The effects of zolpidem treatment on GABA(A) receptors in cultured cerebellar granule cells: changes in functional coupling

AimsHypnotic zolpidem is a positive allosteric modulator of γ-aminobutyric acid (GABA) action, with preferential although not exclusive binding for α1 subunit-containing GABA_A receptors. The pharmacological profile of this drug is different from that of classical benzodiazepines, although it acts through benzodiazepine binding sites at GABA_A receptors. The aim of this study was to further explore the molecular mechanisms of GABA_A receptor induction by zolpidem.Main methodsIn the present study, we explored the effects of two-day zolpidem (10 μM) treatment on GABA_A receptors on the membranes of rat cerebellar granule cells (CGCs) using [³H]flunitrazepam binding and semi-quantitative PCR analysis.Key findingsTwo-day zolpidem treatment of CGCs did not significantly affect the maximum number (B_max) of [³H]flunitrazepam binding sites or the expression of α1 subunit mRNA. However, as shown by decreased GABA [³H]flunitrazepam binding, two-day exposure of CGCs to zolpidem caused functional uncoupling of GABA and benzodiazepine binding sites at GABA_A receptor complexes.SignificanceIf functional uncoupling of GABA and benzodiazepine binding sites at GABA_A receptors is the mechanism responsible for the development of tolerance following long-term administration of classical benzodiazepines, chronic zolpidem treatment may induce tolerance.     

A new plasmid (QpDV) common to Coxiella burnetii isolates associated with acute and chronic Q fever

Abstract Genetic studies of Coxiella burnetii strains suggested the possibility of differentiating new isolates according to their plasmid DNA content. Virulence and/or clinical manifestations ('chronic' and 'acute' Q fever) had been claimed to correlate with this plasmid typing. A new plasmid, named QpDV, was found to be common to C. burnetii isolates obtained from acute and chronic Q fever. According to the results obtained, plasmid usage for detection and differentiation of respective pathovars of C. burnetii and the correlation between gene specificity and pathovar has to be revised. Closer studies suggested a common origin of C. burnetii plasmids, but also showed some differences characteristic for each plasmid, probably reflecting divergent evolution.     

Non-functional Fas ligand increases the formation of cartilage early in the endochondral bone induction by rhBMP-2

It has previously been shown that mice with a defect in Fas ligand-mediated apoptosis have an enhancement of ectopic bone formation. We investigated the expression of bone-related markers--alkaline phosphatase, collagen, bone sialoprotein, osteocalcin, osteopontin, and bone morphogenetic proteins (BMP) -2, -4, and -7; and cytokines interleukin-1alpha (IL-1), IL-1beta, and tumor necrosis factor-alpha (TNF-alpha) in ectopic new bone induced by recombinant human (rh) BMP-2 in mice without functional Fas-ligand (gld mice). At day 6 after rhBMP-2 implantation, gld mice formed more cartilage and mesenchyme compared with their wild type littermates. At later stages, gld mice did not differ from the control mice in the volume of newly formed tissue, expressing higher level of BMP genes and lower levels of genes involved in osteoblast maturation--bone sialoprotein and osteopontin. Differences in the levels of expression of IL-1alpha and TNF-alpha were observed only at day 12 after rhBMP-2 implantation. These results suggest that gld mice have an increased recruitment of cells of mesenchymal origin and an abnormal pattern of differentiation and maturation of the newly formed mesenchymal tissues.     

Presence of Phospholipid-N-Methyltransferases and Base-Exchange Enzymes in Rat Central Nervous System Axolemma-Enriched Fractions

Rat CNS myelinated axons were fractionated by sucrose density gradient centrifugation with a zonal rotor. Fraction VI, obtained at 28-30% sucrose, appeared, on the basis of the presence of related marker enzymes, to be enriched in axolemma. Phospholipid-N-methyltransferases (PMTs) and base-exchange enzymes were associated with fraction VI. PMT activity was significantly stimulated by the addition of either phosphatidylmonomethylethanolamine or phosphatidyldimethylethanolamine but the PMT activity of the homogenate or the myelinated axons was unresponsive. Recoveries of the ethanolamine, serine, and choline base-exchange activities were 14.4%, 13.8%, and 3.4%, respectively, of that present in the myelinated axons. The myelin-rich fraction obtained simultaneously seems contaminated with other membrane fractions.     

Toxocariasis associated with chronic cough in childhood: a longitudinal study in Hungary

Chronic cough lasting 8 weeks or more often seems to be an intractable problem in childhood. Toxocara infection is associated with an increased prevalence of airway symptoms and may be the possible aetiological agent of chronic cough. Of 425 children aged 2-17 years with chronic cough who were investigated for toxocariasis and the distribution of bronchial asthma (BA), cough variant asthma (CVA) and non-asthmatic eosinophilic bronchitis (NAEB), 136 (32%) were seropositive for Toxocara canis antigens. Ninety-three of the 136 were adequately assessed, diagnosed and followed up during 1 year. BA was diagnosed in 40%, CVA in 27% and NAEB in 33% of the children. The eosinophil cell count, serum T. canis IgG levels and symptoms are predictors of the improvement or the decline of the condition. Presuming the aetiopathogenetic role of T. canis in the inflammatory process of chronic cough, we treated the children not only with inhaled corticosteroid (ICS), but also with a 1-week course of anthelminthics. We could significantly decrease the dose of ICS in 23 (62%) of the 37 with BA. The administration of anthelminthics and the avoidance of sensitizers were sufficient for those with NAEB; none needed ICS. ICS therapy could be stopped 2-3 months later in 17 (68%) of the 25 with CVA. We found that 8 of the 25 with CVA (32%) presented asthmatic symptoms at the end of the 1-year period. In Hungary, T. canis may be a potential sensitizer for chronic cough in seropositive children. Deworming therapy will then alleviate the airway symptoms without exacerbation in patients with BA, and have a positive effect on those with NAEB and the majority of those with CVA.     

Depletion of CD4 and CD8 T lymphocytes in mice in vivo enhances 1,25-dihydroxyvitamin D3-stimulated osteoclast-like cell formation in vitro by a mechanism that is dependent on prostaglandin synthesis

To investigate the role of T lymphocytes in osteoclastogenesis, we performed in vivo depletion of CD4 and/or CD8 T lymphocyte subsets and evaluated in vitro osteoclast-like cell (OCL) formation. T lymphocyte depletion (TLD) with mAbs was confirmed 24 h later by flow cytometry. OCL formation was stimulated with 1, 25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)) in bone marrow and with recombinant mouse (rm) receptor activator of NF-kappaB ligand (RANK-L) and rmM-CSF in bone marrow and spleen cell cultures. OCL formation was up to 2-fold greater in 1,25-(OH)(2)D(3)-stimulated bone marrow cultures from TLD mice than in those from intact mice. In contrast, TLD did not alter OCL formation in bone marrow or spleen cell cultures that were stimulated with rmRANK-L and rmM-CSF. The effects of TLD seemed to be mediated by enhanced PG synthesis, because the PGE(2) concentration in the medium of 1, 25-(OH)(2)D(3)-stimulated bone marrow cultures from TLD mice was 5-fold higher than that in cultures from intact mice, and indomethacin treatment abolished the stimulatory effect of TLD on OCL formation. There was a 2-fold increase in RANK-L expression and an almost complete suppression of osteoprotegerin expression in 1, 25-(OH)(2)D(3)-stimulated bone marrow cultures from TLD mice compared with those from intact mice. Although there was a small (20%) increase in IL-1alpha expression in 1, 25-(OH)(2)D(3)-stimulated bone marrow cultures from TLD mice, TLD in mice lacking type I IL-1R and wild-type mice produced similar effects on OCL formation. Our data demonstrate that TLD up-regulates OCL formation in vitro by increasing PG production, which, in turn, produces reciprocal changes in RANK-L and osteoprotegerin expression. These results suggest that T lymphocytes influence osteoclastogenesis by altering bone marrow stromal cell function.     

High-Resolution Linkage Analyses to Identify Genes That Influence Varroa Sensitive Hygiene Behavior in Honey Bees

Varroa mites (V. destructor) are a major threat to honey bees (Apis melilfera) and beekeeping worldwide and likely lead to colony decline if colonies are not treated. Most treatments involve chemical control of the mites; however, Varroa has evolved resistance to many of these miticides, leaving beekeepers with a limited number of alternatives. A non-chemical control method is highly desirable for numerous reasons including lack of chemical residues and decreased likelihood of resistance. Varroa sensitive hygiene behavior is one of two behaviors identified that are most important for controlling the growth of Varroa populations in bee hives. To identify genes influencing this trait, a study was conducted to map quantitative trait loci (QTL). Individual workers of a backcross family were observed and evaluated for their VSH behavior in a mite-infested observation hive. Bees that uncapped or removed pupae were identified. The genotypes for 1,340 informative single nucleotide polymorphisms were used to construct a high-resolution genetic map and interval mapping was used to analyze the association of the genotypes with the performance of Varroa sensitive hygiene. We identified one major QTL on chromosome 9 (LOD score = 3.21) and a suggestive QTL on chromosome 1 (LOD = 1.95). The QTL confidence interval on chromosome 9 contains the gene ‘no receptor potential A’ and a dopamine receptor. ‘No receptor potential A’ is involved in vision and olfaction in Drosophila, and dopamine signaling has been previously shown to be required for aversive olfactory learning in honey bees, which is probably necessary for identifying mites within brood cells. Further studies on these candidate genes may allow for breeding bees with this trait using marker-assisted selection.