| Summary: | 碩士 === 長庚大學 === 基礎醫學研究所 === 94 === Dopamine D3 receptor (D3R) belongs to a family of G protein-coupled receptors (GPCRs), primarily found in limbic brain areas such as the nucleus accumbens (rostral pole and parts of shell), islands of Calleja and olfactory tubercle. Due to its restricted localization, D3 receptor has led to the hypothesis that it may play an important role in emotion, cognition, and drug addiction. Brain-derived neurotrophic factor (BDNF) could mediate activity-dependent plasticity in a local and synapse-specific manner. It also contributes to the survival and differentiation of dopaminergic neurons, and influences the release of dopamine and serotonin, which are closely linked to drug addiction. These evidences indicate that the role of BDNF receptor (trkB) in addictive drug-induced behavioral and/or biochemical/cellular effects could be more complicated, hence requires further characterization. Previously, we had found that behavioral sensitization to methamphetamine (METH) were escalated in dopamine (DA) D3 receptor KO mice. On behalf of existing evidence that trkB receptor and D3R are co-localized in the shell of nucleus accumbens, we intend to investigate the impact of BDNF-trkB on METH sensitization. First, we found basal trkB expression was enhanced in D3KO mice versus wild type in the olfactory tubercle. After treatment of 2 mg/kg METH for 8 days, D3KO mice displayed an even heightened level of trkB than METH-treated wild type. To delineate the possible receptor crosstalk between D3R and trkB, we established a cell line in HEK293 cells which could stably express both DA D3 and trkB receptors. Via receptor binding assay against [3H] 7-OH-DPAT and functional screening to monitor the BDNF-evoked ERK1/2 phosphorylation, we successively selected the double transfected D3R/trkB clone and established HEK293/hD3R and HEK293/trkB controls. We further found in HEK293/D3R-trkB cells, D3R agonist and/or BDNF could induce, dose- and time-dependent, ERK1/2 and JNK as well as Akt phosphorylation. For trkB down-stream signals, ERK1/2 and JNK could not be transactivated through dopamine D3 receptor activation. Besides, internalization may not involve in those intracellular signaling after activation of either dopamine D3 or trkB receptor in HEK-293/D3R-trkB cells. Co-stimulation of dopamine D3 and trkB receptors would produce synergistic effect on ERK1/2, JNK, and AktThr308 phosphorylation, but not on AktSer473 phosphorylation. These results provide a notion that up-regulation of dopamine and BDNF in synapse after chronic psychostimulants (such as amphetamine, methamphetamine or cocaine) could simultaneously activate postsynaptic dopamine D3 and trkB receptors, induce various abused drugs-caused neuroplasticity.
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