stimulate mainly G␣ q/11 and G␣ i/o proteins (1-5). By contrast, G-protein coupling, signaling cascades, and biological effects of AT 2 receptors are less defined. Many in vivo studies with genetic deletion or transgenic overexpression of AT 2 receptors show that the AT 2 receptor inhibits AT 1 receptor-mediated functions (6 -8). The inhibitory effect of the AT 2 receptor has often been attributed to the AT 2 -mediated activation of protein phosphatases (9). However, in several systems AT 2 receptor-mediated signal inhibition persists in the presence of AT 2 -specific antagonists (10, 11) suggesting that the AT 2 receptor mediates additional functions independent of an agonist-AT 2 interaction. We previously demonstrated that AT 1 receptors can form stable heterodimers with another receptor, the bradykinin B 2 receptor (12). This receptor heterodimerization is agonist-independent and modifies AT 1 receptor-mediated responses (12, 13). Therefore we asked whether AT 1 receptors formed heterodimers with AT 2 receptors, and whether AT 1 /AT 2 receptor heterodimerization was responsible for agonist-independent effects of the AT 2 receptor causing inhibition of AT 1 receptor stimulation.We report here that the AT 2 receptor antagonizes AT 1 receptor-mediated responses by direct binding to the AT 1 receptor. Heterodimerization of the AT 1 with the AT 2 receptor was detected on transfected cells expressing AT 1 and AT 2 receptors, on fetal fibroblasts, and on myometrial biopsies from nonpregnant and pregnant women. The AT 2 -mediated antagonism of the AT 1 receptor was released upon decrease of AT 2 receptor expression, or when AT 1 /AT 2 receptor heterodimerization was inhibited. Thus, the AT 2 receptor antagonizes the activation of the AT 1 receptor by direct binding, and is therefore the first identified example of a G-protein-coupled receptor which acts as a receptor-specific antagonist. EXPERIMENTAL PROCEDURESCell Culture and Transfections-PC-12 cells and rat fetal fibroblasts were cultured and transfected as described (13,14).Construction of AT 2 Receptor Mutants-Site-directed mutagenesis of the rat AT 2 receptor-cDNA was performed by polymerase chain reaction similarly as described previously (14).Functional Assays-Inositol phosphate levels of PC-12 cells and of fetal fibroblasts was determined as described previously (14). The cellular content of cAMP was determined by radioimmunoassay (Immunotech) in the presence of 100 M isobutylmethylxanthine (14).Receptor-stimulated Redistribution of G-proteins-The activation of different G-proteins in cells and in tissues was determined by the agonist-stimulated release of G␣ proteins into the cytosol and subsequent detection of the cytosolic G␣ i/o , G␣ q/11 , or G␣ s proteins in immunoblot similarly as described (12, 13). For the detection of cytosolic G␣, proteins were separated on 7.5% SDS-containing polyacrylamide gels under nonreducing conditions. As a control, the amount of G␣ subunits in the total cell lysates was detected in immunoblot after separation of the prote...

No abstract

Parkinson's disease (PD) is a slowly progressive neurodegenerative movement disorder. Apoptosis, neuroinflammation, and oxidative stress are the current hypothesized mechanisms for PD pathogenesis. Tetramethylpyrazine (TMP), the major bioactive component of Ligusticum wallichii Franchat (ChuanXiong), Family Apiaceae, reportedly has anti-apoptotic, anti-inflammatory and antioxidant effects. This study investigated the role of 'TMP' in preventing rotenone-induced neurobiological and behavioral sequelae. A preliminary dose-response study was conducted where rats received TMP (10, 20, and 40 mg/kg, i.p.) concomitantly with rotenone (2 mg/kg, s.c.) for 4 weeks. Catalepsy, locomotor activity, striatal dopamine content, and tyrosine hydroxylase "TH" and α-synuclein immunoreactivity were evaluated. The selected TMP dose (20 mg/kg) was used for western blot analysis of Bax, Bcl2, and DJ-1, immunohistochemical detection of nuclear factor kappa B (NF-кB), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX2), and glial fibrillary acidic protein (GFAP) expression, in addition to biochemical analysis of caspase-3 activity, nuclear factor erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1) levels. Results showed that TMP (20 mg/kg) significantly improved midbrain and striatal TH expression and striatal dopamine content as well as the motor deficits, compared to rotenone-treated group. These results were correlated with reduction in caspase-3 activity and α-synuclein expression, along with improvement of midbrain and striatal Bax/Bcl2 ratio compared to rotenone-treated group. TMP also attenuated rotenone-induced upregulation of Nrf2/HO-1 pathway. Furthermore, TMP downregulated rotenone-induced neuroinflammation markers: NF-кB, iNOS, COX2, and GFAP expression in both the midbrain and striatum. Taken together, the current study suggests that TMP is entitled to, at least partially, preventing PD neurobiological and behavioral deficits by virtue of its anti-apoptotic, anti-inflammatory, and antioxidant actions.

No abstract

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT• Albendazole is used in several anthelminthic drug programs.• It may be necessary to include sectors of the community (such as breastfeeding women) not previously involved in community or clinical programmes (e.g. filariasis, hydatid disease) to achieve a reasonable level of elimination of the worm.• However, there are no studies in which the amount of the drug and/or its metabolites in the milk of lactating women was analysed, so as to assess the possible exposure of the breastfed infant. WHAT THIS STUDY ADDS• In the present study, the concentrations of albendazole and its active metabolite (albendazole sulphoxide) were analysed in the milk of lactating women through 36 h after administration of a single (400 mg) oral dose of albendazole.• Albendazole was barely secreted in milk as such.• On the other hand, albendazole sulphoxide was analysed through the 36 h of the study, and it was concluded that albendazole and albendazole sulphoxide attain levels in breast milk that are unlikely to be considered harmful for the breastfed infant.• These findings would help in deciding whether to involve breastfeeding mothers in albendazole mass drug administration programmes. AIMSAlbendazole (ABZ) is used in several anthelminthic drug programmws. ABZ side-effects are generally mild, but ABZ-induced pancytopenia may be serious. In filariasis programmes, it may be necessary to administer ABZ to breastfeeding women. Few data are available on safety of ABZ for breastfed infants. In addition, the pharmacokinetics of ABZ and its metabolites in human milk is insufficiently investigated. The aim was to study pharmacokinetics of ABZ and its metabolites [ABZ sulphoxide (ABSX) and ABZ sulphone] in the breast milk lactating women after one single oral dose of ABZ. METHODSThirty-three lactating women (age 18-40 years) participated in the study. They received a single oral 400-mg dose of ABZ. Five milk samples were taken at 0, 6, 12, 24 and 36 h. One serum sample was taken after 6 h. Samples were analysed using high-performance liquid chromatography and pharmacokinetic analysis was performed. RESULTSABZ was detectable in milk samples 6 h after the oral dose. The mean concentration of serum ABZ was 63.7 Ϯ 11.9 ng ml -1. The pharmacokinetic parameters for ABSX were calculated as follows: 351.9 Ϯ 32.4 ng ml for Cmax, Tmax, t1/2 and AUC0-36, respectively. The milk-to-serum ratios (range) for ABZ and ABSX were 0.9 (0.2-6.5) and 0.6 (0.1-1.5), respectively. CONCLUSIONSAfter an oral dose of 400 mg, ABZ and ABSX attain low concentrations in breast milk that are unlikely to be considered harmful for the breastfed infant.