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1.
Summary Morphologic findings of widely dilated intercellular spaces in fluid transporting epithelia have been claimed as evidence for the existence of an epithelial compartment in which the coupling between solute and water fluxes takes place. The validity of using epithelial geometry in sectioned material as an argument can be questioned. The present report describes the morphological appearance of frog gallbladder epithelium — normal and ouabain-treated — in the living state in vitro and after fixation, dehydration and embedding. Gallbladder segments were photographed in the living state and at the end of each step of the preparative procedure. Direct observations of whole-mounted gallbladder segments were carried out, taking advantage of the possibility of optical sectioning and high resolution by Nomarski-microscopy. The same specimens were then sectioned and examined by conventional light and electron microscopy. The observations were quantitated and showed that the epithelial cells of normal and ouabain-treated gallbladders experienced an average linear shrinkage down to 70% of their length in Ringer's solution, which corresponds to a volume shrinkage down to 35%. Moreover, dilated lateral intercellular spaces appeared during the dehydration and embedding procedure in normal but only very moderately or not at all in ouabain-treated gallbladder specimens.  相似文献   

2.
The ion activities in the lateral spaces of the unilateral preparation of the gallbladder of Rana catesbiana were measured by double-barrelled ion-selective microelectrodes. The bladders were bathed in a saline solution with a low osmolarity (62 mOsm) containing, in mM: 27 Na+, 27 Cl-, 2 K+, 1 Ca++, 4 HCO3-. Working at reduced osmolarities had the advantage of an increased volume transport and of widened intercellular spaces. The reference barrel recorded an electrical potential of +2.7 mV in the spaces; they contained a solution similar to the external solution. The electrodes recorded a Na+ concentration of 27 mM, a K+ concentration of 1.7 mM, a Ca++ concentration of 0.69 mM and a Cl- concentration of 28.5 mM. In the spaces there was a lower resistance between the tip of the electrode and the serosal bath than that recorded with the tip in the lumen, and injection of fluorescent dye (11 A diameter) via the electrodes did not stain the cells. The concentrations in the secretion were similar to those in the spaces. The intracellular compartment had an apparent K+ concentration of 95 mM, and the concentrations of Na+ and Cl- were both about 5 mM. These data indicate that when the gallbladder is bathed with hypotonic solutions and is transporting fluid at approximately three or four times the normal rate, there are no significant osmotic gradients between the lumen and the lateral spaces. It is suggested that transcellular transport of water is implemented by a combination of high osmotic permeabilities across both mucosal and serosal cell membranes and low reflection coefficients (for K+ salts) at the serosal cell membranes.  相似文献   

3.
Summary The functional role of cytokeratin intermediate filaments in the translocation of asymmetric membrane plaques between cytoplasm and surface of apical urothelial cells was investigated during contraction and expansion of rat urinary bladders. A stereological investigation of electron micrographs provided estimations of surface area, volume, and number of discoidal vesicles and infoldings per unit volume of urothelial apical cell cytoplasm. Contracted and distended bladders incubated in 0.01 M sodium bicarbonate were compared to identical preparations experimentally incubated in 5 mM thioglycolic acid. The latter reagent disrupts the intermediate filament network by reducing sulfhydryl bridges. Densities of discoidal vesicles in cells contracted after incubation in thioglycolate were similar to density estimations in cells expanded under control conditions. Similarly, densities of vesicles in cells expanded after exposure to thioglycolate were comparable in number to those in normally contracted cells. Thus, membrane translocation to and from the luminal surface was blocked by thioglycolate treatment. The lack of normal membrane transfer at the luminal surface induces apical cells exposed to experimental conditions to undergo extraordinary adjustments in response to external pressures of bladder contraction and distension. During contraction, the apical-intermediate cell interface unfolded while the luminal surface ballooned out into the lumen. In distended bladders, large intercellular spaces formed between apical cells along their lateral margins. The results support a model published earlier implicating the filament network as a critical mediator of membrane translocation.  相似文献   

4.
Summary Protein uptake from cerebral ventricles into the epithelium of the choroid plexus, and transport across the epithelium were studied ultrastructurally in rats. Horseradish peroxidase (HRP, MW 40,000) was used as protein tracer. Steady-state ventriculo-cisternal perfusion with subatmospheric pressure (-10cm of water) in the ventricular system was applied. HRP dissolved in artificial CSF was perfused from the lateral ventricles to cisterna magna for various times, and ventriculo-cisternal perfusion, vascular perfusion or immersion fixation with a formaldehyde-glutaraldehyde solution was performed.Coated micropinocytic vesicles containing HRP were seen both connected with the apical, lateral and basal epithelial surface and within the cells. Heavily HRP-labeled vesicles were often fused with the lining membrane of slightly labeled or unlabeled intercellular spaces. Since the apical tight junctions of the epithelium never appeared open or never contained HRP in the spaces between the fusion points, and since the intercellular spaces between adjacent epithelial cells below the junctions only infrequently contained tracer after 5 min, by increasing amounts after 15–60 min of HRP perfusion, a vesicular transport of HRP from the apical epithelial surface to the intercellular spaces, bypassing the tight junctions, is suggested.In addition to the transepithelial transport, micropinocytic vesicles also transported HRP to the lysosomal apparatus of the epithelial cells. With increasing length of exposure to HRP, a sequence of HRP-labeled structures could be evaluated, from slightly labeled apical vacuoles and multivesicular bodies to very heavily labeled dense bodies.  相似文献   

5.
PATHS OF TRANSTUBULAR WATER FLOW IN ISOLATED RENAL COLLECTING TUBULES   总被引:11,自引:4,他引:7       下载免费PDF全文
The cells of perfused rabbit collecting tubules swell and the intercellular spaces widen during osmotic flow of water from lumen to bath induced by antidiuretic hormone (ADH). Ouabain had no influence on these changes. In the absence of net water flow intercellular width was unaffected when tubules were swollen in hypotonic external media. Therefore, during ADH-induced flow widening of intercellular spaces is not a consequence of osmotic swelling of a closed intercellular compartment containing trapped solutes, but rather is due to flow of solution through the channel. Direct evidence of intercellular flow was obtained. Nonperfused tubules swollen in hypotonic media were reimmersed in isotonic solution with resultant entry of water into intercellular spaces. The widened spaces gradually collapsed completely. Spaces enlarged in this manner could be emptied more rapidly by increasing the transtubular hydrostatic pressure difference. In electron micrographs a path of exit of sufficient width to accommodate the observed rate of fluid flow was seen at the base of the intercellular channel. It is concluded that the intercellular spaces communicate with the external extracellular fluid and that water, having entered the cells across the luminal plasma membrane in response in ADH, leaves the cells by osmosis across both the lateral and basilar surface membranes.  相似文献   

6.
Summary We recently described a method by which the resistance to water flow of the luminal membrane of ADH-stimulated toad bladder can be quantitatively distinguished from that of barriers lying in series with it. This method requires estimates of both total bladder water permeability (assessed by transbladder osmotic water flow at constant gradient) and luminal membrane water permeability (assessed by quantitation of the frequency of ADH-induced luminal membrane particle aggregates). In the present study we examined the effect of bladder distension on transepithelial osmotic water flow before and during maximal ADH stimulation. Base-line water flow was unaffected by bladder distension, but hormonally stimulated flow increased systematically as bladders became more distended. Distension had no effect on the frequency of ADH-induced intramembranous particle aggregates. By comparing the relationships between aggregate frequency and hormonally induced water permeability in distended and undistended bladders, we found that distension appeared to enhance ADH-stimulated water flow by decreasing the resistance of the series permeability barrier while the apparent water permeability associated with each single luminal membrane aggregate was unaffected. In that bladder distension causes tissue thinning, the series resistance limiting ADH-stimulated water flow appears to be accounted for by deformable barriers within the bladder tissue itself, probably unstirred layers of water.  相似文献   

7.
The volume of the cells and lateral intercellular spaces were measured in living Necturus gallbladder epithelium. Under control conditions, the volume of the lateral spaces was 9% of the cell volume. Replacement of mucosal NaCl by sucrose or tetramethylammonium chloride (TMACl) caused intercellular spaces to collapse. During mucosal NaCl replacement, cell volume decreased to 79% of its control value. When NaCl was reintroduced into the mucosal bath, the intercellular spaces reopened and the cells returned to control volume. The NaCl active transport rate, calculated from the rate of cell volume decrease, was 266 pM/cm2.s, close to the observed rate of transepithelial salt transport. It was calculated from the decrease in cell volume that all of the intracellular NaCl was transported out of the cell during removal of mucosal NaCl. The flux of salt across the apical membrane, calculated from the rate of cell volume increase upon reintroducing mucosal NaCl, was 209 pM/cm2.s, in good agreement with estimates by other methods. The electrical resistance of the tight junctions was estimated to be 83.9% of the total tissue resistance in control conditions, suggesting that the lateral intercellular spaces normally offer only a small resistance to electrolyte movement.  相似文献   

8.
Summary The purpose of these experiments was to determine the effect of osmotic gradients on the permeability of the rabbit gallbladder. Increasing the tonicity of the mucosal solution reduced the permeability of the gallbladder to both ions and nonelectrolytes, whereas there was no significant effect when the serosal solution was made hypertonic. These results cannot be explained by solvent/solute interactions in either the epithelial membranes or the unstirred layers. Associated with the changes in permeability was the appearance of the transport number effect and current induced resistance changes. Morphological studies of the gallbladder under these conditions showed that the extracellular spaces of the epithelium and the rest of the wall dilate in the presence of osmotic flow to the serosa, but that the spaces collapse when the flow is in the opposite direction. Reconstruction of the permeability changes from the dimensions of the tissue show that all the physiological phenomena are accounted for by the changes in morphology, the dominant effect being in the lateral intercellular spaces. These results suggest that the lateral spaces are a common pathway shared by all solutes crossing the epithelium, and that diffusion along these spaces becomes rate limiting as the spaces collapse.  相似文献   

9.
Summary Gallbladders transport isotonically over a wide range of osmolarities. This ability has been assumed to depend on the geometry of the lateral intercellular spaces. We report that this geometry in theNecturus gallbladder varies extensively with the external osmolarity and dependsin vitro on the integrity of the subepithelial tissues. The structure of the living epithelium was studied by Nomarski light microscopy while ultrastructural effects were revealed by electron microscopy. The short-term effects (<60 min) of low external osmolarities were: 1) the cells became bell-shaped with an increased cell height measured centrally, 2) lateral intercellular spaces lost their convoluted character; and 3) numerous membrane-bound cavities appeared in the cells. Furthermore, long-term exposure to the low external osmolarities caused an uneven density of epithelial cells. With subepithelial tissues intact, blistering of the epithelium cell layer was evident. Qualitative electron-microscopic data indicate that the membrane of the cavities was recruited from the basolateral cell membrane. This agrees well with light-microscopic observation that the cavities were initiated as invaginations of this cell membrane.  相似文献   

10.
The relationship between epithelial fluid transport, standing osmotic gradients, and standing hydrostatic pressure gradients has been investigated using a perturbation expansion of the governing equations. The assumptions used in the expansion are: (a) the volume of lateral intercellular space per unit volume of epithelium is small; (b) the membrane osmotic permeability is much larger than the solute permeability. We find that the rate of fluid reabsorption is set by the rate of active solute transport across lateral membranes. The fluid that crosses the lateral membranes and enters the intercellular cleft is driven longitudinally by small gradients in hydrostatic pressure. The small hydrostatic pressure in the intercellular space is capable of causing significant transmembrane fluid movement, however, the transmembrane effect is countered by the presence of a small standing osmotic gradient. Longitudinal hydrostatic and osmotic gradients balance such that their combined effect on transmembrane fluid flow is zero, whereas longitudinal flow is driven by the hydrostatic gradient. Because of this balance, standing gradients within intercellular clefts are effectively uncoupled from the rate of fluid reabsorption, which is driven by small, localized osmotic gradients within the cells. Water enters the cells across apical membranes and leaves across the lateral intercellular membranes. Fluid that enters the intercellular clefts can, in principle, exit either the basal end or be secreted from the apical end through tight junctions. Fluid flow through tight junctions is shown to depend on a dimensionless parameter, which scales the resistance to solute flow of the entire cleft relative to that of the junction. Estimates of the value of this parameter suggest that an electrically leaky epithelium may be effectively a tight epithelium in regard to fluid flow.  相似文献   

11.
Summary The experiments described herein use an in vitro preparation of choroid plexus to demonstrate that it is a vasopressin-responsive organ by morphologic criteria. Choroid plexus from rats was incubated for one hour in graded concentrations of arginine vasopressin (AVP). Within physiologic range of molar concentration, incubation in vasopressin induced a decrease in basal and lateral spaces in choroid plexus epithelial cells as well as an increase in number of dark cells. The number of cells with basal spaces decreased significantly from 82.7±9.2 in control tissue to 19±18 in tissue incubated in 10-12 M AVP; similarly, the number with lateral cellular spaces decreased from 20±8.8 to 7.6±2.2 cells in 10-10 M AVP. Dark cells increased in number from 3.8±2.6 in control conditions to 49±4 with 10-9 M vasopressin. These data suggest important effects of arginine vasopressin in cerebrospinal fluid (CSF) on choroid plexus, compatible with enhanced fluid transport across choroid epithelial cells.  相似文献   

12.
Summary In order to assess the contribution of transcellular water flow to isosmotic fluid transport acrossNecturus gallbladder epithelium, we have measured the water permeability of the epithelial cell membranes using a nuclear magnetic resonance method. Spin-lattice (T 1) relaxation of water protons in samples of gallbladder tissue where the extracellular fluid contained 10 to 20mm Mn2+ showed two exponential components. The fraction of the total water population responsible for the slower of the two was 24±2%. Both the size of the slow component, and the fact that it disappeared when the epithelial layer was removed from the tissue, suggest that it was due to water efflux from the epithelial cells. The rate constant of efflux was estimated to be 15.6±1.0 sec1 which would be consistent with a diffusive membrane water permeabilityP d of 1.6×103 cm sec1 and an osmotic permeabilityP os of between 0.3×104 and 1.4×104 cm sec1 osmolar1. Using these data and a modified version of the standing-gradient model, we have reassessed the adequacy of a fluid transport theory based purely on transcellular osmotic water flow. We find that the model accounts satisfactorily for near-isosmotic fluid transport by the unilateral gallbladder preparation, but a substantial serosal diffusion barrier has to be included in order to account for the transport of fluid against opposing osmotic gradients.  相似文献   

13.
Large quantities of colloidal particles were rapidly transported around the junctional complex into the lateral intercellular spaces by flounder renal epithelial cells. Large invaginations containing particles developed in the apical cytoplasm of cells when tracer particles were injected into the tubular lumens. Some membranebounded profiles containing particles appeared close to the lateral intercellular spaces. Particles were then found in the lateral intercellular spaces, between the basal plasmalemma and the basement membrane, and within the basement membrane. It is suggested that this transport might operate in situ and provide a morphological mechanism to explain a type of protein transport noted in the renal tubules of another flounder species by Maack and Kinter ('67). It is interesting to consider that perhaps a similar mechanism for the transport of intact proteins might also operate in mammalian nephrons as well.  相似文献   

14.
Ionic lanthanum has been used to study transepithelial ion permeation in in vitro rabbit gallbladder and intestine (ileum) by adding 1 mM La3+ to only the mucosal bathing solution. Transepithelial fluid transport electrical potential differences (p.d.), and resistances were measured. During La3+ treatment the gallbladder''s rate of active solute-coupled fluid transport remained constant, the resistance increased, and the 2:1 NaCl diffusion p.d. decreased. Mucosa-to-serosa fluxes of 140La3+ were measured and indicate a finite permeability of the gallbladder to La3+. La3+ also increased the transepithelial resistance and p d. of ileum. Electron microscopic examination of La3+-treated gallbladder showed: (a) good preservation of the fine structure, (b) electron-opaque lanthanum precipitates in almost every lateral intercellular space, most frequently near the apical end of the lateral spaces close to or within the junctional complex, (c) lanthanum among the subjacent muscle and connective tissue layers, and (d) lanthanum filling almost the entire length of so-called "tight" junctions. No observations were made which unequivocally showed the penetration of lanthanum into the gallbladder cells. Electron micrographs of similar La3+-treated ilea showed lanthanum deposits penetrating the junctional complexes. These results coupled with other physiological studies indicate that the low resistance pathway for transepithelial ion permeation in gallbladder and ileum is through the tight junctions A division of salt-transporting epithelia into two main groups, those with "leaky" junctional complexes and those with tight junctional complexes, has been proposed.  相似文献   

15.
Summary The sheep parotid is a compound tubular gland; its ultrastructure reflects the function of this gland to secrete large amounts of fluid with very little protein. The cells of the secretory tubules possess extensively folded lateral plasma membranes and a fairly large number of mitochondria. Rapid equilibration of water across the epithelium is assured by the close proximity over large areas of intercellular spaces and the wide secretion canaliculi. Numerous long microvilli extend into the latter. Although secretion granules may be quite numerous, there is evidence that many of these granules are not discharged but undergo degradation by lysosomal enzymes. The intercalated ducts are often dilated but excessive distension is probably prevented by bundles of microfilaments in their epithelial cells.  相似文献   

16.
Summary Human gall-bladder epithelium obtained straight from the operating theatre was incubated in an Ussing chamber with the fluid phase marker, horseradish peroxidase (HRP), for up to 60 min. When the marker was presented on the apical surface, within 30 min it had moved readily across the apical cytoplasm in transport vesicles to receptosomes and into the lateral intercellular space, extending across the basement membrane into the lamina propria. When HRP was presented at the basal aspect, within 30 min it had moved through the lamina propria, across the basement membrane and into the lateral intercellular space. By 60 min, only small amounts had been taken up by the epithelial cells and transported to receptosomes. These data indicate a rapid transmucosal endocytotic pathway for blood-or bile-borne macromolecules.  相似文献   

17.
Summary Horseradish peroxidase (HRP) was administered intravenously to mice by bolus injection. The subsequent uptake and fate of the HRP by the lateral and basal cell surfaces of resting and stimulated gallbladder epithelial cells was followed by light and electron microscopy. At 10 min after injection, HRP was visible in the lamina propria of the gallbladder and within 20 min of injection, HRP had permeated the basement membrane and had entered the lateral intercellular space, extending as far as the apical tight junction. Over the following 30 min, there was evidence of vesicular epithelial HRP uptake and 1 h after injection, HRP was visible in epithelial secretory granules within the lumen of the gallbladder and apical transport vesicles. These data provide evidence of a blood-to-bile transport pathway which could represent an important route of entry to bile by various blood-borne macromolecules.  相似文献   

18.
Experiments were done for indentification and localization of certain structural changes at different levels of jejunal villus of the hamster during positive and negative water transport across the intestine in vivo and in vitro. Positive transport occurred when the mucosal surface of the intestine was bathed (in vitro experiments) or perfused (in vivo experiments) with isotonic Krebs-Ringer bicarbonate solution containing 10 mM glucose, and negative water transport was achieved by rendering this solution hypertonic with 150 mM mannitol. Results indicate that during positive net water transport the intestine in vivo transported more fluid and exhibited a more conspicuous dilatation of the lateral intercellular spaces (L.I.S.) than did the in vitro preparation. Dilatation of the L.I.S. in both preparations was present only in the apical part of the villus, suggesting that this is the principal site of water absorption. When the mucosal solution was made hypertonic with mannitol, the L.I.S. in the in vivo intestine totally collapsed, whereas in the in vitro intestine these spaces remained open very slightly. These morphological changes correspond well with our finding that in the presence of the hypertonic mucosal solution there was a greater net negative water transport in vivo than in vitro. Incubation of the intestine in the isotonic mucosal solution produced subnuclear swelling of the mid-villus epithelial cells, and this morphological change was associated with an increase in the water content of the tissue. Perfusion of the in vivo intestine with the isotonic solution produced neither the swellings nor the increase in water content of the tissue. In the presence of hypertonic mucosal solution there was a water loss from the tissue both in vivo and in vitro, and these swellings were not observed. These results are discussed in relation to intestinal sugar transport and to the maturity of the epithelial cells, and it is concluded that transport studies on in vitro preparations may provide valid information on a qualitative basis, if not on a strictly quantitative basis.  相似文献   

19.
The Ultrastructural Route of Fluid Transport in Rabbit Gall Bladder   总被引:18,自引:5,他引:13  
The route of fluid transport across the wall of the rabbit gall bladder has been examined by combined physiological and morphological techniques. Fluid transport was either made maximal or was inhibited by one of six physiological methods (metabolic inhibition with cyanide-iodoacetate, addition of ouabain, application of adverse osmotic gradients, low temperature, replacement of Cl by SO4, or replacement of NaCl by sucrose). Then the organ was rapidly fixed and subsequently embedded, sectioned, and examined by light and electron microscopy. The structure of the gall bladder is presented with the aid of electron micrographs, and changes in structure are described and quantitated. The most significant morphological feature seems to be long, narrow, complex channels between adjacent epithelial cells; these spaces are closed by tight junctions at the luminal surface of the epithelium but are open at the basal surface. They are dilated when maximal fluid transport occurs, but are collapsed under all the conditions which inhibit transport. Additional observations and experiments make it possible to conclude that this dilation is the result of fluid transport through the spaces. Evidently NaCl is constantly pumped from the epithelial cells into the spaces, making them hypertonic, so that water follows osmotically. It is suggested that these spaces may represent a "standing-gradient flow system," in which osmotic equilibration takes place progressively along the length of a long channel.  相似文献   

20.
K+ and Cl--selective double-barreled microelectrodes were used to study the effect of changes in external K+ concentration on intracellular Cl- activity (aiCl) in epithelial cells of Necturus gallbladder. Decreasing the K+ concentration simultaneously in both bathing solutions produced a decrease in aiCl. Steady-state values of aiCl were related to the values of the chemical potential gradient for K+ (delta microK) across either the apical or the basolateral cell membrane. A similar dependence between aiCl and delta microK appeared when the K+ concentration was changed in the serosal solution only. This indicates that aiCl depends on delta microK across the basolateral membrane. aiCl was virtually independent of the membrane potential. This supports the idea that both the mucosal and the basolateral membranes of Necturus gallbladder cells have very low passive permeabilities to Cl-. These results indicate that the exit of Cl- from Necturus gallbladder cells is driven by delta microK across the basolateral membrane, and suggest that KCl electroneutral coupled mechanism in this membrane plays an important role in transcellular Cl- transport.  相似文献   

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