A comparison of anionic sites in the glomerular basement membranes from different classes of fishes

Summary Cationized ferritin was injected into the circulatory system of teleosts, the sea raven and Atlantic eelpout, and into elasmobranchs, the spiny dogfish and the skate, to determine if the glomerular basement membranes (GBM) from these different groups of fishes possess anionic binding sites similar to those present in the GBM of mammals. The distribution of cationized ferritin was the same in all fishes listed. Cationized ferritin was localized only in the GBM and the mesangial matrix. The regular distribution of cationized ferritin within the laminae rarae (60 nm intervals) was taken as evidence of the presence of anionic binding sites. Cationized ferritin did not bind to the glomerular capillary endothelium, nor was any of it localized at the base of the slit diaphragms of the foot processes of the podocytes. The distribution of binding sites in the GBM of these fishes is similar to that in another teleost, the winter flounder, and in a cyclostome, the hagfish.     

A general method for the induction and screening of antisera for cDNA-encoded polypeptides: antibodies specific for a coronavirus putative polymerase-encoding gene

A prokaryotic vector, pGE374, containing the recA and lacZ genes, out-of-frame, was used for the expression of cDNA derived from the putative polymerase-encoding gene of the coronavirus mouse hepatitis virus strain A59 (MHV-A59). The pGE374/viral recombinant vector generates a tripartite bacterial/viral protein composed of a segment of the RecA protein at the N terminus, the coronaviral sequences in the middle, and an enzymatically active beta-galactosidase at the C terminus. Rabbits immunized with such recombinant proteins generated antibodies to the MHV-A59 portion of the tripartite protein. Because the MHV-A59 polymerase proteins have been difficult to identify during infection, we used a novel method to demonstrate the viral specificity of the antiserum. The viral cDNA was excised from the expression vector, and transferred to a pGem vector, downstream from and in-frame with a portion of the cat gene. This construct contained a bacteriophage RNA polymerase promoter that enabled the cell-free synthesis of a fusion protein that was used to verify that antibodies were generated to the expressed viral DNA. This strategy was shown to successfully result in the specific generation of antibodies to the encoded information of the viral cDNA. Furthermore, this method has general applicability in the generation and characterization of antibodies directed against proteins encoded in cDNAs.     

Antifreeze glycopeptide adsorption on single crystal ice surfaces using ellipsometry

Antarctic fishes synthesise antifreeze proteins which can effectively inhibit the growth of ice crystals. The mechanism relies on adsorption of these proteins to the ice surface. Ellipsometry has been used to quantify glycopeptide antifreeze adsorption to the basal and prism faces of single ice crystals. The rate of accumulation was determined as a function of time and at concentrations between 0.0005 and 1.2 mg/ml. Estimates of packing density at saturation coverage have been made for the basal and prism faces.     

Isolation and characterization of axolemma-enriched fractions from rabbit and bovine peripheral nerve

Axolemma-enriched fractions were isolated from bovine spinal accessory nerves, bovine intradural dorsal roots, and rabbit sciatic nerve by differential centrifugation and separation on a linear 10–40% sucrose (w/w) gradient. The fractions were enriched 4 to 10 fold in acetylcholinesterase, a biochemical marker for axolemma. Axolemma-enriched fractions isolated from uniformly well-myelinated fibers (bovine spinal accessory nerve) contained lower CNPase activity and higher acetylcholinesterase activity than comparable fractions isolated from variably myelinated fibers (rabbit sciatic nerve and bovine intradural roots). Separation by polyacrylamide electrophoresis showed that the molecular weight distribution of all peripheral nerve axolemma-enriched fractions was similar and ranged from 20 to over 150 kilodaltons. All axolemma-enriched fractions appeared to contain a small but variable amount of myelin-specific proteins. Based on biochemical properties, peripheral nerves containing uniformly well-myelinated fibers yield an axolemma-enriched fraction which is least contaminated with myelin-related membranes.     

CYCLIC NUCLEOTIDE LEVELS IN NORMAL AND BIOLOGICALLY FRACTIONATED MOUSE RETINA: EFFECTS OF LIGHT AND DARK ADAPTATION

Abstract— The content of cyclic AMP and cyclic GMP was measured in whole eyes and in normal retinas from C57BL(6)J mice, in receptorless retinas from congenic mice homozygous for the receptor dystrophy gene (rd/rd), and in retinas from mice treated postnatally with monosodium glutamate. Normal retinas contain approx 320 μg of protein: dystrophic (rd/rd) retinas contain approx 110μg of protein, lack rods but possess some surviving cone somata and terminals: glutamate-modified retinas contain approx 200 μg of protein and have both a reduced area and thickness with a marked deficiency of ganglion cells and amacrine cells. In normal mice, more than 90% of the cyclic GMP, but only 607, of the cyclic AMP of the whole eye was in the retina. In normal dark-adapted retinas isolated under dim red light cyclic AMP and cyclic GMP content was 4.1 and 20.2pmol/retina, respectively. The content of both cyclic AMP and cyclic GMP was 40% less, 2.5 and 11.5pmol/retina, respectively, in light-adapted retinas. In dark-adapted retinas isolated under infra-red light, cyclic AMP content was 40%, higher than that in retinas isolated under dim red light; cyclic GMP content was the same under these two conditions. Receptorless retinas contained approx 50% as much cyclic AMP and only 1-2% as much cyclic GMP as normal retinas. Although glutamate-modified retinas also had approx 50% as much cyclic AMP, they contained 60-85%, as much cyclic GMP as normal retinas. Light decreased by 30-50% levels of both cyclic AMP and cyclic GMP in glutamate-modified retinas, but only reduced cyclic nucleotide levels in receptorless retinas by 20%.
These data indicate that 95% or more of the cyclic GMP is in photoreceptor cells, whereas cyclic AMP is more evenly distributed throughout the retina. In addition, both cyclic AMP and cyclic GMP levels are influenced by light- and dark-adaptation.     

Diaminopimelic Acid Synthesis in Cultures of an Escherichia coli Double Auxotroph: Effects of Cultural Conditions

The metabolic response to L-lysine of Escherichia coli ATCC 13002, a lysine-histidine double auxotroph, has been examined in a synthetic medium containing sucrose. In shaken cultures largest amounts of extracellular DAP were produced with an initial lysine concentration of 7·5 mg/1 and in static cultures of 2·5 mg/1. Considerably smaller amounts of DAP accumulated under stationary conditions. In cultures shaken for 20 and 43 h there was an overall decrease in the yields of DAP, expressed in terms of cell biomass and of sucrose consumed, as the initial concentration of lysine was increased from 0·75 mg/1 in steps up to 25 mg/1. The regulatory effect of lysine on DAP production was also observed when lysine was supplied to cultures at a constant rate employing diffusion capsules.     

Expression of a mutation affecting F incompatibility in the integrated but not the autonomous state of F.

Previously we have described a mutant Hfr strain in which incompatibility between the integrated F factor and an autonomous F-prime (F') factor was abolished. The mutation (inc) was located in the integrated F factor. F-prime factors isolated from the mutant Hfr strain have the same incompatibility behavior as those isolated from normal Hfr strains. Reintegration of these F' factors into the chromosome restores the Inc- phenotype characteristic of the mutant Hfr. The inc mutation thus affects incompatibility between integrated F and autonomous F(Fi-Fa incompatibility) but not incompatibility between two autonomous F factors (Fa-Fa incompatibility). The implications of this finding for the mechanism of plasmid incompatibility are discussed.     

Adaptations for cryopelagic life in the antarctic notothenioid fish Pagothenia borchgrevinki

Summary The cryopelagic circumantarctic notothenioid fish Pagothenia borchgrevinki lives near the undersurface of the sea ice in McMurdo Sound, Antarctica. When compared to closely related benthic species (especially Trematomus bernacchii), Pagothenia exhibited substantial morphological differences in a variety of organ systems. The values of the fineness ratio and the indices of trunk shape and flatness suggested streamlining and drag reduction, adaptations to life in the water column. Pagothenia also lacked substrate contact adaptations in the pelvic and anal fins. Silvery reflective layers (strata argentea) beneath the skin and in the iris and choroid of the eye provided camouflage when Pagothenia were viewed against a background of platelet ice. The retina had many cones indicating the eye was adapted to both diurnal and nocturnal vision. During the austral spring Pagothenia fed exclusively on nektonic organisms near the ice-water interface. Dietary diversity was low; copepods and amphipods were the most frequently occurring and amphipods were the most frequently occurring taxa. In conclusion, Pagothenia appear specialized for life in the water column.     

Phagocytosis of degenerating myelin in transected feline optic nerve: an immunohistochemical study

The phagocytic activity of neuroglial cells in adult feline degenerating optic nerve was investigated by immunocytochemistry at both light and electron microscopy levels. Degeneration was initiated by unilateral eye enucleation and the segment distal to the transection showing true Wallerian degeneration was examined. Following enucleation, twelve adult domestic cats were examined over a period of seven to 215 days. All cases showed slow clearance of myelin debris and absence of proliferating monocytes throughout the post-enucleation period. All phagocytic cells present were neuroglial cells, and many of these cells expressed oligodendroglial antigens. These findings demonstrate the persistence of an active population of oligodendrocytes that might play an additional functional role during Wallerian degeneration of feline optic nerve.