RESUMEN
Persistent cancer cells are the discrete and usually undetected cells that survive cancer drug treatment and constitute a major cause of treatment failure. These cells are characterized by their slow proliferation, highly flexible energy consumption, adaptation to their microenvironment, and phenotypic plasticity. Mechanisms that underlie their persistence offer highly coveted and sought-after therapeutic targets, and include diverse epigenetic, transcriptional, and translational regulatory processes, as well as complex cell-cell interactions. Although the successful clinical targeting of persistent cancer cells remains to be realized, immense progress has been made in understanding their persistence, yielding promising preclinical results.
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Neoplasias/patología , Animales , Supervivencia Celular , Metabolismo Energético , Transición Epitelial-Mesenquimal , Humanos , Mitocondrias/metabolismo , Neoplasias/terapia , Microambiente TumoralRESUMEN
Small molecule neurotensin receptor 1 (NTSR1) agonists have been pursued for more than 40 years as potential therapeutics for psychiatric disorders, including drug addiction. Clinical development of NTSR1 agonists has, however, been precluded by their severe side effects. NTSR1, a G protein-coupled receptor (GPCR), signals through the canonical activation of G proteins and engages ß-arrestins to mediate distinct cellular signaling events. Here, we characterize the allosteric NTSR1 modulator SBI-553. This small molecule not only acts as a ß-arrestin-biased agonist but also extends profound ß-arrestin bias to the endogenous ligand by selectively antagonizing G protein signaling. SBI-553 shows efficacy in animal models of psychostimulant abuse, including cocaine self-administration, without the side effects characteristic of balanced NTSR1 agonism. These findings indicate that NTSR1 G protein and ß-arrestin activation produce discrete and separable physiological effects, thus providing a strategy to develop safer GPCR-targeting therapeutics with more directed pharmacological action.
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Conducta Adictiva/metabolismo , Receptores de Neurotensina/metabolismo , beta-Arrestinas/metabolismo , Regulación Alostérica/efectos de los fármacos , Regulación Alostérica/fisiología , Animales , Conducta Adictiva/tratamiento farmacológico , Línea Celular , Femenino , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Animales , Receptores Acoplados a Proteínas G/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Bibliotecas de Moléculas Pequeñas/farmacologíaRESUMEN
The κ-opioid receptor (KOP) mediates the actions of opioids with hallucinogenic, dysphoric, and analgesic activities. The design of KOP analgesics devoid of hallucinatory and dysphoric effects has been hindered by an incomplete structural and mechanistic understanding of KOP agonist actions. Here, we provide a crystal structure of human KOP in complex with the potent epoxymorphinan opioid agonist MP1104 and an active-state-stabilizing nanobody. Comparisons between inactive- and active-state opioid receptor structures reveal substantial conformational changes in the binding pocket and intracellular and extracellular regions. Extensive structural analysis and experimental validation illuminate key residues that propagate larger-scale structural rearrangements and transducer binding that, collectively, elucidate the structural determinants of KOP pharmacology, function, and biased signaling. These molecular insights promise to accelerate the structure-guided design of safer and more effective κ-opioid receptor therapeutics.
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Simulación del Acoplamiento Molecular , Receptores Opioides kappa/química , Analgésicos/química , Analgésicos/farmacología , Animales , Sitios de Unión , Células HEK293 , Humanos , Simulación de Dinámica Molecular , Morfinanos/química , Morfinanos/farmacología , Unión Proteica , Estabilidad Proteica , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/metabolismo , Células Sf9 , SpodopteraRESUMEN
The super elongation complex (SEC) is required for robust and productive transcription through release of RNA polymerase II (Pol II) with its P-TEFb module and promoting transcriptional processivity with its ELL2 subunit. Malfunction of SEC contributes to multiple human diseases including cancer. Here, we identify peptidomimetic lead compounds, KL-1 and its structural homolog KL-2, which disrupt the interaction between the SEC scaffolding protein AFF4 and P-TEFb, resulting in impaired release of Pol II from promoter-proximal pause sites and a reduced average rate of processive transcription elongation. SEC is required for induction of heat-shock genes and treating cells with KL-1 and KL-2 attenuates the heat-shock response from Drosophila to human. SEC inhibition downregulates MYC and MYC-dependent transcriptional programs in mammalian cells and delays tumor progression in a mouse xenograft model of MYC-driven cancer, indicating that small-molecule disruptors of SEC could be used for targeted therapy of MYC-induced cancer.
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Antineoplásicos/farmacología , Neoplasias Experimentales/tratamiento farmacológico , Factor B de Elongación Transcripcional Positiva/metabolismo , Proteínas Represoras/metabolismo , Elongación de la Transcripción Genética/efectos de los fármacos , Factores de Elongación Transcripcional/metabolismo , Animales , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Drosophila , Femenino , Células HCT116 , Células HEK293 , Respuesta al Choque Térmico , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Unión Proteica/efectos de los fármacos , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo , ARN Polimerasa II/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacologíaRESUMEN
KRASG12C was recently identified to be potentially druggable by allele-specific covalent targeting of Cys-12 in vicinity to an inducible allosteric switch II pocket (S-IIP). Success of this approach requires active cycling of KRASG12C between its active-GTP and inactive-GDP conformations as accessibility of the S-IIP is restricted only to the GDP-bound state. This strategy proved feasible for inhibiting mutant KRAS in vitro; however, it is uncertain whether this approach would translate to in vivo. Here, we describe structure-based design and identification of ARS-1620, a covalent compound with high potency and selectivity for KRASG12C. ARS-1620 achieves rapid and sustained in vivo target occupancy to induce tumor regression. We use ARS-1620 to dissect oncogenic KRAS dependency and demonstrate that monolayer culture formats significantly underestimate KRAS dependency in vivo. This study provides in vivo evidence that mutant KRAS can be selectively targeted and reveals ARS-1620 as representing a new generation of KRASG12C-specific inhibitors with promising therapeutic potential.
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Antineoplásicos/farmacología , Neoplasias Experimentales/tratamiento farmacológico , Piperazinas/farmacología , Proteínas Proto-Oncogénicas p21(ras)/antagonistas & inhibidores , Quinazolinas/farmacología , Animales , Antineoplásicos/química , Antineoplásicos/uso terapéutico , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Femenino , Células HCT116 , Células HEK293 , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Simulación del Acoplamiento Molecular , Mutación , Piperazinas/química , Piperazinas/uso terapéutico , Unión Proteica , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Quinazolinas/química , Quinazolinas/uso terapéuticoRESUMEN
Acute myeloid leukemia with KMT2A (MLL) rearrangements is characterized by specific patterns of gene expression and enhancer architecture, implying unique core transcriptional regulatory circuitry. Here, we identified the transcription factors MEF2D and IRF8 as selective transcriptional dependencies of KMT2A-rearranged AML, where MEF2D displays partially redundant functions with its paralog, MEF2C. Rapid transcription factor degradation followed by measurements of genome-wide transcription rates and superresolution microscopy revealed that MEF2D and IRF8 form a distinct core regulatory module with a narrow direct transcriptional program that includes activation of the key oncogenes MYC, HOXA9, and BCL2. Our study illustrates a mechanism of context-specific transcriptional addiction whereby a specific AML subclass depends on a highly specialized core regulatory module to directly enforce expression of common leukemia oncogenes.
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Leucemia Mieloide Aguda , Proteína de la Leucemia Mieloide-Linfoide , Reordenamiento Génico , Humanos , Factores Reguladores del Interferón/genética , Factores Reguladores del Interferón/metabolismo , Leucemia Mieloide Aguda/genética , Proteína de la Leucemia Mieloide-Linfoide/genética , Proteína de la Leucemia Mieloide-Linfoide/metabolismo , Oncogenes/genéticaRESUMEN
The transformed state in acute leukemia requires gene regulatory programs involving transcription factors and chromatin modulators. Here, we uncover an IRF8-MEF2D transcriptional circuit as an acute myeloid leukemia (AML)-biased dependency. We discover and characterize the mechanism by which the chromatin "reader" ZMYND8 directly activates IRF8 in parallel with the MYC proto-oncogene through their lineage-specific enhancers. ZMYND8 is essential for AML proliferation in vitro and in vivo and associates with MYC and IRF8 enhancer elements that we define in cell lines and in patient samples. ZMYND8 occupancy at IRF8 and MYC enhancers requires BRD4, a transcription coactivator also necessary for AML proliferation. We show that ZMYND8 binds to the ET domain of BRD4 via its chromatin reader cassette, which in turn is required for proper chromatin occupancy and maintenance of leukemic growth in vivo. Our results rationalize ZMYND8 as a potential therapeutic target for modulating essential transcriptional programs in AML.
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Factores Reguladores del Interferón/metabolismo , Leucemia Mieloide Aguda/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Línea Celular Tumoral , Proliferación Celular/genética , Cromatina/genética , Elementos de Facilitación Genéticos/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Factores Reguladores del Interferón/genética , Leucemia Mieloide Aguda/genética , Proteínas Nucleares/metabolismo , Regiones Promotoras Genéticas/genética , Proto-Oncogenes Mas , Factores de Transcripción/metabolismo , Transcripción Genética/genética , Proteínas Supresoras de Tumor/genéticaRESUMEN
Punishment such as electric shock or physical discipline employs a mixture of physical pain and emotional distress to induce behavior modification. However, a neural circuit that produces behavior modification by selectively focusing the emotional component, while bypassing the pain typically induced by peripheral nociceptor activation, is not well studied. Here, we show that genetically silencing the activity of neurons expressing calcitonin gene-related peptide (CGRP) in the parabrachial nucleus blocks the suppression of addictive-like behavior induced by footshock. Furthermore, activating CGRP neurons suppresses not only addictive behavior induced by self-stimulating dopamine neurons but also behavior resulting from self-administering cocaine, without eliciting nocifensive reactions. Moreover, among multiple downstream targets of CGRP neurons, terminal activation of CGRP in the central amygdala is effective, mimicking the results of cell body stimulation. Our results indicate that unlike conventional electric footshock, stimulation of CGRP neurons does not activate peripheral nociceptors but effectively curb addictive behavior.
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Conducta Adictiva , Péptido Relacionado con Gen de Calcitonina , Neuronas , Núcleos Parabraquiales , Animales , Núcleos Parabraquiales/metabolismo , Núcleos Parabraquiales/fisiología , Péptido Relacionado con Gen de Calcitonina/metabolismo , Ratones , Neuronas/metabolismo , Neuronas/fisiología , Conducta Adictiva/metabolismo , Masculino , Neuronas Dopaminérgicas/metabolismo , Neuronas Dopaminérgicas/fisiología , Cocaína/farmacología , Conducta Animal/fisiologíaRESUMEN
ΔfosB is an alternatively spliced product of the FosB gene that is essential for dopamine-induced reward pathways and that acts as a master switch for addiction. However, the molecular mechanisms of its generation and regulation by dopamine signaling are unknown. Here, we report that dopamine D1 receptor signaling synergizes with the activin/ALK4/Smad3 pathway to potentiate the generation of ΔFosB mRNA in medium spiny neurons (MSNs) of the nucleus accumbens (NAc) via activation of the RNA-binding protein PCBP1, a regulator of mRNA splicing. Concurrent activation of PCBP1 and Smad3 by D1 and ALK4 signaling induced their interaction, nuclear translocation, and binding to sequences in exon-4 and intron-4 of FosB mRNA. Ablation of either ALK4 or PCBP1 in MSNs impaired ΔFosB mRNA induction and nuclear translocation of ΔFosB protein in response to repeated co-stimulation of D1 and ALK4 receptors. Finally, ALK4 is required in NAc MSNs of adult mice for behavioral sensitization to cocaine. These findings uncover an unexpected mechanism for ΔFosB generation and drug-induced sensitization through convergent dopamine and ALK4 signaling.
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Cocaína , Proteínas de Unión al ADN/metabolismo , Proteínas de Unión al ARN/metabolismo , Receptores de Activinas Tipo I/metabolismo , Empalme Alternativo , Animales , Cocaína/metabolismo , Cocaína/farmacología , Dopamina/metabolismo , Ratones , Ratones Endogámicos C57BL , Núcleo Accumbens , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , ARN Mensajero/metabolismo , Receptores de Dopamina D1/genética , Receptores de Dopamina D1/metabolismoRESUMEN
The escalating drug addiction crisis in the United States underscores the urgent need for innovative therapeutic strategies. This study embarked on an innovative and rigorous strategy to unearth potential drug repurposing candidates for opioid and cocaine addiction treatment, bridging the gap between transcriptomic data analysis and drug discovery. We initiated our approach by conducting differential gene expression analysis on addiction-related transcriptomic data to identify key genes. We propose a novel topological differentiation to identify key genes from a protein-protein interaction network derived from DEGs. This method utilizes persistent Laplacians to accurately single out pivotal nodes within the network, conducting this analysis in a multiscale manner to ensure high reliability. Through rigorous literature validation, pathway analysis and data-availability scrutiny, we identified three pivotal molecular targets, mTOR, mGluR5 and NMDAR, for drug repurposing from DrugBank. We crafted machine learning models employing two natural language processing (NLP)-based embeddings and a traditional 2D fingerprint, which demonstrated robust predictive ability in gauging binding affinities of DrugBank compounds to selected targets. Furthermore, we elucidated the interactions of promising drugs with the targets and evaluated their drug-likeness. This study delineates a multi-faceted and comprehensive analytical framework, amalgamating bioinformatics, topological data analysis and machine learning, for drug repurposing in addiction treatment, setting the stage for subsequent experimental validation. The versatility of the methods we developed allows for applications across a range of diseases and transcriptomic datasets.
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Reposicionamiento de Medicamentos , Transcriptoma , Estados Unidos , Reposicionamiento de Medicamentos/métodos , Reproducibilidad de los Resultados , Perfilación de la Expresión Génica , Biología Computacional/métodosRESUMEN
Epigenetic control of regulatory networks is only partially understood. Expression of Insulin-like growth factor-II (IGF2) is controlled by genomic imprinting, mediated by silencing of the maternal allele. Loss of imprinting of IGF2 (LOI) is linked to intestinal and colorectal cancers, causally in murine models and epidemiologically in humans. However, the molecular underpinnings of the LOI phenotype are not clear. Surprisingly, in LOI cells, we find a reversal of the relative activities of two canonical signaling pathways triggered by IGF2, causing further rebalancing between pro- and anti-apoptotic signaling. A predictive mathematical model shows that this network rebalancing quantitatively accounts for the effect of receptor tyrosine kinase inhibition in both WT and LOI cells. This mechanism also quantitatively explains both the stable LOI phenotype and the therapeutic window for selective killing of LOI cells, and thus prevention of epigenetically controlled cancers. These findings suggest a framework for understanding epigenetically modified cell signaling.
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Neoplasias Colorrectales/genética , Epigénesis Genética/genética , Impresión Genómica/genética , Factor II del Crecimiento Similar a la Insulina/genética , Animales , Apoptosis/genética , Línea Celular Tumoral , Neoplasias Colorrectales/patología , Regulación Neoplásica de la Expresión Génica , Humanos , Ratones , Fenotipo , Transducción de SeñalRESUMEN
Bacterial toxin-antitoxin (TA) modules are abundant genetic elements that encode a toxin protein capable of inhibiting cell growth and an antitoxin that counteracts the toxin. The majority of toxins are enzymes that interfere with translation or DNA replication, but a wide variety of molecular activities and cellular targets have been described. Antitoxins are proteins or RNAs that often control their cognate toxins through direct interactions and, in conjunction with other signaling elements, through transcriptional and translational regulation of TA module expression. Three major biological functions of TA modules have been discovered, post-segregational killing ("plasmid addiction"), abortive infection (bacteriophage immunity through altruistic suicide), and persister formation (antibiotic tolerance through dormancy). In this review, we summarize the current state of the field and highlight how multiple levels of regulation shape the conditions of toxin activation to achieve the different biological functions of TA modules.
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Antitoxinas/metabolismo , Bacterias/metabolismo , Proteínas Bacterianas/metabolismo , Toxinas Bacterianas/metabolismo , ARN Bacteriano/metabolismo , Antitoxinas/química , Antitoxinas/genética , Bacterias/genética , Bacterias/inmunología , Bacterias/patogenicidad , Proteínas Bacterianas/química , Proteínas Bacterianas/genética , Toxinas Bacterianas/química , Toxinas Bacterianas/genética , Farmacorresistencia Bacteriana/genética , Evolución Molecular , Regulación Bacteriana de la Expresión Génica , Inmunidad Innata , Viabilidad Microbiana , Modelos Moleculares , Conformación de Ácido Nucleico , Conformación Proteica , Procesamiento Postranscripcional del ARN , ARN Bacteriano/química , ARN Bacteriano/genética , Relación Estructura-Actividad , Transcripción GenéticaRESUMEN
Substance use disorder is a major concern, with few therapeutic options. Heparan sulfate (HS) and chondroitin sulfate (CS) interact with a plethora of growth factors and their receptors and have profound effects on cellular signaling. Thus, targeting these dynamic interactions might represent a potential novel therapeutic modality. In the present study, we performed mass spectrometry-based glycomic and proteomic analysis to understand the effects of cocaine and methamphetamine (METH) on HS, CS, and the proteome of two brain regions critically involved in drug addiction: the lateral hypothalamus and the striatum. We observed that cocaine and METH significantly alter HS and CS abundances as well as sulfate contents and composition. In particular, repeated METH or cocaine treatments reduced CS 4-O-sulfation and increased CS 6-O-sulfation. Since C4S and C6S exercise differential effects on axon growth, regeneration, and plasticity, these changes likely contribute to drug-induced neural plasticity in these brain regions. Notably, we observed that restoring these alterations by increasing CS 4-0 levels in the lateral hypothalamus by adeno-associated virus delivery of an shRNA to arylsulfatase B (N-acetylgalactosamine-4-sulfatase) ameliorated anxiety and prevented the expression of preference for cocaine in a novelty induced conditioned place preference test during cocaine withdrawal. Finally, proteomics analyses revealed a number of aberrant proteins in METH- and cocaine-treated versus saline-treated mice, including myelin proteolipid protein, calcium/calmodulin-dependent protein kinase type II subunit alpha, synapsin-2, tenascin-R, calnexin, annexin A7, hepatoma-derived growth factor, neurocan, and CSPG5, and oxidative phosphorylation among the top perturbed pathway. Taken together, these data support the role of HS, CS, and associated proteins in stimulants abuse and suggest that manipulation of HSPGs can represent a novel therapeutic strategy.
RESUMEN
Activating JAK2 point mutations are implicated in the pathogenesis of myeloid and lymphoid malignancies, including high-risk B-cell acute lymphoblastic leukemia (B-ALL). In preclinical studies, treatment of JAK2 mutant leukemias with type I JAK2 inhibitors (e.g., Food and Drug Administration [FDA]-approved ruxolitinib) provided limited single-agent responses, possibly due to paradoxical JAK2Y1007/1008 hyperphosphorylation induced by these agents. To determine the importance of mutant JAK2 in B-ALL initiation and maintenance, we developed unique genetically engineered mouse models of B-ALL driven by overexpressed Crlf2 and mutant Jak2, recapitulating the genetic aberrations found in human B-ALL. While expression of mutant Jak2 was necessary for leukemia induction, neither its continued expression nor enzymatic activity was required to maintain leukemia survival and rapid proliferation. CRLF2/JAK2 mutant B-ALLs with sustained depletion or pharmacological inhibition of JAK2 exhibited enhanced expression of c-Myc and prominent up-regulation of c-Myc target genes. Combined indirect targeting of c-Myc using the BET bromodomain inhibitor JQ1 and direct targeting of JAK2 with ruxolitinib potently killed JAK2 mutant B-ALLs.
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Janus Quinasa 2/genética , Janus Quinasa 2/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/fisiopatología , Animales , Antineoplásicos/farmacología , Azepinas/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Humanos , Masculino , Ratones , Mutación , Nitrilos , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamiento farmacológico , Pirazoles/farmacología , Pirazoles/uso terapéutico , Pirimidinas , Interferencia de ARN , Receptores de Citocinas/genética , Transcriptoma , Triazoles/farmacologíaRESUMEN
Opioid receptors (ORs) are undisputed targets for the treatment of pain. Unfortunately, targeting these receptors therapeutically poses significant challenges including addiction, dependence, tolerance, and the appearance of side effects, such as respiratory depression and constipation. Moreover, misuse of prescription and illicit narcotics has resulted in the current opioid crisis. The mu-opioid receptor (MOR) is the cellular mediator of the effects of most commonly used opioids, and is a prototypical G protein-coupled receptor (GPCR) where new pharmacological, signalling and cell biology concepts have been coined. This review summarises the knowledge of the life cycle of this therapeutic target, including its biogenesis, trafficking to and from the plasma membrane, and how the regulation of these processes impacts its function and is related to pathophysiological conditions.
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Analgésicos Opioides , Receptores Opioides , Analgésicos Opioides/farmacología , Animales , Tolerancia a Medicamentos , Estadios del Ciclo de VidaRESUMEN
Cocaine use disorder is a significant public health issue without an effective pharmacological treatment. Successful treatments are hindered in part by an incomplete understanding of the molecular mechanisms that underlie long-lasting maladaptive plasticity and addiction-like behaviors. Here, we leverage a large RNA sequencing dataset to generate gene coexpression networks across six interconnected regions of the brain's reward circuitry from mice that underwent saline or cocaine self-administration. We identify phosphodiesterase 1b (Pde1b), a Ca2+/calmodulin-dependent enzyme that increases cAMP and cGMP hydrolysis, as a central hub gene within a nucleus accumbens (NAc) gene module that was bioinformatically associated with addiction-like behavior. Chronic cocaine exposure increases Pde1b expression in NAc D2 medium spiny neurons (MSNs) in male but not female mice. Viral-mediated Pde1b overexpression in NAc reduces cocaine self-administration in female rats but increases seeking in both sexes. In female mice, overexpressing Pde1b in D1 MSNs attenuates the locomotor response to cocaine, with the opposite effect in D2 MSNs. Overexpressing Pde1b in D1/D2 MSNs had no effect on the locomotor response to cocaine in male mice. At the electrophysiological level, Pde1b overexpression reduces sEPSC frequency in D1 MSNs and regulates the excitability of NAc MSNs. Lastly, Pde1b overexpression significantly reduced the number of differentially expressed genes (DEGs) in NAc following chronic cocaine, with discordant effects on gene transcription between sexes. Together, we identify novel gene modules across the brain's reward circuitry associated with addiction-like behavior and explore the role of Pde1b in regulating the molecular, cellular, and behavioral responses to cocaine.
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Trastornos Relacionados con Cocaína , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 1 , Redes Reguladoras de Genes , Ratones Endogámicos C57BL , Núcleo Accumbens , Caracteres Sexuales , Animales , Masculino , Femenino , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 1/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 1/metabolismo , Ratones , Trastornos Relacionados con Cocaína/genética , Trastornos Relacionados con Cocaína/metabolismo , Redes Reguladoras de Genes/efectos de los fármacos , Redes Reguladoras de Genes/genética , Núcleo Accumbens/efectos de los fármacos , Núcleo Accumbens/metabolismo , Ratas , Cocaína/farmacología , RecompensaRESUMEN
The initiation of abstinence after chronic drug self-administration is stressful. Cocaine-seeking behavior on the first day of the absence of the expected drug (Extinction Day 1, ED1) is reduced by blocking 5-HT signaling in dorsal hippocampal cornu ammonis 1 (CA1) in both male and female rats. We hypothesized that the experience of ED1 can substantially influence later relapse behavior and that dorsal raphe (DR) serotonin (5-HT) input to CA1 may be involved. We inhibited 5-HT1A/1B receptors (WAY-100635 plus GR-127935), or DR input (chemogenetics), in CA1 on ED1 to test the role of this pathway on cocaine-seeking persistence 2â weeks later. We also inhibited 5-HT1A or 5-HT1B receptors in CA1 during conditioned place preference (CPP) for cocaine, to examine mechanisms involved in the persistent effects of ED1 manipulations. Inhibition of DR inputs, or 5-HT1A/1B signaling, in CA1 decreased drug seeking on ED1 and decreased cocaine seeking 2â weeks later revealing that 5-HT signaling in CA1 during ED1 contributes to persistent drug seeking during abstinence. In addition, 5-HT1B antagonism alone transiently decreased drug-associated memory performance when given prior to a CPP test, whereas similar antagonism of 5-HT1A alone had no such effect but blocked CPP retrieval on a test 24â h later. These CPP findings are consistent with prior work showing that DR inputs to CA1 augment recall of the drug-associated context and drug seeking via 5-HT1B receptors and prevent consolidation of the updated nondrug context via 5-HT1A receptors. Thus, treatments that modulate 5-HT-dependent memory mechanisms in CA1 during initial abstinence may facilitate later maintenance of abstinence.
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Cocaína , Comportamiento de Búsqueda de Drogas , Oxadiazoles , Serotonina , Animales , Masculino , Comportamiento de Búsqueda de Drogas/fisiología , Comportamiento de Búsqueda de Drogas/efectos de los fármacos , Ratas , Serotonina/metabolismo , Femenino , Cocaína/administración & dosificación , Cocaína/farmacología , Hipocampo/metabolismo , Hipocampo/efectos de los fármacos , Piridinas/farmacología , Antagonistas de la Serotonina/farmacología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Piperazinas/farmacología , Ratas Sprague-Dawley , Trastornos Relacionados con Cocaína/metabolismo , Trastornos Relacionados con Cocaína/psicología , Autoadministración , Extinción Psicológica/efectos de los fármacos , Extinción Psicológica/fisiología , Receptor de Serotonina 5-HT1B/metabolismo , Región CA1 Hipocampal/efectos de los fármacos , Región CA1 Hipocampal/metabolismoRESUMEN
The ventral pallidum (VP) is a central hub in the reward circuitry with diverse projections that have different behavioral roles attributed mostly to the connectivity with the downstream target. However, different VP projections may represent, as in the striatum, separate neuronal populations that differ in more than just connectivity. In this study, we performed in mice of both sexes a multimodal dissection of four major projections of the VP-to the lateral hypothalamus (VPâLH), ventral tegmental area (VPâVTA), lateral habenula (VPâLHb), and mediodorsal thalamus (VPâMDT)-with physiological, anatomical, genetic, and behavioral tools. We also tested for physiological differences between VP neurons receiving input from nucleus accumbens medium spiny neurons (MSNs) that express either the D1 (D1-MSNs) or the D2 (D2-MSNs) dopamine receptor. We show that each VP projection (1) when inhibited during a cocaine conditioned place preference (CPP) test affects performance differently, (2) receives a different pattern of inputs using rabies retrograde labeling, (3) shows differentially expressed genes using RNA sequencing, and (4) has projection-specific characteristics in excitability and synaptic input characteristics using whole-cell patch clamp. VPâLH and VPâVTA projections have different effects on CPP and show low overlap in circuit tracing experiments, as VPâVTA neurons receive more striatal input, while VPâLH neurons receive more olfactory input. Additionally, VPâVTA neurons are less excitable, while VPâLH neurons are more excitable than the average VP neuron, a difference driven mainly by D2-MSN-responding neurons. Thus, VPâVTA and VPâLH neurons may represent largely distinct populations of VP neurons.
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Prosencéfalo Basal , Cocaína , Vías Nerviosas , Recompensa , Animales , Ratones , Prosencéfalo Basal/fisiología , Masculino , Cocaína/farmacología , Cocaína/administración & dosificación , Femenino , Vías Nerviosas/fisiología , Ratones Endogámicos C57BL , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D1/genética , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D2/genética , Área Tegmental Ventral/fisiología , Área Tegmental Ventral/citologíaRESUMEN
Drugs of abuse induce neuroadaptations, including synaptic plasticity, that are critical for transition to addiction, and genes and pathways that regulate these neuroadaptations are potential therapeutic targets. Tropomodulin 2 (Tmod2) is an actin-regulating gene that plays an important role in synapse maturation and dendritic arborization and has been implicated in substance abuse and intellectual disability in humans. Here, we mine the KOMP2 data and find that Tmod2 knock-out mice show emotionality phenotypes that are predictive of addiction vulnerability. Detailed addiction phenotyping shows that Tmod2 deletion does not affect the acute locomotor response to cocaine administration. However, sensitized locomotor responses are highly attenuated in these knock-outs, indicating perturbed drug-induced plasticity. In addition, Tmod2 mutant animals do not self-administer cocaine indicating lack of hedonic responses to cocaine. Whole-brain MR imaging shows differences in brain volume across multiple regions, although transcriptomic experiments did not reveal perturbations in gene coexpression networks. Detailed electrophysiological characterization of Tmod2 KO neurons showed increased spontaneous firing rate of early postnatal and adult cortical and striatal neurons. Cocaine-induced synaptic plasticity that is critical for sensitization is either missing or reciprocal in Tmod2 KO nucleus accumbens shell medium spiny neurons, providing a mechanistic explanation of the cocaine response phenotypes. Combined, these data, collected from both males and females, provide compelling evidence that Tmod2 is a major regulator of plasticity in the mesolimbic system and regulates the reinforcing and addictive properties of cocaine.
Asunto(s)
Cocaína , Cuerpo Estriado , Ratones Noqueados , Plasticidad Neuronal , Animales , Cocaína/farmacología , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Ratones , Masculino , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/metabolismo , Ratones Endogámicos C57BL , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/fisiología , Femenino , Trastornos Relacionados con Cocaína/fisiopatología , Trastornos Relacionados con Cocaína/genética , Proteínas de Microfilamentos/metabolismo , Proteínas de Microfilamentos/genética , Excitabilidad Cortical/efectos de los fármacos , Inhibidores de Captación de Dopamina/farmacología , Inhibidores de Captación de Dopamina/administración & dosificaciónRESUMEN
Much remains unknown about the etiology of compulsion-like alcohol drinking, where consumption persists despite adverse consequences. The role of the anterior insula (AIC) in emotion, motivation, and interoception makes this brain region a likely candidate to drive challenge-resistant behavior, including compulsive drinking. Indeed, subcortical projections from the AIC promote compulsion-like intake in rats and are recruited in heavy-drinking humans during compulsion for alcohol, highlighting the importance of and need for more information about AIC activity patterns that support aversion-resistant responding. Single-unit activity was recorded in the AIC from 15 male rats during alcohol-only and compulsion-like consumption. We found three sustained firing phenotypes, sustained-increase, sustained-decrease, and drinking-onset cells, as well as several firing patterns synchronized with licking. While many AIC neurons had session-long activity changes, only neurons with firing increases at drinking onset had greater activity under compulsion-like conditions. Further, only cells with persistent firing increases maintained activity during pauses in licking, suggesting roles in maintaining drive for alcohol during breaks. AIC firing was not elevated during saccharin drinking, similar to lack of effect of AIC inhibition on sweet fluid intake in many studies. In addition, we observed subsecond changes in AIC neural activity tightly entrained to licking. One lick-synched firing pattern (determined for all licks in a session) predicted compulsion-like drinking, while a separate lick-associated pattern correlated with greater consumption across alcohol intake conditions. Collectively, these data provide a more integrated model for the role of AIC firing in compulsion-like drinking, with important relevance for how the AIC promotes sustained motivated responding more generally.