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1.
Int J Mol Sci ; 22(15)2021 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-34361094

RESUMO

Protein-protein interactions between G protein-coupled receptors (GPCRs) can augment their functionality and increase the repertoire of signaling pathways they regulate. New therapeutics designed to modulate such interactions may allow for targeting of a specific GPCR activity, thus reducing potential for side effects. Dopamine receptor (DR) heteromers are promising candidates for targeted therapy of neurological conditions such as Parkinson's disease since current treatments can have severe side effects. To facilitate development of such therapies, it is necessary to identify the various DR binding partners. We report here a new interaction partner for DRD2 and DRD3, the orphan receptor G protein-coupled receptor 143 (GPR143), an atypical GPCR that plays multiple roles in pigment cells and is expressed in several regions of the brain. We previously demonstrated that the DRD2/ DRD3 antagonist pimozide also modulates GPR143 activity. Using confocal microscopy and two FRET methods, we observed that the DRs and GPR143 colocalize and interact at intracellular membranes. Furthermore, co-expression of wildtype GPR143 resulted in a 57% and 67% decrease in DRD2 and DRD3 activity, respectively, as determined by ß-Arrestin recruitment assay. GPR143-DR dimerization may negatively modulate DR activity by changing affinity for dopamine or delaying delivery of the DRs to the plasma membrane.


Assuntos
Dopamina/metabolismo , Proteínas do Olho/metabolismo , Glicoproteínas de Membrana/metabolismo , Domínios e Motivos de Interação entre Proteínas , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , beta-Arrestinas/metabolismo , Proteínas do Olho/genética , Humanos , Glicoproteínas de Membrana/genética , Mutação , Ligação Proteica , Receptores de Dopamina D2/genética , Receptores de Dopamina D3/genética , Transdução de Sinais
2.
Int J Mol Sci ; 22(16)2021 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-34445358

RESUMO

The human dopamine receptors D2S and D3 belong to the group of G protein-coupled receptors (GPCRs) and are important drug targets. Structural analyses and development of new receptor subtype specific drugs have been impeded by low expression yields or receptor instability. Fusing the T4 lysozyme into the intracellular loop 3 improves crystallization but complicates conformational studies. To circumvent these problems, we expressed the human D2S and D3 receptors in Escherichia coli using different N- and C-terminal fusion proteins and thermostabilizing mutations. We optimized expression times and used radioligand binding assays with whole cells and membrane homogenates to evaluate KD-values and the number of receptors in the cell membrane. We show that the presence but not the type of a C-terminal fusion protein is important. Bacteria expressing receptors capable of ligand binding can be selected using FACS analysis and a fluorescently labeled ligand. Improved receptor variants can thus be generated using error-prone PCR. Subsequent analysis of clones showed the distribution of mutations over the whole gene. Repeated cycles of PCR and FACS can be applied for selecting highly expressing receptor variants with high affinity ligand binding, which in the future can be used for analytical studies.


Assuntos
Escherichia coli/genética , Engenharia de Proteínas/métodos , Receptores Dopaminérgicos/genética , Calibragem , Membrana Celular/metabolismo , Clonagem Molecular/métodos , Escherichia coli/metabolismo , Biblioteca Gênica , Humanos , Mutação , Organismos Geneticamente Modificados , Engenharia de Proteínas/normas , Receptores Dopaminérgicos/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/genética , Receptores de Dopamina D3/metabolismo , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transformação Bacteriana , Transgenes
3.
J Med Chem ; 64(12): 8684-8709, 2021 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-34110814

RESUMO

3-(2-Amino-4-methylthiazol-5-yl)propyl-substituted carbamoylguanidines are potent, subtype-selective histamine H2 receptor (H2R) agonists, but their applicability as pharmacological tools to elucidate the largely unknown H2R functions in the central nervous system (CNS) is compromised by their concomitant high affinity toward dopamine D2-like receptors (especially to the D3R). To improve the selectivity, a series of novel carbamoylguanidine-type ligands containing various heterocycles, spacers, and side residues were rationally designed, synthesized, and tested in binding and/or functional assays at H1-4 and D2long/3 receptors. This study revealed a couple of selective candidates (among others 31 and 47), and the most promising ones were screened at several off-target receptors, showing good selectivities. Docking studies suggest that the amino acid residues (3.28, 3.32, E2.49, E2.51, 5.42, and 7.35) are responsible for the different affinities at the H2- and D2long/3-receptors. These results provide a solid base for the exploration of the H2R functions in the brain in further studies.


Assuntos
Guanidinas/farmacologia , Agonistas dos Receptores Histamínicos/farmacologia , Receptores Histamínicos H2/metabolismo , Tiazóis/farmacologia , Animais , Sítios de Ligação , Guanidinas/síntese química , Guanidinas/metabolismo , Cobaias , Células HEK293 , Agonistas dos Receptores Histamínicos/síntese química , Agonistas dos Receptores Histamínicos/metabolismo , Humanos , Camundongos , Simulação de Acoplamento Molecular , Estrutura Molecular , Ratos , Receptores de Dopamina D2/química , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/química , Receptores de Dopamina D3/metabolismo , Receptores Histamínicos H2/química , Células Sf9 , Relação Estrutura-Atividade , Tiazóis/síntese química , Tiazóis/metabolismo
4.
Biomolecules ; 11(5)2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-34064602

RESUMO

Biomolecules has launched a Special Issue entitled "Dopamine D3 Receptor: Contemporary Views of Its Function and Pharmacology for Neuropsychiatric Diseases [...].


Assuntos
Transtornos Mentais/tratamento farmacológico , Neuropsiquiatria/métodos , Receptores de Dopamina D3/metabolismo , Animais , Humanos , Transtornos Mentais/metabolismo , Publicações/estatística & dados numéricos
5.
J Chem Inf Model ; 61(6): 2829-2843, 2021 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-33988991

RESUMO

Targeting the D3 dopamine receptor (D3R) is a promising pharmacotherapeutic strategy for the treatment of many disorders. The structure of the D3R is similar to the D2 dopamine receptor (D2R), especially in the transmembrane spanning regions that form the orthosteric binding site, making it difficult to identify D3R selective pharmacotherapeutic agents. Here, we examine the molecular basis for the high affinity D3R binding and D3R vs D2R binding selectivity of substituted phenylpiperazine thiopheneamides. We show that removing the thiophenearylamide portion of the ligand consistently decreases the affinity of these ligands at D3R, while not affecting their affinity at the D2R. Our long (>10 µs) molecular dynamics simulations demonstrated that both dopamine receptor subtypes adopt two major conformations that we refer to as closed or open conformations, with D3R sampling the open conformation more frequently than D2R. The binding of ligands with conjoined orthosteric-allosteric binding moieties causes the closed conformation to populate more often in the trajectories. Also, significant differences were observed in the extracellular loops (ECL) of these two receptor subtypes leading to the identification of several residues that contribute differently to the ligand binding for the two receptors that could potentially contribute to ligand binding selectivity. Our observations also suggest that the displacement of ordered water in the binding pocket of D3R contributes to the affinity of the compounds containing an allosteric binding motif. These studies provide a better understanding of how a bitopic mode of engagement can determine ligands that bind selectively to D2 and D3 dopamine receptor subtypes.


Assuntos
Receptores de Dopamina D3 , Ligantes , Conformação Molecular , Ligação Proteica , Receptores de Dopamina D3/metabolismo , Relação Estrutura-Atividade
6.
J Med Chem ; 64(11): 7778-7808, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34011153

RESUMO

The need for safer pain-management therapies with decreased abuse liability inspired a novel drug design that retains µ-opioid receptor (MOR)-mediated analgesia, while minimizing addictive liability. We recently demonstrated that targeting the dopamine D3 receptor (D3R) with highly selective antagonists/partial agonists can reduce opioid self-administration and reinstatement to drug seeking in rodent models without diminishing antinociceptive effects. The identification of the D3R as a target for the treatment of opioid use disorders prompted the idea of generating a class of ligands presenting bitopic or bivalent structures, allowing the dual-target binding of the MOR and D3R. Structure-activity relationship studies using computationally aided drug design and in vitro binding assays led to the identification of potent dual-target leads (23, 28, and 40), based on different structural templates and scaffolds, with moderate (sub-micromolar) to high (low nanomolar/sub-nanomolar) binding affinities. Bioluminescence resonance energy transfer-based functional studies revealed MOR agonist-D3R antagonist/partial agonist efficacies that suggest potential for maintaining analgesia with reduced opioid-abuse liability.


Assuntos
Antagonistas de Dopamina/química , Ligantes , Receptores de Dopamina D3/metabolismo , Receptores Opioides mu/metabolismo , Analgésicos Opioides/uso terapêutico , Animais , Sítios de Ligação , Compostos de Bifenilo/química , Compostos de Bifenilo/metabolismo , Compostos de Bifenilo/uso terapêutico , Modelos Animais de Doenças , Antagonistas de Dopamina/metabolismo , Antagonistas de Dopamina/uso terapêutico , Desenho de Fármacos , Transferência Ressonante de Energia de Fluorescência , Camundongos , Simulação de Acoplamento Molecular , Transtornos Relacionados ao Uso de Opioides/tratamento farmacológico , Dor/tratamento farmacológico , Manejo da Dor , Receptores de Dopamina D3/agonistas , Receptores de Dopamina D3/antagonistas & inibidores , Receptores Opioides mu/agonistas , Relação Estrutura-Atividade
7.
Biomolecules ; 11(3)2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33799860

RESUMO

Dopamine receptors (DRs) are generally considered as mediators of vasomotor functions. However, when used in pharmacological studies, dopamine and/or DR agonists may not discriminate among different DR subtypes and may even stimulate alpha1 and beta-adrenoceptors. Here, we tested the hypothesis that D2R and/or D3R may specifically induce vasoconstriction in isolated mouse aorta. Aorta, isolated from wild-type (WT) and D3R-/- mice, was mounted in a wire myograph and challenged with cumulative concentrations of phenylephrine (PE), acetylcholine (ACh), and the D3R agonist 7-hydrxy-N,N-dipropyl-2-aminotetralin (7-OH-DPAT), with or without the D2R antagonist L741,626 and the D3R antagonist SB-277011-A. The vasoconstriction to PE and the vasodilatation to ACh were not different in WT and D3R-/-; in contrast, the contractile responses to 7-OH-DPAT were significantly weaker in D3R-/-, though not abolished. L741,626 did not change the contractile response induced by 7-OH-DPAT in WT or in D3R-/-, whereas SB-277011-A significantly reduced it in WT but did not in D3R-/-. D3R mRNA (assessed by qPCR) was about 5-fold more abundant than D2R mRNA in aorta from WT and undetectable in aorta from D3R-/-. Following transduction with lentivirus (72-h incubation) delivering synthetic microRNAs to specifically inactivate D2R (LV-miR-D2) or D3R (LV-miR-D3), the contractile response to 7-OH-DPAT was unaffected by LV-miR-D2, while it was significantly reduced by LV-miR-D3. These data indicate that, at least in mouse aorta, D3R stimulation induces vasoconstriction, while D2R stimulation does not. This is consistent with the higher expression level of D3R. The residual vasoconstriction elicited by high concentration D3R agonist in D3R-/- and/or in the presence of D3R antagonist is likely to be unrelated to DRs.


Assuntos
Aorta/fisiologia , Receptores de Dopamina D3/metabolismo , Vasoconstrição/genética , Animais , Aorta/efeitos dos fármacos , Indóis/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nitrilas/farmacologia , Piperidinas/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Dopamina D2/genética , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/agonistas , Receptores de Dopamina D3/antagonistas & inibidores , Receptores de Dopamina D3/genética , Tetra-Hidroisoquinolinas/farmacologia , Tetra-Hidronaftalenos/farmacologia , Vasoconstrição/efeitos dos fármacos
8.
Biomolecules ; 11(4)2021 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-33924613

RESUMO

The dopamine D2/D3 receptor (D2R/D3R) agonists are used as therapeutics for Parkinson's disease (PD) and other motor disorders. Selective targeting of D3R over D2R is attractive because of D3R's restricted tissue distribution with potentially fewer side-effects and its putative neuroprotective effect. However, the high sequence homology between the D2R and D3R poses a challenge in the development of D3R selective agonists. To address the ligand selectivity, bitopic ligands were designed and synthesized previously based on a potent D3R-preferential agonist PF592,379 as the primary pharmacophore (PP). This PP was attached to various secondary pharmacophores (SPs) using chemically different linkers. Here, we characterize some of these novel bitopic ligands at both D3R and D2R using BRET-based functional assays. The bitopic ligands showed varying differences in potencies and efficacies. In addition, the chirality of the PP was key to conferring improved D3R potency, selectivity, and G protein signaling bias. In particular, compound AB04-88 exhibited significant D3R over D2R selectivity, and G protein bias at D3R. This bias was consistently observed at various time-points ranging from 8 to 46 min. Together, the structure-activity relationships derived from these functional studies reveal unique pharmacology at D3R and support further evaluation of functionally biased D3R agonists for their therapeutic potential.


Assuntos
Agonistas de Dopamina/farmacologia , Receptores de Dopamina D3/metabolismo , Aminopiridinas/química , Aminopiridinas/farmacologia , Sítios de Ligação , Agonistas de Dopamina/síntese química , Transferência de Energia , Células HEK293 , Humanos , Luminescência , Morfolinas/química , Morfolinas/farmacologia , Ligação Proteica , Receptores de Dopamina D2/química , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/química , Estereoisomerismo , Relação Estrutura-Atividade
9.
Biomolecules ; 11(4)2021 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-33918451

RESUMO

[18F]Fallypride and [18F]Fluortriopride (FTP) are two different PET radiotracers that bind with sub-nanomolar affinity to the dopamine D3 receptor (D3R). In spite of their similar D3 affinities, the two PET ligands display very different properties for labeling the D3R in vivo: [18F]Fallypride is capable of binding to D3R under "baseline" conditions, whereas [18F]FTP requires the depletion of synaptic dopamine in order to image the receptor in vivo. These data suggest that [18F]Fallypride is able to compete with synaptic dopamine for binding to the D3R, whereas [18F]FTP is not. The goal of this study was to conduct a series of docking and molecular dynamic simulation studies to identify differences in the ability of each molecule to interact with the D3R that could explain these differences with respect to competition with synaptic dopamine. Competition studies measuring the ability of each ligand to compete with dopamine in the ß-arrestin assay were also conducted. The results of the in silico studies indicate that FTP has a weaker interaction with the orthosteric binding site of the D3R versus that of Fallypride. The results of the in silico studies were also consistent with the IC50 values of each compound in the dopamine ß-arrestin competition assays. The results of this study indicate that in silico methods may be able to predict the ability of a small molecule to compete with synaptic dopamine for binding to the D3R.


Assuntos
Ligantes , Receptores de Dopamina D3/química , Benzamidas/química , Benzamidas/metabolismo , Sítios de Ligação , Humanos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Tomografia por Emissão de Pósitrons , Compostos Radiofarmacêuticos/química , Compostos Radiofarmacêuticos/metabolismo , Receptores de Dopamina D3/metabolismo , Termodinâmica
10.
Int J Mol Sci ; 22(6)2021 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-33803578

RESUMO

Relapse in the seeking and intake of cocaine is one of the main challenges when treating its addiction. Among the triggering factors for the recurrence of cocaine use are the re-exposure to the drug and stressful events. Cocaine relapse engages the activity of memory-related nuclei, such as the basolateral amygdala (BLA) and the hippocampal dentate gyrus (DG), which are responsible for emotional and episodic memories. Moreover, D3 receptor (D3R) antagonists have recently arisen as a potential treatment for preventing drug relapse. Thus, we have assessed the impact of D3R blockade in the expression of some dopaminergic markers and the activity of the mTOR pathway, which is modulated by D3R, in the BLA and DG during the reinstatement of cocaine-induced conditioned place preference (CPP) evoked by drug priming and social stress. Reinstatement of cocaine CPP paralleled an increasing trend in D3R and dopamine transporter (DAT) levels in the BLA. Social stress, but not drug-induced reactivation of cocaine memories, was prevented by systemic administration of SB-277011-A (a selective D3R antagonist), which was able, however, to impede D3R and DAT up-regulation in the BLA during CPP reinstatement evoked by both stress and cocaine. Concomitant with cocaine CPP reactivation, a diminution in mTOR phosphorylation (activation) in the BLA and DG occurred, which was inhibited by D3R blockade in both nuclei before the social stress episode and only in the BLA when CPP reinstatement was provoked by a cocaine prime. Our data, while supporting a main role for D3R signalling in the BLA in the reactivation of cocaine memories evoked by social stress, indicate that different neural circuits and signalling mechanisms might mediate in the reinstatement of cocaine-seeking behaviours depending upon the triggering stimuli.


Assuntos
Complexo Nuclear Basolateral da Amígdala/metabolismo , Cocaína/farmacologia , Condicionamento Clássico , Giro Denteado/metabolismo , Receptores de Dopamina D3/metabolismo , Estresse Psicológico/metabolismo , Animais , Comportamento Animal/efeitos dos fármacos , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Nitrilas/administração & dosagem , Nitrilas/farmacologia , Fosforilação/efeitos dos fármacos , Receptores de Dopamina D3/antagonistas & inibidores , Derrota Social , Serina-Treonina Quinases TOR/metabolismo , Tetra-Hidroisoquinolinas/administração & dosagem , Tetra-Hidroisoquinolinas/farmacologia
11.
Biomed Pharmacother ; 138: 111517, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33773463

RESUMO

Several brain neurotransmitters, including histamine (HA), acetylcholine (ACh), and dopamine (DA) are suggested to be involved in several brain disorders including cognitive deficits, depression, schizophrenia, anxiety, and narcolepsy, all of which are comorbid with Autism spectrum disorder (ASD). Therefore, the ameliorative effects of the novel multiple-active compound ST-713 with high binding affinities at histamine H3 receptor (H3R), dopamine D2sR and D3R on ASD-like behaviors in male BTBR T+tf/J mice model were assessed. ST-713 (3-(2-chloro-10H-phenothiazin-10-yl)-N-methyl-N-(4-(3-(piperidin-1-yl)propoxy)benzyl)propan-1-amine; 2.5, 5, and 10 mg/kg, i.p.) ameliorated dose-dependently social deficits, and significantly alleviated the repetitive/compulsive behaviors of BTBR mice (all P < 0.05). Moreover, ST-713 modulated disturbed anxiety levels, but failed to obliterate increased hyperactivity of tested mice. Furthermore, ST-713 (5 mg/kg) attenuated the increased levels of hippocampal and cerebellar protein expressions of NF-κB p65, COX-2, and iNOS in BTBR mice (all P < 0.05). The ameliorative effects of ST-713 on social parameters were entirely reversed by co-administration of the H3R agonist (R)-α-methylhistamine or the anticholinergic drug scopolamine. The obtained results demonstrate the potential of multiple-active compounds for the therapeutic management of neuropsychiatric disorders, e.g. ASD.


Assuntos
Transtorno Autístico/tratamento farmacológico , Antagonistas de Dopamina/uso terapêutico , Antagonistas dos Receptores de Dopamina D2/uso terapêutico , Antagonistas dos Receptores Histamínicos/uso terapêutico , Receptores de Dopamina D3/antagonistas & inibidores , Receptores Histamínicos H3 , Animais , Transtorno Autístico/genética , Transtorno Autístico/metabolismo , Cerebelo/efeitos dos fármacos , Cerebelo/metabolismo , Antagonistas de Dopamina/farmacologia , Antagonistas dos Receptores de Dopamina D2/farmacologia , Relação Dose-Resposta a Droga , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Antagonistas dos Receptores Histamínicos/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Receptores Histamínicos H3/metabolismo
12.
Int J Mol Sci ; 22(4)2021 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-33669336

RESUMO

Autism spectrum disorder (ASD) is a complex heterogeneous neurodevelopmental disorder characterized by social and communicative impairments, as well as repetitive and restricted behaviors (RRBs). With the limited effectiveness of current pharmacotherapies in treating repetitive behaviors, the present study determined the effects of acute systemic treatment of the novel multi-targeting ligand ST-2223, with incorporated histamine H3 receptor (H3R) and dopamine D2/D3 receptor affinity properties, on ASD-related RRBs in a male Black and Tan BRachyury (BTBR) mouse model of ASD. ST-2223 (2.5, 5, and 10 mg/kg, i.p.) significantly mitigated the increase in marble burying and self-grooming, and improved reduced spontaneous alternation in BTBR mice (all p < 0.05). Similarly, reference drugs memantine (MEM, 5 mg/kg, i.p.) and aripiprazole (ARP, 1 mg/kg, i.p.), reversed abnormally high levels of several RRBs in BTBR (p < 0.05). Moreover, ST-2223 palliated the disturbed anxiety levels observed in an open field test (all p < 0.05), but did not restore the hyperactivity parameters, whereas MEM failed to restore mouse anxiety and hyperactivity. In addition, ST-2223 (5 mg/kg, i.p.) mitigated oxidative stress status by decreasing the elevated levels of malondialdehyde (MDA), and increasing the levels of decreased glutathione (GSH), superoxide dismutase (SOD), and catalase (CAT) in different brain parts of treated BTBR mice (all p < 0.05). These preliminary in vivo findings demonstrate the ameliorative effects of ST-2223 on RRBs in a mouse model of ASD, suggesting its pharmacological prospective to rescue core ASD-related behaviors. Further confirmatory investigations on its effects on various brain neurotransmitters, e.g., dopamine and histamine, in different brain regions are still warranted to corroborate and expand these initial data.


Assuntos
Transtorno do Espectro Autista/tratamento farmacológico , Encéfalo/metabolismo , Antagonistas dos Receptores de Dopamina D2/administração & dosagem , Asseio Animal/efeitos dos fármacos , Antagonistas dos Receptores Histamínicos H3/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Receptores de Dopamina D3/antagonistas & inibidores , Animais , Ansiedade/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Modelos Animais de Doenças , Antagonistas dos Receptores de Dopamina D2/metabolismo , Células HEK293 , Antagonistas dos Receptores Histamínicos H3/metabolismo , Humanos , Ligantes , Locomoção/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Receptores Histamínicos H3/metabolismo
13.
Neurobiol Aging ; 99: 65-78, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33422895

RESUMO

Mutations in the leucine-rich repeat kinase 2 (LRRK2) are the most common genetic determinants of Parkinson's disease (PD), with the G2019S accounting for about 3% of PD cases. LRRK2 regulates various cellular processes, including vesicle trafficking that is crucial for receptor localization at the plasma membrane. In this study, induced pluripotent stem cells derived from 2 PD patients bearing the G2019S LRRK2 kinase activating mutation were used to generate neuronal cultures enriched in dopaminergic neurons. The results show that mutant LRRK2 prevents the membrane localization of both the dopamine D3 receptors (D3R) and the nicotinic acetylcholine receptors (nAChR) and the formation of the D3R-nAChR heteromer, a molecular unit crucial for promoting neuronal homeostasis and preserving dopaminergic neuron health. Interestingly, D3R and nAChR as well as the corresponding heteromer membrane localization were rescued by inhibiting the abnormally increased kinase activity. Thus, the altered membrane localization of the D3R-nAChR heteromer associated with mutation in LRRK2 might represent a pre-degenerative feature of dopaminergic neurons contributing to the special vulnerability of this neuronal population.


Assuntos
Membrana Celular/metabolismo , Neurônios Dopaminérgicos/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Mutação , Doença de Parkinson/genética , Receptores de Dopamina D3/metabolismo , Receptores Nicotínicos/metabolismo , Humanos
14.
Mol Psychiatry ; 26(6): 2038-2047, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33420479

RESUMO

Research using rodent models has established a relationship between the steroid hormone estrogen and dopamine function, by revealing changes throughout the estrous cycle and by directly manipulating neuroendocrine signaling through ovariectomy and administration of estrogen. However, a direct link between estrogen levels and dopamine signaling had not been established in humans. The goal of this study, therefore, was to assess the relationship between circulating 17ß-estradiol and dopamine signaling in the human brain by testing for a relationship between two proxies for these variables: peripheral 17ß-estradiol and striatal dopamine D2-type receptor availability, measured with [18F]fallypride and positron emission tomography (PET). Sixteen (23-45 years of age) women were tested on 2 days of the menstrual cycle estimated prospectively to occur during (a) the early follicular phase, when estrogen levels are near their nadir, and (b) the periovulatory phase, when estrogen levels peak. PET scans with [18F]fallypride were performed on these 2 days, and serum 17ß-estradiol was measured using radioimmunoassay. Dopamine D2-type receptor availability did not differ significantly in the whole striatum or the caudate, putamen, or accumbens subregions during the high-estrogen vs. the low-estrogen phases of the menstrual cycle. We conclude that circulating estrogen levels do not affect dopamine D2-type receptor availability in the human striatum although other indices of dopaminergic function may be affected.


Assuntos
Dopamina , Receptores de Dopamina D2 , Corpo Estriado/metabolismo , Estradiol , Feminino , Humanos , Tomografia por Emissão de Pósitrons , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo
15.
Drug Alcohol Depend ; 220: 108514, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33454626

RESUMO

BACKGROUND: Positron emission tomography (PET) work with the dopamine D3 receptor (D3R) preferring ligand [11C]PHNO in obese individuals has demonstrated higher binding and positive correlations with body mass index (BMI) in otherwise healthy individuals. These findings implicated brain reward areas including the substantia nigra/ventral tegmental area (SN/VTA) and pallidum. In cocaine use disorder (CUD), similar SN/VTA binding profiles have been found compared to healthy control subjects. This study investigates whether BMI-[11C]PHNO relationships are similar in individuals with CUD. METHODS: Non-obese CUD subjects (N = 12) were compared to age-matched obese CUD subjects (N = 14). All subjects underwent [11C]PHNO acquisition using a High Resolution Research Tomograph PET scanner. Parametric images were computed using the simplified reference tissue model with cerebellum as the reference region. [11C]PHNO measures of receptor availability were calculated and expressed as non-displaceable binding potential (BPND). RESULTS: In between-group analyses, D2/3R availability in non-obese and obese CUD groups was not significantly different overall. BMI was inversely correlated withBPND in the SN/VTA (r = -0.45, p = 0.02 uncorrected) in all subjects. CONCLUSION: These data suggest that obesity in CUD was not associated with significant differences in D2/3R availability. This in contrast to previous findings in non-CUD individuals that found increased availability of D3Rs in the SN/VTA associated with obesity. These findings could potentially reflect dysregulation of D3R in CUD, impacting how affected individuals respond to natural stimuli such as food.


Assuntos
Transtornos Relacionados ao Uso de Cocaína/patologia , Obesidade/patologia , Tomografia por Emissão de Pósitrons , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Adulto , Índice de Massa Corporal , Globo Pálido/diagnóstico por imagem , Humanos , Ligantes , Pessoa de Meia-Idade , Substância Negra/diagnóstico por imagem , Área Tegmentar Ventral/diagnóstico por imagem
16.
Biomolecules ; 11(1)2021 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-33466844

RESUMO

Dopamine (DA), as one of the major neurotransmitters in the central nervous system (CNS) and periphery, exerts its actions through five types of receptors which belong to two major subfamilies such as D1-like (i.e., D1 and D5 receptors) and D2-like (i.e., D2, D3 and D4) receptors. Dopamine D3 receptor (D3R) was cloned 30 years ago, and its distribution in the CNS and in the periphery, molecular structure, cellular signaling mechanisms have been largely explored. Involvement of D3Rs has been recognized in several CNS functions such as movement control, cognition, learning, reward, emotional regulation and social behavior. D3Rs have become a promising target of drug research and great efforts have been made to obtain high affinity ligands (selective agonists, partial agonists and antagonists) in order to elucidate D3R functions. There has been a strong drive behind the efforts to find drug-like compounds with high affinity and selectivity and various functionality for D3Rs in the hope that they would have potential treatment options in CNS diseases such as schizophrenia, drug abuse, Parkinson's disease, depression, and restless leg syndrome. In this review, we provide an overview and update of the major aspects of research related to D3Rs: distribution in the CNS and periphery, signaling and molecular properties, the status of ligands available for D3R research (agonists, antagonists and partial agonists), behavioral functions of D3Rs, the role in neural networks, and we provide a summary on how the D3R-related drug research has been translated to human therapy.


Assuntos
Pesquisa Biomédica , Doenças do Sistema Nervoso Central/metabolismo , Neurônios/metabolismo , Receptores de Dopamina D3/metabolismo , Pesquisa Médica Translacional , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Humanos , Receptores de Dopamina D3/antagonistas & inibidores , Receptores de Dopamina D3/química
17.
Eur J Nucl Med Mol Imaging ; 48(4): 1103-1115, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-32995944

RESUMO

PURPOSE: Pridopidine is an investigational drug for Huntington disease (HD). Pridopidine was originally thought to act as a dopamine stabilizer. However, pridopidine shows highest affinity to the sigma-1 receptor (S1R) and enhances neuroprotection via the S1R in preclinical studies. Using [18F] fluspidine and [18F] fallypride PET, the purpose of this study was to assess in vivo target engagement/receptor occupancy of pridopidine to the S1R and dopamine D2/D3 receptor (D2/D3R) at clinical relevant doses in healthy volunteers (HVs) and as proof-of-concept in a small number of patients with HD. METHODS: Using [18F] fluspidine PET (300 MBq, 0-90 min), 11 male HVs (pridopidine 0.5 to 90 mg; six dose groups) and three male patients with HD (pridopidine 90 mg) were investigated twice, without and 2 h after single dose of pridopidine. Using [18F] fallypride PET (200 MBq, 0-210 min), four male HVs were studied without and 2 h following pridopidine administration (90 mg). Receptor occupancy was analyzed by the Lassen plot. RESULTS: S1R occupancy as function of pridopidine dose (or plasma concentration) in HVs could be described by a three-parameter Hill equation with a Hill coefficient larger than one. A high degree of S1R occupancy (87% to 91%) was found throughout the brain at pridopidine doses ranging from 22.5 to 90 mg. S1R occupancy was 43% at 1 mg pridopidine. In contrast, at 90 mg pridopidine, the D2/D3R occupancy was only minimal (~ 3%). CONCLUSIONS: Our PET findings indicate that at clinically relevant single dose of 90 mg, pridopidine acts as a selective S1R ligand showing near to complete S1R occupancy with negligible occupancy of the D2/D3R. The dose S1R occupancy relationship suggests cooperative binding of pridopidine to the S1R. Our findings provide significant clarification about pridopidine's mechanism of action and support further use of the 45-mg twice-daily dose to achieve full and selective targeting of the S1R in future clinical trials of neurodegenerative disorders. Clinical Trials.gov Identifier: NCT03019289 January 12, 2017; EUDRA-CT-Nr. 2016-001757-41.


Assuntos
Dopamina , Doença de Huntington , Benzamidas , Benzofuranos , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Voluntários Saudáveis , Humanos , Doença de Huntington/diagnóstico por imagem , Masculino , Piperidinas , Tomografia por Emissão de Pósitrons , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo
18.
Eur J Pharmacol ; 890: 173635, 2021 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-33065094

RESUMO

F17464 (N-(3-{4-[4-(8-Oxo-8H-[1,3]-dioxolo-[4,5-g]-chromen-7-yl)-butyl]-piperazin-1-yl}-phenyl)-methanesulfonamide, hydrochloride) is a new potential antipsychotic with a unique profile. The compound exhibits high affinity for the human dopamine receptor subtype 3 (hD3) (Ki = 0.17 nM) and the serotonin receptor subtype 1a (5-HT1a) (Ki = 0.16 nM) and a >50 fold lower affinity for the human dopamine receptor subtype 2 short and long form (hD2s/l) (Ki = 8.9 and 12.1 nM, respectively). [14C]F17464 dynamic studies show a slower dissociation rate from hD3 receptor (t1/2 = 110 min) than from hD2s receptor (t1/2 = 1.4 min) and functional studies demonstrate that F17464 is a D3 receptor antagonist, 5-HT1a receptor partial agonist. In human dopaminergic neurons F17464 blocks ketamine induced morphological changes, an effect D3 receptor mediated. In vivo F17464 target engagement of both D2 and 5-HT1a receptors is demonstrated in displacement studies in the mouse brain. F17464 increases dopamine release in the rat prefrontal cortex and mouse lateral forebrain - dorsal striatum and seems to reduce the effect of MK801 on % c-fos mRNA medium expressing neurons in cortical and subcortical regions. F17464 also rescues valproate induced impairment in a rat social interaction model of autism. All the neurochemistry and behavioural effects of F17464 are observed in the dose range 0.32-2.5 mg/kg i.p. in both rats and mice. The in vitro - in vivo pharmacology profile of F17464 in preclinical models is discussed in support of a therapeutic use of the compound in schizophrenia and autism.


Assuntos
Antipsicóticos/farmacologia , Antagonistas de Dopamina/farmacologia , Receptores de Dopamina D3/antagonistas & inibidores , Animais , Antipsicóticos/uso terapêutico , Transtorno Autístico/induzido quimicamente , Transtorno Autístico/tratamento farmacológico , Comportamento Animal/efeitos dos fármacos , Monoaminas Biogênicas/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Catalepsia/tratamento farmacológico , Células Cultivadas , Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Feminino , Genes fos/efeitos dos fármacos , Masculino , Camundongos , Plasticidade Neuronal/efeitos dos fármacos , Prolactina/sangue , Ratos Sprague-Dawley , Receptores de Dopamina D3/metabolismo , Ácido Valproico/toxicidade
19.
Psychopharmacology (Berl) ; 238(5): 1343-1350, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33180175

RESUMO

RATIONALE: Unlike other antipsychotics, our previous positron emission tomography (PET) study demonstrated that a single dose of blonanserin occupied dopamine D3 as well as dopamine D2 receptors in healthy subjects. However, there has been no study concerning the continued use of blonanserin. OBJECTIVES: We examined D2 and D3 receptor occupancies in patients with schizophrenia who had been treated with blonanserin. METHODS: Thirteen patients with schizophrenia participated. PET examinations were performed on patients treated with clinical dosage of blonanserin or olanzapine alone. A crossover design was used in which seven patients switched drugs after the first scan, and PET examinations were conducted again. D2 and D3 receptor occupancies were evaluated by [11C]-(+)-PHNO. We used nondisplaceable binding potential (BPND) of 6 healthy subjects which we previously reported as baseline. To consider the effect of upregulation of D3 receptor by continued use of antipsychotics, D3 receptor occupancy by blonanserin in seven subjects who completed 2 PET scans were re-analyzed by using BPND of olanzapine condition as baseline. RESULTS: Average occupancy by olanzapine (10.8 ± 6.0 mg/day) was as follows: caudate 32.8 ± 18.3%, putamen 26.3 ± 18.2%, globus pallidus - 33.7 ± 34.9%, substantia nigra - 112.8 ± 90.7%. Average occupancy by blonanserin (12.8 ± 5.6 mg/day) was as follows: caudate 61.0 ± 8.3%, putamen 55.5 ± 9.5%, globus pallidus 48.9 ± 12.4%, substantia nigra 34.0 ± 20.6%. EC50 was 0.30 ng/mL for D2 receptor for caudate and putamen (df = 19, p < 0.0001) and 0.70 ng/mL for D3 receptor for globus pallidus and substantia nigra (df = 19, p < 0.0001). EC50 for D3 receptor of blonanserin changed to 0.22 ng/mL (df = 13, p = 0.0041) when we used BPND of olanzapine condition as baseline. CONCLUSIONS: Our study confirmed that blonanserin occupied both D2 and D3 receptors in patients with schizophrenia.


Assuntos
Piperazinas/farmacologia , Piperidinas/farmacologia , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Esquizofrenia/tratamento farmacológico , Adulto , Idoso , Antipsicóticos/uso terapêutico , Estudos Cross-Over , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Olanzapina/uso terapêutico , Tomografia por Emissão de Pósitrons/métodos , Putamen/metabolismo , Adulto Jovem
20.
Ann Clin Transl Neurol ; 8(1): 224-237, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33348472

RESUMO

OBJECTIVE: Dopamine D2-like receptors - mainly dopamine D2 receptors (D2R) and dopamine D3 receptors (D3R) - are believed to be greatly involved in the pathology of Parkinson disease (PD) progression. However, these receptors have not been precisely examined in PD patients. Our aim was to quantitatively calculate the exact densities of dopamine D1 receptors (D1R), D2R, and D3R in control, Alzheimer disease (AD), and Lewy body disease (LBD) patients (including PD, Dementia with Lewy bodies, and Parkinson disease dementia); and analyze the relationship between dopamine receptors and clinical PD manifestations. METHODS: We analyzed the densities of D1R, D2R, and D3R in the striatum and substantia nigra (SN) using a novel quantitative autoradiography procedure previously developed by our group. We also examined the expression of D2R and D3R mRNA in the striatum by in situ hybridization. RESULTS: The results showed that although no differences of striatal D1R were found among all groups; D2R was significantly decreased in the striatum of PD patients when compared with control and AD patients. Some clinical manifestations: age of onset, PD stage, dopamine responsiveness, and survival time after onset; showed a better correlation with striatal D1R + D3R densities combined compared to D1R or D3R alone. INTERPRETATION: There is a possibility that we may infer the results in diagnosis, treatment, and prognosis of PD by detecting D1R + D3R as opposed to using dopamine D1 or D3 receptors alone. This is especially true for elderly patients with low D2R expression as is common in this disease.


Assuntos
Encéfalo/metabolismo , Doença de Parkinson/metabolismo , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo , Receptores de Dopamina D3/metabolismo , Idoso , Idoso de 80 Anos ou mais , Autorradiografia/métodos , Encéfalo/patologia , Feminino , Humanos , Masculino , Doença de Parkinson/patologia
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