The osmotic/calcium stress theory of brain damage: Are free radicals involved?

This overview presents data showing that glucose use increases and that excitatory amino acids (i.e., glutamate, aspartate), taurine and ascorbate increase in the extracellular fluid during seizures. During the cellular hyperactive state taurine appears to serve as an osmoregulator and ascorbate may serve as either an antioxidant or as a pro-oxidant. Finally, a unifying hypothesis is given for seizure-induced brain damage. This unifying hypothesis states that during seizures there is a release of excitatory amino acids which act on glutamatergic receptors, increasing neuronal activity and thereby increasing glucose use. This hyperactivity of cells causes an influx, of calcium (i.e. calcium stress) and water movements (i.e., osmotic stress) into the cells that culminate in brain damage mediated by reactive oxygen species.Special issue dedicated to Dr. Frederick E. Samson     

Sigma receptors in schizophrenic cerebral cortices

Antipsychotics represent high affinity for sigma receptors and sigma-like drugs often have the psychotomimetic properties. Besides, the receptors are unevenly distributed in human brain. These findings suggest that sigma receptors might be involved in the pathophysiology of schizophrenia. Sigma receptors in rat and human brain were measured with [³H]-1, 3, di-o-tolylguanidine (DTG) and non-specific binding of [³H]DTG was determined in the presence of 10^–5M haloperidol. Monovalent and divalent cations strongly inhibited [³H]DTG binding. Glutamate, aspartate and glycine also decreased the binding to human cerebral membranes. With post-mortem brain samples from 12 schizophrenics and 10 controls, sigma receptors were measured in 17 areas of cerebral cortex. Sigma receptors binding showed the regional differences in the cortex, but no significant differences between schizophrenics and controls were observed except the superior parietal cortex where the binding significantly increased in the schizophrenic group. These results suggest that sigma receptors in cerebral cortices might not be directly concerned with the pathophysiological role in schizophrenia.Dedicated to Dr. Morris Aprison. Received too late for publication in special issue.     

Structure of the Bombyx mori fibroin light-chain-encoding gene: upstream sequence elements common to the light and heavy chain.

Two overlapping genomic clones containing the fibroin light-chain (Fib-L)-encoding gene (Fib-L) were obtained from the cosmid library of the silkworm, Bombyx mori J-139, by hybridization with the Fib-L cDNA clone. Sequencing of the 14.6-kb region revealed that Fib-L was 13472 bp long containing seven exons, and that the gene contained a large first intron which occupied about 60% of the gene. Comparison of restriction patterns of the J-139 Fib-L with those of eight other B. mori breeds producing normal-level fibroin demonstrated that considerable restriction-fragment length polymorphisms were present in regions containing the first intron and the 3′-flanking sequence. However, sizes of the Fib-L mRNA and the Fib-L polypeptide were very similar among the nine breeds tested, suggesting that the exon sequences and the splice signals were all well conserved. 5′-Flanking regions of Fib-L and the fibroin heavy-chain (Fib-H)-encoding gene (Fib-H) compared in this study contained three 18-30-bp sequences of high similarity and many 8-10-bp common elements, six of which coincided with the binding sites of homeodomain proteins. Gel retardation assays with the nuclear extracts of the posterior and middle silk glands suggested that protein factors present in the posterior silk-gland nuclei could bind to a set of those common upstream elements.     

Three-dimensional structure of the quinoprotein methylamine dehydrogenase from Paracoccus denitrificans determined by molecular replacement at 2.8 A resolution.

The three-dimensional structure of the quinoprotein methylamine dehydrogenase from Paracoccus dentrificans (PD-MADH) has been determined at 2.8 A resolution by the molecular replacement method combined with map averaging procedures, using data collected from an area detector. The structure of methylamine dehydrogenase from Thio-bacillus versutus, which contains an "X-ray" sequence, was used as the starting search model. MADH consists of 2 heavy (H) and 2 light (L) subunits related by a molecular 2-fold axis. The H subunit is folded into seven four-stranded beta segments, forming a disk-shaped structure, arranged with pseudo-7-fold symmetry. A 31-residue elongated tail exists at the N-terminus of the H subunit in MADH from T. versutus but is partially digested in this crystal form of MADH from P. denitrificans, leaving the H subunit about 18 residues shorter. Each L subunit contains 127 residues arranged into 10 beta-strands connected by turns. The active site of the enzyme is located in the L subunit and is accessible via a hydrophobic channel between the H and L subunits. The redox cofactor of MADH, tryptophan tryptophylquinone is highly unusual. It is formed from two covalently linked tryptophan side chains at positions 57 and 107 of the L subunit, one of which contains an orthoquinone.     

The effect of the enantiomers of ibuprofen and flurbiprofen on the beta-oxidation of palmitate in the rat.

The effects of the enantiomers of ibuprofen (0.25 and 0.50 mmol/kg b.w.) and flurbiprofen (0.01, 0.03, and 0.06 mmol/kg b.w.) on the beta-oxidation of palmitate were investigated in the rat. The mean cumulative exhalation of 14CO2 after ip administration of [U-14C]palmitic acid was significantly reduced over 6 h by ibuprofen at the higher dose but not at the lower dose for either enantiomer. There was no difference between the enantiomers, the reduction over 6 h being 31.3 and 33.0% for (R)- and (S)-ibuprofen, respectively. There was also a significant inhibition of beta-oxidation by flurbiprofen at all 3 doses. Again, there was no stereoselectivity evident in this inhibition. Flurbiprofen was much more potent than ibuprofen in eliciting this effect, the 0.01mmol/kg dose giving a similar reduction in beta-oxidation as observed for the 0.50 mmol/kg dose of ibuprofen. The data support the hypothesis that inhibition of the in vivo beta-oxidation of palmitate by ibuprofen and flurbiprofen is primarily via a nonstereoselective noncoenzyme A-dependent mechanism.     

Reconstitution of the tonoplast amino-acid carrier into liposomes

The tonoplast amino-acid transporter of barley (Hordeum vulgare L.) mesophyll cells was functionally reconstituted by incorporating solubilized tonoplast membranes, vacuoplast membranes or tonoplast-enriched microsomal vesicles into phosphatidylcholine liposomes. (i) Time-, concentration- and ATP-dependence of amino-acid uptake were similar to results with isolated vacuoles. Although the orientation of incorporation could not be controlled, the results indicate that the transporter functions as a uniport system which allows regulated equilibration by diffusion between the cytosolic and vacuolar amino-acid pools. (ii) The ATP-modulated amino-acid carrier was also successfully reconstituted from barley epidermal protoplasts and Valerianella or Tulipa vacuoplasts, indicating its general occurrence. (iii) Fractionation of solubilized tonoplasts by size-exclusion chromatography followed by reconstitution of the fractions for glutamine transport gave two activity peaks: the first eluted in the region of high-molecular-mass vesicles and the second at a size of 300 kDa for the Triton-protein micelle.Abbreviation SDS-PAGE sodium dodecyl sulfate-polyacryl-amide gel electrophoresis This work was part of our research efforts within the Sonderforschungsbereich 176 of the University. We gratefully acknowledge experimental support by Marion Betz and valuable discussions with Professors U. Heber and U.-I. Flügge and Dr. Armin Gross (University of Würzburg) and Dr. E. Martinoia (ETH, Zürich, Switzerland).     

Microalgae aquaculture feeds

Microalgae feeds are currently used in relatively small amounts in aquaculture, mainly for the production of larvae and juvenile shell- and finfish, as well as for raising the zooplankton required for feeding of juvenile animals. The blue-green algaSpirulina is used in substantial amounts (over 100 t y^–1) as a fish and shrimp feed, and even larger markets can be projected if production costs could be reduced. Another potential large-scale application of microalgae is the cultivation ofHaematococcus for the production of the carotenoid astaxanthin, which gives salmon flesh its reddish color. In the long-term microalgae biomass high in lipids (omega-3 fatty acids) may be developed as substitutes for fish oil-based aquaculture feeds. In shrimp ponds the indigenous algal blooms supply a part of the dietary requirements of the animals, but it is difficult to maximize algal productivities. A separate algal production system could feed the shrimps and minimize the need for added feed. Bivalves feed essentially exclusively on marine microalgae throughout their life cycle. The development of cultivation technologies for such microalgae would allow the onshore production of these animals, with greatly improved product quality and safety.This paper was presented at the Symposium on Applied Phycology at the Fourth International Phycological Congress, Duke University.     

Interactions between central glutamatergic,catecholaminergic and cholinergic systems with regard to locomotor activity in monoamine-depleted mice

Summary Following injection of 5µg of the competitive NMDA receptor antagonist AP-5 into the nucleus accumbens, but not following injection of the same dose into the dorsal striatum, a pronounced locomotor stimulation in monoamine-depleted mice was produced; the-adrenoceptor agonist clonidine (1 mg/kg) administered ip caused a marked potentiation of an intraaccumbens AP-5 (2.5µg) injection.On the other hand, 10µg of AP-5 combined with an ip injection of clonidine (1 mg/kg) caused a marked locomotor stimulation following local application into the dorsal striatum but not following application into the prefrontal cortex. Likewise, in combination with systemically administered clonidine, a substantial locomotor stimulation was observed after application of the muscarine receptor antagonist methscopolamine (62µg) into the dorsal striatum but not into the prefrontal cortex.This study suggests that NMDA receptors in the nucleus accumbens exert an inhibitory influence on locomotor activity. The dorsal striatum may also be involved in such control via NMDA and muscarinic receptors.