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R K Crouch

Publications and source records attributed to R K Crouch.

At least 91 records · Page 5Linked to original sources
PubMed1990

Hydrogen peroxide is the most toxic oxygen species for Onchocerca cervicalis microfilariae.

The toxicity of the active oxygen species hydrogen peroxide, superoxide radical, hydroxyl radical and singlet oxygen to microfilariae (mf) has been studied in vitro, using active oxygen-generating systems and scavengers/inhibitors. Mf viability was monitored by uptake of the radiolabel, [3H]2-deoxy-D-glucose. Hydrogen peroxide and singlet oxygen, but not superoxide radical or hydroxyl radical, are toxic for mf. Hydrogen peroxide was toxic for mf within 2 h at concentrations as low as 5 microM, an amount eosinophils have been shown to release in vitro (Weiss et al. 1986). Catalase and thiourea, but not inactivated catalase, superoxide dismutase (SOD), singlet oxygen scavengers, or hydroxyl radical scavengers, protected mf. Mf have relatively high levels of endogenous SOD but no measurable glutathione peroxidase and low levels of catalase when compared with other parasites (Callahan, Crouch & James, 1988). The low levels of hydrogen peroxide-scavenging enzymes correlate well with mf sensitivity to hydrogen peroxide and the protective effect of exogenous catalase.

Animals↗
PubMed1990

Sensitization of bleached rod photoreceptors by 11-cis-locked analogues of retinal.

Photoactivation of rhodopsin initiates both excitation and adaptation in vertebrate rod photoreceptors. Bleaching of rhodopsin to free opsin and all-trans-retinal in isolated rods produces a stable desensitization (bleaching adaptation) that is much larger than expected from pigment depletion alone. In our experiments, a 93% bleach produced a 500-fold increase in the light intensity required for saturation of the light response. This component of adaptation was 32-fold larger than the 16-fold increase expected from pigment depletion alone. 11-cis-Retinal, when delivered to isolated rods from liposomes, combines with free opsin to form a bleachable photopigment that fully restores sensitivity. 11-cis-Locked analogues of retinal combine with opsin to form unbleachable pigments in isolated bleached rods from the tiger salamander. They restore sensitivity to a substantial (16- to 25-fold) but incomplete extent. The analogues apparently relieve a stable component of adaptation when they interact with opsin. Because these analogues do not detectably excite rods, the structural requirements of both retinal and opsin for the relief of adaptation are different from those of excitation. The biochemical basis of light adaptation resulting from pigment bleaching and the minimum structural requirements of retinal for its relief remain to be determined.

Ambystoma↗
PubMed1989

Analogue pigment studies of chromophore-protein interactions in metarhodopsins.

Several analogue pigments have been prepared containing retinals altered at the cyclohexyl ring or proximal to the aldehyde group in order to examine the role of the chromophore in the formation of the metarhodopsin I and II states of visual pigments. Deletion of the 13-methyl group on the isoprenoid chain did not affect metarhodopsin formation. However, analogue pigments containing chromophores with modified rings did not show the typical absorption changes associated with the metarhodopsin transitions of native or regenerated rhodopsins. In particular, 4-hydroxyretinal pigments did not show clear transitions between the metarhodopsin I and metarhodopsin II states. Pigment formed with an acyclic retinal showed no evidence by absorption spectroscopy of metarhodopsin formation. A retinal altered by substitution of a five-membered ring containing a nitroxide required a more acidic pH than the native pigment for formation of the metarhodopsin II state. ESR data suggest that the ring remains buried within the protein through the metarhodopsin II state. However, the Schiff base linkage is susceptible to hydrolysis of hydroxylamine in the metarhodopsin II state. These data indicate that (1), in the transition from rhodopsin to metarhodopsin II, major protein conformational changes are occurring near the lysine-retinal linkage whereas the ring portion of the chromophore remains deeply buried within the protein and (2) pigment absorptions characteristic of the metarhodopsin I and II states may be due to specific protein-chromophore interactions near the region of the chromophore ring.

Animals↗
PubMed1989

Spin trapping of the superoxide anion: complications in the use of the water-soluble nitroso-aromatic reagent DBNBS.

Sodium 3,5-dibromo-4-nitrosobenzenesulfonate (DBNBS) is reported to be a useful spin trap for the measurement of superoxide anions in aqueous solution. However, the signal observed arises from interaction of the spin trap with some species other than the superoxide radical or hydrogen peroxide, a product of its dismutation, as the addition of both superoxide dismutase and catalase to a superoxide generating system failed to attenuate the signal. Therefore caution must be employed in the interpretation of results obtained using this spin trap.

Benzenesulfonates↗
PubMed1989

Retinoid requirements for recovery of sensitivity after visual-pigment bleaching in isolated photoreceptors.

After visual-pigment bleaching, single isolated rod photoreceptors of Ambystoma tigrinum recover their sensitivity to light when supplied with 11-cis-retinal from liposomes or with 11-cis-retinal bound to interphotoreceptor retinoid-binding protein. Bleached rods do not recover sensitivity, or do so only very slowly, after exposure to 11-cis-retinol. The latter retinoid is "toxic" in that rods actually lose sensitivity in its presence. In contrast, bleached isolated cone cells recover sensitivity when either retinoid is supplied. It is suggested that the major pathway for rhodopsin regeneration during dark adaptation in the intact eye is transport of 11-cis-retinal from the pigment epithelium to the retina. The results also suggest that there may be separate pathways for visual-pigment regeneration in rods and cones during dark adaptation.

Ambystoma↗
PubMed1989

An in vitro radiolabel uptake viability assay for Onchocerca microfilariae.

A radiolabel uptake viability assay for Onchocerca cervicalis using [3H]2-deoxy-D-glucose in Hanks' balanced salt solution, pH 7.5, at 30 C is described and compared to the traditional visual motility assay. A correlation of r = 0.92 between the assays was found, with the radiolabel uptake method apparently a more sensitive indicator of microfilarial viability.

Animals↗
PubMed1988

Helminth anti-oxidant enzymes: a protective mechanism against host oxidants?

Highly reactive oxygen species potentially represent a powerful effector mechanism against parasites. They are produced during normal cellular metabolism, especially by activated phagocytes, and also by some anti parasitic drugs. From studies to date, all protozoan and helminth parasites appear to have one or more anti-oxidant enzymes able to scavenge or quench the reactive oxygen species, and there is strong evidence that such enzymes play a crucial role in protecting against the host response. This detailed review, which summarizes studies on the major anti-oxidant enzymes of helminths, clearly illustrates that methods to block or overcome anti-oxidant protection may be a fertile field in the search for improved ways to inhibit parasite survival.

Journal Article↗
PubMed1988

Lack of interaction of rhodopsin chromophore with membrane lipids. An electron-electron double resonance study using 14N:15N pairs.

Electron-electron double resonance (ELDOR) has been applied to the study of specific interactions of 15N-spin-labeled stearic acid with the retinal chromophore of a rhodopsin analogue containing a 14N spin-labeled retinal. Both the 5 and 16 spin-labeled stearic acids were incorporated into the lipid bilayer of rod outer segment membranes containing the spin-labeled pigment. No interaction between the 15N and 14N spin-labels was observed in rhodopsin or the metarhodopsin II state with either of these labeled stearic acids. Therefore in this system the ring portion of the chromophore must be highly sequestered from the phospholipid bilayer in both the rhodopsin and metarhodopsin II forms.

Animals↗
PubMed1988

Fenretinide does not block visual pigment formation in the rat.

Abnormal retinal function has been reported in patients receiving the synthetic retinoid fenretinide. In this study, the vitamin A deprived rat was used as a model to test the possibility that fenretinide is interfering with the vitamin A transport system and/or blocking the normal uptake of 11-cis retinal into the visual pigment. Assessing retinal function by electroretinogram measurements and visual pigment formation by competition studies, our data demonstrate normal uptake of vitamin A and normal visual pigment formation in the rat after treatment with the drug.

Animals↗
PubMed1988

Catalase pretreatment attenuates oleic acid-induced edema in isolated rabbit lung.

Because reactive O2 metabolites have been demonstrated to be potent mediators of vascular dysfunction and are synthesized by lung tissue, their involvement as mediators of oleic acid (OA)-induced pulmonary edema in the isolated Krebs-perfused rabbit lung was assessed. Injection of OA (0.1 ml) into the pulmonary artery after vehicle pretreatment induced marked increases in lung weight [50.4 +/- 13.9 vs. 4.2 +/- 2.0 (SE) g 45 min after OA or vehicle, respectively, P less than 0.05], an index of pulmonary edema, and airway pressure. OA also caused a significant though minimal increase in pulmonary arterial pressure. Pretreatment with catalase (1,000 U/ml), a scavenger of H2O2, significantly (P less than 0.05, Friedman's) attenuated the increases in lung weight (50.4 +/- 13.9 vs. 15.1 +/- 4.9 g), airway pressure, and pulmonary arterial pressure. In contrast to catalase, pretreatment with Cu-tryptophan (40 microM), a lipid-soluble scavenger of superoxide, provided no protective effect by itself, nor was there any potentiation of protection when combined with catalase. Further evidence implicating O2 metabolites in OA-induced edema was obtained by electron paramagnetic resonance (EPR) spectroscopy of perfusate samples to which the spin trap, sodium 3,5-dibromo-4-nitrosobenzenesulfonate (10 mM), was added. Analysis of these samples revealed the presence of free radicals after OA. Pretreatment with catalase (1,000 U/ml) and superoxide dismutase (250 U/ml) attenuated the EPR signal, indicating that proximal formation of O2 free radicals was in part responsible for the signal. These results suggest that reactive O2 metabolites are mediators of OA-induced pulmonary edema in the isolated perfused rabbit lung.

Animals↗
PubMed1986

Effect of variation of retinal polyene side-chain length on formation and function of bacteriorhodopsin analogue pigments.

The effect of the length of the retinal polyene side chain on bacterioopsin pigment formation and function has been investigated with two series of synthetic retinal analogues. Cyclohexyl derivatives with polyene chains one carbon longer and one or more carbons shorter than retinal and linear polyenes with no ring have been synthesized and characterized. Compounds of six carbons or less in the polyene chain form pigments very poorly or not at all with bacterioopsin. Compounds containing at least seven carbons in the chain are found to form reasonably stable bacterioopsin pigments that show a small shift in absorbance on irradiation. However, photocycling and proton photorelease are not detected. The analogue with nine carbons in the polyene chain (one less than retinal) forms a stable pigment with an M-type intermediate but demonstrates reduced amounts of photocycling and light-activated proton release. The analogue with a polyene chain identical with that of retinal, but containing no ring, forms a pigment that shows both an efficient light-activated proton photocycle and release. The pigment containing the chromophore with the polyene chain one carbon longer than retinal is likewise fully active. We thus conclude that the length of the polyene chain must be at least 9 carbons for the formation of a stable pigment that photocycles and must be 10 carbons for both the photocycle and light-activated proton release to have a high quantum efficiency.

Bacteriorhodopsins↗
PubMed1986

The immunocytochemical demonstration of copper-zinc superoxide dismutase in the brain.

Copper-zinc superoxide dismutase (CuZn-SOD) has been localized in formalin-fixed, paraffin-embedded sections of both canine and rat brains. Staining with an immunoenzyme bridge sequence revealed CuZn-SOD in all regions of the brains examined. Specific sites of localization included cerebral cortical pyramidal cells, cerebellar Purkinje cells, neurons in 'subcortical nuclei', and oligodendrocytes throughout the brain. Similar sites of CuZn-SOD localization were identified in both species. These results are compared with reports by various investigators of SOD bioactivity in the brain.

Animals↗
PubMed1986

Mechanism of the protective activity of ICRF-187 against alloxan-induced diabetes in mice.

ICRF-187, (+)-1,2-bis(3,5-dioxopiperazine-1-yl)propane, has been shown to protect against alloxan diabetes (el-Hage et al., 1981). Since alloxan-induced pancreatic beta cell damage is thought to be mediated through the generation of highly reactive oxygen radicals by a metal catalyzed reaction involving both superoxide anion and hydrogen peroxide, in the present study the protective activity of ICRF-187 was compared with that of free radical scavengers, microsomal enzyme inhibitors and chelating agents. The free radical scavengers DMSO, vitamin E and WR2721 markedly reduced alloxan-induced hyperglycemia. ICRF-187 was found not to interact with superoxide anions, and there is no evidence to indicate that any of the known biological effects of ICRF-187 are mediated through free radical scavenging activity. SKF-525 and cimetidine, known inhibitors of drug metabolizing enzymes, also protected against the diabetogenic action of alloxan. Since it was found that ICRF-187 did not alter hexobarbital sleeping time, this compound must protect by a mechanism other than microsomal enzyme inhibition. Since the chelating agents EDTA and DETAPAC were found to protect against alloxan diabetes, ICRF-187 or its hydrolytic products, which are structurally similar to EDTA, could function as chelating agents. Transitional metals such as iron, zinc and copper were found to bind preferentially to a hydrolysis product of ICRF-187. Chelation of iron by ICRF-187 or its hydrolytic products could decrease in vivo formation of reactive oxygen radicals and provide a means for protecting against chronic anthracycline cardiotoxicity and alloxan diabetes.

Alloxan↗
PubMed1985

The protective role of copper-zinc superoxide dismutase against alloxan-induced diabetes: morphological aspects.

Copper-zinc superoxide dismutase is known to protect pancreatic beta-cell function from the damage of alloxan. A morphological investigation of copper-zinc superoxide dismutase prophylaxis against alloxan was undertaken in rats to investigate the mechanism of this protective action. Exogenous copper-zinc superoxide dismutase reproducibly protected the morphological features of pancreatic beta cells against damage by alloxan as determined by light microscopic immunostaining for insulin and by ultrastructural examination. By the same criteria, alpha and delta cells appeared unaffected by administration of a combination of alloxan and copper-zinc superoxide dismutase or either agent alone. Autoradiography after injection of 125I labelled copper-zinc superoxide dismutase into normal rats showed no evidence that the enzyme enters viable islet cells, suggesting an extracellular site of protection against alloxan.

Alloxan↗
PubMed1985

Immunolocalization of copper-zinc superoxide dismutase. II. Rat.

Copper-zinc superoxide dismutase (CuZn SOD) has been localized in formalin-fixed rat tissues. Staining with a modified immunoenzyme bridge technique using the avidin-biotin-peroxidase complex revealed abundant endogenous CuZn SOD in cells that function in transporting ions, either cellularly, as in the case of tracheal, bronchiolar, and colonic epithelial cells, gastric oxyntic cells, and cells lining the salivary ducts and proximal convoluted tubules in the nephron, or intracellularly, as exemplified by skeletal muscle and neurons. Additionally, the enzyme was consistently demonstrable in hepatocytes, endocrine cells of the islets of Langerhans, and the highly membranous oligodendrocytes in the central nervous system. Cellular processes that maintain high ionic gradients appear especially vulnerable to the superoxide anion, thus necessitating the presence of CuZn SOD to scavenge toxic free radicals of oxygen. Comparison of these observations with other immunocytochemical reports indicates that the cellular distribution of CuZn SOD varies between different species.

Animals↗