Expression of aquaporin-4 water channels in rat cholangiocytes

Abstract

We recently reported that secretin induces the exocytic insertion of functional aquaporin-1 water channels (AQP1) into the apical membrane of cholangiocytes and proposed that this was a key process in ductal bile secretion. Because AQP1 is present on the basolateral cholangiocyte membrane in low amounts, we hypothesized that another AQP must be expressed at this domain to facilitate transbasolateral water movement. Thus, we investigated the expression, subcellular localization, possible regulation by secretin, and functional activity of AQP4, a mercury-insensitive water channel expressed in other fluid transporting epithelia. Using reverse transcription-polymerase chain reaction (RT-PCR) on RNA prepared from purified rat cholangiocytes, we amplified a product of 311 bp that was 100% homologous to the reported AQP4 sequence. RNase protection assay confirmed the presence of an appropriate size transcript for AQP4 in cholangiocytes. Immunoblotting detected a band of approximately 31 kd corresponding to AQP4 in basolateral but not apical membranes of cholangiocytes. Secretin did not alter the amount of plasma membrane AQP4 but, as expected, induced AQP1 redistribution from intracellular to apical plasma membranes. Functional studies showed that AQP4 accounts for about 15% of total cholangiocyte membrane water permeability. Our results indicate that: (1) cholangiocytes express AQP4 messenger RNA (mRNA) and protein and (2) in contrast to AQP1, which is targeted to the apical cholangiocyte membrane by secretin, AQP4 is constitutively expressed on the basolateral cholangiocyte membrane and is secretin unresponsive. The data suggest that AQP4 facilitates the basolateral transport of water in cholangiocytes, a process that could be relevant to ductal bile formation. (HEPATOLOGY 2000;31:1313-1317.)Cholangiocytes, or bile duct epithelial cells, account for secretin-induced ductal bile secretion. Cholangiocytes express aquaporin-1 (AQP1), a plasma membrane channel protein through which water moves passively in response to osmotic gradients. 1 We recently found that secretin induces the exocytic insertion of AQP1 specifically into the apical plasma membrane domain of cholangiocytes, a mechanism that seems to be key in ductal bile secretion 2-4 and is similar to the antidiuretic hormone (ADH)-induced insertion of AQP2 into the apical plasma membrane of the epithelial cells lining the kidney collecting duct. 5 The fact that AQP1 is present in the basolateral cholangiocyte membrane at only very low amounts 4 suggested to us that another member of the aquaporin family of proteins is expressed in the basolateral cholangiocyte domain. The basolateral expression of a water channel protein would facilitate the transbasolateral transport of water during bile secretion, fluid presumably derived from the closely apposed peribiliary vascular plexus. One candidate protein for this role is AQP4, initially called MIWC (for mercurial insensitive water channel). This water channel has been localized to the basolateral membranes of several flu...

The stem cell niche of human livers: Symmetry between development and regeneration

“…By contrast, the large ones express the secretin receptor, cytochrome P450 2E1 and the ClϪ/HC03Ϫ exchanger, but do not express BCL-2. We propose that the findings of La Russo 39,40 and those of Alpini and associates 38 are indicative of a maturational lineage of biliary cells with subpopulations of small intrahepatic biliary epithelia representing early lineage stage(s), and with the extrahepatic biliary cells representing later lineage stages. Our working model of the early stages of the liver's maturational lineages is presented in Fig.…”

Expression and Localization of Aquaporin Water Channels in Rat Hepatocytes

“…AQP5 in the salivary gland, which is normally sequestered in intracellular vesicles, redistributes to the apical plasma membrane upon stimulation by the neurotransmitter, epinephrine (24). Finally, we have demonstrated that AQP4 is constitutively expressed on the basolateral membrane of cholangiocytes, whereas AQP1, normally residing in intracellular vesicles, undergoes secretin-induced exocytic insertion into the apical cholangiocyte membrane (15,25,26). Thus, a generalizable model accounting for the physiological role of AQPs in water-transporting epithelia is developing in which multiple, differentially localized, constitutively expressed or trafficked AQPs function together to facilitate regulated transcellular water movement in water-transporting epithelia in both the basal and stimulated states.…”

The Water Channel Aquaporin-8 Is Mainly Intracellular in Rat Hepatocytes, and Its Plasma Membrane Insertion Is Stimulated by Cyclic AMP

“…aquaporin-1, located mainly in the apical (luminal) membrane domain and in intracellular vesicles (11,24,25), and aquaporin-4, present only in the basolateral membrane domain (26). The cAMP-dependent hormone secretin regulates the subcellular distribution of aquaporin-1 by stimulating its exocytic insertion exclusively in the apical plasma membrane domain of cholangiocytes (25), a mechanism that seems to be involved in ductal bile formation.…”

“…The cAMP-dependent hormone secretin regulates the subcellular distribution of aquaporin-1 by stimulating its exocytic insertion exclusively in the apical plasma membrane domain of cholangiocytes (25), a mechanism that seems to be involved in ductal bile formation. In contrast, aquaporin-4 in cholangiocytes is constitutively expressed in the basolateral plasma membrane domain (26). In hepatocytes, cyclic AMP-regulated targeting of membrane transporters has been reported to in- volve both the apical as well as the basolateral plasma membrane domains (15)(16)(17)(18)(19)(20).…”