Electrical stimulation of the medial forebrain bundle increases 32 P incorporation into striatal tyrosine hydroxylase (TH) at Ser 19 , Ser 31 , and Ser 40 . In the present studies, the effects of acute haloperidol and related drugs on sitespecific TH phosphorylation stoichiometry (PS) in the nigrostriatal and mesolimbic systems were determined by quantitative blot immunolabeling using phosphorylation statespecific antibodies. The striatum (Str), substantia nigra (SN), nucleus accumbens (NAc), and ventral tegmental area (VTA) from Sprague-Dawley rats were harvested 30 -40 min after a single injection of either vehicle, haloperidol (2 mg/kg), raclopride (2 mg/kg), clozapine (30 mg/kg), or SCH23390 (0.5 mg/kg). In vehicle-injected control rats, Ser 19 PS was 1.5-to 2.5-fold lower in Str and NAc than in SN and VTA, Ser 31 PS was two-to fourfold higher in Str and NAc than in SN and VTA, and Ser 40 PS was similar between the terminal field and cell body regions. After haloperidol, Ser 40 PS increased twofold in Str and NAc, whereas a smaller increase in SN and VTA was observed. The effects of haloperidol on Ser 19 PS were similar to those on Ser 40 in each region; however, haloperidol treatment increased Ser 31 PS at least 1.6-fold in all regions. The effects of raclopride on TH PS were comparable to those of haloperidol, whereas clozapine treatment increased TH PS at all sites in all regions. By contrast, the effects of SCH23390 on TH PS were relatively small and restricted to the NAc. The stoichiometries of site-specific TH phosphorylation in vivo are presented for the first time. The nigrostriatal and mesolimbic systems have common features of TH PS, distinguished by differences in TH PS between the terminal field and cell body regions and by dissimilar increases in TH PS in the terminal field and cell body regions after acute haloperidol.

Nanog is involved in controlling pluripotency and differentiation of stem cells in vitro. However, its function in vivo has been studied only in mouse embryos and various reports suggest that Nanog may not be required for the regulation of differentiation. To better understand endogenous Nanog function, more animal models should be introduced to complement the murine model. Here, we have identified the homolog of the mammalian Nanog gene in teleost fish and describe the endogenous expression of OlNanog mRNA and protein during medaka (Oryzias latipes) embryonic development and in the adult gonads. Using medaka fish as a vertebrate model to study Nanog function, we demonstrate that Ol-Nanog is necessary for Sphase transition and proliferation in the developing embryo. Moreover, inhibition or overexpression of OlNanog does not affect gene expression of various pluripotency and differentiation markers, suggesting that this transcription factor may not play a direct role in embryonic germ layer differentiation.

A detailed phylogenetic analysis of tetraspanins from 10 fully sequenced metazoan genomes and several fungal and protist genomes gives insight into their evolutionary origins and organization. Our analysis suggests that the superfamily can be divided into four large families. These four families-the CD family, CD63 family, uroplakin family, and RDS family-are further classified as consisting of several ortholog groups. The clustering of several ortholog groups together, such as the CD9/Tsp2/CD81 cluster, suggests functional relatedness of those ortholog groups. The fact that our studies are based on whole genome analysis enabled us to estimate not only the phylogenetic relationships among the tetraspanins, but also the first appearance in the tree of life of certain tetraspanin ortholog groups. Taken together, our data suggest that the tetraspanins are derived from a single (or a few) ancestral gene(s) through sequence divergence, rather than convergence, and that the majority of tetraspanins found in the human genome are vertebrate (21 instances), tetrapod (4 instances), or mammalian (6 instances) inventions.

In the CNS, the regulators of G-protein signaling (RGS) proteins belonging to the Rz subfamily, RGS19 (Ga interacting protein (GAIP)) and RGS20 (Z1), control the activity of opioid agonists at m but not at d receptors. Rz proteins show high selectivity in deactivating Gaz-GTP subunits. After reducing the expression of RGSZ1 with antisense oligodeoxynucleotides ( Knockdown of GAIP and of the GAIP interacting protein C-terminus (GIPC) led to changes in agonist effects at m but not at d receptors. The impairment of RGSZ1 extended the duration of morphine analgesia by at least 1 h beyond that observed in control animals. CTOP (Cys 2 , Tyr 3 , Orn 5 , Pen 7 -amide) antagonized morphine analgesia when given during the period in which the effect of morphine was enhanced by RGSZ1 knockdown. Thus, in naive mice, morphine tachyphylaxis originated in the presence of the opioid agonist and during the analgesia time course. The knockdown of RGSZ1 facilitated the development of tolerance to a single dose of morphine and accelerated tolerance to continuous delivery of the opioid. These results indicate that m but not d receptors are linked to Rz regulation. The m receptor-mediated activation of Gz proteins is effective at recruiting the adaptive mechanisms leading to the development of opioid desensitization.