Isotonic Water Transport in Secretory Epithelia ,

The model proposed by Diamond and Bossert [1] for isotonic water transport has received wide acceptance in recent years. It assumes that the local driving force for water transport is a standing osmotic gradient produced in the lateral intercellular spaces of the epithelial cell layer by active solute transport. While this model is based on work done in absorptive epithelia where the closed to open direction of the lateral space and the direction of net transport are the same, it has been proposed that the lateral spaces could also serve as the site of the local osmotic gradients for water transport in secretory epithelia, where the closed to open direction of the lateral space and net transport are opposed, by actively transporting solute out of the space rather than into it. Operation in the backward direction, however, requires a lower than ambient hydrostatic pressure within the lateral space which would seem more likely to cause the space to collapse with loss of function. On the other hand, most secretory epithelia are characterized by transport into a restricted ductal system which is similar to the lateral intercellular space in the absorptive epithelia in that its closed to open direction is the same as that of net transport. In vitro micropuncture studies on the exocrine pancreas of the rabbit indicate the presence of a small but statistically significant increase in juice osmolality, 6 mOsm/kg H₂O, at the site of electrolyte and water secretion in the smallest extralobular ducts with secretin stimulation which suggests that the ductal system in the secretory epithelia rather than the lateral intercellular space is the site of the local osmotic gradients responsible for isotonic water transport.     

Optimizing automated characterization of liver fibrosis histological images by investigating color spaces at different resolutions

Background

7D-cadherins like LI-cadherin are cell adhesion molecules and represent exceptional members of the cadherin superfamily. Although LI-cadherin was shown to act as a functional Ca²⁺-dependent adhesion molecule, linking neighboring cells together, and to be dysregulated in a variety of diseases, the physiological role is still enigmatic. Interestingly 7D-cadherins occur only in the lateral plasma membranes of cells from epithelia of water transporting tissues like the gut, the liver or the kidney. Furthermore LI-cadherin was shown to exhibit a highly cooperative Ca²⁺-dependency of the binding activity. Thus it is tempting to assume that LI-cadherin regulates the water transport through the epithelium in a passive fashion by changing its binding activity in dependence on the extracellular Ca²⁺.

Results

We developed a simple mathematical model describing the epithelial lining of a lumen with a content of variable osmolarity covering an interstitium of constant osmolarity. The width of the lateral intercellular cleft was found to influence the water transport significantly. In the case of hypertonic luminal content a narrow cleft is necessary to further increase concentration of the luminal content. If the cleft is too wide, the water flux will change direction and water is transported into the lumen. Electron microscopic images show that in fact areas of the gut can be found where the lateral intercellular cleft is narrow throughout the lateral cell border whereas in other areas the lateral intercellular cleft is widened.

Conclusions

Our simple model clearly predicts that changes of the width of the lateral intercellular cleft can regulate the direction and efficiency of water transport through a simple epithelium. In a narrow cleft the cells can increase the concentration of osmotic active substances easily by active transport whereas if the cleft is wide, friction is reduced but the cells can hardly build up high osmotic gradients. It is now tempting to speculate that 7D-cadherins, owing to their location and their Ca²⁺-dependence, will adapt their binding activity and thereby the width of the lateral intercellular cleft automatically as the Ca²⁺-concentration is coupled to the overall electrolyte concentration in the lateral intercellular cleft. This could provide a way to regulate the water resorption in a passive manner adapting to different osmotic conditions.     

The role of intercellular channels in the transepithelial transfer of water and sodium in the frog urinary bladder

Summary Epithelial cells of frog urinary bladders fixed in different physiological states were examined by electron microscopy. It was shown: (1) that when bladders incubated with a hypotonic mucosal medium are water-permeabilized with oxytoxin, arginine-vasotocin, cyclic 3′,5′-AMP and theophylline, this leads to a cellular swelling and the opening of intercellular channels; (2) that these effects are not observed when the transepithelial net water flow is suppressed by abolishing the external osmotic driving force; and (3) that modifications in the rate of active sodium transport do not change the morphological appearance of intercellular channels. These results are expecially discussed with respect to the localization to the intracellular site of the final effect of antidiuretic hormone on water permeability, and to the role of intercellular channels in the transepithelial transfer of water and sodium.     

Apoplasmic and Protoplasmic Water Transport through the Parenchyma of the Potato Storage Organ

Stationary volume fluxes through living and denatured parenchyma slices of the potato (Solanum tuberosum L.) storage organ were studied to estimate the hydraulic conductivity of the cell wall and to evaluate the significance of water transport through protoplasts, cell walls, and intercellular spaces. Slices were placed between liquid compartments, steady-state fluxes induced by pressure or concentration gradients of low- and high-molecular-mass osmotica were measured, and water transport pathways were distinguished on the basis of their difference in limiting pore size. The protoplasts were the dominating route for osmotically driven water transport through living slices, even in the case of a polymer osmoticum that is excluded from cell walls. The specific hydraulic conductivity of the cell wall matrix is too small to allow a significant contribution of the narrow cell wall bypass to water flow through the living tissue. This conclusion is based on (a) ultrafilter coefficients of denatured parenchyma slices, (b) the absence of a significant difference between ultrafilter coefficients of the living tissue slices for osmotica with low and high cell wall reflection coefficients, and (c) the absence of a significant interaction (solvent drag) between apoplasmic permeation of mannitol and the water flux caused by a concentration difference of excluded polyethylene glycol. Liquid-filled intercellular spaces were the dominating pathways for pressure-driven volume fluxes through the parenchyma tissue.     

Ultrastructural markers of tubular transport within the rat nephron in experimental diabetes insipidus

Using stereological methods mitochondrial energy states and intercellular spaces with basal infolded channels were evaluated in proximal and distal tubules in rats. The studies were performed on animals with experimental diabetes insipidus and on control rats by means of electron microscopy. No significant differences were found in mitochondrial energy states and sizes of intercellular spaces with basal infolded channels in the proximal tubules, which indicates undisturbed transport in this nephron segment. However, significant differences of these parameters were found in the distal tubules. In diabetes insipidus mitochondrial energy states approximated the condensed state, while in the control animals they were similar to the orthodox state. Intercellular spaces became significantly narrowed in diabetes insipidus in comparison with these in the controls. These observations suggest that mitochondrial energy states may be considered as ultrastructural markers of active tubular transport, while intercellular spaces with basal infolded channels may reflect ultrastructural counterparts of water transport.     

The correlation of ethanol-induced depression of glucose and water transport with morphological changes in the hamster jejunum in vivo

Glucose and water transport is depressed in the hamster jejunum in vivo by ethanol (4.8%) which also produced fluid-filled blebs at the tips of the villi. The epithelial cells over the blebs appeared stretched and cuboidal, the lateral intercellular spaces (LIS) were no longer recognizable, and the lacteals were closed. Forty-five minutes after discontinuation of the ethanol, water transport returned to normal while glucose transport remained depressed. At this time the villus structure had returned to normal. The blebs had disappeared, the LIS were again recognizable, and their appearance and number were similar to those in the control animals. Thus, the depression of water transport correlated with the obvious structural changes caused by ethanol; however, the depression of glucose absorption is associated with some effect of ethanol not evident by routine light microscopy.     

ULTRASTRUCTURAL STUDIES OF VASOPRESSIN EFFECT ON ISOLATED PERFUSED RENAL COLLECTING TUBULES OF THE RABBIT

Isolated cortical collecting tubules from rabbit kidney were studied during perfusion with solutions made either isotonic or hypotonic to the external bathing medium. Examination of living tubules revealed a reversible increase in thickness of the cellular layer, prominence of lateral cell membranes, and formation of intracellular vacuoles during periods of vasopressin-induced osmotic water transport. Examination in the electron microscope revealed that vasopressin induced no changes in cell structure in collecting tubules in the absence of an osmotic difference and significant bulk water flow across the tubule wall. In contrast, tubules fixed during vasopressin-induced periods of high osmotic water transport showed prominent dilatation of lateral intercellular spaces, bulging of apical cell membranes into the tubular lumen, and formation of intracellular vacuoles. It is concluded that the ultrastructural changes are secondary to transepithelial bulk water flow and not to a direct effect of vasopressin on the cells, and that vasopressin induces osmotic flow by increasing water permeability of the luminal cell membrane. The lateral intercellular spaces may be part of the pathway for osmotically induced transepithelial bulk water flow.     

Coupled water transport in standing gradient models of the lateral intercellular space.

A standing gradient model of the lateral intercellular space is presented which includes a basement membrane of finite solute permeability. The solution to the model equations is estimated analytically using the "isotonic convection approximation" of Segel. In the case of solute pumps uniformly distributed along the length of the channel, the achievement of isotonic transport depends only on the water permeability of the cell membranes. The ability of the model to transport water against an adverse osmotic gradient is the sum of two terms: The first term is simply that for a well-stirred compartment model and reflects basement membrane solute permeability. The second term measures the added strength due to diffusion limitation within the interspace. It is observed, however, that the ability for uphill water transport due to diffusion limitation is diminished by high cell membrane water permeability. For physiologically relevant parameters, it appears that the high water permeability required for isotonic transport renders the contribution of the standing gradient relatively ineffective in transport against an osmotic gradient. Finally, when the model transports both isotonically and against a gradient, it is shown that substantial intraepithelial solute polarization effects are unavoidable. Thus, the measured epithelial water permeability will grossly underestimate the water permeability of the cell membranes. The accuracy of the analytic approximation is demonstrated by numerical solution of the complete model equations.     

Redox states of plastids and mitochondria differentially regulate intercellular transport via plasmodesmata

Recent studies suggest that intercellular transport via plasmodesmata (PD) is regulated by cellular redox state. Until now, this relationship has been unclear, as increased production of reactive oxygen species (ROS) has been associated with both increased and decreased intercellular transport via PD. Here, we show that silencing two genes that both increase transport via PD, INCREASED SIZE EXCLUSION LIMIT1 (ISE1) and ISE2, alters organelle redox state. Using redox-sensitive green fluorescent proteins targeted to the mitochondria or plastids, we show that, relative to wild-type leaves, plastids are more reduced in both ISE1- and ISE2-silenced leaves, whereas mitochondria are more oxidized in ISE1-silenced leaves. We further show that PD transport is positively regulated by ROS production in mitochondria following treatment with salicylhydroxamic acid but negatively regulated by an oxidative shift in both chloroplasts and mitochondria following treatment with paraquat. Thus, oxidative shifts in the mitochondrial redox state positively regulate intercellular transport in leaves, but oxidative shifts in the plastid redox state counteract this effect and negatively regulate intercellular transport. This proposed model reconciles previous contradictory evidence relating ROS production to PD transport and supports accumulating evidence that mitochondria and plastids are crucial regulators of PD function.     

Path of Downward Oxygen Transport from Aerial to Subterranean Parts in Intact Seedlings of Rice and Other Plants

There are two opposite opinions as regards the mechanism and the path of downward oxygen transport in rice and other higher plants. Van Raalte (1940), Yamada (1952), and others maintain that an oxygen pressure gradient of decreasing magnitude from the stem base down to the root tip exists in the intercellular air spaces, which are interconnected throughout the cortex, and the oxygen transported therein is in free molecular form and moves about by diffusion along its own gradient. Recent diffusion experiments in plants by Barber (1962), employing radioactive O15 as indicator, gave direct confirmation of this hypothesis. The opposite view is held by Brown (1947), Soldatenkov (1963) and others. They consider that the passive diffusion of oxygen along its own gradient is inadequate to account for the actual amount transported downwards. The fact that downward oxygen transport in roots comes almost to a standstill, once the aerial part is removed while the cut end of the short stump is still left in air, casts doubts as to the validity of the diffusion hypothesis; and is in favour of their claim that in addition to, or in placement of, diffusion, active participation of living tissues in shoot is necessary to drive enough oxygen to meet the demand of roots. The oxygen in active transport is no longer in free gaseous state but is in dissolved or combined form (as in peroxides) and moves presumably along the vascular bundles in a way which is hitherto unrevealed but is apparently dependent upon the physiological activity of the conducting tissue. In our previous report (Lou et al 1964), we gave data based on quantitative measurement of the amount of oxygen transported downwards from aerial to submerged parts in intact seedlings with the respiratory hydrometer specially designed for the purpose. In seedlings of marshy plants (e.g. rice), it amounts to about 50% of the total oxygen absorbed by the aerial part; in water cultured seedlings of ordinary land plants (e.g. pea), 20%–30%. By deliberately blocking the alternative paths of oxygen transport in seedlings, one at a time, and measuring the downward oxygen transport accordingly in the same way as before, we should be able to decide which one of the two paths is mainly responsible for the transport. The blocking can be conveniently carried out at the upper end of the radical in a pea (or broadbean) seedling by surgical treatment (see Fig.1); either by ringing off the peripheral cortex where most of the air spaces reside; or by piercing through the central cylinder, within which the vascular bundles are confined. The treated radical is then submerged in water and ready for measurement. Without recourse to surgical treatment and mechanical injury, the air space in the cortex can also be blocked by displacing its air content with water through vacuum infiltration. The present investigation has shown that when the intercellular spaces in the cortex of the radical are blocked either by ringing or by infiltration, the aerial part of the treated seedling absorbs much less oxygen than the control as though its radical were completely severed (Table 2); or, in other words, the downward oxygen transport is effectively stopped by such a means. On the other hand, interruption of vascular bundles in the central cylinder only reduces the amount of oxygen in transport to less than one half, which can be accounted for by the combined effect of the reduced root activity due to shortage of food supply and the unavoidable partial disruption of the peripheral cortex. Besides taking actual measurement, downward oxygen transport in intact pea (or broadbean) seedlings can also be detected by simply noticing the growth rate of its radical. As is shown in this investigation, the radical ceases growing in still water, if the oxygen supply from its aerial part is interrupted. As a result of oxygen deficiency, the radical tip deteriorates in a few days. These effects can be easily realized by ringing off the cortex or by infiltrating its air spaces with water. That the peripheral ringing of the radical does no harm to its growth process is revealed by the fact that if air is bubbled through the water culture steadily, normal growth ensues. The above results leave no doubt that in seedlings of rice, pea, and broadbean, downward oxygen transport mainly takes place in the intercellular spaces in the radical cortex, and seems to have no concern with the activities of vascular bundle and cortex. Although there are evidence that rice roots may actively secrete oxygen in the form of peroxides to its immediate neighborhood (the rhizosphere), the actual amount and the distance traversed in such an active transport however, is very much limited and is insignificant as compared with that taking place in the intercellular spaces.     

Relationship Between Downward Oxygen Transport and Intercellular Spaces in Root Cortex in Water Cultured and Moist Cultured Seedlings

In the present investigation, seedlings of rice, pea, sorghum, and maize are raised both in water culture and moist culture. The former culture is to provide the roots with an oxygen deficient condition; while the latter, a direct access to air. The amount of oxygen transported downwards in the seedlings varies not only with the nature of plants but also with the way how they are raised: More oxygen is transported downwards in marsh plant (rice) than in land plants (pea, sorghum, maize); and, in case the same plant is concerned, more in water cultured seedlings than in moist cultured ones. Downward oxygen transport in the various seedlings is intimately correlated with the relative volume of the intercellular spaces in the root: the more the downward transport, the larger the air spaces in the cortex. The fractional volume of the intercellular spaces in a small plant segment can be conveniently estimated by determining the specific gravities of the fresh turgescent segment before and after it is filled with water by vaccum infiltration. The difference between the two consecutive measurements in specific gravity times 100 gives directly the percentage of the volume occupied by air spaces. When large root segments are used, the relative volume can also be determined by weighing before and after vaccum infiltration. To test whether oxygen diffusion in the intercellular spaces of roots could actually account for its downward transport, a model is built of capillary tubings with dimensions and oxygen pressure gradients similar to those found in roots. The amount of oxygen diffused in such a model is measured with a respiratory hydrometer (see Fig. 1) and fits closely that measured in roots. By comparing the amount of oxygen transported downwards in a seedling with that consumed by its excised roots in air, it can be shown that, in case of rice, it could meet (and at times may even exceed) 100% of that consumed by roots in water cultured seedlings, but is less in moist cultured ones. In land plants (pea, sorghum, and maize), however, the downward oxygen supply is far below its requirement, being 80%–100% in water cultured seedlings and 30%–60% in moist cultured ones. The above results, together with those obtained in previous communications, support the view that adaptation of a plant to flooded condition is primarily achieved by its capacity of providing adequate intercellular spaces for downward oxygen diffusion. The capacity depends not only upon the phylogeny of the plant concerned but also upon its ontogenic development.     

Fine structural organization of the rectum in the blowfly, Calliphora erythrocephala (Meig.) with special reference to connective tissue, tracheae and neurosecretory innervation in the rectal papillae

The rectal epithelium of Calliphora is made up of three quite distinct cell types: rectal, cortical and junctional cells. The thin wall of the rectal pouch is made up of rectal cells which are relatively simple and unspecialized; their general structure does not suggest any direct participation in ion transport. A function of ion and water transport can probably be ascribed to the cortical cells, which are arranged in the form of four cones which project into the rectal lumen. The cavity of each cone is filled up with tracheae, nerve and neurosecretory terminals, and connective tissue to form medulla. The medulla and cortex are separated from each other by deeply staining bridges or trabeculae to form an infundibular space. The most conspicuous feature of the cortex is the presence of an extensive intercellular sinus formed by complex infoldings of the lateral plasma-membranes. It is postulated that fluid, which is absorbed from the rectal lumen, is transported into the intercellular sinus and finally reaches the haemolymph via the infundibular space. The actual site of ion transport is probably the stacks of lateral plasma-membrane which are closely associated with mitochondria. The junctional cells, which are packed with microtubules, form a collar around the base of the papillae at the point of their insertion into the rectal wall. It is suggested that the neurosecretory terminals present in the medulla might release a hormone which controls rate of ion and water reabsorption by the papillae cells.     

Aluminum-induced 1-->3-beta-D-glucan inhibits cell-to-cell trafficking of molecules through plasmodesmata. A new mechanism of aluminum toxicity in plants

Symplastic intercellular transport in plants is achieved by plasmodesmata (PD). These cytoplasmic channels are well known to interconnect plant cells to facilitate intercellular movement of water, nutrients, and signaling molecules including hormones. However, it is not known whether Al may affect this cell-to-cell transport process, which is a critical feature for roots as organs of nutrient/water uptake. We have microinjected the dye lucifer yellow carbohydrazide into peripheral root cells of an Al-sensitive wheat (Triticum aestivum cv Scout 66) either before or after Al treatment and followed the cell-to-cell dye-coupling through PD. Here we show that the Al-induced root growth inhibition is closely associated with the Al-induced blockage of cell-to-cell dye coupling. Immunofluorescence combined with immuno-electron microscopic techniques using monoclonal antibodies against 1-->3-beta-D-glucan (callose) revealed circumstantial evidence that Al-induced callose deposition at PD may responsible for this blockage of symplastic transport. Use of 2-deoxy-D-glucose, a callose synthesis inhibitor, allowed us to demonstrate that a reduction in callose particles correlated well with the improved dye-coupling and reduced root growth inhibition. While assessing the tissue specificity of this Al effect, comparable responses were obtained from the dye-coupling pattern in tobacco (Nicotiana tabacum) mesophyll cells. Analyses of the Al-induced expression of PD-associated proteins, such as calreticulin and unconventional myosin VIII, showed enhanced fluorescence and co-localizations with callose deposits. These results suggest that Al-signal mediated localized alterations to calcium homeostasis may drive callose formation and PD closure. Our data demonstrate that extracellular Al-induced callose deposition at PD could effectively block symplastic transport and communication in higher plants.     

Symplastic Transport of Carboxyfluorescein in Staminal Hairs of Setcreasea purpurea Is Diffusive and Includes Loss to the Vacuole

The kinetics of symplastic transport in staminal hairs of Setcreasea purpurea was studied. The tip cell of a staminal hair was microinjected with carboxyfluorescein (CF) and the symplastic transport of this CF was videotaped and the digital data analyzed to produce kinetic curves. Using a finite difference equation for diffusion between cells and for loss of dye into the vacuole, kinetic curves were calculated and fitted to the observed data. These curves were matched with data from actual microinjection experiments by adjusting K (the coefficient of intercellular junction diffusion) and L (the coefficient of intracellular loss) until a minimum in the least squares difference between the curves was obtained. (a) Symplastic transport of CF was governed by diffusion through intercellular pores (plasmodesmata) and intracellular loss. Diffusion within the cell cytoplasm was never limiting. (b) Each cell and its plasmodesmata must be considered as its own diffusion system. Therefore, a diffusion coefficient cannot be calculated for an entire chain of cells. (c) The movement through plasmodesmata in either direction was the same since the data are fit by a diffusion equation. (d) Diffusion through the intercellular pores was estimated to be slower than diffusion through similar pores filled with water.     

Homeostasis in the vertebrate lens: mechanisms of solute exchange

The eye lens is avascular, deriving nutrients from the aqueous and vitreous humours. It is, however, unclear which mechanisms mediate the transfer of solutes between these humours and the lens' fibre cells (FCs). In this review, we integrate the published data with the previously unpublished ultrastructural, dye loading and magnetic resonance imaging results. The picture emerging is that solute transfer between the humours and the fibre mass is determined by four processes: (i) paracellular transport of ions, water and small molecules along the intercellular spaces between epithelial and FCs, driven by Na(+)-leak conductance; (ii) membrane transport of such solutes from the intercellular spaces into the fibre cytoplasm by specific carriers and transporters; (iii) gap-junctional coupling mediating solute flux between superficial and deeper fibres, Na(+)/K(+)-ATPase-driven efflux of waste products in the equator, and electrical coupling of fibres; and (iv) transcellular transfer via caveoli and coated vesicles for the uptake of macromolecules and cholesterol. There is evidence that the Na(+)-driven influx of solutes occurs via paracellular and membrane transport and the Na(+)/K(+)-ATPase-driven efflux of waste products via gap junctions. This micro-circulation is likely restricted to the superficial cortex and nearly absent beyond the zone of organelle loss, forming a solute exchange barrier in the lens.     

Apparent Transport of Water by Insect Excretory Systems

Insects are capable of producing strongly hyperosmotic urinebut most species do not possess the anatomical equivalent ofthe mammalian kidney's couiitercurrent system. Concentrationof the excreta occurs in the rectum where water is absorbedagainst increasing osmotic gradients without strict dependenceon simultaneous absorption of solute. Properties of this processare reviewed. It is currently postulated that this apparenttransport of water is driven by local transport and recyclingof solute within the lateral intercellular spaces of the epitheliumof the rectal pad. The most concentrated excreta so far reported are those of themealworm, Tenebrio molitor. This species possesses a cryptonephridialcomplex in which the posterior end of the malpighian tubulesis closely applied to the rectum and both are enclosed withina complex membranous sheath. Active transport of potassium chlorideby the malpighian tubules into the complex creates a local highosmotic pressure within the complex which is responsible, inpart if not completely, for removal of water from the rectallumen. This system bears some resemblance to the countercurrentsystem of the mammalian kidney.     

Uptake of horseradish peroxidase from CSF into the choroid plexus of the rat,with special reference to transepithelial transport

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.     

A mechanism for rapid transport of colloidal particles by flounder renal epithelium

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.     

Standing-gradient osmotic flow. A mechanism for coupling of water and solute transport in epithelia

At the ultrastructural level, epithelia performing solute-linked water transport possess long, narrow channels open at one end and closed at the other, which may constitute the fluid transport route (e.g., lateral intercellular spaces, basal infoldings, intracellular canaliculi, and brush-border microvilli). Active solute transport into such folded structures would establish standing osmotic gradients, causing a progressive approach to osmotic equilibrium along the channel's length. The behavior of a simple standing-gradient flow system has therefore been analyzed mathematically because of its potential physiological significance. The osmolarity of the fluid emerging from the channel's open end depends upon five parameters: channel length, radius, and water permeability, and solute transport rate and diffusion coefficient. For ranges of values of these parameters encountered experimentally in epithelia, the emergent osmolarity is found by calculation to range from isotonic to a few times isotonic; i.e., the range encountered in epithelial absorbates and secretions. The transported fluid becomes more isotonic as channel radius or solute diffusion coefficient is decreased, or as channel length or water permeability is increased. Given appropriate parameters, a standing-gradient system can yield hypertonic fluids whose osmolarities are virtually independent of transport rate over a wide range, as in distal tubule and avian salt gland. The results suggest that water-to-solute coupling in epithelia is due to the ultrastructural geometry of the transport route.     

Effect of acetylcholine on the pituitrin induced osmotic flow of water through the wall of the frog urinary bladder]

The role of intercellular pathways in the ADH-dependent water transport was studied on the frog urinary bladder by means of acetylcholine (AC) and other cholinergic compounds. AC (10(-3) M) was found to cause a strong suppression of the pituitrin-stimulated water flow. Analogous effect was produced by AC on the osmotic flow stimulated by cyclic adenosine monophosphate (cAMP) and theolin. The antipituitrin effect was not reproduced either by nicotine, nor by potent M-cholinomimetic agents (methylfurmetide and F-2268), and was not prevented by M- and N-cholynolytic drugs (atropine, metacin, flaxedil, hexamethonium). However, the antipituitrin effect of AC was completely removed by the anticholinesterase drugs with different mode of action (eserine, proserine, armin, acridine iodmethylate, GD-42) in concentrations of 10(-6)--10(-3) M. It was concluded that the smooth muscles contraction with the subsequent closure of the intercellular spaces was not responsible for the antipituitrinic action of AC. This effect appears to be connected with cholinesterase activation. A possible role of the phosphoinositides in the water permeability regulation of the urinary bladder wall is discussed.