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Biomedical subjects

D L Edwards

Publications and source records attributed to D L Edwards.

At least 37 records · Page 2Linked to original sources
PubMed1986

Intraocular tetrodotoxin reduces axonal transport and transcellular transfer of adenosine and other nucleosides in the visual system of goldfish.

Intraocular injection of tetrodotoxin (TTX) in goldfish, which abolishes physiological activity in the optic axons, decreased by up to about 30% the amount of radioactively labeled adenosine, uridine and guanosine (and their nucleotide derivatives) that was axonally transported in the optic nerve. The amount of labeled nucleoside that reached the optic tectum and became incorporated into RNA in the postsynaptic tectal neurons and glial cells was reduced by up to about 50%. There was no change, however, in the amount of transported nucleoside that became incorporated into RNA in the optic nerve glia. The TTX-induced changes were eliminated when axonal transport was blocked with vincristine, indicating that this change did not involve material moving along the nerve by diffusion. If the TTX injection was delayed until several hours after labeling of the transported materials, the transported labeled nucleoside in the nerve was reduced very little, but the RNA labeling in the tectum was reduced just as much as when TTX was given prior to labeling. This indicates that the labeling of the tectal cells was affected more by the level of activity in the pathway than by the amount of transported nucleoside reaching the optic nerve terminals. It appears likely, therefore, that the process most affected by the decrease in physiological activity is the release of nucleoside from the terminals of the presynaptic neurons or its uptake into postsynaptic tectal neurons and glia. The fact that physiological activity may modify the amount of axonally transported nucleosides made available for metabolism (including RNA synthesis) in postsynaptic neurons may provide an explanation for activity-linked neurotrophic effects.

Adenosine↗
PubMed1985

The effect of conformity and plastic thickness on contact stresses in metal-backed plastic implants.

Surface damage in polyethylene components for total joint replacement is associated with large contact stresses. An elasticity solution and finite element analyses were used to determine the influence of design parameters on the stresses due to contact in metal-backed components. For nearly conforming contact surfaces, it was found that the stresses in the plastic are very sensitive to clearance, that minimum plastic thickness of 4-6 mm should be maintained for metal-backed components, and that bonding the plastic to the metal backing reduces tensile stresses in the plastic at the edge of the contact zone.

Elasticity↗
PubMed1984

Intraocular injection of tetrodotoxin in goldfish decreases fast axonal transport of [3H]glucosamine-labeled materials in optic axons.

When physiological activity in goldfish visual system was abolished by repeated intraocular injection of tetrodotoxin (TTX), the fast axonal transport of radioactive amino acid-labeled protein in the optic axons was unaltered. However, the TTX treatment reduced the amount of [3H]glucosamine-labeled glycolipids that were axonally transported to the optic tectum, and may have decreased their rate of turnover in the tectum. A similar though smaller effect was observed for glucosamine-containing glycoproteins. These alterations in axonal transport may be the basis for at least some of the deleterious effects of TTX on axonal regeneration in this system.

Animals↗
PubMed1984

Antibodies against filamentous components in discrete cell types of the mouse retina.

Three monoclonal antibodies have been raised against the ganglion cell layer of the adult mouse retina. The first antibody, R3, labeled optic axons in the inner retina, and with colchicine pretreatment somata and dendrites of large ganglion cells could be seen. A small number of other processes, including fibers projecting to the retina from elsewhere (efferent fibers), were also labeled in the inner retina. In the outer plexiform layer R3 stained the axonless class of horizontal cells. R3 recognized a 185,000- to 200,000-dalton polypeptide which is most probably the heaviest of the neurofilament subunits. Antibodies R4 and R5 labeled filamentous components mainly in glia and cells of mesenchymal origin. The antigens appeared in most but not quite all locations morphologically closely related to the intermediate filament protein vimentin. In the retina both antibodies labeled strongly the regularly spaced Müller glia. The astroglia of the optic fiber layer was stained with R5 but not R4. Although the two antigens were in general not expressed in neurons, they were both present in axonless horizontal cells in the outer plexiform layer, coexisting with neurofilaments in this neuron.

Animals↗
PubMed1983

Intraocular tetrodotoxin in goldfish hinders optic nerve regeneration.

Repeated intraocular injection of tetrodotoxin (TTX) was used to produce a maintained block of neural activity in goldfish optic axons which were regenerating following a crush of the optic nerve. The recovery of visual function was delayed in the TTX-treated fish, as demonstrated by delays in the return of the startle reaction to sudden illumination, the dorsal light reflex and food pellet localization. A single injection of TTX at the time of optic nerve crush delayed recovery of the startle reaction but not of food localization. There was no loss of ganglion cells in the TTX-treated animals, but the number of regenerated axons detectable by light microscopy was reduced. Also, axonal transport of radioactively labeled protein in some of the regenerating axons showed a deficit at 21-28 days after the lesion, i.e., during innervation of the tectum. Incorporation of labeled amino acid into protein in the retinal ganglion cells was depressed during the same period, but both the transport and incorporation had returned to normal by 36 days after the lesion. These results, together with the results of the accompanying electrophysiological analysis of the effects of TTX58,59, suggest that TTX treatment interferes with two separate events in regeneration, one occurring soon after the nerve lesion and the other during innervation of the tectum. During at least the first of these events the effect of TTX treatment may be to reduce the number of size of the regenerating axon branches. We propose that the TTX effect may be mediated by a reduction in the axonal transport of certain materials, including gangliosides, nucleosides, or proteins specifically involved in regeneration.

Animals↗
PubMed1983

The re-establishment of synaptic transmission by regenerating optic axons in goldfish: time course and effects of blocking activity by intraocular injection of tetrodotoxin.

Intraocular injections of tetrodotoxin were used to block activity for 27 days in normal fish and for the first 27 or 31 days of regeneration in fish with one optic nerve crushed. Synaptic activity was then assessed by a current source-density analysis of field potentials evoked by optic nerve shock at different times following the TTX treatment. In normal fish, the lack of activity for 4 weeks had no significant effect on the maintenance of synaptic strength. Likewise, in fish with nerve crush, lack of activity did not prevent the regenerating optic fibers from forming synapses that were nearly as effective as those formed in controls injected with the citrate buffer vehicle. The earliest synapses were formed at the rostromedial corner of the tectum (where the tract enters) at 20 days after nerve crush, when fibers had not yet reached the caudal areas. By 28 days synaptic potentials could be recorded everywhere on the surface of the tectum in both controls and TTX injected fish. However, the latency of the responses with TTX were longer, suggesting a smaller caliber of fiber, which is consistent with an earlier finding of decreased axonal transport in TTX fish. Maturation of the regenerating fibers proceeded slowly in both TTX and control fish. After more than 5 months, the projections were nearly normal but still not completely normal.

Animals↗
PubMed1983

Activity sharpens the map during the regeneration of the retinotectal projection in goldfish.

In the regenerating retinotectal projection of goldfish, we have used intraocular injections of tetrodotoxin (TTX) to determine whether activity plays a role in organizing or refining the retinotopic map. Repeated injections produced a continuous 27-day block without producing extraocular effects or causing deleterious effects in the retinal ganglion cells. The retinotectal maps regenerated in the TTX fish were normally organized but the multiunit receptive fields were grossly enlarged. In control regenerates, 1-3 units (arbors of retinal ganglion cell axons) were simultaneously recorded at each penetration and their combined receptive field averaged 11-12 degrees, nearly the same as for single units. In TTX fish each penetration yielded at least 5-10 units whose receptive fields were clustered over a wider area averaging 27 degrees across. Individual ganglion cell receptive fields were assessed both by tectal and by intraretinal recording and were not enlarged. Many fish were recorded up to 4 months after the release from TTX block, but no further refinement of the maps occurred. If the nerve was recrushed and regenerated a second time without TTX, a normal map was formed, ruling out any permanent changes in the retinal ganglion cells or in the tectum. Blocks during various portions of the regeneration process showed that lack of activity during the process of axonal elongation (first 2 weeks) does not cause enlargement of the multiunit receptive fields, but lack of activity during the period of synapse formation and maturation (14-34 days) does. The results are discussed in terms of an activity-dependent stabilization of synapses. Neighboring retinal ganglion cells are known to fire in a statistically correlated fashion and this could help in their elimination of incorrect branches following an early period of diffuse connections.

Animals↗
PubMed1983

Neurofilaments contain alpha-melanocyte-stimulating hormone (alpha-MSH)-like immunoreactivity.

An antiserum to alpha-melanocyte-stimulating hormone (alpha-MSH) was found to contain antibodies to at least two types of determinants on the alpha-MSH peptide: one is present only on the free peptide, the other is shared with neurofilaments. Immunoblots from mouse brain showed the neurofilament crossreactivity to be located on proteins in the Mr 140,000 range. The neurofilament-crossreactive portion of the antiserum could be selectively absorbed out with a cytoskeletal preparation, which abolished all affinity of the antiserum to the retina but did not affect the labeling pattern in the pituitary. Absorptions with desacetyl-alpha-MSH and corticotropin seemed to indicate that the determinant shared with neurofilaments is not located at either end of the alpha-MSH peptide, but somewhere in between. The immunohistochemical labeling of the retina with the alpha-MSH antiserum was compared to the labeling with monoclonal antibodies against Mr 200,000 neurofilaments. In the adult retina the alpha-MSH-like immunoreactivity was found to be slightly more widespread; most consistently it was detectable in cell bodies of large ganglion cells, whereas the heavy neurofilament subunit was absent from somata and proximal axons of these cells. In the developing mouse brain, expression of the heavy subunit was found to lag 2-3 wk behind expression of the Mr 140,000 proteins. This confirms previous reports of a more restricted distribution and late expression of high molecular weight neurofilaments as compared to the lower subunits.

Animals↗
PubMed1979

Selection of succinic dehydrogenase mutants of Neurospora crassa.

A method is described which permits the selection of mutants of Neurospora crassa that are deficient in succinic dehydrogenase activity. The method relies on the observation that succinic dehydrogenase-deficient strains fail to reduce the dye nitrotetrazolium blue when overlaid with the dye in the presence of succinate and phenazine methosulfate. Wild-type colonies reduced the dye and turned blue, whereas mutant colonies remained colorless. In this communication we present studies of a mutant, SDH-1, isolated by this method. The mutant had 18% of the succinic dehydrogenase activity of the parent strain used in the mutation experiments as determined from the ratio of Vmax activities obtained from Lineweaver-Burk plots. The SDH-1 mutant segregated in a Mendelian manner when back-crossed to its parent strain. Succinate oxidase activity in SDH-1 was low and was markedly inhibited by adenosine 5'-diphosphate. The succinate oxidase activity of the parent strain was high and was not affected by the presence of adenosine 5'-diphosphate.

Mutation↗
PubMed1979

Antibody-dependent cellular cytotoxicity effector cell capability among normal individuals.

Nonadherent peripheral blood lymphocytes (PBL) from 110 normal individuals were evaluated for their capability to mediate antibody-dependent cellular cytotoxicity (ADCC) against rabbit antibody-sensitized chicken erythrocytes (CRBC). Titration of PBL against a constant number of chromium-51 labeled CRBC allowed for construction of dose response cytotoxicity curves where percent chromium release was plotted against the number of PBL. From these curves the maximum degree of lymphocyte-mediated cytotoxicity and the number of lymphoid cells required for 50% of the maximum lymphocyte-mediated cytotoxicity were determined. The results suggested that in this system the cytotoxic capability of PBL from normal individuals was independent of the age and sex of the donor. The cytotoxic capability of PBL was also found to be constant in individuals tested sequentially. However, significant differences were found to exist between different individuals in their capability of mediating ADCC in this system.

Adult↗
PubMed1978

Nuclear mutations conferring oligomycin resistance in Neurospora crassa.

Mutants of Neurospora crassa have been isolated that are highly resistant to inhibition by oligomycin, an inhibitor of mitochondrial ATPase activity. Dixon plots (Dixon, M., and Webb, E.C. (1964) Enzymes, 2nd Ed, pp. 328-330, Academic Press, New York) of oligomycin inhibition curves of the parent strain and the resistant mutants are linear, indicating that oligomycin interacts at a single site within the ATPase complex. The Ki values obtained from the mutants vary from 150 to 900 times greater than the Ki obtained for the parent strain. The parent strain and the oligomycin-resistant mutants are also inhibited by bathophenanthroline, a lipophilic chelating agent that inhibits F1 ATPase activity. Dixon plots of bathophenanthroline inhibition curves are also linear and Ki values obtained are all approximately equal. Crosses of the oligomycin-resistant mutants to the oligomycin-sensitive parent strain show a mendelian segregation of the resistance characteristic. These data show that mutations leading to oligomycin resistance in Neurospora are due to alterations in nuclear genes.

Adenosine Triphosphatases↗
PubMed1978

Electron transport in the cni-1 mutant of Neurospora crassa.

1. Mitochondria from the nuclear mutant cni-1 have no optically detectable cytochrome aa3 in early log phase growth. These mitochondria have a high level of respiration that is not inhibited by cyanide but is inhibited by salicylhydroxamic acid. They also show a substantial amount of cyanide-sensitive respiration. 2. As cultures of mutant cni-1 age, flux through the hydroxamate-sensitive pathway decreases markedly while flux through the cytochrome chain remains constant. 3. Growth studies with mutant cni-1 indicate that the cytochrome chain in this mutant is more important in supporting growth than the hydroxamate-sensitive pathway. 4. Measurements of the steady-state level of reduction of cytochrome c in mutant cni-1 indicate that the rate-limiting step in the cytochrome chain is at the position occupied by cytochrome oxidase. 5. Electron spin resonance studies with cni-1 mitochondria show normal cytochrome oxidase signals in the g approximately 6 region although there is little or no optically detectable cytochrome aa3.

Cyanides↗
PubMed1978

Cyanide- and hydroxamate-resistant respiration in Neurospora crassa.

Strain inl-89601 of Neurospora crassa respires exclusively by means of the mitochondrial cytochrome chain. The respiration of this strain is entirely inhibited by cyanide or antimycin A, the classical inhibitors of cytochrome chain respiration. When this strain was grown in the presence of chloramphenicol, however, two additional terminal oxidases were detected. One of these oxidases is inhibited by substituted hydroxamic acids and has been described previously. The second oxidase was not inhibited by cyanide or hydroxamic acid but was inhibited by azide in the presence of both cyanide and hydroxamic acid. This azide-sensitive respiration was due to a single respiratory pathway with a Ki for azide of 200 micrometer. A small amount of azide-sensitive respiration was detected in mitochondrial fractions obtained from chloramphenicol-treated cells, and it is likely that the azide-sensitive oxidase is localized in the mitochondrion. The determinants for the azide-sensitive and hydroxamate-sensitive oxidases segregate in a Mendelian manner in crosses and are either unlinked or not closely linked to each other.

Azides↗
PubMed1976

Electron spin resonance investigations of mitochondrial electron transport in Neurospora crassa. Characterization of paramagnetic intermediates in a standard strain.

1. Submitochondrial particles from Neurospora strain inl-89601 have been analyzed by electron spin resonance spectroscopy (ESR). Numerous signals due to iron-sulfur proteins are observed at low temperatures. Analysis of these ESR signals at various temperatures allows the assignment of resonances to iron-sulfur centers 1-5 that have been described in other organisms. There are no discrepancies between the signals seen in Neurospora and those described in other organisms and it is likely that Neurospora mitochondria contain the same iron-sulfur centers that are observed elsewhere. 2. NADPH and NADH act to reduce the iron-sulfur centers of respiratory complex I. 3. The drug pyrrolnitrin [3-chloro-4-(2'-nitro-3'-chlorphenyl)pyrrole] is an effective inhibitor of both NADH-supported and succinate-supported electron transport in Neurospora. 4. Analysis of pyrrolnitrin inhibition curves, respiration studies, ESR spectra, and the steady-state level of reduction of cytochrome b in the presence and absence of the drug shows that pyrrolnitrin acts to inhibit electron transport in Neurospora mitochondria at multiple sites in the region between ubiquinone and cytochrome b.

Depression, Chemical↗
PubMed1976

Regulation of cyanide-insensitive respiration in Neurospora.

1. Inhibition of either mitochondrial transcription or translation in Neurospora crassa results in the rapid production of the cyanide-insensitive pathway of mitochondrial respiration. Protein synthesis on cytoplasmic ribosomes is required for the appearance of cyanide-insensitive respiration in the culture. 2. Removal of the inhibition of transcription of translation results in a rapid return to cyanide-sensitive respiration. Additional protein synthesis in the cytoplasm is required for the loss of cyanide-insensitive respiratory activity while additional mitochondrial protein synthesis has no effect. 3. These studies indicate that a mitochondrial gene product is involved in a negative manner in the regulation of cyanide-insensitive respiration. When the mitochondrial product is present, the pathway is not expressed, when the mitochondrial product is absent, the pathway is expressed. 4. Studies with forced heterokaryons formed from respiratory-deficient mutants having cyanide-insensitive respiration and respiratory-competent auxotrophs indicate that the site of action of the mitochondrial gene product is external to the mitochondrion.

Chloramphenicol↗