Unstirred layer effects in osmotic water flow across gallbladder epithelium

Summary The standard one-dimensional model of the unstirred layer is applied in a re-examination of the experimental results of Wright, Smulders and Tormey (Wright, E.M., Smulders, A.P., Tormey, J. McD., 1972,J. Membrane Biol. 7:198) who reported large transients in the osmotic flux of water from the serosal to the mucosal side of rabbit gallbladder epithelium. They initiated osmosis by the addition of sucrose to the mucosal bathing solution (initially, approximately 300mOsm NaCl) and observed that the initial flux was more than ten times its eventual steady-state value; they interpreted this as a consequence of the piling-up of NaCl in the unstirred tissue layer on the serosal side of the epithelium. The present analysis (both steady-state and unsteady) shows that if measured values of layer thickness are used, together with reasonable values of the reduced diffusivity of NaCl in the tissue and of the fraction of tissue available for water flow, then one would predict a discrepancy of only about 10%, not tenfold, between the initial and final values of the flux. Thus the standard model is inconsistent with the observations. Furthermore, Wright et al's results cannot be used to infer that the osmotic permeability of epithelial cell membranes is much larger than steadystate measurements on whole epithelia would indicate. Mucosal-to-serosal flow is also analyzed, and in this case a considerably greater osmotic permeability is predicted; this result is consistent with the observed changes in structure of the lateral intercellular spaces when the direction of flow is reversed.     

The role of membrane in osmotic flow

Osmosis as a phenomenon caused by internal forces goes on without the necessity for the presence of any external forces. Therefore pressure gradient plays no special role in osmotic flow. Membrane as a component of solution with its molecules possessing some kind of mobility ceases to be a passive obstacle to the flow of other components, but becomes also a co-determining factor in osmotic flow. This has been shown by using the methods of irreversible thermodynamics. In the state of osmotic equilibrium osmosis does not occur. So also the mobility of water molecules which may then be found in tracer experiment does not determine the osmotic permeability coefficient. The coefficientsσ andω as defined by the parameters of the system under the condition of zero volume flow are not directly connected to L_p.     

Volume flows across gallbladder epithelium induced by small hydrostatic and osmotic gradients

Summary The hydraulic conductivity of rabbit gallbladder epithelium has been studied using a continuous volumetric method based on capacitance measurements. The time resolution for measuring osmotic flows is in the range of seconds. Volume flows have been induced by osmotic gradients between 0 and 100 mosmol. In this range the flow-force relation is linear and theP _f value is 9.3×10^–3 cm/sec. After correction for solute polarization effects, theP _f value amounts to 0.05 cm/sec. The observed flow is constant between 5 sec up to 20 min after a sudden increase in the osmolarity of the mucosal solution. The wet weight of the gallbladder tissue decreases by 22% and increases by 30% during osmotic flows from serosa to mucosa and from mucosa to serosa, respectively. Volume flows induced by hydrostatic pressure gradients on the mucosal surface are linearly related to the driving forces between 0 and 40 mbar. TheP _f value is 0.15 cm/sec. The volume flows are constant between 2 sec and 15 min after pressure application. The flow-force relation for pressure gradients on the serosal surface is markedly nonlinear for gradients greater than 5 mbar. Below 5 mbar theP _f value is 4.5 cm/sec. From electrical measurements, e.g., resistance and streaming potentials, and from flux studies with inulin and polyethylene glycol 4000, it is concluded that hydrostatic and osmotic gradients are not comparable when they are applied to gallbladder epithelium. They induce volume flows across different pathways, e.g., osmosis predominantly across the cellular route and pressure filtration predominantly across paracellular routes.     

Brush cells of rodent gallbladder and stomach epithelia express neurofilaments.

It has been suggested that brush cells (BCs), a distinct type of cell occurring in various epithelia of the respiratory and gastrointestinal tracts, may function as receptor cells. The major characteristics of BCs are a prominent brush border and an unusually highly ordered arrangement of cytoskeletal elements (F-actin, microtubules, and intermediate filaments). In this study we aimed to characterize the nature of the intermediate filaments in BCs by light and electron microscopic immunostaining. Gallbladder and stomach specimens from mice and rats, respectively, were fixed in various solutions, embedded either in paraffin or epoxy resin, and processed for immunodetection. Commercially available, well-characterized antibodies against neurofilaments, peripherin, and cytokeratin peptide 18 were used. The polyclonal antiserum cocktail to neurofilaments was applied as a supplement in a double-labeling procedure with anti-actin and anti-cytokeratin 18 antibodies. The results demonstrate that the BCs of both organs express two types of intermediate filaments, i.e., neurofilaments and cytokeratin 18 filaments, and that these have a compartmentalized distribution in the cytoplasm. BCs do not express peripherin. The immunodetection of intermediate filaments distinctive for mature neurons in BCs supports their putative receptor function. The co-expression of neurofilaments and cytokeratins is shown for the first time in healthy tissues.     

Mechanism of osmotic flow in a periodic fiber array

The classic analysis by Anderson and Malone (Biophys J 14: 957-982, 1974) of the osmotic flow across membranes with long circular cylindrical pores is extended to a fiber matrix layer wherein the confining boundaries are the fibers themselves. The equivalent of the well-known result for the reflection coefficient sigma0 = (1 - phi)2, where phi is the partition coefficient, is derived for a periodic fiber array of hexagonally ordered core proteins. The boundary value problem for the potential energy function describing the solute distribution surrounding each fiber is solved by defining an equivalent fluid annulus in which the pressures and osmotic forces are determined. This model is of special interest in the osmotic flow of water across a capillary wall, where recent experimental studies suggest that the endothelial glycocalyx is a quasiperiodic fiber array that serves as the primary molecular sieve for plasma proteins. Results for the reflection coefficient are presented in terms of two dimensionless numbers, alpha = a/R and beta = b/R, where a and b are the solute and fiber radii, respectively, and R is the outer radius of the fluid annulus. In general, the results differ substantially from the classic expression for a circular pore because of the large difference in the shape of the boundary along which the osmotic force is generated. However, as in circular pore theory, one finds that the reflection coefficients for osmosis and filtration are the same.     

Barrier components acting against osmotic water flow inNitella

In order to check whether or not the layer of chloroplasts densely arranged in the cortical gel of aNitella internode offers substantial resistance to osmotic water flow, a material was prepared which had the cortical gel layer freed from chloroplasts by centrifugation either longitudinal or lateral to the cell axis. The water permeability of the cell remained the same as normal even though the chloroplasts were exfoliated from the cell cortex to the extent of 50% of the total area, showing that the chloroplast layer plays hardly any significant part as a barrier to osmotic flow. Since it is known that the layer of the streaming endoplasm is also negligible as resistance against osmotic water flow (Tasawa and Kamiya, 1965), it is concluded that the major barrier components against osmotic flow in theNitella internode are the cell wall, plasmalemma and/or tonoplast.     

The development of osmotic flow through an unstirred layer

We investigate the errors involved in estimating the osmotic permeability of a semi-permeable membrane, from the measured osmotic flow and the difference in concentration of osmotically active solute across it, without taking account of the unstirred layer in the solution next to the membrane. In the problem solved, this layer is represented as a region of thickness δ at the far side of which a solute concentration C_b is imposed for time . The initial diffusion of solute towards the membrane causes the concentration at the membrane C_m to rise, generating an osmotic flow of water, J, whose convective effect opposes the diffusion. The problem is made non-linear by the dependence of J on C_m. Ultimately a steady state is set up, in which C_m is less than C_b. The solution is shown to depend on a single parameter β, equal to (L_pRT) δ C_b/D, where L_pRT is the osmotic permeability of the membrane and D is the diffusivity of solute. Solution of the steady state leads to a prediction of C_m/C_b as a function of β, and analysis of the decay of transient terms leads to a prediction of the decay time π, also as a function of β. Numerical data for membranes with a wide range of osmotic permeabilites, and for a reasonable range of solute, i.e. sucrose, concentrations, suggest that values of β can range from 0.001 or below to 7.5 or above. The former value implies negligible error in neglecting the unstirred layer, while the latter implies a 79%. error. For β = 0.1 and for δ = 2 × 10⁻⁴ m, π is predicted to be around 74 s. This decreases as β increases (for fixed δ); for values of β above about 27, the decay of transients is no longer monotonic but takes the form of damped oscillations.     

Immunological sensitization of opossum gallbladder by naturally acquired stomach roundworm infection.

1. The opossum gallbladder develops an immediate type hypersensitivity as a result of chronic infection with the gastric nematode, Physaloptera turgida. 2. Hypersensitization is evident from a change in net transepithelial ion transport in response to parasite-derived antigen. 3. The antigen-induced change in ion transport involves mediation by histamine and possibly prostaglandins and is independent of intrinsic neural modulation. 4. The functions of the gallbladder epithelium are influenced by reactions of local immune elements that are part of the common mucosal immune system.     

Changes in the intramural neural apparatus of the stomach and gallbladder in the late periods after vagotomy

By means of morphological methods changes in the wall of the stomach central part and in that of the bile bladder have been studied in 15 patients after a remote vagotomy (in 7-17 years). Material of biopsies and resections has been investigated. In the wall of the organs in question focal and diffuse mono- and plasmocytic infiltrates, leucocytic invasion of the mucous membrane epithelium, microerosions, microfocal hemorrhages in the external layers of the muscular sheath have been revealed. Inflammatory-degenerative and dystrophic changes are observed in the intramural ganglia, in large and small fasciculi of the muscular-intestinal nervous plexuses. In the tissues of the organs studied there are no myelin fibers, that are ++pre-ganglial and receptor conductors. The degeneration of these fibers after vagotomy and loss of connections in the organs investigated with the CNS are supposed to result in essential changes not only of the nervous trophic of tissues in all membranes and sheaths, but bring about changes in the intramural nervous apparatus itself. These changes, in their turn, cause imbalance + in the neurogumoral regulation and can be considered as the base of a number of postvagotomic structural-functional disturbances.     

Identification of serotonin from rabbit upper stomach as a stimulant of in vitro gallbladder contraction

Using an in vitro rabbit gallbladder bioassay, the distribution and identification of bioactive substances in rabbit gastrointestinal tract were investigated. Comparison of the bioactivities of tissue extracts before and after cholecystokinin was removed by affinity chromatography demonstrated that the distributions of cholecystokinin and non-cholecystokinin substances were different. While cholecystokinin bioactivity per g of tissue was highest in the duodenum, non-cholecystokinin bioactivity was greatest in the upper stomach. The biochemical properties of the non-cholecystokinin substance in the upper stomach could not be distinguished from those of serotonin. These included molecular weights of 176, identical ultraviolet spectra, similar nuclear magnetic resonance spectra, and co-chromatography in HPLC. By weight, serotonin had 1/6th of the bioactivity of cholecystokinin octapeptide. We conclude that the principal gallbladder-contracting substance in rabbit upper stomach is serotonin.     

Distribution of mucus glycoprotein in four compartments of lamb and sheep stomach

The regional distribution of mucus glycoprotein in lamb before weaning and sheep was studied. Mucus glycoprotein was present in the forestomach region of lamb and glandular stomach (abomasum) of lamb and sheep. Ester sulfates were found in all mucus glycoproteins obtained.     

The unstirred layer during osmotic flow into a tubule

The interaction between osmotic inflow through the wall of a narrow tubule and bulk flow in the tubule is described. Solution are found by a finite difference method, and two approximate analytic solutions are given. Results given here enable more accurate estimates of osmotic permeability to be obtained for the tubule wall. The theory predicts the behaviour of unstirred layers as experimental parameters are varied and enables tubule experiments to be designed so as to reduce unwanted unstirred layer effects.     

Effect of distension on ADH-induced osmotic water flow in toad urinary bladder

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.