RESUMO
BACKGROUND: Diverse antidepressants were recently described to bind to TrkB (tyrosine kinase B) and drive a positive allosteric modulation of endogenous BDNF (brain-derived neurotrophic factor). Although neurotrophins such as BDNF can bind to p75NTR (p75 neurotrophin receptor), their precursors are the high-affinity p75NTR ligands. While part of an unrelated receptor family capable of inducing completely opposite physiological changes, TrkB and p75NTR feature a crosslike conformation dimer and carry a cholesterol-recognition amino acid consensus in the transmembrane domain. As such qualities were found to be crucial for antidepressants to bind to TrkB and drive behavioral and neuroplasticity effects, we hypothesized that their effects might also depend on p75NTR. METHODS: Enzyme-linked immunosorbent assay-based binding and nuclear magnetic resonance spectroscopy were performed to assess whether antidepressants would bind to p75NTR. HEK293T cells and a variety of in vitro assays were used to investigate whether fluoxetine (FLX) or ketamine (KET) would trigger any α- and γ-secretase-dependent p75NTR proteolysis and lead to p75NTR nuclear localization. Ocular dominance shift was performed with male and female p75NTR knockout mice to study the effects of KET and FLX on brain plasticity, in addition to pharmacological interventions to verify how p75NTR signaling is important for the effects of KET and FLX in enhancing extinction memory in male wild-type mice and rats. RESULTS: Antidepressants were found to bind to p75NTR. FLX and KET triggered the p75NTR proteolytic pathway and induced p75NTR-dependent behavioral/neuroplasticity changes. CONCLUSIONS: We hypothesize that antidepressants co-opt both BDNF/TrkB and proBDNF/p75NTR systems to induce a more efficient activity-dependent synaptic competition, thereby boosting the brain's ability for remodeling.
RESUMO
Brain-derived neurotrophic factor (BDNF) acting upon its receptor Neurotrophic tyrosine kinase receptor 2 (NTRK2, TRKB) plays a central role in the development and maintenance of synaptic function and activity- or drug-induced plasticity. TRKB possesses an inverted cholesterol recognition and alignment consensus sequence (CARC), suggesting this receptor can act as a cholesterol sensor. We have recently shown that antidepressant drugs directly bind to the CARC domain of TRKB dimers, and that this binding as well as biochemical and behavioral responses to antidepressants are lost with a mutation in the TRKB CARC motif (Tyr433Phe). However, it is not clear if this mutation can also compromise the receptor function and lead to behavioral alterations. Here, we observed that Tyr433Phe mutation does not alter BDNF binding to TRKB, or BDNF-induced dimerization of TRKB. In this line, primary cultures from embryos of heterozygous Tyr433Phe mutant mice (hTRKB.Tyr433Phe) are responsive to BDNF-induced activation of TRKB, and samples from adult mice do not show any difference on TRKB activation compared to wild-type littermates (TRKB.wt). The behavioral phenotype of hTRKB.Tyr433Phe mice is indistinguishable from the wild-type mice in cued fear conditioning, contextual discrimination task, or the elevated plus maze, whereas mice heterozygous to BDNF null allele show a phenotype in context discrimination task. Taken together, our results indicate that Tyr433Phe mutation in the TRKB CARC motif does not show signs of loss-of-function of BDNF responses, while antidepressant binding to TRKB and responses to antidepressants are lost in Tyr433Phe mutants, making them an interesting mouse model for antidepressant research.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Receptor trkB , Camundongos , Animais , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Receptor trkB/metabolismo , Antidepressivos/farmacologia , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais , Mutação/genéticaRESUMO
Psychedelics produce fast and persistent antidepressant effects and induce neuroplasticity resembling the effects of clinically approved antidepressants. We recently reported that pharmacologically diverse antidepressants, including fluoxetine and ketamine, act by binding to TrkB, the receptor for BDNF. Here we show that lysergic acid diethylamide (LSD) and psilocin directly bind to TrkB with affinities 1,000-fold higher than those for other antidepressants, and that psychedelics and antidepressants bind to distinct but partially overlapping sites within the transmembrane domain of TrkB dimers. The effects of psychedelics on neurotrophic signaling, plasticity and antidepressant-like behavior in mice depend on TrkB binding and promotion of endogenous BDNF signaling but are independent of serotonin 2A receptor (5-HT2A) activation, whereas LSD-induced head twitching is dependent on 5-HT2A and independent of TrkB binding. Our data confirm TrkB as a common primary target for antidepressants and suggest that high-affinity TrkB positive allosteric modulators lacking 5-HT2A activity may retain the antidepressant potential of psychedelics without hallucinogenic effects.
Assuntos
Antidepressivos , Alucinógenos , Dietilamida do Ácido Lisérgico , Psilocibina , Receptor trkB , Alucinógenos/metabolismo , Humanos , Células HEK293 , Sítios de Ligação , Simulação de Dinâmica Molecular , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Transdução de Sinais , Receptor trkB/metabolismo , Plasticidade Neuronal/efeitos dos fármacos , Antidepressivos/metabolismo , Regulação Alostérica , Masculino , Feminino , Animais , Camundongos , Camundongos Endogâmicos C57BL , Embrião de Mamíferos/citologia , Neurônios/efeitos dos fármacos , Dietilamida do Ácido Lisérgico/química , Dietilamida do Ácido Lisérgico/metabolismo , Dietilamida do Ácido Lisérgico/farmacologia , Psilocibina/química , Psilocibina/metabolismo , Psilocibina/farmacologiaRESUMO
Ketamine has been described as a fast-acting antidepressant, exerting effects in depressed patients and in preclinical models with a rapid onset of action. The typical antidepressant fluoxetine is known to induce plasticity in the adult rodent visual cortex, as assessed by a shift in ocular dominance, a classical model of brain plasticity, and a similar effect has been described for ketamine and its metabolite 2R,6R-hydroxynorketamine (R,R-HNK). Here, we demonstrate that ketamine (at 3 or 20 mg/kg) and R,R-HNK facilitated the shift in ocular dominance in monocularly deprived mice, after three injections, throughout the 7-day monocular deprivation regimen. Notably, the comparison between the treatments indicates a higher effect size of R,R-HNK compared with ketamine. Treatment with ketamine or R,R-HNK failed to influence the levels of perineuronal nets (PNNs) surrounding parvalbumin-positive interneurons. However, we observed in vitro that both ketamine and R,R-HNK are able to disrupt the tropomyosin-related kinase B (TRKB) interaction with the protein tyrosine phosphatase sigma (PTPσ), which upon binding to PNNs dephosphorylates TRKB. These results support a model where diverse drugs promote the reinstatement of juvenile-like plasticity by directly binding TRKB and releasing it from PTPσ regulation, without necessarily reducing PNNs deposits.
Assuntos
Ketamina , Animais , Camundongos , Antidepressivos/farmacologia , Depressão/metabolismo , Dominância Ocular , Interneurônios/metabolismo , Ketamina/farmacologia , Parvalbuminas , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores , TropomiosinaRESUMO
Nitric oxide (NO) has been long recognized as an important modulator of neural plasticity, but characterization of the molecular mechanisms involved - specially the guanylyl cyclase-independent ones - has been challenging. There is evidence that NO could modify BDNF-TRKB signaling, a key mediator of neuronal plasticity. However, the mechanism underlying the interplay of NO and TRKB remains unclear. Here we show that NO induces nitration of the tyrosine 816 in the TRKB receptor in vivo and in vitro, and that post-translational modification inhibits TRKB phosphorylation and binding of phospholipase Cγ1 (PLCγ1) to this same tyrosine residue. Additionally, nitration triggers clathrin-dependent endocytosis of TRKB through the adaptor protein AP-2 and ubiquitination, thereby increasing translocation of TRKB away from the neuronal surface and directing it towards lysosomal degradation. Accordingly, inhibition of nitric oxide increases TRKB phosphorylation and TRKB-dependent neurite branching in neuronal cultures. In vivo, chronic inhibition of neuronal nitric oxide synthase (nNOS) dramatically reduced TRKB nitration and facilitated TRKB signaling in the visual cortex, and promoted a shift in ocular dominance upon monocular deprivation - an indicator of increased plasticity. Altogether, our data describe and characterize a new molecular brake on plasticity, namely nitration of TRKB receptors.
Assuntos
Fator Neurotrófico Derivado do Encéfalo , Óxido Nítrico , Humanos , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Plasticidade Neuronal/fisiologia , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo I , Receptor trkB/metabolismo , TirosinaRESUMO
Cholesterol is an essential constituent of cell membranes. The discovery of cholesterol-recognition amino acid consensus (CRAC) motif in proteins indicated a putative direct, non-covalent interaction between cholesterol and proteins. In the present study, we evaluated the presence of a CRAC motif and its inverted version (CARC) in the transmembrane region (TMR) of the tyrosine kinase receptor family (RTK) in several species using in silico methods. CRAC motifs were found across all species analyzed, while CARC was found only in vertebrates. The tropomyosin-related kinase B (TRKB), a member of the RTK family, through interaction with its endogenous ligand brain-derived neurotrophic factor (BDNF) is a core participant in the neuronal plasticity process and exhibits a CARC motif in its TMR. Upon identifying the conserved CARC motif in the TRKB, we performed molecular dynamics simulations of the mouse TRKB.TMR. The simulations indicated that cholesterol interaction with the TRKB CARC motif occurs mainly at the central Y433 residue. Our binding assay suggested a bell-shaped effect of cholesterol on BDNF interaction with TRKB receptors, and our results suggest that CARC/CRAC motifs may play a role in the function of the RTK family TMR.
Assuntos
Colesterol , Receptores Proteína Tirosina Quinases , Animais , Fator Neurotrófico Derivado do Encéfalo , Membrana Celular , Humanos , Ligantes , Camundongos , Domínios Proteicos , Receptor trkBRESUMO
It is unclear how binding of antidepressant drugs to their targets gives rise to the clinical antidepressant effect. We discovered that the transmembrane domain of tyrosine kinase receptor 2 (TRKB), the brain-derived neurotrophic factor (BDNF) receptor that promotes neuronal plasticity and antidepressant responses, has a cholesterol-sensing function that mediates synaptic effects of cholesterol. We then found that both typical and fast-acting antidepressants directly bind to TRKB, thereby facilitating synaptic localization of TRKB and its activation by BDNF. Extensive computational approaches including atomistic molecular dynamics simulations revealed a binding site at the transmembrane region of TRKB dimers. Mutation of the TRKB antidepressant-binding motif impaired cellular, behavioral, and plasticity-promoting responses to antidepressants in vitro and in vivo. We suggest that binding to TRKB and allosteric facilitation of BDNF signaling is the common mechanism for antidepressant action, which may explain why typical antidepressants act slowly and how molecular effects of antidepressants are translated into clinical mood recovery.
Assuntos
Antidepressivos/farmacologia , Receptor trkB/metabolismo , Animais , Antidepressivos/química , Antidepressivos/metabolismo , Sítios de Ligação , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Linhagem Celular , Colesterol/metabolismo , Embrião de Mamíferos , Fluoxetina/química , Fluoxetina/metabolismo , Fluoxetina/farmacologia , Hipocampo/metabolismo , Humanos , Camundongos , Modelos Animais , Simulação de Dinâmica Molecular , Domínios Proteicos , Ratos , Receptor trkB/química , Córtex Visual/metabolismoRESUMO
Heightened neuronal plasticity expressed during early postnatal life has been thought to permanently decline once critical periods have ended. For example, monocular deprivation is able to shift ocular dominance in the mouse visual cortex during the first months of life, but this effect is lost later in life. However, various treatments, such as the antidepressant fluoxetine, can reactivate a critical period-like plasticity in the adult brain. When monocular deprivation is supplemented with chronic fluoxetine administration, a major shift in ocular dominance is produced after the critical period has ended. In the current study, we characterized the temporal patterns of fluoxetine-induced plasticity in the adult mouse visual cortex, using in vivo optical imaging. We found that artificially induced plasticity in ocular dominance extended beyond the duration of the naturally occurring critical period and continued as long as fluoxetine was administered. However, this fluoxetine-induced plasticity period ended as soon as the drug was not given. These features of antidepressant-induced plasticity may be useful when designing treatment strategies involving long-term antidepressant treatment in humans.
Assuntos
Antidepressivos/farmacologia , Fluoxetina/farmacologia , Plasticidade Neuronal , Córtex Visual/fisiologia , Animais , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Tempo , Córtex Visual/efeitos dos fármacosRESUMO
Extensive loss of noradrenaline-containing neurons and fibers is a nearly invariant feature of Alzheimer's Disease (AD). However, the exact noradrenergic contribution to cognitive and histopathological changes in AD is still unclear. Here, this issue was addressed following selective lesioning and intrahippocampal implantation of embryonic noradrenergic progenitors in developing rats. Starting from about 3 months and up to 12 months post-surgery, animals underwent behavioral tests to evaluate sensory-motor, as well as spatial learning and memory, followed by post-mortem morphometric analyses. At 9 months, Control, Lesioned and Lesion + Transplant animals exhibited equally efficient sensory-motor and reference memory performance. Interestingly, working memory abilities were seen severely impaired in Lesion-only rats and fully recovered in Transplanted rats, and appeared partly lost again 2 months after ablation of the implanted neuroblasts. Morphological analyses confirmed the almost total lesion-induced noradrenergic neuronal and terminal fiber loss, the near-normal reinnervation of the hippocampus promoted by the transplants, and its complete removal by the second lesion. Notably, the noradrenergic-rich transplants normalized also the nuclear expression of the transactive response DNA-binding protein 43 (TDP-43) in various hippocampal subregions, whose cytoplasmic (i.e., pathological) occurrence appeared dramatically increased as a result of the lesions. Thus, integrity of ascending noradrenergic inputs to the hippocampus may be required for the regulation of specific aspects of learning and memory and to prevent TDP-43 tissue pathology.