RESUMEN
Alcohol use disorder (AUD) is characterized by cycles of abuse, withdrawal, and relapse. Neuroadaptations in the basal ganglia are observed in AUD; specifically in the putamen, globus pallidus (GP), and ventral pallidum (VP). These regions are associated with habit formation, drug-seeking behaviors, and reward processing. While previous studies have shown the crucial role of glial cells in drug seeking, it remains unknown whether glial cells in the basal ganglia are altered in AUD. Glial cells in the putamen, GP, and VP were examined in human post-mortem tissue of AUD and alcohol remission cases. Immunohistochemistry was performed to analyze cell count, staining intensity, and morphology of microglia and astrocytes, using markers Iba-1 and GFAP. Morphological analysis revealed a significant decrease in microglia cell size and process retraction, indicating activation or a dystrophic microglia phenotype in individuals with AUD compared to controls. Microglia staining intensity was also higher in the GP and VP in AUD cases, whereas microglia staining intensity and cell size in remission cases were not different to control cases. In contrast, no astrocyte changes were observed in examined brain regions for both AUD and remission cases compared to controls. These results suggest alcohol exposure alters microglia, potentially contributing to dysfunctions in the basal ganglia that maintain addiction, and abstinence from alcohol may reverse microglia changes and associated dysfunctions. Overall, this study further characterizes AUD neuropathology and implicates microglia in the putamen, GP, and VP as a potential target for therapy.
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Alcoholismo , Humanos , Alcoholismo/patología , Microglía , Ganglios Basales/patología , Etanol , EncéfaloRESUMEN
The subthalamic nucleus (STN) is a major neuromodulation target for the alleviation of neurological and neuropsychiatric symptoms using deep brain stimulation (DBS). STN-DBS is today applied as treatment in Parkinson´s disease, dystonia, essential tremor, and obsessive-compulsive disorder (OCD). STN-DBS also shows promise as a treatment for refractory Tourette syndrome. However, the internal organization of the STN has remained elusive and challenges researchers and clinicians: How can this small brain structure engage in the multitude of functions that renders it a key hub for therapeutic intervention of a variety of brain disorders ranging from motor to affective to cognitive? Based on recent gene expression studies of the STN, a comprehensive view of the anatomical and cellular organization, including revelations of spatio-molecular heterogeneity, is now possible to outline. In this review, we focus attention to the neurobiological architecture of the STN with specific emphasis on molecular patterns discovered within this complex brain area. Studies from human, non-human primate, and rodent brains now reveal anatomically defined distribution of specific molecular markers. Together their spatial patterns indicate a heterogeneous molecular architecture within the STN. Considering the translational capacity of targeting the STN in severe brain disorders, the addition of molecular profiling of the STN will allow for advancement in precision of clinical STN-based interventions.
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Enfermedad de Parkinson , Núcleo Subtalámico , Animales , Humanos , Encéfalo , Enfermedad de Parkinson/terapia , InvestigadoresRESUMEN
The ubiquitin kinase-ligase pair PINK1-PRKN recognizes and transiently labels damaged mitochondria with ubiquitin phosphorylated at Ser65 (p-S65-Ub) to mediate their selective degradation (mitophagy). Complete loss of PINK1 or PRKN function unequivocally leads to early-onset Parkinson disease, but it is debated whether impairments in mitophagy contribute to disease later in life. While the pathway has been extensively studied in cell culture upon acute and massive mitochondrial stress, basal levels of activation under endogenous conditions and especially in vivo in the brain remain undetermined. Using rodent samples, patient-derived cells, and isogenic neurons, we here identified age-dependent, brain region-, and cell type-specific effects and determined expression levels and extent of basal and maximal activation of PINK1 and PRKN. Our work highlights the importance of defining critical risk and therapeutically relevant levels of PINK1-PRKN signaling which will further improve diagnosis and prognosis and will lead to better stratification of patients for future clinical trials.
RESUMEN
Parkinson's disease (PD) is a progressive neurodegenerative disorder, pathologically hallmarked by the loss of dopamine neurons in the substantia nigra (SN) and alpha-synuclein aggregation. Deep brain stimulation (DBS) of the subthalamic nucleus (STN) is a common target to treat the motor symptoms in PD. However, we have less understanding of the cellular changes in the STN during PD, and the impact of DBS on the STN and SN is limited. We examined cellular changes in the SN and STN in PD patients with and without STN-DBS treatment. Post-mortem brain tissues from 6 PD non-STN-DBS patients, 5 PD STN-DBS patients, and 6 age-matched controls were stained with markers for neurodegeneration (tyrosine hydroxylase, alpha-synuclein, and neuronal loss) and astrogliosis (glial fibrillary acidic protein). Changes were assessed using quantitative and semi-quantitative microscopy techniques. As expected, significant neuronal cell loss, alpha-synuclein pathology, and variable astrogliosis were observed in the SN in PD. No neuronal cell loss or astrogliosis was observed in the STN, although alpha-synuclein deposition was present in the STN in all PD cases. DBS did not alter neuronal loss, astrogliosis, or alpha-synuclein pathology in either the SN or STN. This study reports selective pathology in the STN with deposits of alpha-synuclein in the absence of significant neuronal cell loss or inflammation in PD. Despite being effective for the treatment of PD, this small post-mortem study suggests that DBS of the STN does not appear to modulate histological changes in astrogliosis or neuronal survival, suggesting that the therapeutic effects of DBS mechanism may transiently affect STN neural activity.
RESUMEN
Parkinson's disease (PD) is a neurodegenerative disorder characterized by a range of motor symptoms. Treatments are focused on dopamine replacement therapy or deep brain stimulation (DBS). The subthalamic nucleus (STN) is a common target for DBS treatment of PD. However, the function of the STN in normal conditions and pathology is poorly understood. Here, we show in rats that optogenetic modulation of STN neuronal activity exerts bidirectional control of motor function, where inhibition of the STN increases movement and STN activation decreases movement. We also examined the effect of bidirectional optogenetic manipulation STN neuronal activity under dopamine depleted condition using the bilateral rodent 6-hydroxydopamine (6-OHDA) model of Parkinson's disease. Optogenetic inhibition of the STN in the absence of dopamine had no impact on motor control yet STN excitation led to pronounced abnormal involuntary movement. Administration of levodopa rescued the abnormal involuntary movements induced by STN excitation. Although dopamine and STN dysfunction are well established in PD pathology, here we demonstrate simultaneous STN over activity and loss of dopamine lead to motor deficits. Moreover, we show the dysfunction of the STN is dependent on dopamine. This study provides evidence that the loss of dopamine and the over activity of the STN are key features of PD motor deficits. These results provide insight into the STN pathology in PD and therapeutic mechanism of targeting the STN for the treatment for PD.
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Clinical studies provide fundamental knowledge of substance use behaviors (substance of abuse, patterns of use, relapse rates). The combination of neuroimaging approaches reveal correlation between substance use disorder (SUD) and changes in neural structure, function, and neurotransmission. Here, we review these advances, placing special emphasis on sex specific findings from structural neuroimaging studies of those dependent on alcohol, nicotine, cannabis, psychostimulants, or opioids. Recent clinical studies in SUD analyzing sex differences reveal neurobiological changes that are differentially impacted in common reward processing regions such as the striatum, hippocampus, amygdala, insula, and corpus collosum. We reflect on the contribution of sex hormones, period of drug use and abstinence, and the potential impact of these factors on the interpretation of the reported findings. With the overall recognition that SUD impacts the brains of females and males differentially, it is of fundamental importance that future research is designed with sex as a variable of study in this field. Improved understanding of neurobiological changes in males and females in SUD will advance knowledge underlying sex-specific susceptibility and the neurobiological impact in these disorders. Together these findings will inform future treatments that are tailor designed for improved efficacy in females and males with SUD.
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The ventral pallidum (VP) is central to the reinforcing effects across a variety of drugs and relapse to drug seeking. Emerging studies from animal models of reinstatement reveal a complex neurobiology of the VP that contributes to different aspects of relapse to drug seeking. This review builds on classical understanding of the VP as part of the final common pathway of relapse but also discusses the properties of the VP as an independent structure. These include VP neural anatomical subregions, cellular heterogeneity, circuitry, neurotransmitters and peptides. Collectively, this review provides a current understanding of the VP from molecular to circuit level architecture that contributes to both the appetitive and aversive symptoms of drug addiction. We show the complex neurobiology of the VP in drug seeking, emphasizing its critical role in addiction, and review strategic approaches that target the VP to reduce relapse rates.
Asunto(s)
Prosencéfalo Basal , Preparaciones Farmacéuticas , Animales , Comportamiento de Búsqueda de Drogas , Neuronas , RecurrenciaRESUMEN
Parkinson's disease (PD) is a neurological disorder with motor dysfunction and a number of psychiatric symptoms. Symptoms such as anxiety and cognitive deficits emerge prior to motor symptoms and persist over time. There are limited treatments targeting PD psychiatric symptoms. Emerging studies reveal that the gut microbe is altered in PD patients. Here we assessed the effect of a probiotic treatment in a rat model of PD. We used the neurotoxin (6-hydroxydopamine, 6-OHDA) in a preclinical PD model to examine the impact of a probiotic treatment (Lacticaseibacillus rhamnosus HA-114) on anxiety and memory. Rats underwent either sham surgery or received 6-OHDA bilaterally into the striatum. Three weeks post-surgery, rats were divided into three experimental groups: a sham group that received probiotics, a 6-OHDA group that received probiotics, and the third group of 6-OHDA received the placebo formula. All rats had access to either placebo or probiotics formula for 6 weeks. All groups were assessed for anxiety-like behaviour using the elevated plus maze. Cognition was assessed for both non-hippocampal and hippocampal dependent tasks using the novel object recognition and novel place recognition. We report that the 6-OHDA lesion induced anxiety-like behaviour and deficits in hippocampal dependent cognition. Interestingly, the probiotics treatment had no impact on anxiety-like behaviour but selectively improved hippocampal dependent cognition deficits. Together, the results presented here highlight the utility of animal models in examining the neuropsychiatric symptoms of PD and the potential of probiotics as adjunctive treatment for non-motor symptoms of PD.
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The mesolimbic dopamine system comprises distinct compartments supporting different functions in learning and motivation. Less well understood is how complex addiction-related behaviors emerge from activity patterns across these compartments. Here we show how different forms of relapse to alcohol-seeking in male rats are assembled from activity across the VTA and the nucleus accumbens. First, we used chemogenetic approaches to show a causal role for VTA TH neurons in two forms of relapse to alcohol-seeking: renewal (context-induced reinstatement) and reacquisition. Then, using gCaMP fiber photometry of VTA TH neurons, we identified medial and lateral VTA TH neuron activity profiles during self-administration, renewal, and reacquisition. Next, we used optogenetic inhibition of VTA TH neurons to show distinct causal roles for VTA subregions in distinct forms of relapse. We then used dLight fiber photometry to measure dopamine binding across the ventral striatum (medial accumbens shell, accumbens core, lateral accumbens shell) and showed complex and heterogeneous profiles of dopamine binding during self-administration and relapse. Finally, we used representational similarity analysis to identify mesolimbic dopamine signatures of self-administration, extinction, and relapse. Our results show that signatures of relapse can be identified from heterogeneous activity profiles across the mesolimbic dopamine system and that these signatures are unique for different forms of relapse.SIGNIFICANCE STATEMENT It is axiomatic that the actions of dopamine are critical to drug addiction. Yet how relapse to drug-seeking is assembled from activity across the mesolimbic dopamine system is poorly understood. Here we show how relapse to alcohol-seeking relates to activity in specific VTA and accumbens compartments, how these change for different forms of relapse, and how relapse-associated activity relates to activity during self-administration and extinction. We report the mesolimbic dopamine activity signatures for relapse and show that these signatures are unique for different forms of relapse.
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Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/fisiología , Comportamiento de Búsqueda de Drogas/fisiología , Etanol/administración & dosificación , Núcleo Accumbens/efectos de los fármacos , Núcleo Accumbens/fisiología , Área Tegmental Ventral/efectos de los fármacos , Área Tegmental Ventral/fisiología , Animales , Conducta Adictiva/fisiopatología , Condicionamiento Operante/efectos de los fármacos , Condicionamiento Operante/fisiología , Dopamina/metabolismo , Masculino , Potenciales de la Membrana , Optogenética , Ratas Long-Evans , Recurrencia , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Alcohol-use disorders are chronically relapsing conditions characterized by cycles of use, abstinence and relapse. The ventral pallidum (VP) is a key node in the neural circuits controlling relapse to alcohol seeking and a key target of pharmacotherapies for relapse prevention. There has been a significant increase in our understanding of the molecular, anatomical, pharmacological and functional properties of the ventral pallidum, laying foundations for a new understanding of its role in relapse to alcohol seeking and motivation. Here we review these advances, placing special emphasis on how advances in understanding in the cellular and circuit architectures of ventral pallidum contributes to the relapse to alcohol seeking. We show how this knowledge improves mechanistic understanding of current relapse prevention pharmacotherapies, how it may be used to tailor these against different forms of relapse and how it may help provide insights into the mental health problems frequently co-morbid with alcohol-use disorders.
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Prosencéfalo Basal , Comportamiento de Búsqueda de Drogas , Etanol , Humanos , RecurrenciaRESUMEN
The ventral pallidum (VP) is a key node in the neural circuits controlling relapse to drug seeking. How this role relates to different VP cell types and their projections is poorly understood. Using male rats, we show how different forms of relapse to alcohol-seeking are assembled from VP cell types and their projections to lateral hypothalamus (LH) and ventral tegmental area (VTA). Using RNAScope in situ hybridization to characterize activity of different VP cell types during relapse to alcohol-seeking provoked by renewal (context-induced reinstatement), we found that VP Gad1 and parvalbumin (PV), but not vGlut2, neurons show relapse-associated changes in c-Fos expression. Next, we used retrograde tracing, chemogenetic, and electrophysiological approaches to study the roles of VPGad1 and VPPV neurons in relapse. We show that VPGad1 neurons contribute to contextual control over relapse (renewal), but not to relapse during reacquisition, via projections to LH, where they converge with ventral striatal inputs onto LHGad1 neurons. This convergence of striatopallidal inputs at the level of individual LHGad1 neurons may be critical to balancing propensity for relapse versus abstinence. In contrast, VPPV neurons contribute to relapse during both renewal and reacquisition via projections to VTA. These findings identify a double dissociation in the roles for different VP cell types and their projections in relapse. VPGad1 neurons control relapse during renewal via projections to LH. VPPV neurons control relapse during both renewal and reacquisition via projections to VTA. Targeting these different pathways may provide tailored interventions for different forms of relapse.SIGNIFICANCE STATEMENT Relapse to drug or reward seeking after a period of extinction or abstinence remains a key impediment to successful treatment. The ventral pallidum, located in the ventral basal ganglia, has long been recognized as an obligatory node in a 'final common pathway' for relapse. Yet how this role relates to the considerable VP cellular and circuit heterogeneity is not well understood. We studied the cellular and circuit architecture for VP in relapse control. We show that different forms of relapse have complementary VP cellular and circuit architectures, raising the possibility that targeting these different neural architectures may provide tailored interventions for different forms of relapse.
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Prosencéfalo Basal/fisiología , Comportamiento de Búsqueda de Drogas/fisiología , Área Hipotalámica Lateral/fisiología , Neuronas/fisiología , Área Tegmental Ventral/fisiología , Animales , Condicionamiento Operante/efectos de los fármacos , Condicionamiento Operante/fisiología , Etanol/administración & dosificación , Masculino , Vías Nerviosas/fisiología , Ratas , Ratas Sprague-Dawley , Recurrencia , RecompensaRESUMEN
Contexts exert bi-directional control over relapse to drug seeking. Contexts associated with drug self-administration promote relapse, whereas contexts associated with the absence of self-administration protect against relapse. The nucleus accumbens shell (AcbSh) is a key brain region determining these roles of context. However, the specific cell types, and projections, by which AcbSh serves these dual roles are unknown. Here, we show that contextual control over relapse and abstinence is embedded within distinct output circuits of dopamine 1 receptor (Drd1) expressing AcbSh neurons. We report anatomical and functional segregation of Drd1 AcbSh output pathways during context-induced reinstatement and extinction of alcohol seeking. The AcbShâventral tegmental area (VTA) pathway promotes relapse via projections to VTA Gad1 neurons. The AcbShâlateral hypothalamus (LH) pathway promotes extinction via projections to LH Gad1 neurons. Targeting these opposing AcbSh circuit contributions may reduce propensity to relapse to, and promote abstinence from, drug use.
Asunto(s)
Consumo de Bebidas Alcohólicas/metabolismo , Condicionamiento Operante/fisiología , Comportamiento de Búsqueda de Drogas/fisiología , Núcleo Accumbens/metabolismo , Consumo de Bebidas Alcohólicas/prevención & control , Consumo de Bebidas Alcohólicas/psicología , Animales , Condicionamiento Operante/efectos de los fármacos , Comportamiento de Búsqueda de Drogas/efectos de los fármacos , Etanol/administración & dosificación , Masculino , Vías Nerviosas/química , Vías Nerviosas/fisiología , Núcleo Accumbens/química , Núcleo Accumbens/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Ratas Transgénicas , Recurrencia , AutoadministraciónRESUMEN
Ventral pallidum (VP) is a well-established locus for the reinforcing effects of drugs of abuse and reinstatement of drug seeking. However, VP neurons are at the origin of multiple output pathways, with strong projections to ventral tegmental area (VTA), subthalamic nucleus (STN), lateral hypothalamus, among others, and the roles of these VP output pathways in reinstatement of drug seeking remain poorly understood. Here we addressed these issues using a combination of neuroanatomical tracing and chemogenetic approaches. First, using dual-retrograde tracing, we show that VP neurons projecting to either VTA or STN are recruited during context-induced reinstatement of extinguished alcohol seeking in rats. Then, using chemogenetics, we show modulation of context-induced reinstatement and reacquisition of alcohol seeking via designer receptors exclusively activated by designer drugs excitation or inhibition of the VP. To determine the causal roles of VP â VTA and VP â STN pathways in context-induced reinstatement and reacquisition we used a chemogenetic disconnection approach and show that silencing either the VP â VTA or VP â STN pathways is sufficient to reduce both reinstatement and reacquisition of alcohol seeking. Moreover, these disconnections also each reduced responding and motivation during a progressive ratio test but had no effect on locomotor activity. Together, these results show that multiple ventral pallidal output pathways contribute to relapse to alcohol seeking. SIGNIFICANCE STATEMENT: Ventral pallidum (VP) serves important roles in reward and motivation and is a critical node in the neural circuitry for reinstatement of drug seeking. Despite being a common locus for different forms of reinstatement, fundamental aspects of neural circuitry for these VP contributions to reinstatement of drug seeking remain unknown. Here we used a combination of neuroanatomical tracing and chemogenetic approaches to map the VP output pathways for context-induced reinstatement and reacquisition of alcohol seeking. We show that VP output pathways to the subthalamic nucleus and also to the ventral tegmental area are necessary for these forms of reinstatement.
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Prosencéfalo Basal/fisiología , Condicionamiento Operante/fisiología , Comportamiento de Búsqueda de Drogas/fisiología , Vías Eferentes/fisiología , Etanol/administración & dosificación , Extinción Psicológica/fisiología , Animales , Prosencéfalo Basal/citología , Prosencéfalo Basal/efectos de los fármacos , Condicionamiento Operante/efectos de los fármacos , Comportamiento de Búsqueda de Drogas/efectos de los fármacos , Vías Eferentes/citología , Vías Eferentes/efectos de los fármacos , Extinción Psicológica/efectos de los fármacos , Masculino , Red Nerviosa/citología , Red Nerviosa/efectos de los fármacos , Red Nerviosa/fisiología , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Ratas , Ratas Sprague-Dawley , RecurrenciaRESUMEN
The striatum is a large brain nucleus with an important role in the control of movement and emotions. Medium spiny neurons (MSNs) are striatal output neurons forming prominent descending axon tracts that target different brain nuclei. However, how MSN axon tracts in the forebrain develop remains poorly understood. Here, we implicate the Wnt binding receptor Frizzled3 in several uncharacterized aspects of MSN pathway formation [i.e., anterior-posterior guidance of MSN axons in the striatum and their subsequent growth into the globus pallidus (GP), an important (intermediate) target]. In Frizzled3 knock-out mice, MSN axons fail to extend along the anterior-posterior axis of the striatum, and many do not reach the GP. Wnt5a acts as an attractant for MSN axons in vitro, is expressed in a posterior high, anterior low gradient in the striatum, and Wnt5a knock-out mice phenocopy striatal anterior-posterior defects observed in Frizzled3 mutants. This suggests that Wnt5a controls anterior-posterior guidance of MSN axons through Frizzled3. Axons that reach the GP in Frizzled3 knock-out mice fail to enter this structure. Surprisingly, entry of MSN axons into the GP non-cell-autonomously requires Frizzled3, and our data suggest that GP entry may be contingent on the correct positioning of "corridor" guidepost cells for thalamocortical axons by Frizzled3. Together, these data dissect MSN pathway development and reveal (non)cell-autonomous roles for Frizzled3 in MSN axon guidance. Further, they are the first to identify a gene that provides anterior-posterior axon guidance in a large brain nucleus and link Frizzled3 to corridor cell development. SIGNIFICANCE STATEMENT: Striatal axon pathways mediate complex physiological functions and are an important therapeutic target, underscoring the need to define how these connections are established. Remarkably, the molecular programs regulating striatal pathway development remain poorly characterized. Here, we determine the embryonic ontogeny of the two main striatal pathways (striatonigral and striatopallidal) and identify novel (non)cell-autonomous roles for the axon guidance receptor Frizzled3 in uncharacterized aspects of striatal pathway formation (i.e., anterior-posterior axon guidance in the striatum and axon entry into the globus pallidus). Further, our results link Frizzled3 to corridor guidepost cell development and suggest that an abnormal distribution of these cells has unexpected, widespread effects on the development of different axon tracts (i.e., striatal and thalamocortical axons).
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Axones/fisiología , Polaridad Celular/genética , Cuerpo Estriado/citología , Receptores Frizzled/metabolismo , Vías Nerviosas/embriología , Neuronas/citología , Animales , Células Cultivadas , Cuerpo Estriado/embriología , Embrión de Mamíferos , Femenino , Receptores Frizzled/genética , Globo Pálido/citología , Células HEK293 , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Vías Nerviosas/metabolismo , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
The ventral pallidum (VP) is a key component of the neural circuitry mediating relapse to drug seeking, but the critical afferent pathways to VP recruited during relapse remain poorly understood. We studied the role of the nucleus accumbens core (AcbC) â VP pathway in ABA renewal and reacquisition of alcohol seeking. Rats received application of adenoviral vectors encoding eYFP, ChR2(H134R), or eNpHr3.0 to AcbC and implantation of fiber optic cannulas into VP to permit photostimulation of AcbC terminals there. Rats were then trained to self-administer alcoholic beer in 1 context (A), extinguished in a second context (B), tested in the extinction (ABB) and training (ABA) contexts, and were then tested for reacquisition of alcoholic beer seeking. There was ABA renewal of alcohol seeking, but neither optogenetic excitation nor inhibition of the AcbC â VP pathway affected this renewal. In contrast, optogenetic inhibition of the AcbC â VP striatopallidal pathway reduced reacquisition of alcohol seeking-measured either by the number of active nosepokes emitted or by the number of alcohol rewards earned and consumed. Moreover, optogenetic excitation of the AcbC â VP striatopallidal pathway increased magazine entries during reacquisition test. This finding shows the importance of the AcbC â VP pathway in controlling relapse when the drug reinforcer is present on test and is consistent with a role for the AcbC â VP pathway in regulating the hedonic or incentive motivational properties of drug reinforcers.
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Prosencéfalo Basal/fisiología , Condicionamiento Operante/fisiología , Comportamiento de Búsqueda de Drogas/fisiología , Núcleo Accumbens/fisiología , Animales , Cerveza , Etanol/administración & dosificación , Extinción Psicológica/fisiología , Masculino , Vías Nerviosas/fisiología , Optogenética , Ratas , Ratas Sprague-Dawley , Recurrencia , AutoadministraciónRESUMEN
Two experiments used vivo morpholinos to assess the role of orexin/hypocretin in ABA renewal of extinguished alcohol seeking. Rats were trained to respond for alcoholic beer in a distinctive context, A, and then extinguished in a second distinctive context, B. When rats were tested in the extinction context, ABB, responding was low but when they were tested in the training context, ABA, responding was significantly higher. Microinjection of an orexin/hypocretin antisense vivo morpholino into LH significantly reduced orexin/hypocretin protein expression but had no effect on the ABA renewal of alcohol seeking (Experiment 1). Microinjection of a higher dose of the antisense vivo morpholino into LH also significantly reduced orexin/hypocretin protein expression but this was not selective and yielded significant reduction in melanin-concentrating hormone (MCH) protein expression. This non-selective knockdown did significantly reduce ABA renewal as well as reduce the reacquisition of alcohol seeking. Taken together, these findings show an important role for LH in the ABA renewal of alcohol seeking but that orexin/hypocretin is not necessary for this renewal.
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Alcoholismo/genética , Alcoholismo/fisiopatología , Comportamiento de Búsqueda de Drogas/fisiología , Área Hipotalámica Lateral/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neuropéptidos/metabolismo , Análisis de Varianza , Animales , Cerveza , Técnicas de Silenciamiento del Gen , Hormonas Hipotalámicas/metabolismo , Inmunohistoquímica , Masculino , Melaninas/metabolismo , Microinyecciones , Morfolinos/administración & dosificación , Morfolinos/genética , Orexinas , Hormonas Hipofisarias/metabolismo , Ratas , Ratas Long-EvansRESUMEN
Ryk is an atypical transmembrane receptor tyrosine kinase that has been shown to play multiple roles in development through the modulation of Wnt signaling. Within the developing ventral midbrain (VM), Wnts have been shown to contribute to the proliferation, differentiation, and connectivity of dopamine (DA) neurons; however, the Wnt-related receptors regulating these events remain less well described. In light of the established roles of Wnt5a in dopaminergic development (regulating DA differentiation as well as axonal growth and repulsion), and its interaction with Ryk elsewhere within the central nervous system, we investigated the potential role of Ryk in VM development. Here we show temporal and spatial expression of Ryk within the VM, suggestive of a role in DA neurogenesis and axonal plasticity. In VM primary cultures, we show that the effects of Wnt5a on VM progenitor proliferation, DA differentiation, and DA axonal connectivity can be inhibited using an Ryk-blocking antibody. In support, Ryk knockout mice showed reduced VM progenitors and DA precursor populations, resulting in a significant decrease in DA cells. However, Ryk(-/-) mice displayed no defects in DA axonal growth, guidance, or fasciculation of the MFB, suggesting other receptors may be involved and/or compensate for the loss of this receptor. These findings identify for the first time Ryk as an important receptor for midbrain DA development.
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Neuronas Dopaminérgicas/metabolismo , Mesencéfalo/citología , Neurogénesis , Proteínas Tirosina Quinasas Receptoras/fisiología , Proteínas Wnt/metabolismo , Animales , Axones/fisiología , Regulación del Desarrollo de la Expresión Génica , Células HEK293 , Humanos , Mesencéfalo/embriología , Ratones , Ratones Noqueados , Morfogénesis , Células-Madre Neurales/fisiología , Prosencéfalo/citología , Prosencéfalo/embriología , Ratas , Ratas Sprague-Dawley , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Técnicas de Cultivo de Tejidos , Proteína Wnt-5aRESUMEN
Monoaminergic neurons [serotonergic (5-HT) and dopaminergic (mdDA)] in the brainstem project axons along the anterior-posterior axis. Despite their important physiological functions and implication in disease, the molecular mechanisms that dictate the formation of these projections along the anterior-posterior axis remain unknown. Here we reveal a novel requirement for Wnt/planar cell polarity signaling in the anterior-posterior organization of the monoaminergic system. We find that 5-HT and mdDA axons express the core planar cell polarity components Frizzled3, Celsr3, and Vangl2. In addition, monoaminergic projections show anterior-posterior guidance defects in Frizzled3, Celsr3, and Vangl2 mutant mice. The only known ligands for planar cell polarity signaling are Wnt proteins. In culture, Wnt5a attracts 5-HT but repels mdDA axons, and Wnt7b attracts mdDA axons. However, mdDA axons from Frizzled3 mutant mice are unresponsive to Wnt5a and Wnt7b. Both Wnts are expressed in gradients along the anterior-posterior axis, consistent with their role as directional cues. Finally, Wnt5a mutants show transient anterior-posterior guidance defects in mdDA projections. Furthermore, we observe during development that the cell bodies of migrating descending 5-HT neurons eventually reorient along the direction of their axons. In Frizzled3 mutants, many 5-HT and mdDA neuron cell bodies are oriented abnormally along the direction of their aberrant axon projections. Overall, our data suggest that Wnt/planar cell polarity signaling may be a global anterior-posterior guidance mechanism that controls axonal and cellular organization beyond the spinal cord.
Asunto(s)
Tronco Encefálico/fisiología , Polaridad Celular/fisiología , Dopamina/fisiología , Proteínas Proto-Oncogénicas/fisiología , Serotonina/fisiología , Transducción de Señal/fisiología , Proteínas Wnt/fisiología , Animales , Axones , Tronco Encefálico/citología , Señales (Psicología) , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Ratones Transgénicos , Vías Nerviosas/citología , Vías Nerviosas/fisiología , Proteínas Proto-Oncogénicas/deficiencia , Proteínas Proto-Oncogénicas/genética , Transducción de Señal/genética , Proteínas Wnt/deficiencia , Proteínas Wnt/genética , Proteína Wnt-5aRESUMEN
Dopaminergic neurons in the mesodiencephalon (mdDA neurons) make precise synaptic connections with targets in the forebrain via the mesostriatal, mesolimbic, and mesoprefrontal pathways. Because of the functional importance of these remarkably complex ascending axon pathways and their implication in human disease, the mechanisms underlying the development of these connections are of considerable interest. Despite extensive in vitro studies, the molecular determinants that ensure the perfect formation of these pathways in vivo remain mostly unknown. Here, we determine the embryonic origin and ontogeny of the mouse mesoprefrontal pathway and use these data to reveal an unexpected requirement for semaphorin 3F (Sema3F) and its receptor neuropilin-2 (Npn-2) during mdDA pathway development using tissue culture approaches and analysis of sema3F(-/-), npn-2(-/-), and npn-2(-/-);TH-Cre mice. We show that Sema3F is a bifunctional guidance cue for mdDA axons, some of which have the remarkable ability to regulate their responsiveness to Sema3F as they develop. During early developmental stages, Sema3F chemorepulsion controls previously uncharacterized aspects of mdDA pathway development through both Npn-2-dependent (axon fasciculation and channeling) and Npn-2-independent (rostral growth) mechanisms. Later on, chemoattraction mediated by Sema3F and Npn-2 is required to orient mdDA axon projections in the cortical plate of the medial prefrontal cortex. This latter finding demonstrates that regulation of axon orientation in the target field occurs by chemoattractive mechanisms, and this is likely to also apply to other neural systems. In all, this study provides a framework for additional dissection of the molecular basis of mdDA pathway development and disease.