ABSTRACT
Cell adhesion molecule close homolog of L1 (CHL1) and the dopamine receptor D2 (DRD2) are associated with psychiatric and mental disorders. We here show that DRD2 interacts with CHL1 in mouse brain, as evidenced by co-immunostaining, proximity ligation assay, co-immunoprecipitation, and pull-down assay with recombinant extracellular CHL1 domain fused to Fc (CHL1-Fc). Direct binding of CHL1-Fc to the first extracellular loop of DRD2 was shown by ELISA. Using HEK cells transfected to co-express CHL1 and the short (DRD2-S) or long (DRD2-L) DRD2 isoforms, co-localization of CHL1 and both isoforms was observed by immunostaining and proximity ligation assay. Moreover, CHL1 inhibited agonist-triggered internalization of DRD2-S. Proximity ligation assay showed close interaction between CHL1 and DRD2 in neurons expressing dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP32) or tyrosine hydroxylase (TH) in tissue sections of adult mouse striatum. In cultures of striatum or ventral midbrain, CHL1 was also closely associated with DRD2 in DARPP32- or TH-immunopositive cells, respectively. In the dorsal striatum of CHL1-deficient mice, lower levels of DRD2 and phosphorylated TH were observed, when compared to wild-type littermates. In the ventral striatum of CHL1-deficient mice, levels of phosphorylated DARPP32 were reduced. We propose that CHL1 regulates DRD2-dependent presynaptic and postsynaptic functions.
Subject(s)
Cell Adhesion Molecules/metabolism , Receptors, Dopamine D2/metabolism , Animals , Binding Sites , Brain/cytology , Brain/metabolism , Cell Adhesion Molecules/chemistry , Cell Adhesion Molecules/genetics , Dopamine and cAMP-Regulated Phosphoprotein 32/genetics , Dopamine and cAMP-Regulated Phosphoprotein 32/metabolism , HEK293 Cells , Humans , Mice , Mice, Inbred C57BL , Neurons/metabolism , Protein Binding , Receptors, Dopamine D2/chemistry , Receptors, Dopamine D2/genetics , Tyrosine 3-Monooxygenase/genetics , Tyrosine 3-Monooxygenase/metabolismABSTRACT
The repeated intake of cocaine evokes oxidative stress that is present even during drug withdrawal. Recent studies demonstrate that cocaine-induced oxidative and/or endoplasmic reticulum stress can affect mitochondrial function and dynamics as well as the expression of mitochondrial and nuclear genes. These alterations in mitochondrial function may determine synaptic and behavioral plasticity. Mitochondria and mitochondrial DNA (mtDNA) seem to play an important role in the initiation of drug addiction. We used a microarray approach to investigate the expression patterns of nuclear-encoded genes relevant for mitochondrial functions and quantitative real-time PCR assays to determine the numbers of copies of mtDNA and of mRNAs corresponding to two mitochondrial proteins in the prefrontal cortex and hippocampus of rats during early cocaine abstinence. We found a significant elevation in the copy number of mtDNA and concomitant increased expression of mitochondrial genes. Moreover, microarray analysis revealed changes in the transcription of nuclear genes engaged in mtDNA replication, nucleoid formation, the oxidative phosphorylation pathway, and mitochondrial fission and fusion. Finally, we observed the upregulation of endoplasmic reticulum stress-induced genes. Cocaine self-administration influences the expression of both nuclear and mitochondrial genes as well as mtDNA replication. To determine whether these alterations serve as compensatory mechanisms to help maintain normal level of ATP production, further studies are necessary.
Subject(s)
Brain/metabolism , Cocaine/administration & dosage , DNA Copy Number Variations/physiology , Genes, Mitochondrial/physiology , Mitochondria/metabolism , Animals , Brain/drug effects , Conditioning, Operant/drug effects , Conditioning, Operant/physiology , DNA Copy Number Variations/drug effects , Genes, Mitochondrial/drug effects , Male , Mitochondria/drug effects , Rats , Rats, Wistar , Self AdministrationABSTRACT
The serotonergic system plays important roles in multiple functions of the nervous system and its malfunctioning leads to neurological and psychiatric disorders. Here, we show that the cell adhesion molecule close homolog of L1 (CHL1), which has been linked to mental disorders, binds to a peptide stretch in the third intracellular loop of the serotonin 2c (5-HT2c) receptor through its intracellular domain. Moreover, we provide evidence that CHL1 deficiency in mice leads to 5-HT2c-receptor-related reduction in locomotor activity and reactivity to novelty, and that CHL1 regulates signaling pathways triggered by constitutively active isoforms of the 5-HT2c receptor. Furthermore, we found that the 5-HT2c receptor and CHL1 colocalize in striatal and hippocampal GABAergic neurons, and that 5-HT2c receptor phosphorylation and its association with phosphatase and tensin homolog (PTEN) and ß-arrestin 2 is regulated by CHL1. Our results demonstrate that CHL1 regulates signal transduction pathways through constitutively active 5-HT2c receptor isoforms, thereby altering 5-HT2c receptor functions and implicating CHL1 as a new modulator of the serotonergic system.
