RESUMO
The dopamine D4 receptor (D4R) is a candidate gene for attention deficit/hyperactivity disorder (ADHD) based on genetic studies reporting that particular polymorphisms are present at a higher frequency in affected children. However, the direct participation of the D4R in the onset or progression of ADHD has not been tested. Here, we generated a mouse model with high face value to screen candidate genes for the clinical disorder by neonatal disruption of central dopaminergic pathways with 6-hydroxydopamine (6-OHDA). The lesioned mice exhibited hyperactivity that waned after puberty, paradoxical hypolocomotor responses to amphetamine and methylphenidate, poor behavioral inhibition in approach/avoidance conflict tests and deficits in continuously performed motor coordination tasks. To determine whether the D4R plays a role in these behavioral phenotypes, we performed 6-OHDA lesions in neonatal mice lacking D4Rs (Drd4(-/-)). Although striatal dopamine contents and tyrosine hydroxylase-positive midbrain neurons were reduced to the same extent in both genotypes, Drd4(-/-) mice lesioned with 6-OHDA did not develop hyperactivity. Similarly, the D4R antagonist PNU-101387G prevented hyperactivity in wild-type 6-OHDA-lesioned mice. Furthermore, wild-type mice lesioned with 6-OHDA showed an absence of behavioral inhibition when tested in the open field or the elevated plus maze, while their Drd4(-/-) siblings exhibited normal avoidance for the unprotected areas of these mazes. Together, our results from a combination of genetic and pharmacological approaches demonstrate that D4R signaling is essential for the expression of juvenile hyperactivity and impaired behavioral inhibition, relevant features present in this ADHD-like mouse model.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade/genética , Transtorno do Deficit de Atenção com Hiperatividade/fisiopatologia , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/fisiologia , Anfetamina/farmacologia , Animais , Animais não Endogâmicos , Transtorno do Deficit de Atenção com Hiperatividade/tratamento farmacológico , Comportamento Animal/fisiologia , Estimulantes do Sistema Nervoso Central/farmacologia , Corpo Estriado/citologia , Corpo Estriado/fisiopatologia , Denervação , Modelos Animais de Doenças , Masculino , Metilfenidato/farmacologia , Camundongos , Camundongos Knockout , Atividade Motora/fisiologia , Vias Neurais , Oxidopamina , Fenótipo , Receptores de Dopamina D4 , Substância Negra/citologia , Substância Negra/fisiopatologia , SimpatolíticosRESUMO
The dopamine D(4) receptor (D(4)R) is predominantly expressed in the frontal cortex (FC), a brain region that receives dense input from midbrain dopamine (DA) neurons and is associated with cognitive and emotional processes. However, the physiological significance of this dopamine receptor subtype has been difficult to explore because of the slow development of D(4)R agonists and antagonists the selectivity and efficacy of which have been rigorously demonstrated in vivo. We have attempted to overcome this limitation by taking a multidimensional approach to the characterization of mice completely deficient in this receptor subtype. Electrophysiological current and voltage-clamp recordings were performed in cortical pyramidal neurons from wild-type and D(4)R-deficient mice. The frequency of spontaneous synaptic activity and the frequency and duration of paroxysmal discharges induced by epileptogenic agents were increased in mutant mice. Enhanced synaptic activity was also observed in brain slices of wild-type mice incubated in the presence of the selective D(4)R antagonist PNU-101387G. Consistent with greater electrophysiological activity, nerve terminal glutamate density associated with asymmetrical synaptic contacts within layer VI of the motor cortex was reduced in mutant neurons. Taken together, these results suggest that the D(4)R can function as an inhibitory modulator of glutamate activity in the FC.
Assuntos
Córtex Cerebral/fisiopatologia , Receptores de Dopamina D2/deficiência , Convulsões/fisiopatologia , 4-Aminopiridina/farmacologia , Animais , Bicuculina/farmacologia , Córtex Cerebral/efeitos dos fármacos , Convulsivantes/farmacologia , Dopamina/metabolismo , Antagonistas dos Receptores de Dopamina D2 , Relação Dose-Resposta a Droga , Ácido Glutâmico/metabolismo , Imuno-Histoquímica , Técnicas In Vitro , Potenciais da Membrana/efeitos dos fármacos , Camundongos , Camundongos Mutantes Neurológicos , Córtex Motor/efeitos dos fármacos , Córtex Motor/metabolismo , Córtex Motor/fisiopatologia , Inibição Neural/efeitos dos fármacos , Inibição Neural/genética , Técnicas de Patch-Clamp , Piperazinas/farmacologia , Terminações Pré-Sinápticas/metabolismo , Células Piramidais/efeitos dos fármacos , Células Piramidais/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D4 , Convulsões/induzido quimicamente , Sulfonamidas/farmacologiaRESUMO
The orphan nuclear receptor Nurr1 is critical for the survival of mesencephalic dopaminergic precursor neurons. Little is known about the mechanisms that regulate Nurr1 expression in vivo. Other members of this receptor family have been shown to be activated by dopamine. We sought to determine if Nurr1 expression is also regulated by endogenous dopamine through dopamine receptors. Consequently, we investigated the expression of Nurr1 mRNA in genetically modified mice lacking both functional copies of the D2 dopamine receptor gene and in their congenic siblings. Quantitative in situ hybridization demonstrated a significant increased expression of Nurr1 mRNA in the substantia nigra pars compacta and the ventral tegmental area of D2 dopamine receptor -/- mice. No change in Nurr1 expression was detected in other brain regions, such as the habenular nuclei and temporal cortex. Among the cell groups studied, mesencephalic dopaminergic neurons are unique in that they express both Nurr1 and the D2 dopamine receptor, and synthesize dopamine. Thus, it seems plausible that the selective increase in Nurr1 expression observed in D2 receptor-deficient mice is the consequence of an impaired dopamine autoreceptor function.
Assuntos
Proteínas de Ligação a DNA , Mesencéfalo/metabolismo , Proteínas do Tecido Nervoso/genética , Neurônios/metabolismo , RNA Mensageiro/metabolismo , Receptores de Dopamina D2/fisiologia , Fatores de Transcrição/genética , Animais , Hibridização In Situ , Camundongos , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares , Receptores de Dopamina D2/biossíntese , Receptores de Dopamina D3 , Substância Negra/citologiaRESUMO
The D4 receptor (D4R), a member of the dopamine D2-like receptor family, has been implicated in the pathophysiology of several diseases and has been the target of various investigations regarding its distribution and quantification. The brain distribution of the D4R has been well described in various species, but the quantification is still an issue of controversy, because no specific ligand is commercially available. To circumvent this difficulty we have performed a biochemical and autoradiographical study in brain samples obtained from mice lacking D4Rs and their wild-type siblings; comparison of their binding parameters allows a more accurate quantification of the members of the D2-like receptor family (D2, D3, and D4 receptors). We found that the distribution of D2-like receptors in mouse brain is similar to that of rat brain, i.e., caudate putamen, nucleus accumbens, olfactory tubercle, and hippocampus. The contribution of the D4R to the overall population of D2-like receptors is 17% in nucleus accumbens, 21% in caudate putamen and olfactory tubercle, and 40% in hippocampus. Based on our study we conclude that nemonapride probably binds to nondopaminergic sites that if not properly blocked may lead to overestimations of D4R levels. We observed that the experimental condition that better estimates the density of D4 receptors is the displacement of D2 and D3 [3H]nemonapride binding sites with cold raclopride.
Assuntos
Química Encefálica/genética , Característica Quantitativa Herdável , Receptores de Dopamina D2/genética , Animais , Autorradiografia , Benzamidas/metabolismo , Benzamidas/farmacologia , Sítios de Ligação , Antagonistas de Dopamina/metabolismo , Antagonistas de Dopamina/farmacologia , Feminino , Ligantes , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Racloprida/metabolismo , Racloprida/farmacologia , Ensaio Radioligante , Receptores de Dopamina D2/química , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D4 , Sinaptossomos/química , Sinaptossomos/metabolismo , TrítioRESUMO
The human dopamine D4 receptor (D4R) has received considerable attention because of its high affinity for the atypical antipsychotic clozapine and the unusually polymorphic nature of its gene. To clarify the in vivo role of the D4R, we produced and analyzed mutant mice (D4R-/-) lacking this protein. Although less active in open field tests, D4R-/- mice outperformed wild-type mice on the rotarod and displayed locomotor supersensitivity to ethanol, cocaine, and methamphetamine. Biochemical analyses revealed that dopamine synthesis and its conversion to DOPAC were elevated in the dorsal striatum from D4R-/- mice. Based on these findings, we propose that the D4R modulates normal, coordinated and drug-stimulated motor behaviors as well as the activity of nigrostriatal dopamine neurons.