ABSTRACT
Previous data showed that cell surface expression of the glutamate transporters GLT1a and excitatory amino acid carrier 1 (EAAC1), localized in glia and neurons of the CNS, can be regulated by protein kinase C (PKC). Regulation and physiological importance of GLT1b, a splice variant of GLT1a, is not understood. In the present study we used cultured cerebellar granule cells (CGCs) from mice to investigate PKC dependent trafficking of GLT1b in comparison to GLT1a and EAAC1 using immunohistochemistry and subcellular fractionation followed by Western blotting. In neurites of CGCs, GLT1b and EAAC1 were localized to different aggregates of vesicles that were different from vesicle aggregates containing vesicular glutamate transporters. In CGCs cultured with low-potassium medium, stimulation of PKC by phorbol ester enhanced the formation of large varicosities in neurites that exhibited immunoreactivity for GLT1a, GLT1b, and EAAC1. Stimulation of PKC leads to a significant increase of GLT1b and EAAC1 in the plasma membrane whereas GLT1a in the plasma membrane was decreased. Following PKC stimulation, also a significant increase of transporter-mediated glutamate uptake representing sodium dependent glutamate uptake, was observed. Similarly, the fraction of glutamate uptake, that was sensitive to the inhibitor WAY-213613 and represents uptake by GLT1a and GLT1b, was increased after stimulation by PKC. The findings suggest that PKC is similarly involved in regulation of surface trafficking of GLT1b and EAAC1 and that PKC stimulated increase in surface location of GLT1b and EAAC1 in glutamatergic CGCs.
Subject(s)
Cerebellum/metabolism , Excitatory Amino Acid Transporter 2/metabolism , Excitatory Amino Acid Transporter 3/metabolism , Neurons/metabolism , Protein Kinase C/metabolism , Animals , Cell Membrane/drug effects , Cell Membrane/metabolism , Cells, Cultured , Cerebellum/drug effects , Cerebellum/enzymology , Enzyme Activators/administration & dosage , Excitatory Amino Acid Transporter 2/antagonists & inhibitors , Glutamic Acid/metabolism , Mice , Mice, Inbred C57BL , Neurons/drug effects , Neurons/enzymology , Phorbol Esters/administration & dosage , Potassium/metabolism , Protein Transport , Subcellular Fractions/metabolism , Synaptic Vesicles/metabolismABSTRACT
We describe an unusual clinical strain of catalase-negative methicillin-resistant Staphylococcus aureus sensu stricto. Sequence analysis of its catalase gene showed 99.60% identities to the catalase genes of the reference strains. A 5-base deletion, however, led to a shift of the nucleotide reading frame and a loss of the enzymatic activity.
Subject(s)
Catalase/genetics , Catalase/metabolism , Methicillin Resistance , Staphylococcal Infections/microbiology , Staphylococcus aureus/drug effects , Staphylococcus aureus/enzymology , Aged , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Humans , Male , Molecular Sequence Data , Sequence Analysis, DNA , Staphylococcus aureus/isolation & purificationABSTRACT
The biological evaluation of hypericin in various test models is hampered by its poor water solubility. In former studies we have shown that the water solubility of hypericin was remarkably enhanced in the presence of the procyanidins or flavonol glycosides of Hypericum extract. The present pharmacokinetic study was designed to find out whether the improved water solubility in the presence of procyanidin B2 or hyperoside is correlated to increased plasma levels of hypericin. Plasma levels of hypericin in rats in the presence and absence of procyanidin B2 or hyperoside were determined by reversed phase HPLC using fluorimetric detection. Both compounds increased the oral bioavailability of hypericin by ca. 58 % (B2) and 34 % (hyperoside). Procyanidin B2 and hyperoside had a different influence on the plasma kinetics of hypericin; median maximal plasma levels of hypericin were detected after 360 min (C max : 8.6 ng/mL) for B2, and after 150 min (C max : 8.8 ng/mL) for hyperoside. It can be speculated that, when administered together with these compounds, a significant accumulation of hypericin in rat plasma in the presence of both polyphenols might be responsible for the observed increased in vivo activity.
Subject(s)
Antidepressive Agents/pharmacokinetics , Biflavonoids , Catechin/pharmacology , Perylene/analogs & derivatives , Perylene/pharmacokinetics , Phytotherapy , Proanthocyanidins , Quercetin/analogs & derivatives , Quercetin/pharmacology , Administration, Oral , Animals , Anthracenes , Antidepressive Agents/administration & dosage , Antidepressive Agents/blood , Biological Availability , Chromatography, High Pressure Liquid , Drug Synergism , Male , Perylene/administration & dosage , Perylene/blood , Plant Extracts/administration & dosage , Plant Extracts/blood , Plant Extracts/pharmacokinetics , Rats , Rats, Sprague-Dawley , SolubilityABSTRACT
A commercially available extract of the aerial parts of Hypericum perforatum, LI 160, showed pronounced activity in selected animal bioassays. These include the forced swimming test (FST) and the tail suspension test, used to determine antidepressant activity, and tests indicating activity on the central nervous system, such as body temperature and ketamine induced sleeping time. The counteracting effects of drugs known to interfere with the central dopaminergic system strongly suggested that dopamine mediated activity is important for the activity of the extract. Dose-response experiments of the total extract and of fractions rich in flavonoids and napthodianthrones produced inverted U-shaped dose response curves.
