RESUMO
Major depressive disorder is typically treated with selective serotonin reuptake inhibitors (SSRIs), however, SSRIs take approximately six weeks to produce therapeutic effects, if any. Not surprisingly, there has been great interest in findings that low doses of ketamine, a non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist, produce rapid and long-lasting antidepressant effects. Preclinical studies show that the antidepressant-like effects of ketamine are dependent upon availability of serotonin, and that ketamine increases extracellular serotonin, yet the mechanism by which this occurs is unknown. Here we examined the role of the high-affinity, low-capacity serotonin transporter (SERT), and the plasma membrane monoamine transporter (PMAT), a low-affinity, high-capacity transporter for serotonin, as mechanisms contributing to ketamine's ability to increase extracellular serotonin and produce antidepressant-like effects. Using high-speed chronoamperometry to measure real-time clearance of serotonin from CA3 region of hippocampus in vivo, we found ketamine robustly inhibited serotonin clearance in wild-type mice, an effect that was lost in mice constitutively lacking SERT or PMAT. As expected, in wild-type mice, ketamine produced antidepressant-like effects in the forced swim test. Mapping onto our neurochemical findings, the antidepressant-like effects of ketamine were lost in mice lacking SERT or PMAT. Future research is needed to understand how constitutive loss of either SERT or PMAT, and compensation that occurs in other systems, is sufficient to void ketamine of its ability to inhibit serotonin clearance and produce antidepressant-like effects. Taken together with existing literature, a critical role for serotonin, and its inhibition of uptake via SERT and PMAT, cannot be ruled out as important contributing factors to ketamine's antidepressant mechanism of action. Combined with what is already known about ketamine's action at NMDA receptors, these studies help lead the way to the development of drugs that lack ketamine's abuse potential but have superior efficacy in treating depression.
Assuntos
Antidepressivos/farmacologia , Proteínas de Transporte de Nucleosídeo Equilibrativas/metabolismo , Ketamina/farmacologia , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Animais , Região CA3 Hipocampal/efeitos dos fármacos , Região CA3 Hipocampal/metabolismo , Proteínas de Transporte de Nucleosídeo Equilibrativas/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Núcleo Accumbens/efeitos dos fármacos , Núcleo Accumbens/metabolismo , Serotonina/metabolismo , Proteínas da Membrana Plasmática de Transporte de Serotonina/genéticaRESUMO
BACKGROUND AND PURPOSE: Tau pathology contributes to a bidirectional relationship between sleep disruption and neurodegenerative disease. Tau transgenic rTg4510 mice model tauopathy symptoms, including sleep/wake disturbances, which manifest as marked hyperarousal. This phenotype can be prevented by early transgene suppression; however, whether hyperarousal can be rescued after onset is unknown. EXPERIMENTAL APPROACH: Three 8-week experiments were conducted with wild-type and rTg4510 mice after age of onset of hyperarousal (4.5 months): (1) Tau transgene suppression with doxycycline (200 ppm); (2) inactive phase rapid eye movement (REM) sleep enhancement with the dual orexin receptor antagonist suvorexant (50 mg·kg-1 ·day-1 ); or (3) Active phase non-NREM (NREM) and REM sleep enhancement using the selective orexin 2 (OX2 ) receptor antagonist MK-1064 (40 mg·kg-1 ·day-1 ). Sleep was assessed using polysomnography, cognition using the Barnes maze, and tau pathology using immunoblotting and/or immunohistochemistry. KEY RESULTS: Tau transgene suppression improved tauopathy and hippocampal-dependent spatial memory, but did not modify hyperarousal. Pharmacological rescue of REM sleep deficits did not improve spatial memory or tau pathology. In contrast, normalising hyperarousal by increasing both NREM and REM sleep via OX2 receptor antagonism restored spatial memory, independently of tauopathy, but only in male rTg4510 mice. OX2 receptor antagonism induced only short-lived hypnotic responses in female rTg4510 mice and did not improve spatial memory, indicating a tau- and sex-dependent disruption of OX2 receptor signalling. CONCLUSIONS AND IMPLICATIONS: Pharmacologically reducing hyperarousal corrects tau-induced sleep/wake and cognitive deficits. Tauopathy causes sex-dependent disruptions of OX2 receptor signalling/function, which may have implications for choice of hypnotic therapeutics in tauopathies.
Assuntos
Doenças Neurodegenerativas , Receptores de Orexina , Transtornos do Sono-Vigília , Tauopatias , Animais , Feminino , Masculino , Camundongos , Cognição , Modelos Animais de Doenças , Hipnóticos e Sedativos/farmacologia , Camundongos Transgênicos , Orexinas , Sono/fisiologia , Tauopatias/tratamento farmacológico , Tauopatias/genética , Tauopatias/patologia , Vigília/fisiologia , Receptores de Orexina/metabolismo , Antagonistas dos Receptores de Orexina/farmacologia , Antagonistas dos Receptores de Orexina/uso terapêuticoRESUMO
BACKGROUND: Efficient DNA repair in response to standard chemo and radiation therapies often contributes to glioblastoma (GBM) therapy resistance. Understanding the mechanisms of therapy resistance and identifying the drugs that enhance the therapeutic efficacy of standard therapies may extend the survival of GBM patients. In this study, we investigated the role of KDM1A/LSD1 in DNA double-strand break (DSB) repair and a combination of KDM1A inhibitor and temozolomide (TMZ) in vitro and in vivo using patient-derived glioma stem cells (GSCs). METHODS: Brain bioavailability of the KDM1A inhibitor (NCD38) was established using LS-MS/MS. The effect of a combination of KDM1A knockdown or inhibition with TMZ was studied using cell viability and self-renewal assays. Mechanistic studies were conducted using CUT&Tag-seq, RNA-seq, RT-qPCR, western blot, homologous recombination (HR) and non-homologous end joining (NHEJ) reporter, immunofluorescence, and comet assays. Orthotopic murine models were used to study efficacy in vivo. RESULTS: TCGA analysis showed KDM1A is highly expressed in TMZ-treated GBM patients. Knockdown or knockout or inhibition of KDM1A enhanced TMZ efficacy in reducing the viability and self-renewal of GSCs. Pharmacokinetic studies established that NCD38 readily crosses the blood-brain barrier. CUT&Tag-seq studies showed that KDM1A is enriched at the promoters of DNA repair genes and RNA-seq studies confirmed that KDM1A inhibition reduced their expression. Knockdown or inhibition of KDM1A attenuated HR and NHEJ-mediated DNA repair capacity and enhanced TMZ-mediated DNA damage. A combination of KDM1A knockdown or inhibition and TMZ treatment significantly enhanced the survival of tumor-bearing mice. CONCLUSIONS: Our results provide evidence that KDM1A inhibition sensitizes GBM to TMZ via attenuation of DNA DSB repair pathways.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Glioma , Animais , Camundongos , Temozolomida/farmacologia , Temozolomida/uso terapêutico , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Lisina/genética , Lisina/farmacologia , Lisina/uso terapêutico , Quebras de DNA de Cadeia Dupla , Espectrometria de Massas em Tandem , Linhagem Celular Tumoral , Glioma/tratamento farmacológico , Reparo do DNA , DNA/farmacologia , DNA/uso terapêutico , Histona Desmetilases/genética , Histona Desmetilases/farmacologia , Histona Desmetilases/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Antineoplásicos Alquilantes/farmacologia , Antineoplásicos Alquilantes/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Selective serotonin reuptake inhibitors (SSRIs) are less efficacious in treating depression in children than in adults. SSRIs block serotonin uptake via the high-affinity, low-capacity serotonin transporter. However, the low-affinity, high-capacity organic cation transporter 3 (OCT3) and plasma membrane monoamine transporter (PMAT) are emerging as important players in serotonin uptake. We hypothesized that OCT3 and/or PMAT are functionally upregulated in juveniles, thereby buffering SSRIs' ability to enhance serotonergic neurotransmission. Unlike in adult mice, we found the OCT/PMAT blocker, decynium-22, to have standalone antidepressant-like effects in juveniles. Using in vivo high-speed chronoamperometry, we found that juveniles clear serotonin from the CA3 region of the hippocampus ~2-fold faster than adult mice. Cell density did not differ between ages, suggesting that faster serotonin clearance in juveniles is unrelated to faster diffusion through the extracellular matrix. Western blot and immunohistochemistry showed that juvenile mice have modestly greater expression of PMAT than adults, whereas OCT3 expression in the CA3 region of the hippocampus was similar between ages. Together, these data suggest that faster serotonin clearance and antidepressant-like effects of decynium-22 in juvenile mice may be due to functionally upregulated PMAT. Faster serotonin clearance via PMAT in juveniles may contribute to reduced therapeutic efficacy of SSRIs in children relative to adults.