Osmotic Water Flow Research Articles

KCl transport mechanism across charged mosaic membrane in KCl–sucrose mixed system

The ion and water transports across charged mosaic membrane were investigated in the presence of nonelectrolyte (sucrose). In addition, according to irreversible thermodynamics, phenomenological equations were developed in the system of KCl–sucrose mixed solution. From the analysis of the volume flux and solute flux based on the developed equations, practical membrane parameters such as σ and ω were estimated and compared with the simplest system (KCl/water system). As a result, water flows through charged mosaic membrane caused by KCl diffusion, and osmotic flow of water caused by sucrose were additive. On the other hand, in spite of prediction due to theoretical consideration, the effect of sucrose on KCl transport was too small to detect experimentally.

Water transport controversies--an overview.

The last 20 years have seen major advances in the understanding of the mechanisms and molecular bases of water transport across cell membranes. Prominent among these advances was the discovery of a family of transmembrane proteins that permeate exclusively or dominantly water (aquaporins), followed by structural studies identifying the intramolecular permeation pathway, as well as by studies directed to a detailed understanding of these molecules in health and disease. Agre and colleagues (Agre et al. 2002) discuss the molecular structure of aquaporins and the physiology and pathophysiology of aquaporins in diverse organs and tissues. Another important advance was the development of methods for rapid measurements of changes in cell volume. This resulted in accurate estimates of cell-membrane water permeability, and in the prediction that very small osmotic gradients would suffice to drive transepithelial transport, without the need for the existence of compartments of much higher osmolality than that of the bathing solution. Around 1990, faced with the question of what pathways and driving forces underlie net water fluxes across cell membranes, most physiologists working on animal cells would have had direct and clear answers. Water moves across the phospholipid moiety of the plasma membrane (although this membrane can be extremely water-tight in certain cases), and through water pores, in cells that express these proteins. The driving force is the difference in effective osmolality across the cell membrane. Enlightened physiologists working on epithelia would add that transepithelial water absorption or secretion is always a passive phenomenon, secondary to net solute transport in the same direction. Further, complicated compartment models, such as the standing osmotic gradient hypothesis, are unnecessary to explain water transport, because very small osmotic gradients, perhaps too small to measure directly, are a sufficient driving force. The question of how water fluxes are partitioned between transcellular and paracellular pathways remained difficult to answer because of the lack of direct measurements. The two central questions in the problem of water transport across simple or complex membranes are the pathway and the mechanism. Considering the cell membrane, one asks whether water permeates the lipid moiety and/or transmembrane proteins. These pathways are expected to have different properties, including the possibility that transmembrane water pores could also be permeable to other molecules. Concerning the driving force for water transport, in principle it could be primary active, secondary active or passive. At the time prior to the discoveries that provide the experimental bases for the Topical Reviews in this Special Issue, the virtually unanimous opinion of the experts would have been that water transport is passive. With respect to transepithelial water transport, the relative contributions of the transcellular and paracellular routes remain to be determined. There is no agreement on the molecular mechanism, i.e. whether there is secondary-active water transport at the cell membrane in addition to simple osmosis, or on the precise nature of osmotic water flow, i.e. is there truly isosmotic transport? This Special Issue highlights the current excitement in this area of physiology. Three current controversies in the field of water transport are discussed. In each case, the ‘controversial’ novel theory is presented (in the form of a Topical Review) and discussed critically in the form of a Perspective or a Research Paper. The three controversies presented in this Special Issue were debated at the XXXIV International Congress of Physiological Science in Christchurch, New Zealand.

Diuretic effect and mechanism of action of parsley

This work provides substantial evidence for the advocated diuretic effect of parsley in folk medicine and determines the mechanism of action of the herb. Rats offered an aqueous parsley seed extract to drink, eliminated a significantly larger volume of urine per 24 h as compared to when they were drinking water. These findings were supported by the results of other experiments using an in situ kidney perfusion technique which demonstrated also a significant increase in urine flow rate with parsley seed extract. This effect was still apparent in presence of amiloride, furosemide and in the absence of sodium, but not in the absence of potassium, suggesting that the diuretic effect of the herb is mediated through an increase in K + retention in the lumen. Parsley extract, was shown on the other hand, to reduce the activity of the Na +–K + ATPase in both cortex and medulla homogenates. Such an inhibition would decrease apical cellular Na + reabsorption, lower K + secretion, increase K + concentration in the intercellular space and consequently would inhibit passive K + influx across the tight junctions. The mechanism of action of parsley seems to be mediated through an inhibition of the Na +–K + pump that would lead to a reduction in Na + and K + reabsorption leading thus to an osmotic water flow into the lumen, and diuresis.

Effects of vasopressin on hyaluronate hydrolase activities and water permeability in the frog urinary bladder.

The involvement of enzymes catabolizing hyaluronic acid (hyaluronidase, beta-glucuronidase, N-acetyl-beta-D-hexosaminidase) in the hydroosmotic effect of vasopressin in the frog (Rana ridibunda) urinary bladder was studied. It was observed that vasopressin (50 nM), an agonist of V2 receptors, L-desamino-8-D-arginine-vasopressin (dDAVP, 1.5 microM) and forskolin (30 microM) activated the enzymes and caused their release into Ringer solution at the mucosal side, together with an increase in osmotic water flow. The effect of AVP on enzyme activity developed 10 min after the hydroosmotic response. Cytochalasin B (a specific inhibitor of actin filament elongation, 50 nM) blocked the hydroosmotic response to AVP; hyaluronate hydrolase activity increased in the bladder tissue but not in Ringer solution. It is suggested that the involvement of hyaluronate hydrolases in AVP's effect is mediated by a cAMP-dependent mechanism and provides favorable conditions for an increase in the permeability of glycosaminoglycan structures adjacent to the apical cell surface.

Hydraulic conductivity of rice roots.

A pressure chamber and a root pressure probe technique have been used to measure hydraulic conductivities of rice roots (root Lp(r) per m(2) of root surface area). Young plants of two rice (Oryza sativa L.) varieties (an upland variety, cv. Azucena and a lowland variety, cv. IR64) were grown for 31-40 d in 12 h days with 500 micromol m(-2) s(-1) PAR and day/night temperatures of 27 degrees C and 22 degrees C. Root Lp(r) was measured under conditions of steady-state and transient water flow. Different growth conditions (hydroponic and aeroponic culture) did not cause visible differences in root anatomy in either variety. Values of root Lp(r) obtained from hydraulic (hydrostatic) and osmotic water flow were of the order of 10(-8) m s(-1) MPa(-1) and were similar when using the different techniques. In comparison with other herbaceous species, rice roots tended to have a higher hydraulic resistance of the roots per unit root surface area. The data suggest that the low overall hydraulic conductivity of rice roots is caused by the existence of apoplastic barriers in the outer root parts (exodermis and sclerenchymatous (fibre) tissue) and by a strongly developed endodermis rather than by the existence of aerenchyma. According to the composite transport model of the root, the ability to adapt to higher transpirational demands from the shoot should be limited for rice because there were minimal changes in root Lp(r) depending on whether hydrostatic or osmotic forces were acting. It is concluded that this may be one of the reasons why rice suffers from water shortage in the shoot even in flooded fields.

Identification and localization of aquaporin water channels in human salivary glands.

Aquaporin (AQP) water channels are expressed in a variety of fluid-transporting epithelia and are likely to play a significant role in salivary secretion. Our aim was to identify and localize the aquaporins expressed in human salivary glands. Total RNA was extracted from human parotid, submandibular, sublingual, and labial glands and from human brain. Expression of aquaporin mRNA was assessed by RT-PCR using specific primers for human AQP1, AQP3, AQP4, and AQP5. All four aquaporins were detected by RT-PCR in all of the glands, and the sequences were confirmed after further amplification with nested primers. Cleaned PCR products were then used as (32)P-labeled cDNA probes in a semiquantitative Northern blot analysis using glyceraldehyde-3-phosphate dehydrogenase as reference. Only AQP1, AQP3, and AQP5 mRNAs were present at significant levels. AQP localization was determined by immunohistochemistry on paraffin sections using affinity-purified primary antibodies and peroxidase-linked secondary antibodies. Each salivary gland type showed a broadly similar staining pattern: AQP1 was localized to the capillary endothelium and myoepithelial cells; AQP3 was present in the basolateral membranes of both mucous and serous acinar cells; AQP4 was not detected; and AQP5 was expressed in the luminal and canalicular membranes of both types of acinar cell. We conclude that AQP3 and AQP5 together may provide a pathway for transcellular osmotic water flow in the formation of the primary saliva.

The Peritoneal Microcirculation in Peritoneal Dialysis

This paper deals with the peritoneal microcirculation and with peritoneal exchange occurring in peritoneal dialysis (PD). The capillary wall is a major barrier to solute and water exchange across the peritoneal membrane. There is a bimodal size-selectivity of solute transport between blood and the peritoneal cavity, through pores of radius ∼40–50 Å as well as through a very low number of large pores of radius ∼250 Å. Furthermore, during glucose-induced osmosis during PD, nearly 40% of the total osmotic water flow occurs through molecular water channels, termed “aquaporin-1.” This causes an inequality between 1−σ and the sieving coefficient for small solutes, which is a key feature of the “threepore model” of peritoneal transport. The peritoneal interstitium, coupled in series with the capillary walls, markedly modifies small-solute transport and makes large-solute transport asymmetric. Thus, although severely restricted in the blood-to-peritoneal direction, the absorption of large solutes from the peritoneal cavity occurs at a high clearance rate (∼1 mL/min), largely independent of molecular radius. True absorption of macromolecules to the blood via lymphatics, however, seems to be occurring at a rate of ∼0.2 mL/min. Several controversial issues regarding transcapillary and transperitoneal exchange mechanisms are discussed in this paper.

Coupling between Na+, sugar, and water transport across the intestine.

Water is absorbed across the small intestine in the absence of external driving forces. However, it has been established that water transport is secondary to active sodium transport. In the upper intestine both sodium and water absorption are largely dependent on the presence of D-glucose. The link between active sodium transport and glucose is the coupled transport of sodium and glucose across the brush border membrane of enterocytes by the Na+/glucose cotransporter (SGLT1). Na+ that enters the cells with glucose is pumped out towards the blood by 3Na+/2K+ pumps on the basolateral membrane, and glucose passes out across the basolateral membrane by facilitated diffusion, the net result being that glucose and sodium are transported across the epithelium. The coupling between Na+, glucose, and water transport is less well understood. It is commonly thought that Na+ transport increases the local osmotic pressure in the lateral intercellular spaces, and that this in turn generates osmotic water flow across the epithelium. Recent work suggests a more direct link between Na+, glucose, and water transport; that is, water is cotransported along with Na+ and sugar through SGLT1. Here we review the evidence for Na+/glucose/water cotransport.

Water Transfer by The Toad Bladder

ABSTRACT. Studies have been made on the isolated urinary bladder of the toad, Bufo marinus, in an attempt to investigate the effect of vasopressin on the permeability of water from mucosal surface to serosal surface of the toad bladder. The method adapted was that described by Bentley ( 1 ). The bilobed bladder of the toad is devided into two separate sacs. Each of the sacs is filled with a dilute Ringers solution and then immersed in aerated isotonic Ringers solution. The rate of water loss along the imposed osmotic gradient is estimated by weighing the sacs in air at 30 minute intervals and nothing the weight loss in that time period. In most studies one bladder sac serves as a control for the contra lateral experimental obtained from the same animal. Osmotic flow of water is negligible in both sacs during the initial control periods. However, the addition of vasopressin to the solution bathing the serosal surface of the membrane result in a market increase in net water movement. The effect is readily reversed by rinsing the bladder and adding hormone free Ringgers solution to the serosal surface. Characteristically no response is elicited by addition of hormone to the mucosal bathing solution.

Blood Flow Does Not Limit Peritoneal Transport

We investigated the assumption that blood flow to the microvessels underlying the peritoneum does not limit solute or water exchange between the blood and the dialysis fluid. Small plastic chambers were affixed to the serosal side of the liver, cecum, stomach, and abdominal wall of anesthetized rats. Solutions that contained labeled solutes or that were made hypertonic were placed into the chambers, which restricted the area of transfer across the tissue to the base of the chamber and which permitted calculation of mass or water transfer rates on the basis of area. The local blood flow was monitored continuously with a laser Doppler flowmeter during three periods of observation: control, after 50%-70% reduction of the blood flow, and postmortem. Urea transfer across all serosa, except for the liver, showed no difference in mean mass transfer coefficient (cm/min) between control (0.0038-0.0046) and after 70% flow reduction (0.0037-0.0040), but demonstrated a significant decrease with blood flow equal to zero (0.0020). These tissues demonstrated small but insignificant decreases in osmotic water flow into the chamber (0.7-0.9 microL/min/cm2 under control conditions versus 0.4-0.7 microL/min/cm2 with reduced blood flow). The liver demonstrated limitations in water and solute transport with a 70% decrease in blood flow. Because the liver makes up a small part of the peritoneal area, we conclude that large drops in blood flow do not limit overall solute or water transfer across the peritoneum during dialysis, and therefore acute peritoneal dialysis may be an appropriate modality for ICU patients in shock and renal failure.

Low Trehalase Activity Is Associated with Abdominal Symptoms Caused by Edible Mushrooms

Background: The purpose of the study was to evaluate whether maldigestion of trehalose causes abdominal symptoms and which available diagnostic method best distinguishes intolerant from tolerant subjects. Methods: A 25-g oral trehalose load test was performed in 64 subjects. The 19 experiencing clear symptoms constituted the trehalose-intolerant subjects. Changes from base-line levels of blood glucose, breath hydrogen, and methane and symptoms were recorded after the test. Trehalase activity was determined in serum and on a duodenal biopsy specimen obtained by endoscopy. Results: Intolerant subjects were best differentiated from tolerant subjects by changes in breath gases (hydrogen and methane) and duodenal trehalase to sucrase ratio. The change in breath gases correlated inversely with duodenal trehalase activity, duodenal trehalase to sucrase ratio, and plasma trehalase activity. The correlation between serum and duodenal trehalase activities was on the order of 0.6. Two subjects were found to have trehalase deficiency. Conclusions: It is obvious that trehalose maldigestion can cause symptoms similar to those of lactose maldigestion and intolerance. Three factors control the genesis of symptoms: 1) the activity of small-bowel trehalase: if it is low, trehalose is maldigested and more trehalose is passed into the colon; 2) the maldigested trehalose, which causes osmotic water flow into the colon, resulting in loose stools and diarrhea; and 3) most importantly, the microflora of the colon, from which symptoms will arise if there are bacteria capable of producing gases from maldigested trehalose. If colonic bacteria cannot produce gases, then distention of the abdomen and intestinal gas expulsion as eructations and flatus will not occur.

Role of aquaporins in renal water handling: physiology and pathophysiology.

the six bilayer spanning domains are connected by fiveloops (A–E in Figure 1). The molecule consists of twoAn essential function of the mammalian kidney is its repeats of three a-helices which are 180°mirror imagesability to conserve or excrete free water independent of each other (Figure 1). Each repeat contains theof changes in solute excretion. Early physiological highly conserved family characteristic asparagine-studies have provided evidence for constitutive absorp- proline-alanine (NPA) sequence in loops B and E.tion of about 90% of the GFR volume in proximal LoopsBandEarepostulatedtofoldbackintothetubules and descending limbs of Henle’s loop, whereas membrane and form the water pore. Recently, the three-facultative water reabsorption occurs in collecting dimensional structure of AQP1 was determined at 6A˚ducts under control of the antidiuretic hormone argin- resolution by cryo-electron crystallography [8]. AQP1ine vasopressin (AVP). Although all cell membranes is a homotetramer containing four independent aqueoushave a surpringly high diVusive water permeability, it channels. It was shown that each monomer is composedis now clear that water transport in the nephron is of six tilted a-helices which form a barrel that enclosesfacilitated by water channels [1]. a central density which is attributed to the functionallyDiscovery of the aquaporin family of water channels important NPA boxes in loops B and E. Given thehas provided insight in the molecular mechanisms by highly homologues primary structures of AQP1 and thewhich rapid osmotic water flow occurs in kidney and other aquaporins, it is likely that all AQPs have a morein other organs. Nine mammalian aquaporins, AQPs, or less similar three-dimensional structure. The lesshave been cloned to date, of which AQP1, 2, 3, 4, 6, conserved region among the aquaporins is the carboxy-7 are expressed in kidney (for review see [1–3], new terminus and this property has been exploited to gener-aquaporins [4–6]). Aquaporins belong to an old ubi- ate highly specific polyclonal antibodies which havequitous major intrinsic protein (MIP) family of trans- been very useful for localization studies.membrane channel proteins of which at present closeto hundred members have been identified [7]. The

Gill blood flow control.

The arrangement of the fish gill vasculature is quite complex, and varies between the different fish groups. The use of vascular casting techniques has greatly enhanced our knowledge of the anatomy of the branchial microcirculation, not least through the contributions of Pierre Laurent and co-workers at Strasbourg. At different physiological situations, the contact surface between water and blood (functional surface area) varies to balance oxygen uptake against osmotic water flow ("respiratory-osmoregulatory compromise"). This is controlled by nerves and by blood-borne or locally released substances that affect blood flow patterns in the gill. Histochemical techniques have been used to demonstrate neurotransmitter substances in the branchial innervation. In combination with physioly-osmoregulatory compromise" at different physiological situations.

Water permeability of the oral epithelial layers of the sea anemone, Anemonia viridis

Water permeability of the oral epithelial layers of the anthozoan Anemonia viridis was studied. We determined a diffusional water permeability of 17 × 10–5 cm s–1 and an osmotic water permeability of 2.5 10–5 cm s–1. These permeability coefficients were independent of the state of contraction of the tentacle. Compared to other tissues, water permeability in sea anemones is relatively low. This result is quite surprising, since we determined in a previous study that the ionic permeabilities of the oral epithelial layers of A. viridis were characteristic of a leaky tissue (Bénazet-Tambutté et al., '96a). The low water permeability of the oral epithelial layers of A. viridis can be considered as an adaptational mechanism for maintaining a hydrostatic skeleton. The ratio Po/Pd was about 14. The osmotic flow was symmetric whatever the direction of the gradient. Temperature did not affect diffusional water flux. The Arrhenius energy of activation for osmotic flux was about 5 kcal mol–1. Osmotic water flow was insensitive to mercury. Our results suggest the possible existence of HgCl2-insensitive water channels. J. Exp. Zool. 279:1–8, 1997.© 1997 Wiley-Liss, Inc.

Water permeability of the oral epithelial layers of the sea anemone,Anemonia viridis

Water permeability of the oral epithelial layers of the anthozoan Anemonia viridis was studied. We determined a diffusional water permeability of 17 × 10–5 cm s–1 and an osmotic water permeability of 2.5 10–5 cm s–1. These permeability coefficients were independent of the state of contraction of the tentacle. Compared to other tissues, water permeability in sea anemones is relatively low. This result is quite surprising, since we determined in a previous study that the ionic permeabilities of the oral epithelial layers of A. viridis were characteristic of a leaky tissue (Benazet-Tambutte et al., '96a). The low water permeability of the oral epithelial layers of A. viridis can be considered as an adaptational mechanism for maintaining a hydrostatic skeleton. The ratio Po/Pd was about 14. The osmotic flow was symmetric whatever the direction of the gradient. Temperature did not affect diffusional water flux. The Arrhenius energy of activation for osmotic flux was about 5 kcal mol–1. Osmotic water flow was insensitive to mercury. Our results suggest the possible existence of HgCl2-insensitive water channels. J. Exp. Zool. 279:1–8, 1997.© 1997 Wiley-Liss, Inc.

Germination of Nosema algerae (Microspora) Spores: Conditional Inhibition by D2O, Ethanol and Hg2+ Suggests Dependence of Water Influx upon Membrane Hydration and Specific Transmembrane Pathways

The germination of microsporidian spores under conditions expected to affect water flow across the plasma membrane-wall complex was studied by assessing their responses to in vitro stimulation with Na+ or K+. Partial or full substitution of common water with D2O, which more effectively coats ions and electrostatically-charged cell surfaces with relatively stable hydration layers, delayed and inhibited spore germination in a concentration-dependent manner; yet, preincubation in 100% D2O did not change the normal response to standard stimulation. Water structure-breaking conditions, such as an increase in temperature (within the 15 degrees C to 40 degrees C range) or in ionic strength (1- to 10-fold normal), opposed the inhibition by D2O and allowed significant stimulation by Li+, the monovalent cation with the largest hydration diameter and a usually weak stimulant action on the spores. Ethanol, known to reduce water permeation across cell membranes and phospholipid bilayers, also caused a powerful and dose-dependent (1% to 4% v/v) inhibition of spore germination, but pretreatment with ethanol did not affect the normal response. HgCl2, an inhibitor of specific water channels, blocked spore germination at just 250 microM in the normal stimulation solution irrespective of the temperature, and permitted only a delayed response in high salt stimulation solutions. However,the inhibition by Hg2+ was abolished by the simultaneous presence of 2-mercaptoethanol in the medium. These results suggest (1) that spore germination is keenly dependent upon the hydration states of both the plasma membrane-wall complex and the stimulant ions, and (2) that osmotic water flows into the spores through specific transmembrane pathways with critical sulfhydryl groups, i.e., analogous to the water channels that facilitate water movements across the plasma membranes of highly permeable cells.

Function and regulation of seed aquaporins

The discovery of water channel proteins named aquaporins has shed new light on the molecular mechanisms of transmembrane water transport in higher plants. As with their animal counterparts, plant aquaporins belong to the large MIP family of transmembrane channels. An increasing number of aquaporins is now being identified on both the vacuolar and plasma membranes of plant cells, but their integrated function remains unclear. Aquaporin α-TIP is specifically expressed in the membrane of protein storage vacuoles in seeds of many plant species. α-TIP was previously shown to undergo phosphorylation in bean seeds. The functional significance of this process was further investigated after heterologous expression of the protein in Xenopus oocytes. Using site-directed mutagenesis of α-TIP and in vitro and in vivo phosphorylation by animal cAMP-dependent protein kinase, it is shown that, in oocytes, direct phosphorylation of α-TIP occurs at three distinct sites and stimulates its water channel activity. In addition to aquaporin phosphorylation, other mechanisms that target aquaporin function are used by living cells to regulate their membrane water permeability. These are the fine control of aquaporin gene expression and, in animal cells only, the regulated trafficking of water channel-containing vesicles. The present work and studies by others on the phosphorylation of nodulin-26, an ion channel protein homologous to α-TIP, provide novel insights into the mechanisms of plant membrane protein regulation. These studies might help identifying and characterizing novel membrane-bound protein kinases and phosphatases. Finally, an integrated function for seed vacuolar aquaporins is discussed. During germination, the rehydration of seed cells, the drastic changes in vacuole morphology, the breakdown and the mobilization of storage products from the vacuole may create osmotic perturbations in the cytoplasm. The fine tuning of TIP aquaporin activity may help control the kinetics and amplitude of osmotic water flows across the tonoplast to achieve proper cytoplasm osmoregulation and control of vacuolar volume.

Characteristics of Hg‐ and Zn‐sensitive Water Channels in the Plasma Membrane of Chara Cells*

Abstract:Hydraulic conductivity (Lp) of the plasma membrane of Chara corallina was inhibited by HgCl2 maximally by about 95%. The inhibition was reversed by 2‐mercaptoethanol, reconfirming the observation obtained by Henzler and Steudle (1995). The results suggest that osmotic water transport through Chara cells occurs mostly via mercury‐sensitive water channels containing thiol groups. ZnCl2 dissolved in APW (pH 5.6) also inhibited Lp by about 80% within 1–2 h, while ZnCl2 dissolved in Hepes‐Tris buffer (pH 7.4) inhibited it by about 90% within several minutes. Inhibition of Lp by ZnCl2 was also reversed by 2‐mercaptoethanol, suggesting that zinc acts also on thiol groups of water channel proteins. Cells from which tonoplast had been removed by ECTA were as sensitive to both HgCl2 and ZnCl2 (pH 7.4) as normal cells. This demonstrates that water channels sensitive to thiol reagents really exist in the plasma membrane. On the other hand, ZnCl2 (pH 5.6) did not inhibit Lp of tonoplast‐free cells. This may be accounted for by assuming first that Hg‐ and Zn‐sensitive thiol groups of water channels may exist on the cytoplasmic side, and second that ZnCl2 in acidic medium may exist in ionized species which can be chelated by EGTA after permeation. The polar water permeability, or the endoosmotic Lp being larger than the exoosmotic one, was not affected by lowering the rate of osmosis by decreasing the osmotic gradient for transcellular osmosis down to 0.02 M sorbitol. The polarity disappeared when osmotic water flow through water channels was completely inhibited by HgCl2. Thus the polarity is assumed to be intrinsic to water channels in the plasma membrane.

Transport in Shales and the Design of Improved Water-Based Shale Drilling Fluids

Summary Transport of water and ions in shales and its impact on shale stability were studied to facilitate the improvement of water-based muds as shale drilling fluids. Transport parameters associated with flows driven by gradients in pressure and chemical potential were quantified in key laboratory and full-scale experiments. The experimental results show that the low-permeability matrices of intact, clay-rich shales can act as imperfect or "leaky" membranes that will sustain osmotic flow of water. Moreover, the ability of shales to act as osmotic membranes is shown to provide a powerful new means for stabilizing these rocks when exposed to water-based drilling fluids. Guidelines are presented for effective exploitation of shale membrane action and induced osmotic flows through optimized water-based drilling fluid formulation. In addition, special attention is given to induced electro-osmotic water flow in shales driven by electric potential gradients, which may provide an exciting, new, environmentally benign means for stabilizing shale formations.

Rat Hepatocytes Transport Water Mainly via a Non-channel-mediated Pathway

During bile formation by the liver, large volumes of water are transported across two epithelial barriers consisting of hepatocytes and cholangiocytes (i.e. intrahepatic bile duct epithelial cells). We recently reported that a water channel, aquaporin-channel-forming integral protein of 28 kDa, is present in cholangiocytes and suggested that it plays a major role in water transport by these cells. Since the mechanisms of water transport across hepatocytes remain obscure, we performed physiological, molecular, and biochemical studies on hepatocytes to determine if they also contain water channels. Water permeability was studied by exposing isolated rat hepatocytes to buffers of different osmolarity and measuring cell volume by quantitative phase contrast, fluorescence and laser scanning confocal microscopy. Using this method, hepatocytes exposed to hypotonic buffers at 23 degrees C increased their cell volume in a time and osmolarity-dependent manner with an osmotic water permeability coefficient of 66.4 x 10(-4) cm/s. In studies done at 10 degrees C, the osmotic water permeability coefficient decreased by 55% (p < 0.001, at 23 degrees C; t test). The derived activation energy from these studies was 12.8 kcal/mol. After incubation of hepatocytes with amphotericin B at 10 degrees C, the osmotic water permeability coefficient increased by 198% (p < 0.001) and the activation energy value decreased to 3.6 kcal/mol, consistent with the insertion of artificial water channels into the hepatocyte plasma membrane. Reverse transcriptase polymerase chain reaction with hepatocyte RNA as template did not produce cDNAs for three of the known water channels. Both the cholesterol content and the cholesterol/phospholipid ratio of hepatocyte plasma membranes were significantly (p < 0.005) less than those of cholangiocytes; membrane fluidity of hepatocytes estimated by measuring steady-state anisotropy was higher than that of cholangiocytes. Our data suggests that the osmotic flow of water across hepatocyte membranes occurs mainly by diffusion via the lipid bilayer (not by permeation through water channels as in cholangiocytes).