrites. The basal dendrites of layer V cells receive an excitatory input from pyramidal cells of the basolateral amygdala (BLA), neurons that are activated by the stress hormone CRF. Here we found that CRF induces EPSCs in PFC layer V cells and that ketamine enhanced this effect through the mammalian target of rapamycin complex 1 synaptogenic pathway; the CRF-induced EPSCs required an intact BLA input and were generated primarily in basal dendrites. In contrast to its detrimental effects on apical dendritic structure and function, chronic stress did not induce a loss of CRF-induced EPSCs in basal dendrites, thereby creating a relative imbalance in favor of amygdala inputs. The effects of ketamine were complex: ketamine enhanced apical EPSC responses in all mPFC subregions, anterior cingulate (AC), prelimbic (PL), and infralimbic (IL) but enhanced CRF-induced EPSCs only in AC and PL-responses were unchanged in IL, a critical area for suppression of stress responses. We propose that by restoring the strength of apical inputs relative to basal amygdala inputs, especially in IL, ketamine would ameliorate the hypothesized disproportional negative influence of the amygdala in chronic stress and major depression.