Evidence that ADH-stimulated intramembrane particle aggregates are transferred from cytoplasmic to luminal membranes in toad bladder epithelial cells

In freeze-fracture (FF) preparations of ADH-stimulated toad urinary bladder, characteristic intramembrane particle (IMP) aggregates are seen on the protoplasmic (P) face of the luminal membrane of granular cells while complementary parallel grooves are found on the exoplasmic (E) face. These IMP aggregates specifically correlate with ADH-induced changes in water permeability. Tubular cytoplasmic structures whose membranes contain IMP aggregates which look identical to the IMP aggregates in the luminal membrane have also been described in granular cells from unstimulated and ADH-stimulated bladders. The diameter of these cytoplasmic structures (0.11 +/- 0.004 micrometers) corresponds to that of tubular invaginations of the luminal membrane seen in thin sections of ADH-treated bladders (0.13 +/- 0.005 micrometers). Continuity between the membranes of these cytoplasmic structures (which are not granules) and the luminal membrane has been directly observed in favorable cross-fractures. In FF preparations of the luminal membrane, these apparent fusion events are seen as round, ice-filled invaginations (0.13 +/- 0.01 micrometer Diam), of which about half have the characteristic ADH-associated aggregates near the point of membrane fusion. They are less numerous than, but linearly related to, the number of aggregates counted in the same preparations (n = 78, r = 0.71, P less than 0.01). These observations suggest that the IMP aggregates seen in luminal membrane after ADH stimulation are transferred preformed by fusion of cytoplasmic with luminal membrane.     

Isolation and characterization of specialized regions of toad urinary bladder apical plasma membrane involved in the water permeability response to antidiuretic hormone

Summary Antidiuretic hormone (ADH) increases the apical (external facing) membrane water permeability of granular cells that line the toad urinary bladder. In response to ADH, cytoplasmic vesicles called aggrephores fuse with the apical plasma membrane and insert particle aggregates which are visualized by freeze-fracture electron microscopy. Aggrephores contain particle aggregates within their limiting membranes. It is generally accepted that particle aggregates are or are related to water channels. High rates of transepithelial water flow during ADH stimulation and subsequent hormone removal decrease water permeability and cause the endocytosis of apical membrane and aggrephores which retrieve particle aggregates. We loaded the particle aggregate-rich endocytic vesicles with horseradish peroxidase (HRP) during ADH stimulation and removal. Epithelial cells were isolated and homogenized, and a subcellular fraction was enriched for sequestered HRP obtained. The HRP-enriched membrane fraction was subjected to a density shifting maneuver (Courtoy et al.,J. Cell Biol. 98:870, 1984), which yielded a purified membrane fraction containing vesicles with entrapped HRP. The density shifted vesicles were composed of approximately 20 proteins including prominent species of 55, 17 and 7 kD. Proteins of these molecular weights appear on the apical surface of ADH-stimulated bladders, but not the apical surface of control bladders. Therefore, we believe these density shifted vesicles contain proteins involved in the ADH-stimulated water permeability response, possibly components of particle aggregates and/or water channels.     

Role of osmotic forces in exocytosis: studies of ADH-induced fusion in toad urinary bladder

Antidiuretic hormone (ADH) treatment of toad urinary bladder activates an exocytotic-like process by which intramembrane particle aggregates are transferred from membranes of elongated cytoplasmic tubules to the luminal-facing plasma membrane. We find that the number of these ADH- induced fusion events, and the number of aggregates appearing in the luminal membrane, are reduced when the luminal bathing medium is made hyperosmotic. As an apparent consequence of the inhibition of their fusion with the luminal membrane, the elongated cytoplasmic tubules become enormously swollen into large, rounded vesicles. These results are consistent with the view that osmotic forces are essential to the basic mechanism of exocytosis.     

Retention of antidiuretic hormone-induced particle aggregates by luminal membranes separated from toad bladder epithelial cells

Aggregates of intramembrane particles appear in the luminal membranes of renal collecting duct and amphibian bladder cells after stimulation by antidiuretic hormone (ADH). We undertook this freeze-fracture study to determine whether particle aggregates, once in place, remain in the luminal membrane of the amphibian bladder after the membrane is physically separated from the rest of the cell. We found that the aggregates do remain in high yield in isolated membranes stabilized with a bifunctional imidoester (DTBP) followed by fixation with glutaraldehyde, or unfixed but stabilized with DTBP. These findings support the view that the particles are intrinsic membrane components and that their organization in the form of aggregates does not depend on the presence of the intact cell. In addition, the availability of isolated membranes containing particle aggregates provides a starting point for the isolation of the water-conducting proteins.     

The water permeability of toad urinary bladder. I. Permeability of barriers in series with the luminal membrane

Antidiuretic hormone (ADH) induces a large increase in the water permeability of the luminal membrane of toad urinary bladder. Measured values of the diffusional water permeability coefficient, Pd(w), are spuriously low, however, because of barriers within the tissue, in series with the luminal membrane, that impede diffusion. We have now determined the water permeability coefficient of these series barriers in fully stretched bladders and find it to be approximately 6.3 X 10(- 4) cm/s. This is equivalent to an unstirred aqueous layer of approximately 400 microns. On the other hand, the permeability coefficient of the bladder to a lipophilic molecule, hexanol, is approximately 9.0 X 10(-4) cm/s. This is equivalent to an unstirred aqueous layer of only 100 microns. The much smaller hindrance to hexanol diffusion than to water diffusion by the series barriers implies a lipophilic component to the barriers. We suggest that membrane-enclosed organelles may be so tightly packed within the cytoplasm of granular epithelial cells that they offer a substantial impediment to diffusion of water through the cell. Alternatively, the lipophilic component of the barrier could be the plasma membranes of the basal cells, which cover most of the basement membrane and thereby may restrict water transport to the narrow spaces between basal and granular cells.     

Evidence that the heads of ADH-sensitive aggrephores are clathrin-coated vesicles: implications for aggrephore structure and function

Antidiuretic hormone (ADH) induces the fusion of long tubular organelles (aggrephores) with the luminal membrane of the receptor cell, and the delivery of particle aggregates to the membrane. Water flow is believed to take place through the particles. Nothing is known about the origin of the particle aggregates, their incorporation into the aggrephores, or the possible relationship of the aggrephores to the vesicular traffic that takes place in the epithelial cell. In the present studies of the ADH-sensitive epithelial cells of the toad urinary bladder, we have found that the spherical heads of the aggrephores appear to be clathrin-coated vesicles. We propose that vesicles originating from sites such as the Golgi or the luminal membrane may be engaged in aggrephore assembly, the resupply of particle aggregates to the aggrephores, and/or the removal of aggregates, and that the aggrephores may be central points in the pattern of vesicular traffic in the cell.     

Vasopressin-induced intramembrane particle aggregates. A dose-response relationship in the isolated cortical collecting duct of the rabbit kidney

The water permeability of collecting ducts is greatly increased by the antidiuretic hormone, vasopressin (VP). Freeze-fracture studies were carried out to test if this permeability increase is associated with the appearance of intramembrane particle (IMP) aggregates and whether increased doses of VP lead to an increase in the number and size of particle aggregates in the luminal membrane of principal cells in the isolated cortical collecting duct. Unstimulated cells expressed 17 +/- 6.5 particle aggregates per 100 microns 2. Stimulation with VP at concentrations of 20 or 200 microU/ml increased the number of particle aggregates significantly to 129 +/- 15.8 and 324 +/- 45.8, respectively. The size of the particle aggregates increased from 0.0012 microns 2 under control conditions to 0.025 microns 2 at 20 microU/ml VP and to 0.063 microns 2 at 200 microU/ml VP. In addition, the total area occupied by the IMP increased from 0.02 microns 2/100 microns 2 (controls) to 3.17% and 20.38% (after 20 and 200 microU ADH/ml, respectively). Particle aggregates were also observed in the luminal plasma membrane of isolated collecting ducts fixed immediately after dissection, resembling the in vivo status. These results demonstrate that a dose-dependent relationship exists between the concentration of the applied VP and the number of particle aggregates, as well as the size of the aggregates. Cytoplasmic tubular vesicles in fusion with the apical membrane were observed.     

Localization of barriers to water flow in toad urinary bladder

Although it is well accepted that vasopressin (ADH) increases the permeability to water of the toad bladder granular cell's luminal membrane, recent studies have suggested that regulation also takes place at an additional "postluminal" site within the epithelial granular cell. These studies are based upon the observation that a number of experimental maneuvers can alter tissue permeability to water, but do not change the number of particle aggregates observed on the protoplasmic face of the granular cell's luminal membrane with freeze-fracture electron microscopy. These aggregates are believed by many investigators to mediate the transport of water across the luminal membrane. The dissociation between permeability and aggregate frequency described above has been variously interpreted as the consequence of changes in the permeability of the aggregates themselves, or of changes in the permeability of a "postluminal" barrier that is functionally in series with the luminal membrane. We attempted to distinguish between these 2 possibilities by studying paired toad bladders during 3 protocols that alter vasopressin-stimulated water flow across the intact tissue without altering aggregate frequency. Estimates of the permeability of postluminal barriers were obtained by exposing the luminal surface to amphotericin B, an antibiotic that forms water-permeant channels in the luminal membrane. Of the 3 protocols, only diminishing bladder filling volume decreased the water flow elicited by luminal amphotericin B, suggesting that only that protocol indeed decreased the permeability of some postluminal barrier. The other 2 protocols, increasing PCO2 and repeatedly stimulating the bladder with vasopressin, did not alter amphotericin B-elicited flow, suggesting that postluminal barriers were not altered by these 2 protocols.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of ADH-induced intramembrane particle aggregates (author's transl)

In certain epithelial tissues, water permeability is markedly increased by antidiuretic hormone. This so-called hydrosmotic effect has been shown to be mediated by 3'-5' cyclic adenosine monophosphate, which, in turn, alters the permeability o the luminal membrane of receptor cells. This review deals wity ultrastructural alterations occurring in the membrane, as observed with freeze-fracture electron microscopy. Basically, these alterations consist of organized particle aggregates which appear in the apical membrane. In all experimental conditions, similar aggregates can be observed in the membrane of cytoplasmic vesicles. ADH stimulation triggers the fusion of these vesicles with the apical membrane resulting in the concomitant transfer of particle aggregates. It has been shown, in a wide range of experimental conditions, that both number and total area of the aggregates are directly proportional to the water permeability of the tissue. It is generally assumed that particle aggregates contain transmembrane channels that are selectively to water.     

Role of nonacidic endosomes in recycling of ADH-sensitive water channel structures.

Toad urinary bladder epithelial cells respond to the hormone ADH by increasing the water permeability of their luminal membrane. This action is mediated by insertion into the apical membrane of specific water channels. In the absence of ADH these channels appear to be present in tubular cytoplasmic vesicles as morphologically distinctive intramembrane structures called particle aggregates. ADH induces these vesicles to fuse with the apical membrane, transferring their aggregate-water channels into the apical membrane. When ADH stimulation is removed (ADH reversal), aggregates and fluid-phase markers from the mucosal bath appear in water-permeable vesicles in the cytoplasm. We have examined the fate of fluid-phase markers and aggregates with time after ADH reversal. Although the fluid-phase markers horseradish peroxidase and colloidal gold are initially found predominantly in tubular vesicles near the apical surface, by 30 min the markers were found in perinuclear multivesicular bodies (MVBs) of heterogeneous size and shape. These MVBs appear to be nonacidic since they fail to accumulate DAMP. Acid phosphatase (AcPase) was undetectable in these structures. After 60 min, labeled MVBs tended to be smaller, and some of these structures displayed DAMP accumulation and AcPase activity. By evaluation of uncleaned replicas it was possible to localize recycled aggregate-water channels with respect to internalized fluid-phase markers. Thirty minutes after retrieval from the apical surface in tubular vesicles, aggregates could be localized to both the central body and tubular projections of labeled MVBs. At 60 min following reversal, most MVBs had a reduced number of aggregates compared with 30 min, and compact structures could be identified that contained markers but no detectable aggregates. These observations show that aggregates and fluid-phase markers enter a nonacidic endosomal compartment with an MVB morphology following ADH reversal. At extended times following reversal, labeled MVBS having lysosomal characteristics and labeled MVBs having no detectable aggregates can be found, suggesting that aggregates are sorted or degraded prior to this stage.     

Fusion images and intramembrane particle aggregates during the action of antidiuretic hormone

Summary Antidiuretic hormone (ADH) causes the appearance of water-conducting particle aggregates in the luminal membrane of receptor cells in amphibian bladder and skin, and in the mammalian collecting duct. The aggregates originate from cytoplasmic tubules that fuse with the luminal membrane during ADH stimulation. We have studied the process of fusion and the structure of the particle aggregates by a rapid-freeze technique that renders chemical fixation and glycerol protection unnecessary. Our findings differ in some important respects from previously published work. Aggregate particles, in our study, partition equally between the external (EF) and protoplasmic (PF) membrane leaflets, rather than remaining in the protoplasmic leaflet exlcusively. By including the entire population of fusion images in our survey, we have found that aggregate delivery in ADH-treated cells proceeds preferentially from small fusion images whose diameter is significantly less than the 0.12 m characteristic of the carrier tubules themselves. We have also found that, even in unstimulated preparations, fusion images are numerous, being mostly of small diameter. ADH stimulation produces a moderate increase in the number of fusion images and a significant increase in fusion-image diameter. These findings indicate that the individual particles are mobile within the membrane, lacking interparticle linkage. In addition, contact of cytoplasmic tubules with the luminal membrane may take place even in the absence of ADH, producing small fusion images which are not associated with aggregate delivery to the luminal membrane.Faculty Scholar, Josiah Macey Jr. Foundation     

Visualization of endocytosed markers in freeze-fracture studies of toad urinary bladder

Antidiuretic hormone (ADH) stimulation increases the apical membrane water permeability of granular cells in toad urinary bladder. This response correlates closely with the fusion of tubular cytoplasmic vesicles with the membrane and delivery of intramembrane particle (IMP) aggregates from the tubules (aggrephores) to the apical membrane. These aggregates are believed to be associated with the channels responsible for the water permeability increase. Removal of ADH triggers apical membrane endocytosis and disappearance of aggregates from the apical membrane. However, it has been unclear whether aggregate disappearance is due to disassembly of aggregates within the apical membrane or to their endocytic retrieval as intact structures. Using colloidal gold and horseradish peroxidase to follow endocytosis from the apical surface after ADH removal, we have directly observed in cross-fractured bladder cells the intramembrane structure of intracellular vesicles that contain these fluid-phase markers. Under these conditions, intact aggregates can be identified in the membrane of tubular endocytosed vesicles. This directly demonstrates that conditions which lower apical membrane water permeability cause the tubular aggrephores to "shuttle" intact aggregates from the apical membrane back into the cytoplasm. An additional population of vesicles with tracer are found which are spherical and display structural features of the apical membrane, as well as occasional aggregates. These vesicles may be responsible for retrieval of aggregates from the surface apical membrane.     

Freeze-Fracture Analysis of Plasma Membranes of CHO Cells Stably Expressing Aquaporins 1-5

Several studies suggest that aquaporin water channels can be identified in membranes by freeze-fracture electron microscopy. For this report, Chinese Hamster ovary cells were stably transfected with cDNAs encoding aquaporins 1–5. Measurement of the osmotic water permeability of the cells confirmed that functional protein was expressed and delivered to the plasma membrane. By freeze-fracture electron microscopy, a 20% increase in intramembrane particle (IMP) density was found in plasma membranes of cells expressing AQP2, 3 and 5, and a 100% increase was measured in AQP1-expressing cells, when compared to mock-transfected cells. On membranes of cells expressing AQP4, large aggregates of IMPs were organized into orthogonal arrays, which occupied 10–20% of the membrane surface. IMP aggregates were never seen in AQP2-transfected cells. Hexagonally packed IMP clusters were detected in ∼5% of the membranes from AQP3-expressing cells. Particle size-distribution analysis of rotary shadowed IMPs showed a significant shift from 13.5 (control cells) to 8.5 nm or less in AQP-expressing cells; size distribution analysis of unidirectionally shadowed IMPs also showed a significant change when compared to control. Some IMPs in AQP expressing cells had features consistent with the idea that aquaporins are assembled as tetramers. The results demonstrate that in transfected CHO cells, AQP transfection modifies the general appearance and number of IMPs on the plasma membrane, and show that only AQP4 assembles into well-defined IMP arrays. Received: 17 March 1998/Revised: 19 June 1998     

Rhombic particle arrays in gill epithelium of a mollusc, Aplysia californica.

Gill epithelia from adult and juvenile Aplysia were examined by conventional thin section and freeze-fracture methods. Freeze-fracture replicas of adult gill epithelium revealed septate and gap junctions, which served as membrane markers for the epithelial cells. In these same cell membranes, non-junctional rhombic arrays of intramembranous particles were observed on prominent ridges on the membrane P fracture face of some epithelial cells. In thin sections of adult epithelium, nerve terminals were observed abutting the lateral plasma membranes near the basal lamina of some epithelial cells. Correlative areas of plasma membrane in freeze-fracture replicas showed a close association between rhombic particle arrays and abutting nerve terminals. In thin sections of juvenile Aplysia, nerve terminals abutting the epithelial cells were not recognizable, and rhombic arrays were not observed in freeze-fracture replicas. This suggested that a developmental association existed between the appearance of rhombic arrays in adult epithelia and their innervation. It is not known with certainty if, in invertebrates, rhombic arrays are an essential structural entity of all innervated cell membranes; however, in the cells thus far studied, there appears to be an associative condition. In the case of the gill epithelium of Aplysia, rhombic arrays are located in the same vicinity as the abutting nerve terminals. Similar arrays of intramembranous particles have been observed in myoneural postjunctional complexes of other invertebrates and have been interpreted to be the morphological expression of neurotransmitter receptors. An analogous explanation is put forth, namely that rhombic arrays may represent the structural correlates of neurotransmitter receptors and/or ionic channels in innervated membranes of invertebrates.     

Regulation of luminal membrane water permeability by water flow in toad urinary bladder

Intramembranous particle aggregates (presumed sites for water flow) which appear in the luminal membrane consequent to ADH treatment are derived from cytoplasmic membrane structures (now termed "aggrephores") which fuse with the luminal membrane. We have previously shown that bladders stimulated in the absence of an osmotic gradient have about twice as many aggregates and about three times as many sites of aggrephore fusion as bladders stimulated with ADH in the presence of a 175 milliosmolal gradient. The present studies show that the frequency of fused aggrephores and luminal membrane aggregates can be modified as a consequence of alterations in transmembrane water flow initiated by changing the transbladder osmotic gradient during hormone stimulation. Bladders treated with ADH for 1 hr without a gradient and then for 1 hr with a gradient had approximately 1/3 as many aggregates and fusion sites as paired bladders treated for 2 hr without a gradient. Conversely, bladders treated with ADH for 1 hr with a gradient and then for 1 hr without a gradient had approximately 2x as many aggregates and fusion sites as bladders treated for 2 hr with a gradient. In other experiments we demonstrate that the time course of hormone washout is greatly accelerated if carried out in the presence of an osmotic gradient. In paired bladders that were first stimulated with ADH for 30 min in the absence of a gradient, aggregates and fusion sites as well as osmotic water permeability determined in fixed bladders, persisted at near maximum levels for 15 min of washout in the absence of a gradient.(ABSTRACT TRUNCATED AT 250 WORDS)     

Orthogonal arrays in normal and injured respiratory airway epithelium

Orthogonal arrays are found on plasma membranes of glial cells, in the central nervous system, on muscle plasma membranes at neuromuscular junctions, and on a variety of epithelial cells. These structures have been correlated with ion flux. With the aid of freeze fracture technique, orthogonal particle arrays were found on plasma membranes on airway epithelial cells of rats and hamsters. They have been found in abundance at the base of secretory cells throughout normal airway epithelium. These structures were found to increase in number during regeneration in response to injury and they were found in great numbers on plasma membranes of all airway cells in response to acute and chronic NO2 exposure. The lateral and basal plasma membranes of the respiratory epithelium are a new source for studying orthogonal arrays. The normal number and distribution of these arrays can be perturbed in response to mechanical and chemical injury.     

Similarity of junctions between plasma membranes and endoplasmic reticulum in muscle and neurons

The structure of membranes at junctions between the plasma membrane and underlying cisterns of endoplasmic reticulum in amphioxus muscle and mouse cerebellar neurons was studied using the freeze-fracture technique. In amphioxus muscle, subsurface cisterns of sarcoplasmic reticulum form junctions with the surface membrane at the level of the sarcomere I bands. On the protoplasmic leaflet of the sarcolemma overlying these junctions were aggregates of large particles. On the protoplasmic leaflet of the membranes of cerebellar basket, stellate and Purkinie cells there were similar aggregates of large particles. In both tissues, the corresponding external membrane halves had arrays of pits apparently complementary to the aggregates of large particles. Cross fractures through junctions showed that the particle aggregates in neuronal and muscle membranes were consistently located over intracellular cisterns closely applied to the plasma membrane. Thus, a similar plasma membrane specialization is found at subsurface cisterns in mammalian neurons and amphioxus muscle. This similarity supports the hypothesis that subsurface cisterns in neurons, like those in muscle, couple some intracellular activity to the electrical activity of the plasma membrane.     

Reorganization of the Golgi apparatus of epithelial cells in the frog bladder in stimulation of water transport by antidiuretic hormone

Ultrastructural changes of Golgi apparatus of frog urinary granular cells at antidiuretic hormone (ADH) stimulation of water transport were studied. During a short-time ADH action (5 min) the fragmentation of the complex on single dictyosomes and dilution of certain cisternae is discovered. A conclusion is made that the granular cell giant vacuoles may originate from the Golgi cisternae. It is suggested that the microtubules may be involved in the translocation of dictyosomes and migration of formed vacuoles. The quantity of microtubules increases during ADH action very significantly. Moreover, the involvement of the Golgi apparatus is shown in the maintenance of the cell membrane balance due to budding of tubular structures from transcisternae and shuttling between luminal and vacuolar membranes.     

76 and 14 kDa polypeptides, two major components released from amphibian urinary bladder epithelium. Localization and potential role

Several experimental conditions such as antidiuretic hormone (ADH) challenge, apical treatment with phorbol myristate acetate (PMA), and mechanical stretching of the tissue are known to increase the insertion of intramembrane particle aggregates and/or granule exocytosis at the apical border of epithelial cells of amphibian urinary bladders. A constant release of 2 peptides of 76 and 14 kDa apparent molecular mass, respectively, was associated with these treatments. The localization of these 2 polypeptides was assessed by immunofluorescence and electron microscopy immunocytochemistry using fluorescent, peroxidase, and colloidal gold probes. The 76 kDa polypeptide appeared to be associated with the cell coat and with the granule content which is released at the apical cell surface. The 14 kDa peptide was also found in the cell coat, and postembedding immunocytochemistry indicates its presence in cytoplasmic subapical vesicles (aggrephores and/or granules). The migration of these 76 and 14 kDa polypeptides in SDS-polyacrylamide gel electrophoresis was modified neither by a treatment at 90 degrees C, nor by the presence or absence of calcium in the medium. Treatment with EGTA did not modify the fluorescence emission of the two peptides and, consequently, they are probably not among the major calcium binding proteins. The addition to the mucosal medium of the stretch extract or of antibodies raised against the 76 and 14 kDa peptides did not modify ADH-induced water permeability. However, a significant decrease of the hydrosmotic response to ADH occurred in subsequent stimulation-washout cycles when the anti-14 kDa peptide antiserum was applied to the mucosal bath. When the bladders were incubated with a stretch extract, we observed a slight alteration of the short-circuit current (Isc), an increase of the basal Na+ transport, and a decrease of the maximal Isc in response to ADH. The 76 kDa protein, released in the apical medium, could play a protective role in the cellular plasma membrane and could participate in the formation of the thick cell coat lining the apical membrane of the granular cells. The 14 kDa protein might be one of the proteins associated with the aggregates, but further studies will be necessary to clarify its exact role in the ADH-induced permeability modifications observed in amphibian urinary bladders.     

FINE STRUCTURE OF THE OCTOPUS RETINA

The fine structure of the visual and the supporting cells and of the blood capillaries in the octopus retina is described. Lamellated structures contained in the proximal segment of the visual cell consist of compact arrays of dense membranes each of which is quintuple-layered and divides at its margins into two thinner sheets or membranes which are connected directly with the agranular or granular endoplasmic reticulum. Proximal to the deeper extremities of the rhabdomeres, the lateral plasma membranes of two adjoining visual cells contact each other forming a quintuple-layered compound membrane, which results in occlusion of the intercellular space. The central layer of the compound membrane is of high density, so that the membrane, as a whole, appears to be a single thick layer at low magnifications. The supporting cells are connected with the neighboring visual cells by two types of junctions. Long slender processes extend from the supporting cells to the surface of the retina through narrow spaces among the distal segments of the visual cells. The capillary endothelial cells are characterized by luminal surfaces irregularly contoured and by lateral surfaces elaborately interdigitated. The functional significance of the close contact between adjoining visual cells is discussed.