Effects of pressure on uptake and release of calcium by brain synaptosomes

Uptake of radioactive calcium from guinea pig brain fractions enriched in synaptosomes could be significantly and reproducibly decreased by exposure to high pressure. Calcium efflux from preloaded synaptosomes was unaffected by pressure exposure. It was hypothesized that the development of pressure-induced encephalopathy may be related to an effect of pressure on the central nervous system calcium transport system.     

Characterization of herpesvirus sylvilagus glycoproteins released into the culture medium of infected cells: antisera to gp13 and gp32 neutralize viral infectivity in vitro and identify antigens on plasma membranes of infected cells.

Polypeptides released into the culture medium of herpesvirus sylvilagus-infected cells were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of extracellular fluid from [35S]methionine- and [3H]glucosamine-labeled cell cultures. Virus-induced glycoproteins 31, 32, and 33 (molecular weights of 62,000, 59,000, and 54,000, respectively) were the most abundant species and appeared predominantly in the culture medium. This observation, together with the known cell-associated nature of herpesvirus sylvilagus, suggested that virus-induced glycoproteins 31, 32, and 33 were specifically released. Immunization of rabbits with virus-induced glycoproteins 13 (molecular weight of 130,000) and 32 resulted in the production of antibodies that neutralized viral infectivity in vitro. Both antiserum to gp13 and antiserum to gp32 immunoprecipitated gp13, gp26, gp33a, gp45, and virus-induced polypeptide 39 (molecular weights of 130,000, 77,000, 49,000, 27,000, and 36,000, respectively) from [35S]methionine-labeled cell extracts as well as virus-induced glycoproteins 31, 32, and 33 from the culture medium. In addition, membrane immunofluorescence assays indicate that an antigen(s) reactive with anti-gp13/32 serum was located on the plasma membrane of infected cells.     

The distribution of repeating [Gal beta 1,4GlcNAc beta 1,3] sequences in asparagine-linked oligosaccharides of the mouse lymphoma cell lines BW5147 and PHAR 2.1

The occurrence and distribution of the repeating disaccharide [Gal beta 1,4GlcNAc beta 1,3] in the different types of Asn-linked oligosaccharides in mouse lymphoma BW5147 cells have been studied. Glycopeptides were prepared from cells grown in medium containing [6-3H]galactose, and the bi-, tri-, and tetraantennary Asn-linked oligosaccharides were fractionated by serial lectin affinity chromatography on concanavalin A-Sepharose, pea lectin -Sepharose, leukoagglutinating phytohemagglutinin-agarose, and Datura stramonium agglutinin-agarose. As described in this report, the latter lectin binds glycopeptides that contain either the repeating N-acetyllactosamine sequence or an outer mannose residue substituted at C-2 and C-6 by N-acetyllactosamine. The isolated glycopeptides were subjected to methylation analysis, specific exoglycosidase treatments, and digestion with Escherichia freundii endo-beta-galactosidase. Our data indicate that approximately two-thirds of the tetraantennary and one-half of the triantennary Asn-linked oligosaccharides contain repeating N-acetyllactosamine sequences in at least one branch. Many of the repeating sequences contain an additional galactose residue linked alpha 1,3 to a penultimate galactose residue. By contrast, less than 10% of the biantennary oligosaccharides contain the repeating disaccharide. The distribution of the repeating N-acetyllactosamine unit was also examined in a cell line ( PHAR 2.1) that is deficient in UDP-GlcNAc:alpha-mannoside beta 1,6-N-acetylglucosaminyltransferase. These cells are unable to synthesize tetraantennary and certain triantennary species and instead accumulate biantennary oligosaccharides. The total content of repeating N-acetyllactosamine units is greatly decreased in this line, and those that are present are found predominantly in triantennary Asn-linked oligosaccharides. These results demonstrate that the repeating N-acetyllactosamine sequence occurs commonly in complex-type Asn-linked oligosaccharides in BW5147 cells but is confined primarily to tri- and teraantennary species.     

Differential enhancement of spontaneous transition mutations in the lacI gene of an Ung- strain of Escherichia coli

In this communication, the contribution of cytosine deamination to spontaneous mutagenesis in the lacI gene of E. coli was examined. In a wild-type strain, 75% of the amber mutations recovered were G:C----A:T transitions and 60% of these were at the 5-methylcytosine spontaneous hotspots Am6, Am15 and Am34. In a strain deficient for uracil-DNA glycosylase (Ung-), 96% of the amber mutations were G:C----A:T transitions while only 15% of these occurred at the hotspot sites. This shift in the mutational distribution demonstrates that cytosine deamination is a potent mutagenic process, which is enhanced in the absence of glycosylase. Moreover, some amber sites were greatly enhanced in the Ung- strain while others were only slightly enhanced. This result suggests that the rate of cytosine deamination at individual sites may be influenced by surrounding base composition. Therefore, we examined the neighboring sequences and found a strong correlation between the fold-increase in mutation and the A/T richness of the surrounding sequence. It is suggested that A/T-rich regions denature more often, forming transient single strands in which cytosine residues would be expected to deaminate more readily.     

Mechanism of differences in pathogenicity between two variants of a laboratory strain of herpes simplex virus type 1

The mechanisms responsible for the difference in neurovirulence to inbred mice between two variants of the Miyama strain of herpes simplex virus type 1 (HSV-1) were studied. After intraperitoneal (i.p.) inoculation, the +GC (LPV) variant reached the spinal cord and the brain, and caused death. Conversely, the -GCr variant lacked the ability to gain access to the central nervous system (CNS) after the same route of infection and failed to kill susceptible mice. The initial virus growth after i.p. inoculation, as indicated by the number of infective centers (ICs) produced by the peritoneal exudate cells (PECs), was compared between these two variants. The virulent +GC (LPV) strain induced much more ICs than the attenuated -GCr variant. When the attenuated variant was preinoculated i.p. 24 hr before the challenge inoculation with the virulent variant by the same route, the production of ICs by the pathogenic variant was highly inhibited, and growth of this variant did not occur in the CNS. Thus, mice were protected from lethal infection by the virulent variant by preinoculation with the attenuated one. Moreover, the ability of mice to resist i.p. infection by HSV-1 was shown to be age-dependent.     

Phosphorylation of AMP by inorganic phosphorylating agents

AMP was phosphorylated by inorganic phosphorylating agents: cyclo-triphosphate and diphosphonate, in aqueous solution (70-80 degrees C, pH 6-12). The molecular structures of phosphorylated products were established by use of phosphorus-31 NMR and high-performance liquid chromatography (HPLC). The OH groups on AMP were phosphorylated by both phosphorylating agents to form 2'- or 3'-phosphate but an OH group on dAMP was not phosphorylated. Phosphorylation of OH group proceeds in two steps: formation of hydrogen bond between OH group and phosphorylating agent; subsequent nucleophilic attack of OH group on a phosphorus atom. Phosphate group on AMP was phosphorylated by diphosphonate but not by cyclo-triphosphate. The difference in the reactivities is explained in terms of charge repulsion between AMP and agents.     

Decompression outcome following saturation dives with multiple inert gases in rats

This investigation examined the question of whether gas mixtures containing multiple inert gases provide a decompression advantage over mixtures containing a single inert gas. Unanesthetized male albino rats, Rattus norvegicus, were subjected to 2-h simulated dives at depths ranging from 145 to 220 fsw. At pressure, the rats breathed various He-N2-Ar-O2 mixtures (79.1% inert gas-20.9% O2); they were then decompressed rapidly (within 10 s) to surface pressures. The probability of decompression sickness (DCS), measured either as severe bends symptoms or death, was related to the experimental variables in a Hill equation model incorporating parameters that account for differences in the potencies of the three gases and the weight of the animal. The relative potencies of the three gases, which affect the total dose of decompression stress, were determined as significantly different in the following ascending order of potency: He less than N2 less than Ar; some of these differences were small in magnitude. With mixtures, the degree of decompression stress diminished as either N2 or Ar was replaced by He. No obvious advantage or disadvantage of mixtures over the least potent pure inert gas (He) was evident, although limits to the expectation of possible advantage or disadvantage of mixtures were defined. Also, model analysis did not support the hypothesis that the outcome of decompression with multiple inert gases in rats under these experimental conditions can be explained totally by the volume of gas accumulated in the body during a dive.     

Anaerobic ureolytic bacteria from caecal content and soft faeces of rabbit

Forty strains of ureolytic bacteria were isolated from the caecal content and soft faeces of seven rabbits by the anaerobic roll tube method and were characterized. The isolates were identified with Clostridium coccoides, Cl. innocuum, Peptostreptococcus productus, P. micros, Peptococcus magnus, Fusobacterium russii and Fusobacterium sp. Urease activity of representative strains of the various species was also determined. The study indicated that strongly-ureolytic anaerobic bacteria are present in the caecum of the rabbit.