An ultrastructural study of the frog retinal rod photoreceptor membranes phagocyted by pigment epithelium cells after aldehyde fixations and organic solvents treatments

Borovjagin, V.L.; Ivanina, T.A.

doi:10.1016/0042-6989(73)90039-4

Cited by 6 publications

(1 citation statement)

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“…The effect of lipid extraction on the structure of membranes as revealed by electron microscopy has been studied in a variety of biological membranes (4,7,10,15,20,27). In some of those studies, a mixture of chloroform-methanol was used to achieve total lipid extraction.…”

Section: Discussionmentioning

confidence: 99%

The Ultrastructure of Lipid-Depleted Rod Photoreceptor Membranes

Nir

Hall

1974

The Journal of Cell Biology

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The structure of lipid-depleted retinal rod photoreceptor membranes was studied by means of electron microscopy. Aldehyde-fixed retinas were exhaustively extracted with acetone, chloroform-methanol, and acidified chloroform-methanol. The effect of prefixation on the extractability of lipids was evaluated by means of thin-layer chromatography and fatty acid analysis. Prefixation with glutaraldehyde rendered 38% of the phospholipids unextractable, while only 7% were unextractable after formaldehyde fixation. Embedding the retina in a lipid-retaining, polymerizable glutaraldehyde-urea mixture allows a comparison of the interaction of OsO4 with lipid-depleted membranes and rod disk membranes which contain all their lipids. A decrease in electron density and a deterioration of membrane fine structure in lipid-depleted tissue are correlated with the extent of lipid extraction. These observations are indicative of the role of the lipid bilayer in the ultrastructural visualization of membrane structure with OsO4. Negatively stained thin sections of extracted tissue reveal substructures in the lipid-depleted rod membranes. These substructures are probably the opsin molecules which are the major protein component of retinal rod photoreceptor membranes.The frog rod photoreceptor membrane consists of about 40% lipid and 60% protein (8,18). Of the protein, about 80-90% is opsin (5, 14). Thus the extraction of lipids from the photoreceptor membrane will result in a special situation in which the remaining components will consist mainly of a single protein--opsin. The ultrastructural investigation of lipid-depleted photoreceptors should therefore reveal the contribution of the opsin molecules to the structural organization of the rod photoreceptor membranes.In order to visualize the hydrophobic opsin molecules in lipid-depleted membranes, we have used a technique of embedment in a water-containing, polymerizable glutaraldehyde-urea mixture. This hydrophilic embedment has been successfully used in combination with ionic stains for the visualization of hydrophobic substructures in membranes (12,21,22,23).The retention of lipids by the glutaraldehydeurea-embedding mixture makes possible the evaluation of the role of OsO4 in the visualization of retinal membranes. During conventional preparation, which involves the use of organic solvents, there is a loss of lipids, even after treatment of the tissue with OsO, (16). Thus, only by comparing lipid-depleted sections with controls in which the membranes retain all their lipids, is it possible to evaluate accurately the role of lipids in the visualization of membranes by OsO~.It was previously reported that depletion of lipids from tissue prefixed with glutaraldehyde did

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Section: Discussionmentioning

confidence: 99%

The Ultrastructure of Lipid-Depleted Rod Photoreceptor Membranes

Nir

Hall

1974

The Journal of Cell Biology

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Structural features of the terminal loop region of frog retinal rod outer segment disk membranes: I. Organization of lipid components

1987

J of Comparative Neurology

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We have applied thin sectioning and freeze-fracture techniques to investigate the terminal loop structure of photoreceptive disks in frog retinal rod outer segments. Our studies of this region demonstrate a highly curved terminal loop bilayer that is continuous with both lamellar bilayers of the disk, and equivalent to them in dimensions and staining properties. Rhodopsin, however, appears to be excluded from this region of high curvature.

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