Perinatal Hypoxia-Ischemia Disrupts Striatal High-Affinity [3H]Glutamate Uptake into Synaptosomes

We examined the impact of hypoxia-ischemia on high-affinity [3H]glutamate uptake into a synaptosomal fraction prepared from immature rat corpus striatum. In 7-day-old pups the right carotid artery was ligated, and pups were exposed to 8% oxygen for 0, 0.5, 1, or 2.5 h, and allowed to recover for up to 24 h before they were killed. High-affinity glutamate uptakes in striatal synaptosomes derived from tissue ipsilateral and contralateral to ligation were compared. After 1 h of hypoxia plus ischemia, high-affinity glutamate uptake in the striatum was reduced by 54 +/- 13% compared with values from the opposite (nonischemic) side of the brain (p less than 0.01, t test versus ligates not exposed to hypoxia). There were similar declines after 2.5 h of hypoxia-ischemia. Activity remained low after a 1 h recovery period in room air, but after 24 h of recovery, high-affinity glutamate uptake was equal bilaterally. Kinetic analysis revealed that loss of activity could be attributed primarily to a 40% reduction in the number of uptake sites. Hypoxia alone had no effect on high-affinity glutamate uptake although it reduced synaptosomal uptake of [3H]3,4-dihydroxyphenylethylamine. Addition of 1 mg/ml of bovine serum albumin to the incubation medium preferentially stimulated high-affinity glutamate uptake in hypoxic-ischemic brain compared with its effects in normal tissue. These studies demonstrate that hypoxia-ischemia reversibly inhibits high-affinity glutamate uptake and this occurs earlier than the time required to produce neuronal damage in the model.     

The effect of catecholamines on ion transport in the toad bladder

Noradrenalin (8 · 10⁻⁶ M) and adrenalin (6 · 10⁻⁶ and 6 · 10⁻⁷ M) were found to cause marked stimulation of short-circuit current (S.C.C.) in isolated toad bladder, but isoprenalin (8 · 10⁻⁷ M) was found to be without effect. The percentage rise in S.C.C. due to noradrenalin was found to be inversely proportional to the initial S.C.C. or total conductance of the bladder. Again in the case of noradrenalin the rise in S.C.C. was almost completely abolished by α-adrenergic blockade but not by β-blockade. This rise in S.C.C. was found not to be significantly different from the rise in net Na⁺ flux. Bidirectional Cl⁻ fluxes were estimated using ⁸²Br as a companion radionuclide to ³⁶Cl. No significant net Cl⁻ flux was apparent, either before or after addition of any of the three catecholamines tested. However, in some cases the unidirectional Cl⁻ fluxes rose markedly following addition of noradrenalin or of adrenalin and this change was not reflected in a change in total conductance. This anomaly was noted to occur in bladders whose initial conductance was of the order of 0.5 kΩ⁻¹ · cm⁻² or greater. The evidence presented suggests that two actions of catecholamines on ion transport in toad bladder are (a) to increase Na⁺ transport via stimulation of α-adrenergic sites and (b) at the concentrations tested to cause an increase in passive Cl permeability in bladders whose initial conductance is high.     

Physical and genetic organization of Petite and Grande yeast mitochondrial DNA. II. DNA-DNA hybridization studies and buoyant density determinations

Buoyant density of mitochondrial DNA from 14 cytoplasmic petite mutants issued from the same grande yeast Saccharomyces cerevisiae was determined. Mutants that have retained the mitochondrial gene conferring resistance to erythromycin displayed higher buoyant density, while mutants that have retained the mitochondrial gene conferring resistance to chloramphenicol displayed lower buoyant density. It is inferred that the segment which carries the E^R gene has a higher G + C content than the segment which carries the C^R gene. DNA-DNA filter hybridizations were carried out systematically in different reciprocal pair-wise combinations between mtDNAs purified from various mutants and from the grande. All petites were found to be deleted in 42 to 93% of the grande sequence, depending on the mutant studied. Sequence homology between petite mtDNAs was greatest in mutants retaining common genetic markers and was least when different genetic markers were retained. Practically no hybridization was found between some C^RE^O and C^OE^R mutants. Correlations established between the extent of DNA-DNA hybridization, kinetic and genetic complexity show that a selective enrichment of gene specific sequences occurs in mtDNA of petites.     

Physical and genetic organization of Petite and Grande yeast mitochondrial DNA. III. High resolution melting and reassociation studies

Mitochondrial DNAs from 13 petite mutants have been analyzed by means of high-resolution melting and reassociation in solution, in an attempt to relate their physical chemical properties to the mitochondrial genotype, which displays various combinations of genetic marker deletions. The kinetic complexities of the petite mtDNAs were found to range from $\text{1}\text{3}$ down to $\text{1}\text{500}$ of that of the grande mtDNA; the loss in sequential complexity undergone by petite mtDNAs parallels the loss in mitochondrial genetic complexity. Melting profiles of petite mtDNAs can be resolved into well-defined peaks. Some of them are specific to the marker genes. The genotypic specificity increases as the sequential complexity of mtDNA decreases. The mtDNA region conferring resistance to erythromycin could in this way be shown to be characterized by two melting peaks, at 72 °C and 75 °C. The results are interpreted in terms of selective enrichment in gene-specific sequences.     

Analysis of a yeast nuclear gene involved in the maturation of mitochondrial pre-messenger RNA of the cytochrome oxidase subunit I

We have analyzed the mitochondrial RNA of a yeast nuclear pet mutant with no cytochrome oxidase activity. The product of the gene affected in this mutant appears to be necessary for the correct maturation of the mitochondrial pre-mRNA of the cytochrome oxidase subunit I. It does not affect, however, the overall splicing of cytochrome b pre-mRNA or the intron excision of the 21S ribosomal RNA precursor. This gene has been isolated by genetic complementation in yeast, and its DNA sequence has been determined. It is transcribed, as detected by S1 mapping experiments, and could encode a protein of 436 amino acids.     

The HLA system and the analysis of multifactorial genetic disease

The human leukocyte antigen (HLA) system comprises closely linked genes controlling highly polymorphic proteins involved in the presentation of peptides to the T-cell receptor. Specific alleles at HLA loci are associated with diseases, often those suspected to be of autoimmune aetiology. Many of these associations result from linkage disequilibrium between the HLA gene studied and other HLA genes or non-HLA gebes close by. Owing to its high level of polymorphism and its candidate role in many diseases, HLA was the first system used in many techniques of genetic mapping, such as affected-sib-pair analysis and association (linkage disequilibrium) studies. Much remains unknown about the reasons why diseases are associated with HLA. Experience gained from HLA has, however, shown how other loci involved in complex traits can be identified by studying families with multiple affected cases or sib pairs, followed by linkage-disequilibrium mapping and then analysis of candidate genes.     

Characterization and Localization of a Phenoloxidase in Mung Bean Hypocotyl Cell Walls

The occurrence of proteins able to oxidize polyphenols even in the absence of H2O2 was recently reported in mung bean (Vigna radiata L.) hypocotyl cell wall extracts (R. Goldberg, A. Chabanet, A.M. Catesson [1993] In K.G. Welinder, S.K. Rasmussen, C. Penel, H. Greppin, eds, Plant Peroxidases: Biochemistry and Physiology, pp. 296-300). Therefore, the possible presence of a laccase in the extracts was investigated using immunocytological and biochemical approaches. An enzyme catalyzing phenol oxidation in the presence of molecular O2 was extracted and purified from the cell walls. This 38-kD cationic protein, like o-diphenoloxidases, was unable to oxidize p-diphenols or p-diamines. However, it crossreacted with an anti-laccase antiserum and, like laccases, its activity was inhibited by N-cetyl-N,N,N-trimethylammonium bromide but not by ferulic acid salts. Immunolabeling data showed that the 38-kD oxidase was absent from all cellulosic cell walls. It was localized only in lignifying and lignified cell walls. This restricted localization suggests that this laccase-like phenoloxidase could participate in the lignification process but not in the primary wall stiffening, which develops in the epidermal and cortical tissues along the mung bean hypocotyl.     

Major differences in isoform composition of starch synthase between leaves and embryos of pea (Pisum sativum L.)

Isoforms of starch synthase (EC 2.4.1.21) in pea (Pisum sativum L.) leaves have been identified and compared with those in developing pea embryos. Purification and immunoprecipitation experiments show that most of the soluble starch synthase activity of the leaf is contributed by a novel isoform (SSIII) that is antigenically related to the major soluble isoform of the potato tuber. The major soluble isoform of the embryo (SSII) is also present in the leaf, but contributes only 15% of the soluble activity. Study of the leaf starch of lam mutant peas, which lack the abundant granule-bound isoform responsible for amylose synthesis in the embryo (GBSSI), indicates that GBSSI is not responsible for the synthesis of amylose-like material in the leaf. Leaves appear to contain a novel granule-bound isoform, antigenically related to GBSSI. The implications of the results for understanding of the role of isoforms of starch synthase are discussed. Received: 13 March 1997 / Accepted: 13 May 1997     

The role of phytoplankton in the removal of arsenic by sedimentation from surface waters

The role of phytoplankton in the removal of arsenic (As) by particle adsorption and sedimentation was investigated in Moira Lake, Canada. Sampling water and suspended particles over one year illustrated significant variation in As partitioning between particulate and aqueous phases, but failed to establish a correlation between the partition coefficient, K _d, and indicators of phytoplankton biomass. A highly significant inverse logarithmic relationship was noted between K _d and the concentration of suspended particles (log K _d = 5.1 – 1.4 log SS; p = 0.0001) in an apparent demonstration of the particle concentration effect (O' Connor & Connolly, 1980).Particle deposition, measured by means of sediment traps, appeared to include a substantial component of resuspended surficial sediment making sediment trap results unreliable for quantifying the removal of substances from the water column. The As concentration of particles from deep traps deployed during late summer and early fall exceeded the As concentrations of suspended particles and surficial sediment, and may indicate that a highly contaminated nepheloid layer acts as a temporary sink for As.