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Abstract: Type 1 inositol 1,4,5-trisphosphate receptors (IP3Rs-1), together with ryanodine receptors, are major calcium channels to regulate intracellular Ca2+ concentration. Although our recent report demonstrates the essential involvement of IP3R-1 up-regulation induced by dopamine D1-like and D2-like receptor (D1 and D2R) stimulation in psychological dependence, exact regulatory mechanisms of IP3R-1 expression by D2Rs have not yet been clarified. Mouse cerebral cortical neurons were treated with inhibitor of Ca2+-related signal transduction pathways coupling to D2Rs and used to analyze the mechanisms of IP3R-1 expression regulated by transcriptional factor. A selective D2R agonist, quinpirole, up-regulated IP3R-1 protein following its mRNA increase, which was significantly inhibited by gallein (a Gβγ modulator), U73122 (a phospholipase C inhibitor), BAPTA-AM (an intracellular calcium chelating reagent), W7 (a calmodulin inhibitor), KN-93 (a calmodulin-dependent protein kinases inhibitor), and FK506 (a calcineurin inhibitor). Immunocytochemical assessment showed that quinpirole increased expression of both cFos and phosphorylated-cJun in nucleus and enhanced translocation of NFATc4 complex to nucleus from cytoplasm. In addition, quinpirole directly recruited bindings between AP-1 and IP3R-1 promoter region and between NFATc4 and IP3R-1 promoter region. These results indicate that D2Rs enhance IP3R-1 gene transcription via increased bindings of AP-1 and NFATc4 to IP3R-1 promoter region after Gβγ activation. [Copyright &y& Elsevier]

Abstract: Little is known about regulatory mechanisms of type 1 inositol-1,4,5-triphosphate receptor (IP3R-1) expression in conditioned place preference by methamphetamine (METH), though significant enhancement of IP3R-1 expression in the mouse frontal cortex and limbic forebrain by intermittent administration of cocaine is reported. The present study investigated the role and regulation of IP3R-1 in mice with METH-induced place preference. Injection of IP3R antagonists with different chemical structures, 2-aminophenoxyethane-borate and xestospongin C, into the mouse nucleus accumbens (NAcc) dose-dependently inhibited METH-induced place preference. The levels of IP3R-1 protein in the NAcc of METH-conditioned mice significantly increased, which was completely abolished by microinjection of SCH23390 and raclopride, selective dopamine D1-like and D2-like receptor (D1 and D2DR) antagonists respectively, into the mouse NAcc. Immunohistochemical assessment revealed co-localization of immunoreactivity for IP3R-1 and those for D1 and D2DRs in the NAcc. These findings suggest that IP3R-1 could be involved in the development of METH-induced place preference and that D1 and D2DRs in the NAcc of mice showing METH-induced place preference play possible regulatory roles in IP3R-1 expression. [Copyright &y& Elsevier]

Abstract: Previous data demonstrate that L-type voltage-gated calcium channel (VGCC) blockers, which bind to α1 subunits of VGCC to suppress Ca2+ entry into cells, inhibit the development of psychological dependence on drugs of abuse, suggesting the upregulation of L-type VGCC in the development of psychological dependence. However, there are few available data on changes of the auxiliary subunit α2/δ modifying L-type VGCC under such conditions. We therefore investigated here the role of α2/δ subunits of VGCCs in the brain of mouse after repeated treatment with morphine. The treatment with morphine increased α2/δ subunit expression in the frontal cortex and the limbic forebrain of mice showing rewarding effect and sensitization to hyperlocomotion by morphine. The morphine-induced behavioral sensitization and place preference were also suppressed by gabapentin, which binds to an exofacial epitope of the α2/δ auxiliary subunits of VGCCs. These findings indicate that the upregulation of α2/δ subunit as well as α1 subunits of VGCC in the frontal cortex and the limbic forebrain plays a critical role in development of morphine-induced rewarding effect and behavioral sensitization following neuronal plasticity. [Copyright &y& Elsevier]

Abstract: Our previous investigation demonstrated that repeated administration of morphine significantly enhanced α 2 /δ-1 subunit expression in the frontal cortex and limbic forebrain of mice as well as morphine-induced place preference. However, little is known about regulatory mechanisms of α 2 /δ-1 subunit expression in conditioned place preference by methamphetamine (METH). In the present study, we investigated the role of α 2 /δ-1 subunit of voltage-gated calcium channels (VGCCs) in the mouse brain under repeated treatment with METH. The level of α 2 /δ-1 subunit increased significantly in the limbic forebrain including the nucleus accumbens and the frontal cortex of mice showing METH-induced sensitization. Under these conditions, the development of behavioral sensitization induced by the intermittent administration of METH was significantly suppressed by the co-administration of gabapentin (GBP) with binding activity to an exofacial epitope of α 2 /δ-1 subunit. Furthermore, GBP administered i.c.v. caused a dose-dependent inhibition of the METH-induced place preference. Chronic GBP treatment at the dose alleviating sensitization and place preference significantly reduced the elevation of α 2 /δ-1 subunit of VGCC induced by the repeated administration of METH in the limbic forebrain and frontal cortex, whereas there were no changes in the increase of α 2 /δ-1 subunit mRNA. These findings indicate that α 2 /δ-1 subunit plays a critical role in the development of METH-induced place preference following neuronal plasticity, and that GBP, which significantly suppressed METH-induced place preference by its possible inhibitory action of α 2 /δ subunit to neuronal membrane, may possibly be used as an alternative drug to treat or prevent drug dependence. [Copyright &y& Elsevier]

Abstract: During endochondral ossification, the production of hyaluronan (HA) is strictly and selectively regulated by chondrocytes, with a temporal peak at the hypertrophic stage. This study was conducted to clarify the effects of HA on expression and activity of runt-related gene 2 (Runx2), a potent transcription factor for chondrocyte differentiation in hypertrophic chondrocytes. Immature chondrocytes from an ATDC5 cell line were cultured and differentiated in DMEM/Ham''s F12 with pre-defined supplements. Using real-time PCR, the gene expressions of type II collagen, MMP-13, HAS2, and Runx2 in cultured chondrocytes were analysed from days 0 to 18 of cell differentiation. The activity and expression of Runx2 in hypertrophic chondrocytes were analysed after the treatment with HA oligosaccharide (HAoligo) using AML-3/Runx2 binding, real-time PCR and Western blot analysis. The effects of pre-incubation of anti-CD44 antibody on Runx2 expression were also examined. Expression of type X collagen and Runx2 mRNAs reached a maximum at the terminal differentiation of chondrocytes. The activity and expression of Runx2 was significantly inhibited in hypertrophic chondrocytes treated with HAoligo compared to the untreated controls. High molecular weight-HA did not affect the expression or activity of Runx2. The expression of Runx2 mRNA was significantly decreased in hypertrophic chondrocytes treated with anti-CD44 antibody. These results suggest that HAoligo may affect the terminal differentiation of chondrocytes during the endochondral ossification by inhibiting the expression and activity of Runx2. [Copyright &y& Elsevier]