Glutamate receptors modulate sodium-dependent and calcium-independent vitamin C bidirectional transport in cultured avian retinal cells

Vitamin C is transported in the brain by sodium vitamin C co‐transporter 2 (SVCT‐2) for ascorbate and glucose transporters for dehydroascorbate. Here we have studied the expression of SVCT‐2 and the uptake and release of [¹⁴C] ascorbate in chick retinal cells. SVCT‐2 immunoreactivity was detected in rat and chick retina, specially in amacrine cells and in cells in the ganglion cell layer. Accordingly, SVCT‐2 was expressed in cultured retinal neurons, but not in glial cells. [¹⁴C] ascorbate uptake was saturable and inhibited by sulfinpyrazone or sodium‐free medium, but not by treatments that inhibit dehydroascorbate transport. Glutamate‐stimulated vitamin C release was not inhibited by the glutamate transport inhibitor l ‐β‐threo‐benzylaspartate, indicating that vitamin C release was not mediated by glutamate uptake. Also, ascorbate had no effect on [³H] d ‐aspartate release, ruling out a glutamate/ascorbate exchange mechanism. 2‐Carboxy‐3‐carboxymethyl‐4‐isopropenylpyrrolidine (Kainate) or NMDA stimulated the release, effects blocked by their respective antagonists 6,7‐initroquinoxaline‐2,3‐dione (DNQX) or (5R,2S)‐(1)‐5‐methyl‐10,11‐dihydro‐5H‐dibenzo[a,d]cyclohepten‐5,10‐imine hydrogen maleate (MK‐801). However, DNQX, but not MK‐801 or 2‐amino‐5‐phosphonopentanoic acid (APV), blocked the stimulation by glutamate. Interestingly, DNQX prevented the stimulation by NMDA, suggesting that the effect of NMDA was mediated by glutamate release and stimulation of non‐NMDA receptors. The effect of glutamate was neither dependent on external calcium nor inhibited by 1,2‐bis (2‐aminophenoxy) ethane‐N′,N′,N′,N′,‐tetraacetic acid tetrakis (acetoxy‐methyl ester) (BAPTA‐AM), an internal calcium chelator, but was inhibited by sulfinpyrazone or by the absence of sodium. In conclusion, retinal cells take up and release vitamin C, probably through SVCT‐2, and the release can be stimulated by NMDA or non‐NMDA glutamate receptors.     

The atomic structure of rice dwarf virus reveals the self-assembly mechanism of component proteins

Rice dwarf virus (RDV), the causal agent of rice dwarf disease, is a member of the genus Phytoreovirus in the family Reoviridae. RDV is a double-shelled virus with a molecular mass of approximately 70 million Dalton. This virus is widely prevalent and is one of the viruses that cause the most economic damage in many Asian countries. The atomic structure of RDV was determined at 3.5 A resolution by X-ray crystallography. The double-shelled structure consists of two different proteins, the core protein P3 and the outer shell protein P8. The atomic structure shows structural and electrostatic complementarities between both homologous (P3-P3 and P8-P8) and heterologous (P3-P8) interactions, as well as overall conformational changes found in P3-P3 dimer caused by the insertion of amino-terminal loop regions of one of the P3 protein into the other. These interactions suggest how the 900 protein components are built into a higher-ordered virus core structure.     

Contributions of cytoplasmic free and membrane-bound ribosomes to the synthesis of NADPH-adrenodoxin reductase and adrenodoxin of bovine adrenal cortex mitochondria

Cytoplasmic free and membrane-bound ribosomes were isolated from bovine adrenal cortex, and characterized. Contributions of free and bound ribosomes to the synthesis of NADPH-adrenodoxin reductase (AdR) and adrenodoxin (Ad) were determined by examining the presence of their nascent peptides on isolated ribosomes. Nascent peptides were released from the ribosomes by [3H]puromycin in a high salt buffer in the presence of a detergent, and the nascent peptides of AdR and Ad were separately isolated by immunoprecipitation using antibodies. AdR nascent peptides were associated with free and loosely-bound ribosomes, whereas Ad nascent peptides were associated with free, loosely-bound and tightly-bound ribosomes. Smaller nascent peptides of AdR were carried by free ribosomes, whereas larger nascent peptides were preferentially carried by loosely-bound ribosomes. In the case of Ad, smaller nascent peptides were more abundant in free ribosomes than in bound ribosomes. The nascent peptides of Ad were released from bound ribosomes of rough microsomes to the aqueous milieu by puromycin treatment, suggesting the release of completed Ad peptides into the cytoplasm in cells.     

Whole-body vibration mediates mechanical hypersensitivity through Aβ-fiber and C-fiber thermal sensation in a chronic pain model

Whole-body vibration (WBV), which is widely used as a type of exercise, involves the use of vibratory stimuli and it is used for rehabilitation and sports performance programmes. This study aimed to investigate the effect of WBV treatment in a chronic pain model after 10 WBV sessions. An animal model (chronic pain) was applied in 60 male Wistar rats (±180 g, 12 weeks old) and the animals were treated with low intensity exercise (treadmill), WBV (vibrating platform), and a combined treatment involving both. The controls on the platform were set to a frequency of 42 Hz with 2 mm peak-to-peak displacement, g ≈ 7, in a spiral mode. Before and after the vibration exposure, sensitivity was determined. Aβ-fibers-mediated mechanical sensitivity thresholds (touch-pressure) were measured using a pressure meter. C-fibers-mediated thermal perception thresholds (hot pain) were measured with a hot plate. After each session, WBV influenced the discharge of skin touch-pressure receptors, reducing mechanical sensitivity in the WBV groups (P < 0.05). Comparing the conditions “before vs. after”, thermal perception thresholds (hot pain) started to decrease significantly after the third WBV session (P < 0.05). WBV decreases mechanical hyperalgesia after all sessions and thermal sensitivity after the third session with the use of WBV.     

Excretory/secretory products from plerocercoids of Spirometra erinaceieuropaei suppress interleukin-1beta gene expression in murine macrophages

The present study shows that ES products from plerocercoids of Spirometra erinaceieuropaei suppressed interleukin-1beta mRNA expression in lipopolysaccharide-stimulated RAW 264.7 macrophages in the absence or presence of a cyclic AMP analogue, dibutyryl cyclic AMP. Investigation using the inhibitors of mitogen-activated protein kinase (MAPK) pathways revealed that extracellular signal-regulated protein kinase 1/2 and p38 mitogen-activated protein kinase pathways are crucial for full induction of interleukin-1beta mRNA expression. ES products additionally suppressed interleukin-1beta mRNA expression in the cells treated with p38 mitogen-activated protein kinase inhibitor (SB203580) or extracellular signal-regulated protein kinase 1/2 inhibitor (PD98059). Western blot analysis showed that dibutyryl cyclic AMP enhanced lipopolysaccharide-induced phosphorylation of extracellular signal-regulated protein kinase 1/2, p38 mitogen-activated protein kinase and cyclic AMP responsive element binding protein (CREB) and, in turn, we demonstrated that ES products reduced the lipopolysaccharide and dibutyryl cyclic AMP-induced phosphorylation of extracellular signal-regulated protein kinase 1/2 and p38 mitogen-activated protein kinase, but not cyclic AMP responsive element binding protein. These data demonstrate that ES products from the plerocercoids of S. erinaceieuropaei may evade induction of interleukin-1beta mRNA by inhibiting extracellular signal-regulated protein kinase 1/2 and p38 mitogen-activated protein kinase pathways in lipopolysaccharide and/or dibutyryl cyclic AMP-stimulated macrophages.     

DNA methylation and histone modification in onion chromosomes

Onion, Allium cepa, is a model plant for experimental observation of somatic cell division, whose mitotic chromosome is extremely large, and contains the characteristic terminal heterochromatin. Epigenetic status of the onion chromosome is a matter of deep interest from a molecular cytogenetic point of view, because epigenetic marks regulate chromatin structure and gene expression. Here we examined chromosomal distribution of DNA methylation and histone modification in A. cepa in order to reveal the chromatin structure in detail. Immunodetection of 5-methylcytosine (5mC) and in situ nick-translation analysis showed that onion genomic DNA was highly methylated, and the methylated CG dinucleotides were distributed in entire chromosomes. In addition, distributions of histone methylation codes, which occur in close association with DNA methylation, were similar to those of other large genome species. From these results, a highly heterochromatic and less euchromatic state of large onion chromosomes were demonstrated at an epigenetic level.     

Synthesis of UV-curable chitosan derivatives and palladium (II) adsorption behavior on their UV-exposed films

Novel chitosan derivatives with UV-curable functional groups, such as 3-methoxy-4-(2-hydroxy-3-methacryloyloxypropoxy)benzyl, 3,4-bis(2-hydroxy-3-methacryloyloxypropoxy)benzyl, 3-methoxy-4-methacryloyloxybenzyl, and 3,5-dimethacryloyloxybenzyl groups, were prepared. Introduction of photosensitive functional groups to chitosan was accomplished by reductive N-alkylation via Schiff’s bases using corresponding photosensitive aldehydes. Compared to starting chitosan, UV-curable chitosan derivatives showed better solubility in several organic solvents, such as DMSO and 70% methacrylic acid. The solubility of these compounds increased with an increase in the degree of substitution of the N-alkyl side chains. After UV irradiation for 20 s under a high-pressure mercury lamp at a distance of 15 cm from the samples, acidic methanol solutions of these derivatives were transformed to gels in the presence of photo-initiator, and their dried films adsorbed palladium (II) at pH 1.1 and pH 5.3. The UV-curable chitosan derivatives were successfully used as coating materials for electroless plating on non-conductive substances.