Vergleichende Untersuchungen an Amoebozyten und Blasenzellen von Cepaea nemoralis L. (Gastropoda,Stylommatophora)

Summary The pecten oculi of the sparrow consists of capillaries, pigment cells and a superficial membrane. Because of the loose structure of the first two components broad intercellular spaces occur in the pecten. The capillary wall consists of endothelial cells and a perivascular membrane. The bodies of the endothelial cells are flattened, while the plasmalemma of both their surfaces (basal and luminal) is strongly folded and forms numerous microfolds with an average thickness of 700 Å. The height of the inner microfolds is 1.4–1.8 m, the outer microfolds measure 1.3–1.6 m. They lie densely packed side by side and are separated by recesses of the capillary lumen ca. 500 Å wide. Due to this the surface of the endothelial cell is increased by approximately 20-fold. The adjoining endothelial cells abut or overlap with margins, and are joined by the zonulae adherentes. Pigment cells form numerous processes and microvilli. Some rest on the capillary walls, while others penetrate the superficial membrane of the pecten or fill the intercellular spaces.     

Fine structure of the conus papillaris in the bobtail goanna (Tiliqua rugosa)

The structure of the conus papillaris in an Australian lizard, the bobtail goanna (Tiliqua rugosa) was investigated by light and electron microscopy. In this strongly diurnal species, the conus papillaris consists of a heavily vascularized and pigmented, finger-like structure about 1 mm in diameter and 3-4 mm in length. It is situated over the optic nerve head and projects into the vitreous chamber. Within the conus are numerous capillaries and larger blood vessels, melanocytes and occasional mast cells. Many of the capillaries display prominent luminal and abluminal microfolds. Other capillaries show no microfolds while still others display an intermediate number of microfolds. The larger blood vessels are usually indistinguishable as to being either arterioles or venules. The endothelial cells of all blood vessels show a population of cytoplasmic granules. The melanocytes are large pleomorphic cells usually rich in microfilaments. Unmyelinated nerve processes are plentiful within the conus and the Schwann cells enclosing these nerve fibres are occasionally seen to be pigmented. The morphology of the conus papillaris indicates a heavy involvement in the transport of materials. It is considered to be homologous to the pecten oculi of the avian eye; to the falciform process of the teleost eye; to the supraretinal vessels of amphibians and to the intraretinal vessels of the mammalian eye.     

Electron microscopic observations on the pecten of the great blue heron (Ardea herodias).

The pecten oculi of the great blue heron (Ardea herodias) has been examined by both light and electron microscopy. In this species the pecten is large and of the pleated type. It consists of 14-15 acordion folds that are joined apically by a more heavily pigmented bridge of tissue which holds the pecten in a fan-like shape widest at its base. As in other species it is situated over the optic nerve head and projects out into the vitreous. Within each fold are numerous capillaries, larger supply and drainage vessels and many melanocytes. The capillaries are extremely specialized vessels which display extensive microfolds on both their luminal and abluminal borders. The endothelial cell bodies are extremely thin with most organelles present in a paranuclear location. The capillaries are surrounded by thick fibrillar basal laminae which are felt to be structurally useful. Pericytes are a common feature of these capillaries. The numerous pleomorphic melanocytes which form an incomplete sheath around the capillaries and other blood vessels are also felt to be important in structural support of the pecten. The morphology of the pecten of the great blue heron is indicative of a heavy involvement in the transport of materials.     

Electron microscopic studies of coated membranes in two types of gill epithelial cells of lamprey

Summary Coated membranes in two types of gill epithelial cell of adult lamprey, Lampetra japonica, were studied by electron microscopy. The type 3 gill epithelial cells possess well-developed microvilli or microfolds, apical vesicles and abundant mitochondria. The cytoplasmic surface of the microvillous plasma membrane is covered by a coat of regularly spaced particles with a center-to-center distance of about 15 nm. Each particle consists of a bulbous free end, about 10 nm in diameter, and a connecting piece, about 5 nm long. Apical vesicles are covered by a surface coat which consists of fine filamentous material but lack any special coating on their cytoplasmic surface.The type 4 cells (chloride cells) are characterized by apical vesicles, abundant mitochondria and cytoplasmic tubules. These tubules possess a coat on their luminal surface which consists of spirally wound parallel rows of electron-dense materials. The rows are about 16 nm apart and wound at a pitch of about 45°. The cytoplasmic surface of these tubules does not display a special coat. These coated membranes are assumed to be the sites of active ion transport across the plasma membrane. In particular, particles in type 3 cells and linear coat materials in chloride cells may be either loci of transport enzymes or energy generating systems. Apical vesicles lack any coating on their cytoplasmic surface but a fine filamentous coat is present on their luminal surface. They contain intraluminal vesicles and are continuous with apical ends of cytoplasmic tubules.     

Constituents of connective tissue around the capillary of chick pecten oculi

The constituents of the connective tissues around the capillary of the chick pecten oculi were examined electron microscopically by HCl-collagenase and HCl-elastase methods. The basal lamina like membrane below the endothelial cell of the pecten capillary was digested by collagenases I, II and IV and elastase, and may be a false basal lamina. The basal lamina of cells with pigment granules which surround the capillary was digested by collagenase IV and elastase, and contained type IV collagen. Fibrils between the basal lamina like membrane of the pecten capillary endothelium and the basal lamina of the cells with pigment granules were digested by collagenases I, II and IV, and elastase. Thus, these fibrils are composed of many kinds of collagen. Elastase may be responsible for the breakdown of most collagens as well as elastin.     

Functional involvement of P-glycoprotein in blood-brain barrier.

P-glycoprotein, an active efflux pump of antitumor agents in multidrug-resistant tumor cells, exists in various normal tissues, including brain capillaries. To study the physiological function of P-glycoprotein expressed in brain capillary endothelium, we established nine mouse brain capillary endothelial cell (MBEC) lines and examined the transport of antitumor agents across the monolayer of MBEC epithelia. In the MBECs, the activities of alkaline phosphatase and gamma-glutamyl transpeptidase, specific markers for brain capillary endothelial cells, were about three times higher than those in other cells including human umbilical vein endothelial cells. By immunoblot analysis, P-glycoprotein was detected in all of the nine MBEC clones. The P-glycoprotein expressed in MBECs specifically bound [125I]iodoaryl azidoprazosin as that in multidrug-resistant cells, and efflux of vincristine was observed in the MBECs. When MBECs were grown on a porous filter membrane, they formed a monolayer of epithelium. By immunoelectron microscopic analysis, P-glycoprotein in MBEC epithelia was shown to be localized to the apical surface of the cells. Moreover, the unidirectional transepithelial transport of vincristine from basal side to apical side was demonstrated in vitro. These observations indicate that P-glycoprotein in brain capillary endothelium prevents vincristine from entering the central nervous system and thus may be one of the functional components of the blood-brain barrier.     

Post-natal maturation of the retina in the albino rat. I. The pigment epithelium

The Authors studied the postnatal development of the retinal pigment epithelium in the albino rat, in order to elucidate its morphological and functional evolution, correlated to the numerous functional roles played in Vertebrates (Scheme 1). At birth, epithelial cells show few cytoplasmic organules and the apical surface provided of small depressions. From the third to the fifth postnatal day the first apical microfolds surround the depressions. From the seventh to the ninth day inner segments develop, whilst the apical surface of the epithelial cells is covered by many finger-like microfolds. During the eleventh postnatal day the buds of the outer segments and many lamellar microfolds can be demonstrated. During the sixteenth day the retina reaches its adult morphology. It is therefore well-evident that birth, similarly to many other Vertebrates, is not the last step, but only a moment, in the development of the retina: this process is completed only during postnatal life, when environmental light is able to stimulate every ocular structure.     

Enzymhistochemische und feinstrukturelle Beobachtungen am Pecten oculi von Taube (Columba livia) und Lachmöwe (Larus ridibundus)

Zusammenfassung Im Endothel der Kapillaren des Pecten der Augen von Haustaube und Lachmöwe (Larus ridibundus) lassen sich lichtmikroskopisch alkalische und saure Phosphatase sowie Aminopeptidase nachweisen. Bernsteinsäuredehydrogenase und verschiedene Esterasen fehlen dagegen.Im Elektronenmikroskop ist das Kapillarendothel des Pecten der Lachmöwe durch dichte apikale und basale Mikrovillisäume, Mitochondrienansammlungen, zahlreiche polymorphe elektronendichte Granula, in reichem Maße vorhandene Ribosomen und multivesikuläre Körper gekennzeichnet. Zwischen Basallamina und Endothelzelle befindet sich ein weiter mit Fibrillen gefüllter Raum. Die Pigmentzellen des Pecten bilden mitochondrienhaltige Füßchen aus, die sich der Basallamina der Kapillaren anlegen. Die Oberfläche des Pecten wird von einer weiteren Basallamina gebildet; auch ihr lagern sich Füßchen der Pigmentzellen an. Die vorliegenden histochemischen und feinstrukturellen Befunde und Angaben aus der Literatur lassen vermuten, daß das Kapillarendothel im Pecten besondere Anpassungen an verstärkt ablaufende Transportvorgänge aufweist.

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Enzyme histochemical and fine-structural observations on the pecten oculi of Birds

Summary The endothelium of the capillaries in the pecten of the eyes of domestic pigeons and blackheaded gulls (Larus ridibundus) gives a positive reaction for alkaline and acid phosphatase, as well as for aminopeptidase. Negative results have been obtained for succinic-acid dehydrogenase and various esterases.In the electron microscope the capillary endothelium in the pecten of the blackheaded gull is characterized by densely packed apical and basal microvilli, accumulations of mitochondria and polymorphic electron-dense granules, numerous ribosomes and relatively common occurrence of multivesicular bodies. The endothelial cells are separated from their basal lamina by wide spaces which are filled by fibrils. The pigment cells of the pecten extend mitochondria-containing processes towards the basal lamina of the capillaries. The surface of the pecten is bound by a further basal lamina, which is underlain by other projections of the pigment cells. The present histochemical and fine-structural findings and evidence derived from the literature suggest that the capillary endothelium of the pecten shows particular adaptations in respect of transport mechanisms.

Mit dankenswerter Unterstützung durch die Deutsche Forschungsgemeinschaft.     

Multiple pathways of exocytosis, endocytosis, and membrane recycling: validation of a Golgi route

A number of pathways for intracellular membrane traffic have been detected in various cell types. The major established routes are: 1) the lysosomal pathway, which is the major route utilized in phagocytic and cultured cells; 2) the transcellular route, which represents the major type of traffic in nonfenestrated, capillary endothelial cells and which also appears to be the preferred route for the transport of immunoglobulins (intact) across cells; 3) the exocytosis pathway, utilized in secretory cells for discharge of secretory products, and which is also believed to be used for delivery of intrinsic membrane glycoproteins; 4) the plasmalemma to Golgi route, also highly developed in secretory cells, which is believed to be utilized for the recycling of secretory granule membranes; and 5) the biosynthetic pathways for transport of secretory products, lysosomal enzymes, and membrane proteins from the endoplasmic reticulum to the Golgi complex and for transport of lysosomal enzymes from the Golgi complex to lysosomes. It has become clear that cells repeatedly reutilize or recycle the membranes used in these various transport operations. Clathrin-coated vesicles have been found to be involved in transport along all these routes, which suggests that there are multiple populations of coated vesicles with different transport functions in every cell. It has become clear that the Golgi complex is the site where the membrane and product traffic converges and is sorted and directed to its correct destinations. The validation of a transport route from the cell surface to the Golgi complex raises the possibility that bound ligands and membrane constituents could be modified or repaired in transit during recycling through the Golgi complex, which is a biosynthetic compartment.     

Involvement of OCTN2 and B0,+ in the transport of carnitine through an in vitro model of the blood-brain barrier

Carnitine is known to accumulate in brain, therefore transport of carnitine through the blood-brain barrier was studied in an in vitro system using bovine brain capillary endothelial cells (BBCEC) grown on filter inserts in a co-culture system with glial cells. Long-term exposure of BBCEC to carnitine resulted in a high accumulation of long-chain acyl carnitines, which decreased dramatically upon removal of carnitine. Kinetic analysis of carnitine accumulation indicated a possibility of functioning of more than one transporter. BBCEC were incubated in the presence of substrates and inhibitors of known carnitine transporters added from either apical or basolateral side. Inhibition by replacement of sodium and expression of OCTN2 (RT-PCR) were in agreement with earlier reports on the functioning of OCTN2 in apical membrane. For the first time, functioning of OCTN2 was demonstrated in the basolateral membrane, as well as functioning in both membranes of a low affinity carnitine transporter B(0,+). Expression of B(0,+) in BBCEC was confirmed by RT-PCR. These results suggest that OCTN2 and B(0,+) could be involved in carnitine transport in both the apical and basolateral membrane.     

Transport of ions across the blood-brain barrier

Capillaries in the brain are formed by a uniquely specialized endothelial cell that regulates the movement of substances between blood and brain. Although they provide an impermeable barrier to some solutes, brain capillary endothelial cells facilitate the transcapillary exchange of others. In addition, they contain specific enzymes that contribute to a metabolic blood-brain barrier by limiting the movement of compounds such as neurotransmitters across the capillary wall. Studies of sodium and potassium transport by brain capillaries indicate that the endothelial cell contains distinct types of ion transport systems on the two sides of the capillary wall, i.e., the luminal and antiluminal membranes of the endothelial cell. As a result, specific solutes can be pumped across the capillary against an electrochemical gradient. These transport systems are likely to play a role in the active secretion of fluid from blood to brain and in maintaining a constant concentration of ions in the brain's interstitial fluid. In this way, the brain capillary endothelium is structurally and functionally related to an epithelium.     

The MAL proteolipid is necessary for the overall apical delivery of membrane proteins in the polarized epithelial Madin-Darby canine kidney and fischer rat thyroid cell lines

The MAL proteolipid has been recently demonstrated as being necessary for correct apical sorting of the transmembrane influenza virus hemagglutinin (HA) in Madin-Darby canine kidney (MDCK) cells. The fact that, in contrast to MDCK cells, Fischer rat thyroid (FRT) cells target the majority of glycosylphosphatidylinositol (GPI)-anchored proteins to the basolateral membrane provides us with the opportunity to determine the role of MAL in apical transport of membrane proteins under conditions in which the majority of GPI-anchored proteins are (MDCK cells) or are not (FRT cells) targeted to the apical surface. Using an antisense oligonucleotide-based strategy to deplete endogenous MAL, we have observed that correct transport of apical transmembrane proteins associated (HA) or not (exogenous neurotrophin receptor and endogenous dipeptidyl peptidase IV) with lipid rafts, as well as that of the bulk of endogenous apical membrane, takes place in FRT cells by a pathway that requires normal MAL levels. Even transport of placental alkaline phosphatase, a GPI-anchored protein that is targeted apically in FRT cells, was dependent on normal MAL levels. Similarly, in addition to the reported effect of MAL on HA transport, depletion of MAL in MDCK cells caused a dramatic reduction in the apical delivery of the GPI-anchored gD1-DAF protein, neurotrophin receptor, and the bulk of membrane proteins. These results suggest that MAL is necessary for the overall apical transport of membrane proteins in polarized MDCK and FRT cells.     

Ca++- and Na+, K+-ATPase activities in the fungiform papilla of the tongue of Rana Esculenta (Anura Ranidae)

In the fungiform papilla of Rana esculenta (Anura Ranidae), the Ca⁺⁺-ATPase is mainly distributed on the basolateral membrane of the sensory area cells (i.e., neuroepithelial, supporting, and mucous cells). Apical membranes of all cells facing the surface present a slight enzymatic activity. Lateral wall cells have a strong Ca⁺⁺-ATPase activity on basolateral and apical membranes. Strong Na⁺, K⁺-ATPase activity occurs on the apical surface of neuroepithelial cells. Ca⁺⁺-ATPase activity is absent on the surface of endothelial cells of the capillaries located under the sensory area. These observations lead us to conclude that the sensory area of fungiform papilla is the selective way for calcium influx. Furthermore the absence of ATPase activity on the surface of the endothelial cells indicates that there is no functional barrier to calcium influx into capillary, and that calcium can be removed by vessels from the sensory area.     

Morphology of the differentiation and maturation of LLC-PK1 epithelia

In the present study, a stereologic approach was utilized to quantitatively assess morphological changes during the differentiation of LLC-PK1 cells into an epithelial membrane. This renal epithelial cell line has been described to undergo morphological changes during differentiation and maturation from subconfluent culture to a confluent epithelial layer. An increase in the number of apical microvilli, interpreted as an areal increase in this membrane domain was reported. This morphological differentiation was found to be accompanied by an increase in the expression of apical Na(+)-dependent hexose transport and the activities of certain brush border enzymes. Since no data are available that quantify the morphologic changes during LLC-PK1 differentiation, a quantitative morphologic-stereologic-investigation was performed for an early (6 days) and a late (12 days) state of confluence of LLC-PK1 monolayer cultures. The following morphological parameters were determined by light and electron microscopic morphometry: volume fractions (Vv) of nuclei, mitochondria, and lysosomes, and surface densities (Sv) of the apical and basolateral cell membrane domains. For the apical membrane surface, the microvillous fraction has been measured separately. Since the stereologic approach used in the present study allows the determination of absolute cell volumes, the absolute measures of organelle volumes (V) and membrane surfaces (S) per average cell can be calculated from volume and surface densities. Although no changes in cell density were found for 6 and 12 day old LLC-PK1 monolayers, indicating ceased cell proliferation due to contact inhibition, remarkable changes were found concerning the absolute cell volume and apical membrane surface. The observed increase in the apical cell surface was exclusively due to the enlarged microvillous surface fraction. This finding is in good agreement with the increased number of Na(+)-dependent hexose transporters as well as with the increased expression of apical membrane marker enzymes observed during the differentiation of LLC-PK1 monolayers.     

Characteristics of the ultrastructure of the epithelium of the esophageal mucosa of essentially healthy subjects

Ultrastructure of epithelial surface of the esophageal mucous membrane has been studied in nine practically healthy persons at the age of 32-46 years. The material has been obtained at prophylactic esophagoscopy. The surface of the epithelial layer cells is covered with microfolds 100-200 nm wide and 200-600 nm tall forming labyrinth-like patterns. According to the ultrastructural character of the apical surface of the cells, it is possible to judge about their functional state. The characteristics of the ultrastructure of the fracture surface of the epithelial layer is presented. The data obtained can serve as a base for estimation and differential diagnosis of pathological processes in the organ.     

The MAL proteolipid is necessary for normal apical transport and accurate sorting of the influenza virus hemagglutinin in Madin-Darby canine kidney cells

The MAL (MAL/VIP17) proteolipid is a nonglycosylated integral membrane protein expressed in a restricted pattern of cell types, including T lymphocytes, myelin-forming cells, and polarized epithelial cells. Transport of the influenza virus hemagglutinin (HA) to the apical surface of epithelial Madin-Darby canine kidney (MDCK) cells appears to be mediated by a pathway involving glycolipid- and cholesterol- enriched membranes (GEMs). In MDCK cells, MAL has been proposed previously as being an element of the protein machinery for the GEM-dependent apical transport pathway. Using an antisense oligonucleotide-based strategy and a newly generated monoclonal antibody to canine MAL, herein we have approached the effect of MAL depletion on HA transport in MDCK cells. We have found that MAL depletion diminishes the presence of HA in GEMs, reduces the rate of HA transport to the cell surface, inhibits the delivery of HA to the apical surface, and produces partial missorting of HA to the basolateral membrane. These effects were corrected by ectopic expression of MAL in MDCK cells whose endogenous MAL protein was depleted. Our results indicate that MAL is necessary for both normal apical transport and accurate sorting of HA.     

Oncostatin M regulates membrane traffic and stimulates bile canalicular membrane biogenesis in HepG2 cells

Hepatocytes are the major epithelial cells of the liver and they display membrane polarity: the sinusoidal membrane representing the basolateral surface, while the bile canalicular membrane is typical of the apical membrane. In polarized HepG2 cells an endosomal organelle, SAC, fulfills a prominent role in the biogenesis of the canalicular membrane, reflected by its ability to sort and redistribute apical and basolateral sphingolipids. Here we show that SAC appears to be a crucial target for a cytokine-induced signal transduction pathway, which stimulates membrane transport exiting from this compartment promoting apical membrane biogenesis. Thus, oncostatin M, an IL-6-type cytokine, stimulates membrane polarity development in HepG2 cells via the gp130 receptor unit, which activates a protein kinase A-dependent and sphingomyelin-marked membrane transport pathway from SAC to the apical membrane. To exert its signal transducing function, gp130 is recruited into detergent-resistant membrane microdomains at the basolateral membrane. These data provide a clue for a molecular mechanism that couples the biogenesis of an apical plasma membrane domain to the regulation of intracellular transport in response to an extracellular, basolaterally localized stimulus.     

Distinct cytoskeletal tracks direct individual vesicle populations to the apical membrane of epithelial cells

A key aspect in the structure of epithelial and neuronal cells is the maintenance of a polarized organization based on highly specific sorting machinery at the exit site of the trans Golgi network (TGN). Epithelial cells sort protein and lipid components into different sets of carriers for the apical or basolateral plasma membrane. The two intestinal proteins lactase-phlorizin hydrolase (LPH) and sucrase-isomaltase (SI) are delivered to the apical plasma membrane of epithelial cells with high fidelity but differ in their affinity to detergent-insoluble, glycolipid-enriched complexes (DIGs). Using a two-color labeling technique, we have recently characterized two post-Golgi vesicle populations that direct LPH and SI separately to the apical cell surface. Here, we investigated the structure and identification of protein components in these vesicle populations and assessed the role of cytoskeletal post-Golgi transport routes for apical cargo. Apart from the central role of microtubules in vesicle transport, we demonstrate that the transport of SI-carrying apical vesicles (SAVs) occurs along actin tracks in the cellular periphery, whereas LPH-carrying apical vesicles (LAVs) are transferred in an actin-independent fashion to the apical membrane. Our data further indicate that myosin 1A is the actin-associated motor protein that drives SAVs along actin filaments to the apical cell surface.     

Transport of influenza HA from the trans-Golgi network to the apical surface of MDCK cells permeabilized in their basolateral plasma membranes: energy dependence and involvement of GTP-binding proteins

A procedure employing streptolysin O to effect the selective permeabilization of either the apical or basolateral plasma membrane domains of MDCK cell monolayers grown on a filter support was developed which permeabilizes the entire monolayer, leaves the opposite cell surface domain intact, and does not abolish the integrity of the tight junctions. This procedure renders the cell interior accessible to exogenous macromolecules and impermeant reagents, permitting the examination of their effects on membrane protein transport to the intact surface. The last stages of the transport of the influenza virus hemagglutinin (HA) to the apical surface were studied in pulse-labeled, virus-infected MDCK cells that were incubated at 19.5 degrees C for 90 min to accumulate newly synthesized HA in the trans-Golgi network (TGN), before raising the temperature to 35 degrees C to allow synchronized transport to the plasma membrane. In cells permeabilized immediately after the cold block, 50% of the intracellular HA molecules were subsequently delivered to the apical surface. This transport was dependent on the presence of an exogenous ATP supply and was markedly inhibited by the addition of GTP-gamma-S at the time of permeabilization. On the other hand, the GTP analogue had no effect when it was added to cells that, after the cold block, were incubated for 15 min at 35 degrees C before permeabilization, even though at this time most HA molecules were still intracellular and their appearance at the cell surface was largely dependent on exogenous ATP. These findings indicate that GTP-binding proteins are involved in the constitutive process that effects vesicular transport from the TGN to the plasma membrane and that they are charged early in this process. Transport of HA to the cell surface could be made dependent on the addition of exogenous cytosol when, after permeabilization, cells were washed to remove endogenous cytosolic components. This opens the way towards the identification of cell components that mediate the sorting of apical and basolateral membrane components in the TGN and their polarized delivery to the cell surface.     

Immunocytochemical localization of Na+-HCO3- cotransporters and carbonic anhydrase dependence of fluid transport in corneal endothelial cells

In corneal endothelium, there is evidence for basolateral entry of HCO(3)(-) into corneal endothelial cells via Na(+)-HCO(3)(-) cotransporter (NBC) proteins and for net HCO(3)(-) flux from the basolateral to the apical side. However, how HCO(3)(-) exits the cells through the apical membrane is unclear. We determined that cultured corneal endothelial cells transport HCO(3)(-) similarly to fresh tissue. In addition, Cl(-) channel inhibitors decreased fluid transport by at most 16%, and inhibition of membrane-bound carbonic anhydrase IV by benzolamide or dextran-bound sulfonamide decreased fluid transport by at most 29%. Therefore, more than half of the fluid transport cannot be accounted for by anion transport through apical Cl(-) channels, CO(2) diffusion across the apical membrane, or a combination of these two mechanisms. However, immunocytochemistry using optical sectioning by confocal microscopy and cryosections revealed the presence of NBC transporters in both the basolateral and apical cell membranes of cultured bovine corneal endothelial cells and freshly isolated rabbit endothelia. This newly detected presence of an apical NBC transporter is consistent with its being the missing mechanism sought. We discuss discrepancies with other reports and provide a model that accounts for the experimental observations by assuming different stoichiometries of the NBC transport proteins at the basolateral and apical sides of the cells. Such functional differences might arise either from the expression of different isoforms or from regulatory factors affecting the stoichiometry of a single isoform.