RESUMEN
The loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) and consequent depletion of striatal dopamine are known to underlie the motor deficits observed in Parkinson's disease (PD). Adaptive changes in dopaminergic terminals and in postsynaptic striatal neurons can compensate for significant losses of striatal dopamine, resulting in preservation of motor behavior. In addition, compensatory changes independent of striatal dopamine have been proposed based on PD therapies that modulate nondopaminergic circuits within the basal ganglia. We used a genetic strategy to selectively destroy dopaminergic neurons in mice during development to determine the necessity of these neurons for the maintenance of normal motor behavior in adult and aged mice. We find that loss of 90% of SNc dopaminergic neurons and consequent depletion of >95% of striatal dopamine does not result in changes in motor behavior in young-adult or aged mice as evaluated by an extensive array of motor behavior tests. Treatment of aged mutant mice with the dopamine receptor antagonist haloperidol precipitated motor behavior deficits in aged mutant mice, indicating that <5% of striatal dopamine is sufficient to maintain motor function in these mice. We also found that mutant mice exhibit an exaggerated response to l-DOPA compared with control mice, suggesting that preservation of motor function involves sensitization of striatal dopamine receptors. Our results indicate that congenital loss of dopaminergic neurons induces remarkable adaptions in the nigrostriatal system where limited amounts of dopamine in the dorsal striatum can maintain normal motor function.
Asunto(s)
Dopamina/metabolismo , Neuronas Dopaminérgicas/patología , Marcha , Animales , Toxina Diftérica/genética , Toxina Diftérica/toxicidad , Dopamina/deficiencia , Antagonistas de Dopamina/farmacología , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/deficiencia , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/genética , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/metabolismo , Haloperidol/farmacología , Levodopa/farmacología , Mesencéfalo/efectos de los fármacos , Mesencéfalo/patología , Ratones , Ratones Transgénicos , MutaciónRESUMEN
Drugs acting at the serotonin-2C (5-HT2C) receptor subtype have shown promise as therapeutics in multiple syndromes including obesity, depression, and Parkinson's disease. While it is established that 5-HT2C receptor stimulation inhibits DA release, the neural circuits and the localization of the relevant 5-HT2C receptors remain unknown. This study used dual-probe in vivo microdialysis to investigate the relative contributions of 5-HT2C receptors localized in the rat substantia nigra (SN) and caudate-putamen (CP) in the control of nigrostriatal DA release. Systemic administration (3.0 mg/kg) of the 5-HT2C receptor selective agonist Ro 60-0175 [(αS)-6-Chloro-5-fluoro-α-methyl-1H-indole-1-ethanamine fumarate] decreased, whereas intrastriatal infusions of the selective 5-HT2C antagonist SB 242084 [6-Chloro-2,3-dihydro-5-methyl-N-[6-[(2-methyl-3-pyridinyl)oxy]-3-pyridinyl]-1H-indole-1-carboxyamide; 1.0 µM] increased, basal DA in the CP. Depending on the site within the SN pars reticulata (SNpr), infusions of SB 242084 had more modest but significant effects. Moreover, infusions of the GABA-A receptor agonist muscimol (10 µM) into the SNpr completely reversed the increases in striatal DA release produced by intrastriatal infusions of SB 242084. These findings suggest a role for 5-HT2C receptors regulating striatal DA release that is highly localized. 5-HT2C receptors localized in the striatum may represent a primary site of action that is mediated by the actions on GABAergic activity in the SN. Dopamine (DA) neurons in the substantia nigra pars compacta (SNpc) project to the caudate-putamen (CP; striatum). This circuitry is implicated in numerous pathologies including Parkinson's disease. Using in vivo microdialysis, we demonstrated that blockade of serotonin (5-HT) 2C receptors in the CP increased nigrostriatal DA release. Infusions of a GABA-A agonist into the substantia nigra pars reticulata (SNpr) blocked this increase. This work indicates that striatal serotonin 2C receptors regulate GABAergic tone in the SNpr, which in turn regulates nigrostriatal DA release.
Asunto(s)
Cuerpo Estriado/metabolismo , Dopamina/metabolismo , Vías Nerviosas/fisiología , Receptor de Serotonina 5-HT2C/metabolismo , Receptores de GABA-A/metabolismo , Sustancia Negra/metabolismo , Aminopiridinas/farmacología , Animales , Cromatografía Líquida de Alta Presión , Cuerpo Estriado/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Etilaminas/farmacología , Glutamato Descarboxilasa/metabolismo , Indoles/farmacología , Masculino , Microdiálisis , Vías Nerviosas/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Serotoninérgicos/farmacología , Sustancia Negra/efectos de los fármacosRESUMEN
Mutations in LRRK2 are thus far the most frequent known cause of autosomal dominant and idiopathic Parkinson's disease (PD) with prevalent mutations being found within the GTPase (R1441C/G) and kinase (G2019S) domains. Previous in vitro studies have revealed that R1441C and G2019S mutations are associated with increased kinase activity. To better understand LRRK2-linked PD pathogenesis in vivo, we have generated transgenic C. elegans overexpressing human LRRK2 wild type, R1441C and G2019S in dopaminergic (DA) neurons. Overexpression of these LRRK2 proteins causes age-dependent DA neurodegeneration, behavioral deficits, and locomotor dysfunction that are accompanied by a reduction of dopamine levels in vivo. In comparison, R1441C and G2019S mutants cause more severe phenotypes than the wild type protein. Interestingly, treatment with exogenous dopamine rescues the LRRK2-induced behavioral and locomotor phenotypes. In contrast, expression of the GTP binding defective mutant, K1347A, or knockout of the C. elegans LRRK2 homolog, LRK-1, prevents the LRRK2-induced neurodegeneration and behavioral abnormalities. Hence, our transgenic LRRK2 C. elegans models recapitulate key features of PD including progressive neurodegeneration, impairment of dopamine-dependent behavior and locomotor function, and reduction in dopamine levels. Furthermore, our findings provide strong support for the critical role of GTPase/kinase activity in LRRK2-linked pathologies. These invertebrate models will be useful for studying pathogenesis of PD and for development of potential therapeutics for the disease.
Asunto(s)
Caenorhabditis elegans/fisiología , Dopamina/fisiología , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/patología , Enfermedades del Sistema Nervioso/metabolismo , Enfermedades del Sistema Nervioso/patología , Enfermedad de Parkinson/metabolismo , Proteínas Serina-Treonina Quinasas/fisiología , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Modelos Animales de Enfermedad , Humanos , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina , Neuronas/metabolismo , Neuronas/patología , Enfermedad de Parkinson/fisiopatología , Proteínas Serina-Treonina Quinasas/genéticaRESUMEN
The central serotonin (5-HT) neurotransmitter system is an important modulator of diverse physiological processes and behaviors; however, the transcriptional mechanisms controlling its development are largely unknown. The Pet-1 ETS factor is a precise marker of developing and adult 5-HT neurons and is expressed shortly before 5-HT appears in the hindbrain. Here we show that in mice lacking Pet-1, the majority of 5-HT neurons fail to differentiate. Remaining ones show deficient expression of genes required for 5-HT synthesis, uptake, and storage. Significantly, defective development of the 5-HT system is followed by heightened anxiety-like and aggressive behavior in adults. These findings indicate that Pet-1 is a critical determinant of 5-HT neuron identity and implicate a Pet-1-dependent program in serotonergic modulation of behavior.
Asunto(s)
Agresión/fisiología , Ansiedad/genética , Proteínas de Transporte de Membrana , Proteínas del Tejido Nervioso , Neuronas/fisiología , Serotonina/fisiología , Factores de Transcripción/genética , Alelos , Animales , Conducta Animal/fisiología , Monoaminas Biogénicas/biosíntesis , Proteínas Portadoras/biosíntesis , Proteínas Portadoras/genética , Diferenciación Celular/fisiología , Cromatografía Líquida de Alta Presión , Inmunohistoquímica , Hibridación in Situ , Glicoproteínas de Membrana/biosíntesis , Glicoproteínas de Membrana/genética , Ratones , Ratones Noqueados , Sistema Nervioso/embriología , Equilibrio Postural/fisiología , Rombencéfalo/fisiología , Serotonina/biosíntesis , Serotonina/metabolismo , Proteínas de Transporte de Serotonina en la Membrana PlasmáticaRESUMEN
Immediate early genes (IEGs) of the early growth response gene (Egr) family are activated in the brain in response to stress, social stimuli, and administration of psycho-active medications. However, little is known about the role of these genes in the biological or behavioral response to these stimuli. Here we show that mice lacking the IEG transcription factor Egr3 (Egr3-/- mice) display increased aggression, and a decreased latency to attack, in response to the stressful social stimulus of a foreign intruder. Together with our findings of persistent and intrusive olfactory-mediated social investigation of conspecifics, these results suggest increased impulsivity in Egr3-/- mice. We also show that the aggression of Egr3-/- mice is significantly inhibited with chronic administration of the antipsychotic medication clozapine. Despite their sensitivity to this therapeutic effect of clozapine, Egr3-/- mice display a marked resistance to the sedating effects of acute clozapine compared with WT littermate controls. This indicates that the therapeutic, anti-aggressive action of clozapine is separable from its sedating activity, and that the biological abnormality resulting from loss of Egr3 distinguishes these different mechanisms. Thus Egr3-/- mice may provide an important tool for elucidating the mechanism of action of clozapine, as well as for understanding the biology underlying aggressive behavior. Notably, schizophrenia patients display a similar decreased susceptibility to the side effects of antipsychotic medications compared to non-psychiatric controls, despite the medications producing a therapeutic response. This suggests the possibility that Egr3-/- mice may provide insight into the neurobiological abnormalities underlying schizophrenia.
Asunto(s)
Agresión/efectos de los fármacos , Agresión/fisiología , Antipsicóticos/farmacología , Clozapina/farmacología , Proteína 3 de la Respuesta de Crecimiento Precoz/genética , Análisis de Varianza , Animales , Conducta Animal , Química Encefálica/efectos de los fármacos , Cromatografía Líquida de Alta Presión/métodos , Dopamina/metabolismo , Relación Dosis-Respuesta a Droga , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Tiempo de Reacción/efectos de los fármacos , Serotonina/metabolismoRESUMEN
Glial cell-line derived neurotrophic factor (GDNF)-mediated RET tyrosine kinase signaling is implicated in the survival of several PNS and CNS neuronal populations that are important in the pathogenesis of several disorders including Parkinson's disease and drug addiction. However, it has been difficult to study these processes and the physiological importance of this pathway in adult mice because of the neonatal lethality of Gdnf and Ret null mice. We report successful creation of RET conditional reporter mice to investigate postnatal physiologic roles of RET and monitor the fate of RET-expressing cell types. To delete RET specifically in dopaminergic neurons and determine the physiologic requirement of RET in the maintenance of substantia nigra compacta (SNC) and ventral tegmental area (VTA), we bred the RET conditional mice with mice that specifically express Cre from the dopamine transporter (Dat) locus. A detailed morphometric and biochemical analysis including dopaminergic neuron number and size in SNC and VTA, and fiber density in the striatum and nucleus accumbens, and dopamine levels indicate that RET is not required for providing global trophic support to midbrain dopaminergic neurons in adult mice. Furthermore, RET deficiency in these neurons does not cause major sensorimotor abnormalities. Hence our results support the idea that RET signaling is not critical for the normal physiology of the SNC and VTA in adult mice.
Asunto(s)
Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/fisiología , Mesencéfalo/fisiología , Neuronas/fisiología , Proteínas Proto-Oncogénicas c-ret/fisiología , Animales , Dopamina/fisiología , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/deficiencia , Proteínas de Transporte de Dopamina a través de la Membrana Plasmática/genética , Factor Neurotrófico Derivado de la Línea Celular Glial/fisiología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Actividad Motora/fisiología , Transducción de Señal/fisiología , Sustancia Negra/citología , Sustancia Negra/fisiología , Área Tegmental Ventral/citología , Área Tegmental Ventral/fisiologíaRESUMEN
The mechanism(s) by which serotonin modulates dopamine release in the medial prefrontal cortex is not known, although studies suggest an involvement of 5-HT2 family receptors. We employed in vivo microdialysis and putatively selective 5-HT2A antagonists (M100907, MDL 11,939, SR46349B) to determine if 5-HT2A receptors are responsible for both drug- and stress-induced DA release in the medial prefrontal cortex. MDL 11,939 and SR46349B receptor-binding studies indicated, for the first time, that only MDL 11,939 had greater selectivity for the 5-HT2A vs the 5-HT2C receptor subtypes similar to M100907, and that both showed low or no affinity for non-5-HT2 receptors. Reverse dialysis with 5-HT2A antagonists had little or no effect on basal dopamine efflux. However, intracortical administration of MDL 11,939 or M100907 attenuated dopamine release induced by systemic administration of the 5-HT2 agonist DOI. Dual-probe microdialysis demonstrated that systemic DOI also increased glutamate concentrations in the ventral tegmental area (VTA). This was blocked by intracortical M100907. Cortical perfusion with M100907, or the atypical antipsychotic drug risperidone, but not the 5-HT2B/C ligand SB 206553, also decreased dopamine release induced physiologically by stress. These results indicate that stimulation of cortical 5-HT2A receptors increases the release of dopamine from the mesocortical system. They suggest that this effect may be mediated by increases in glutamate release from corticotegmental projections to the VTA. Additionally, they indicate that cortical 5-HT2A receptors modulate evoked dopamine release, such as that observed physiologically following mild stress. These findings may have implications for the pharmacological treatment of disorders resulting from or exacerbated by stress.
Asunto(s)
Dopamina/metabolismo , Corteza Prefrontal/efectos de los fármacos , Corteza Prefrontal/metabolismo , Receptor de Serotonina 5-HT2A/fisiología , Estrés Fisiológico/metabolismo , Anfetaminas/farmacología , Análisis de Varianza , Animales , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Cromatografía Líquida de Alta Presión/métodos , Interacciones Farmacológicas , Electroquímica/métodos , Ácido Glutámico/metabolismo , Humanos , Masculino , Ratones , Microdiálisis/métodos , Células 3T3 NIH , Unión Proteica/efectos de los fármacos , Ensayo de Unión Radioligante/métodos , Ratas , Ratas Sprague-Dawley , Antagonistas de la Serotonina/farmacología , Agonistas de Receptores de Serotonina/farmacología , Factores de Tiempo , Transfección/métodos , Área Tegmental Ventral/efectos de los fármacos , Área Tegmental Ventral/metabolismoRESUMEN
Disruption of dopamine homeostasis may lead to dopaminergic neuron degeneration, a proposed explanation for the specific vulnerability of dopaminergic neurons in Parkinson's disease. While expression of human alpha-synuclein in C. elegans results in dopaminergic neuron degeneration, the effects of alpha-synuclein on dopamine homeostasis and its contribution to dopaminergic neuron degeneration in C. elegans have not been reported. Here, we examined the effects of alpha-synuclein overexpression on worm dopamine homeostasis. We found that alpha-synuclein expression results in upregulation of dopamine synthesis and content, and redistribution of dopaminergic synaptic vesicles, which significantly contribute to dopaminergic neuron degeneration. These results provide in vivo evidence supporting a critical role for dopamine homeostasis in supporting dopaminergic neuron integrity.
Asunto(s)
Caenorhabditis elegans/fisiología , Dopamina/metabolismo , Homeostasis/fisiología , Degeneración Nerviosa/metabolismo , alfa-Sinucleína/fisiología , Ácido 3,4-Dihidroxifenilacético/metabolismo , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Femenino , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Masculino , Microscopía Confocal , Actividad Motora/fisiología , Vesículas Sinápticas/metabolismo , Proteínas de Transporte Vesicular de Monoaminas/genética , Proteínas de Transporte Vesicular de Monoaminas/metabolismo , alfa-Sinucleína/genética , alfa-Sinucleína/metabolismoRESUMEN
Previous work has demonstrated that dopamine (DA) transmission is regulated by serotonin-2C (5-HT2C) receptors but the site(s) in the brain where these receptors are localized is not known. The present work utilized in vivo microdialysis to investigate the modulation of DA release by 5-HT2C receptors localized in the nerve terminal regions of the mesocortical and nigrostriatal DA pathways. Microdialysis probes implanted in the striatum or the prefrontal cortex (PFC) measured dialysate DA concentrations, while the selective 5-HT2B/2C inverse agonist SB 206553 was given locally by reverse dialysis into these terminal regions. Additionally, the effects of the 5-HT2C agonist mCPP on striatal DA were measured. Local administration of SB 206553 (0.1-100 microM) into the striatum increased DA efflux in a concentration-dependent manner. Systemic administration of mCPP (1.0 mg/kg i.p.) decreased striatal DA and attenuated the SB 206553-induced increase. In contrast, infusion of SB 206553 (0.1-500 microM) by reverse dialysis into the PFC had no significant effect on basal DA efflux in this region. Additionally, high concentrations of SB 206553 had no effect on high potassium (K(+))-stimulated DA release in the PFC. These data contribute to a body of evidence indicating that 5-HT2C receptors inhibit nigrostriatal dopaminergic transmission. In addition, the results suggest that the nigrostriatal system is regulated by 5-HT2C receptors localized in the dorsal striatum. Elucidating the mechanisms by which serotonin (5-HT) modulates striatal and prefrontocortical DA concentrations may lead to improvements in the treatment of diverse syndromes such as schizophrenia, Parkinson's disease, anxiety, drug abuse, and/or depression.
Asunto(s)
Dopamina/metabolismo , Neostriado/metabolismo , Vías Nerviosas/metabolismo , Receptor de Serotonina 5-HT2C/metabolismo , Serotonina/metabolismo , Sustancia Negra/metabolismo , Animales , Relación Dosis-Respuesta a Droga , Sinergismo Farmacológico , Líquido Extracelular/efectos de los fármacos , Líquido Extracelular/metabolismo , Indoles/farmacología , Masculino , Microdiálisis , Neostriado/efectos de los fármacos , Vías Nerviosas/efectos de los fármacos , Potasio/farmacología , Corteza Prefrontal/efectos de los fármacos , Corteza Prefrontal/metabolismo , Terminales Presinápticos/efectos de los fármacos , Terminales Presinápticos/metabolismo , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptor de Serotonina 5-HT2C/efectos de los fármacos , Antagonistas de la Serotonina/farmacología , Sustancia Negra/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Área Tegmental Ventral/efectos de los fármacos , Área Tegmental Ventral/metabolismoRESUMEN
The nigrostriatal dopamine system of the mammalian brain is necessary for normal voluntary motor activity. Dopamine exerts its effects by acting on two primary receptor subtypes: D1-like (D1 and D5) and D2-like (D2, D3, and D4) receptors. Previous research has indicated that both subtypes are involved in the negative feedback regulation of dopamine release in the brain. However, the role of D1-like receptors localized within the striatum remains controversial. Using in vivo microdialysis, we report that infusions of the D1/D5 antagonist SCH 23390 [R-(+)-7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-1H-3-benzazepine] (5-200 microM) directly into the striatum increased dopamine release in a concentration-dependent manner. Systemic administration of the novel, full D1/D5 agonist A-77636 [(-)-(1R,3S)-3-adamantyl-1-(aminomethyl)-3,4-dihydro-5,6-dihydroxy-1H-2-benzopyran] produced the opposite effect, a dose-dependent (0.75-3.0 mg/kg s.c.) decrease in striatal dopamine efflux. Infusions of SCH 23390 (5.0 microM) attenuated this decrease. These findings suggest that endogenous dopamine acts on D1-like receptors localized within the striatum to decrease nigrostriatal dopamine release. This negative feedback may be due to the activation of an inhibitory long-loop pathway. Knowledge of the circuitry underlying D1-mediated regulation of nigrostriatal neurons may have significance in current research on treatments for Parkinson's disease.