Protamine reversibly decreases paracellular cation permeability inNecturus gallbladder

Fromm, Michael; Palant, Carlos E.; Bentzel, Carl J.; Hegel, Ulrich

doi:10.1007/bf01870660

Cited by 55 publications

(37 citation statements)

References 24 publications

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“…34, No. 3, 2001 Our data also show that the water secretion induced by a hypotonic buffer or a choleretic agent, forskolin, was not inhibited by protamine, an agent that blocks water movement via the paracellular pathway [27][28][29] presumably by increasing the barrier function of tight junctions, an interpretation supported by our own data showing an increase in resistance of NRC in culture exposed to protamine. Similar observations have also been made in other AQP-expressing tissues, such as proximal tubules and lung alveolar epithelium.…”

Section: Discussionsupporting

confidence: 76%

“…The relevance of the paracellular pathway on water movement across the biliary epithelial barrier induced by hypotonic buffer and by forskolin was assessed by pretreatment of the IBDUs with protamine, an agent that increases the barrier function of tight junctions between epithelial cells and thus decreases the water movement across an epithelial layer via the paracellular pathway. [27][28][29] In these experiments, after 15 minutes of incubation in the isotonic KRB buffer, the enclosed and polarized IBDUs were treated with 100 g/mL of protamine chloride. After 10 minutes of incubation, the inhibitor was removed by washing IBDUs with isotonic KRB buffer.…”

Section: Effects Of Protamine On Hypotonicity and Forskolin Induced Wmentioning

confidence: 99%

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Water movement across rat bile duct units is transcellular and channel-mediated

et al. 2001

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In recent studies using freshly isolated rat cholangiocytes, we established that water crosses the cholangiocyte membrane by a channel-mediated mechanism involving aquaporins, a family of water-channel proteins. Our goal was to address the importance of channel-mediated water transport in ductal bile formation by employing a physiologic experimental model, the enclosed, polarized rat intrahepatic bile duct unit (IBDU). Expansion and reduction of luminal areas as a reflection of water movement into and out of IBDUs prepared from livers of normal rats were measured by quantitative computer-assisted image analysis. When enclosed IBDUs were exposed to inward or outward osmotic gradients, their luminal area rapidly increased (approximately 25%) or decreased (approximately 20%) reflecting net water secretion or absorption, respectively. These effects were specifically inhibited by 2 water channel blockers, DMSO and HgCl 2 . In both instances, ␤-mercaptoethanol reversed the inhibitory effects. In the absence of an osmotic gradient, choleretic agents (secretin and forskolin) and a cholestatic hormone (somatostatin) induced a significant increase or decrease of IBDU luminal area by 21% and 22%, respectively. These effects were also inhibited by DMSO and reversed by ␤-mercaptoethanol. Under our experimental conditions, DMSO did not interfere with either forskolin-induced cAMP synthesis or the generation of osmotic driving forces via the apical chloride-bicarbonate exchanger. Protamine, an inhibitor of the paracellular pathway, had no effect on hypotonic or forskolin-induced water secretion in IBDUs. These results in a physiologically relevant model of ductal bile formation provide additional support for the concept that osmotically driven and agoniststimulated water movement into (secretion) and out of (absorption) the biliary ductal lumen is transcellular and water channel-mediated. (HEPATOLOGY 2001;34:456-463.)The liver is composed of a variety of resident cells, including hepatocytes and bile duct epithelial cells, or cholangiocytes. These 2 cell types represent the only epithelial cells in the liver and are involved in both absorptive and secretory activities. Bile, mainly composed of solutes and water, is initially elaborated by hepatocytes and subsequently delivered to the intrahepatic bile ducts. 1-3 Cholangiocytes, which form a 3-dimensional network of interconnecting tubular structures, not only deliver bile to the intestine but also participate in ductal bile formation by integrated absorptive and secretory processes that can contribute up to 40% of the daily output of bile depending on the species. 2,4 It is well known that cholangiocytes are involved in hormone-regulated secretion and absorption of water and solutes; for example, secretin stimulates ductal water secretion whereas somatostatin induces ductal water absorption resulting in choleresis and cholestasis, respectively. 4,5 However, the molecular mechanisms by which water moves across biliary epithelia and how water transport is regulated are unclear.T...

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Section: Discussionsupporting

confidence: 76%

Section: Effects Of Protamine On Hypotonicity and Forskolin Induced Wmentioning

confidence: 99%

Water movement across rat bile duct units is transcellular and channel-mediated

et al. 2001

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“…3D). In addition, preincubation with protamine, (300 Ag/ml), which blocks the paracellular pathway in certain tissues (34), did not significantly effect water movement across BDUs (data not shown). These data show that water moves rapidly across polarized cholangiocytes into the lumen of enclosed BDUs in response to osmotic buffers by a HgCl2-inhibitable, protamineindependent process, thus reflecting a transcellular rather than a paracellular pathway.…”

mentioning

confidence: 92%

Cholangiocytes express the aquaporin CHIP and transport water via a channel-mediated mechanism.

Roberts

Yano

Ueno

et al. 1994

Proc. Natl. Acad. Sci. U.S.A.

109

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Cholangiocytes line the intrahepatic bile ducts and regulate salt and water secretion during bile formation, but the mechanism(s) regulating ductal water movement remains obscure. A water-selective channel, the aquaporin CHIP, was recently described in several epithelia, so we tested the hypothesis that osmotic water movement by cholangiocytes is mediated by CHIP. Isolated rodent cholangiocytes showed a rapid increase in volume in the presence of hypotonic extracellular buffers; the ratio of osmotic to diffusional permeability coefficients was >10. The osmotically induced increase in cholangiocyte volume was inversely proportional to buffer osmolality, independent of temperature, and reversibly blocked by HgCl2. Also, the luminal area of isolated, enclosed bile duct units increased after exposure to hypotonic buffer and was reversibly inhibited by HgCl2. RNase protection assays, anti-CHIP immunoblots, and immunocytochemistry confirmed that CHIP transcript and protein were present in isolated cholangiocytes but not in hepatocytes. These results demonstrate that (0) isolated cholangiocytes and intact, polarized bile duct units manifest rapid, mercury-sensitive increases in cell size and luminal area, respectively, in response to osmotic gradients and (it) isolated cholangiocytes express aquaporin CHIP at both the mRNA and the protein level. The data implicate aquaporin water channels in the transcellular movement of water across cholangiocytes lining intrahepatic bile ducts and provide a plausible molecular explanation for ductal water secretion.Bile formation by the liver involves secretion of bile by hepatocytes and delivery to a network of interconnecting ducts where bile is modified by cholangiocytes, the epithelial cells that line these conduits inside the liver. Bile secretion by cholangiocytes contributes to total bile flow through the spontaneous and agonist-induced secretion of both ions and water (1). While data have been accumulating on the cellular mechanisms regulating ion transport by cholangiocytes (2-4), the mechanisms regulating water movement across biliary epithelia remain undefined (5, 6).Conceptually, water may move across biliary epithelia by two pathways: a paracellular pathway between cholangiocytes or a transcellular pathway across both the apical and basolateral cholangiocyte plasma membranes (5, 7). Further, transcellular water movement may occur by simple diffusion across the lipid bilayer or through discrete membrane proteins that form water channels (8). A family of membrane water channels, referred to as aquaporins, was recently identified (9). The aquaporin CHIP [ghannel-forming integral membrane protein of 28 kDa] is the first characterized molecular water channel (10). When expressed in Xenopus laevis oocytes (11) or reconstituted into proteoliposomes (12), CHIP behaves as an osmotically driven, water-selective pore capable of transporting water across the plasma membrane in a rapid, relatively temperature-independent and mercury-sensitive manner. Moreover, immunohistochem...

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“…It has been reported that protamine increases resistance of paracellular shunt pathway in various epithelia [3, 4, 5, 6, 7, 8]. Koyama et al [6, 7, 8]reported that protamine selectively decreases cation permeability by acting mainly on the paracellular shunt pathway in the nephron segments having leaky epithelia.…”

Section: Discussionmentioning

confidence: 99%

“…Protamine has been used to characterize the paracellular shunt pathway in various epithelia including renal tubules [3, 4, 5, 6, 7, 8]. In almost all the epithelia examined so far, the paracellular shunt pathways are characterized by cation-selective permeability which is inhibited by protamine.…”

Section: Introductionmentioning

confidence: 99%

Effect of Protamine on Ion Selectivity of Superficial and Juxtamedullary Proximal Straight Tubules

Takahashi

Taniguchi²,

Muto³

et al. 1999

Nephron

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Protamine is known to inhibit cation selectivity of the paracellular shunt pathway in several leaky nephron segments. Ion permeability of the superficial (SF) proximal straight tubules (PST) is selective to Cl^–, whereas that of the juxtamedullary (JM) PST is selective to Na⁺. Protamine was used to estimate the contribution of the paracellular shunt pathway to ion selectivity in these segments. PSTs were isolated from the kidneys of Japanese white rabbit and microperfused in vitro. The ratio of Na⁺ to Cl^– permeability (P_Na/P_Cl) was estimated from the diffusion potential (dV_T) generated by NaCl gradients. When 300 µg/ml protamine was added to the lumen, P_Na/P_Cl was decreased from 0.33 to 0.26 in SF-PST and from 1.80 to 1.29 in JM-PST, respectively, as the results of inhibition of cation selectivity. Addition of 30 U/ml heparin to the lumen, which neutralizes protamine, returned the ratios toward the control levels. Protamine exhibited similar effect on P_Na/P_Cl in the presence or the absence of ouabain in the bath, indicating that the observed voltage deflections were independent of the transcellular active transport process. Although P_Na/P_Cl was different between SF- and JM-PST, protamine inhibits Na⁺ permeability in both segments. Preferential Cl^– permeability in the SF-PST may be intrinsic to the paracellular route of this segment. The mechanism of this anion selectivity remains to be established.

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Protamine reversibly decreases paracellular cation permeability inNecturus gallbladder

Cited by 55 publications

References 24 publications

Water movement across rat bile duct units is transcellular and channel-mediated

Water movement across rat bile duct units is transcellular and channel-mediated

Cholangiocytes express the aquaporin CHIP and transport water via a channel-mediated mechanism.

Effect of Protamine on Ion Selectivity of Superficial and Juxtamedullary Proximal Straight Tubules

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