THE STRUCTURE OF THE ZONULA OCCLUDENS

Replicas of freeze-fractured toad urinary bladder and gallbladder were analysed in an attempt to determine the fracturing properties and structure of the zonula occludens (tight junction). Chalcroft and Bullivant have proposed that the junction has a double set of fibrils with one set associated with each of the adjacent cell membranes. However, the fracturing pattern that is observed might also result from only a single set of fibrils which is shared by the adjacent membranes if fracturing occurred around either side of the fibrils. These two models predict quite different structures at regions of the junction where tranl sitions are made between face A and face B. The relative heights of face A and face B and the shape of the transition from face A to face B do not agree with that expected according to the two fibril model but agree exactly with that expected if only a single set of fibrils existed. Further evidence for the single fibril model is derived from fractures of the mucosa membrane which cross the junction to the membrane of the adjacent cell without deflection. Such fractures reveal a single ridge which appears to be identical to the juxtaluminal fibril of the junction. In addition, small ridges are occasionally found in place of the grooves on face B which, although not consistent with the double fibril model, is expected if the single fibril model were correct. Although alternative explanations might account for these observations, we believe that the simplest and most consistent explanation is that the zonula occludens fractures as would be expected of a single set of fibrils shared by adjacent cells.

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Effect of aldosterone on abundance and phosphorylation kinetics of Na-K-ATPase of toad urinary bladder

AJP Renal Physiology ◽

10.1152/ajprenal.1984.246.4.f509 ◽

1984 ◽

Vol 246 (4) ◽

pp. F509-F516

Author(s):

C. S. Park ◽

I. S. Edelman

Keyword(s):

Urinary Bladder ◽

Cell Membranes ◽

Specific Activity ◽

Short Circuit ◽

Fold Increase ◽

Sodium Deoxycholate ◽

Kinetic Properties ◽

Toad Urinary Bladder ◽

Bladder Cell ◽

Phosphorylation Reaction

The possibility was explored that aldosterone-dependent modulation of either the abundance or the kinetic properties of the Na+ pump (Na-K-ATPase) is involved in the mechanism of natriferic action on toad urinary bladder. Cell membranes from the epithelium of urinary bladders of toads were prepared by a sucrose-Ficoll step gradient method, which yielded a 10-fold increase in specific activity of plasma membrane marker enzymes and only minor contamination with other subcellular fractions. Phosphorylation of Na-K-ATPase by gamma-PO4 of [gamma-32P]ATP in the presence of Mg2+ was Na+ dependent, whereas dephosphorylation was K+ dependent. Km for phosphorylation by ATP was 3 X 10(-8) M and K1/2 (half-maximal stimulation) for Na+ was 13 +/- 2 mM. Pretreatment of cell membranes with sodium deoxycholate (DOC) increased the maximal extent of phosphorylation (Nmax) about threefold without changing the Km for ATP. Aldosterone (5 X 10(-8) M) stimulated transepithelial Na+ transport two- to threefold in 5 h but had no significant effect on Km for ATP in the phosphorylation reaction (with or without activation by DOC) or on the abundance of the enzyme inferred from Nmax of phosphorylation in the absence of activation by DOC. After pretreatment with DOC, average Nmax was 13% greater in the aldosterone-treated population. Regression analysis of these responses revealed no significant correlation between increments in short-circuit currents and Nmax values of phosphorylation with or without pretreatment with DOC.(ABSTRACT TRUNCATED AT 250 WORDS)

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Dynamic Interfacial Electrochemical Phenomena at Living Cell Membranes: Application to the Toad Urinary Bladder Membrane System

Journal of The Electrochemical Society ◽

10.1149/1.2133139 ◽

1977 ◽

Vol 124 (11) ◽

pp. 1697-1706 ◽

Cited By ~ 43

Author(s):

Arthur A. Pilla ◽

Gary S. Margules

Keyword(s):

Urinary Bladder ◽

Living Cell ◽

Cell Membranes ◽

Membrane System ◽

Toad Urinary Bladder ◽

Electrochemical Phenomena

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JUNCTIONAL COMPLEXES IN VARIOUS EPITHELIA

The Journal of Cell Biology ◽

10.1083/jcb.17.2.375 ◽

1963 ◽

Vol 17 (2) ◽

pp. 375-412 ◽

Cited By ~ 2243

Author(s):

Marilyn G. Farquhar ◽

George E. Palade

Keyword(s):

Tight Junction ◽

Intercellular Space ◽

Cell Membranes ◽

Amorphous Material ◽

Dense Material ◽

Junctional Complex ◽

Cytoplasmic Matrix ◽

Pancreatic Acini ◽

Zonula Occludens ◽

Adjoining Cell

The epithelia of a number of glands and cavitary organs of the rat and guinea pig have been surveyed, and in all cases investigated, a characteristic tripartite junctional complex has been found between adjacent cells. Although the complex differs in precise arrangement from one organ to another, it has been regularly encountered in the mucosal epithelia of the stomach, intestine, gall bladder, uterus, and oviduct; in the glandular epithelia of the liver, pancreas, parotid, stomach, and thyroid; in the epithelia of pancreatic, hepatic, and salivary ducts; and finally, between the epithelial cells of the nephron (proximal and distal convolution, collecting ducts). The elements of the complex, identified as zonula occludens (tight junction), zonula adhaerens (intermediary junction), and macula adhaerens (desmosome), occupy a juxtaluminal position and succeed each other in the order given in an apical-basal direction. The zonula occludens (tight junction) is characterized by fusion of the adjacent cell membranes resulting in obliteration of the intercellular space over variable distances. Within the obliterated zone, the dense outer leaflets of the adjoining cell membranes converge to form a single intermediate line. A diffuse band of dense cytoplasmic material is often associated with this junction, but its development varies from one epithelium to another. The zonula adhaerens (intermediate junction) is characterized by the presence of an intercellular space (∼200 A) occupied by homogeneous, apparently amorphous material of low density; by strict parallelism of the adjoining cell membranes over distances of 0.2 to 0.5 µ; and by conspicuous bands of dense material located in the subjacent cytoplasmic matrix. The desmosome or macula adhaerens is also characterized by the presence of an intercellular space (∼240 A) which, in this case, contains a central disc of dense material; by discrete cytoplasmic plaques disposed parallel to the inner leaflet of each cell membrane; and by the presence of bundles of cytoplasmic fibrils converging on the plaques. The zonula occludens appears to form a continuous belt-like attachment, whereas the desmosome is a discontinuous, button-like structure. The zomula adhaerens is continuous in most epithelia but discontinuous in some. Observations made during experimental hemoglobinuria in rats showed that the hemoglobin, which undergoes enough concentration in the nephron lumina to act as an electron-opaque mass tracer, does not penetrate the intercellular spaces beyond the zonula occludens. Similar observations were made in pancreatic acini and ducts where discharged zymogen served as a mass tracer. Hence the tight junction is impervious to concentrated protein solutions and appears to function as a diffusion barrier or "seal." The desmosome and probably also the zonula adhaerens may represent intercellular attachment devices.

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FRACTURE FACES OF ZONULAE OCCLUDENTES FROM "TIGHT" AND "LEAKY" EPITHELIA

The Journal of Cell Biology ◽

10.1083/jcb.58.2.390 ◽

1973 ◽

Vol 58 (2) ◽

pp. 390-400 ◽

Cited By ~ 729

Author(s):

Philippa Claude ◽

Daniel A. Goodenough

Keyword(s):

Urinary Bladder ◽

Large Fraction ◽

Distal Tubule ◽

Morphological Data ◽

Physiological Data ◽

Toad Urinary Bladder ◽

Leaky Epithelia ◽

Zonulae Occludentes ◽

Zonula Occludens ◽

Variable Morphology

Epithelia vary with respect to transepithelial permeability. In those that are considered "leaky", a large fraction of the passive transepithelial flux appears to follow the paracellular route, passing across the zonulae occludentes and moving down the intercellular clefts. In "tight" epithelia, the resistance of the paracellular pathway to passive flux is greatly increased. To see whether differences in the morphology of the zonula occludens could contribute to this variability in leakiness among epithelia, replicas of zonulae occludentes in freeze-fractured material from a variety of tight and leaky epithelia were examined. The junctions appear as a branching and anastomosing network of strands or grooves on the A and B membrane fracture faces, respectively. It was found that the zonula occludens from a "very leaky" epithelium, the proximal convoluted tubule of the mouse kidney, is extremely shallow in the apical-basal direction, consisting in most places of only one junctional strand. In contrast, the "very tight" frog urinary bladder exhibits a zonula occludens that is relatively deep (>0.5 µm) in the apical-basal direction, and consists of five or more interconnected junctional strands interposed between luminal and lateral membrane surfaces. Epithelia of intermediate permeabilities exhibited junctions with intermediate or variable morphology. Toad urinary bladder, mouse stomach, jejunum, and distal tubule, rabbit gallbladder, and Necturus kidney and gallbladder were also examined, and the morphological data from these epithelia were compared to physiological data from the literature.

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Role of PKC isozymes in water transport in toad urinary bladder

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100085812 ◽

1991 ◽

Vol 49 ◽

pp. 302-303

Author(s):

A.J. Mia ◽

L.X. Oakford ◽

T. Yorio

Keyword(s):

Urinary Bladder ◽

Apical Membrane ◽

Membrane Surface ◽

Protein A ◽

Cell Types ◽

Leukemic Cells ◽

Toad Urinary Bladder ◽

Pkc Isozymes ◽

Antibody Labeling

Protein kinase C (PKC) isozymes, when activated, are translocated to particulate membrane fractions for transport to the apical membrane surface in a variety of cell types. Evidence of PKC translocation was demonstrated in human megakaryoblastic leukemic cells, and in cardiac myocytes and fibroblasts, using FTTC immunofluorescent antibody labeling techniques. Recently, we reported immunogold localizations of PKC subtypes I and II in toad urinary bladder epithelia, following 60 min stimulation with Mezerein (MZ), a PKC activator, or antidiuretic hormone (ADH). Localization of isozyme subtypes I and n was carried out in separate grids using specific monoclonal antibodies with subsequent labeling with 20nm protein A-gold probes. Each PKC subtype was found to be distributed singularly and in discrete isolated patches in the cytosol as well as in the apical membrane domains. To determine if the PKC isozymes co-localized within the cell, a double immunogold labeling technique using single grids was utilized.

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OSMOTIC INHIBITION OF THE NATRIFERIC RESPONSE OF THE TOAD URINARY BLADDER TO VASOPRESSIN

Journal of Endocrinology ◽

10.1677/joe.0.0670119 ◽

1975 ◽

Vol 67 (1) ◽

pp. 119-125

Author(s):

P. J. BENTLEY

Keyword(s):

Urinary Bladder ◽

Water Movement ◽

Short Circuit ◽

Electrical Potential ◽

Short Circuit Current ◽

Toad Bladder ◽

Toad Urinary Bladder ◽

Electrical Potential Difference ◽

Osmotic Gradient

SUMMARY The electrical potential difference and short-circuit current (scc, reflecting active transmural sodium transport) across the toad urinary bladder in vitro was unaffected by the presence of hypo-osmotic solutions bathing the mucosal (urinary) surface, providing that the transmural flow of water was small. Vasopressin increased the scc across the toad bladder (the natriferic response), but this stimulation was considerably reduced in the presence of a hypo-osmotic solution on the mucosal side, conditions under which water transfer across the membrane was also increased. This inhibition of the natriferic response did not depend on the direction of the water movement, for if the osmotic gradient was the opposite way to that which normally occurs, the response to vasopressin was still reduced. The natriferic response to cyclic AMP was also inhibited in the presence of an osmotic gradient. Aldosterone increased the scc and Na+ transport across the toad bladder but this response was not changed when an osmotic gradient was present. The physiological implications of these observations and the possible mechanisms involved are discussed.

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