Tetraethylammonium block of water flux in Aquaporin-1 channels expressed in kidney thin limbs of Henle's loop and a kidney-derived cell line.
© Yool et al; licensee BioMed Central Ltd. 2002
Received: 22 October 2001
Accepted: 15 March 2002
Published: 15 March 2002
Aquaporin-1 (AQP1) channels are constitutively active water channels that allow rapid transmembrane osmotic water flux, and also serve as cyclic-GMP-gated ion channels. Tetraethylammonium chloride (TEA; 0.05 to 10 mM) was shown previously to inhibit the osmotic water permeability of human AQP1 channels expressed in Xenopus oocytes. The purpose of the present study was to determine if TEA blocks osmotic water flux of native AQP1 channels in kidney, and recombinant AQP1 channels expressed in a kidney derived MDCK cell line. We also demonstrate that TEA does not inhibit the cGMP-dependent ionic conductance of AQP1 expressed in oocytes, supporting the idea that water and ion fluxes involve pharmacologically distinct pathways in the AQP1 tetrameric complex.
TEA blocked water permeability of AQP1 channels in kidney and kidney-derived cells, demonstrating this effect is not limited to the oocyte expression system. Equivalent inhibition is seen in MDCK cells with viral-mediated AQP1 expression, and in rat renal descending thin limbs of Henle's loops which abundantly express native AQP1, but not in ascending thin limbs which do not express AQP1. External TEA (10 mM) does not block the cGMP-dependent AQP1 ionic conductance, measured by two-electrode voltage clamp after pre-incubation of oocytes in 8Br-cGMP (10–50 mM) or during application of the nitric oxide donor, sodium nitroprusside (2–4 mM).
TEA selectively inhibits osmotic water permeability through native and heterologously expressed AQP1 channels. The pathways for water and ions in AQP1 differ in pharmacological sensitivity to TEA, and are consistent with the idea of independent solute pathways within the channel structure. The results confirm the usefulness of TEA as a pharmacological tool for the analysis of AQP1 function.
Tetraethylammonium is known as a pore-occluding blocker of voltage-gated potassium channels , but it also blocks other cationic channels such as calcium-dependent K+ channels [2, 3] and the nicotinic acetylcholine receptor . TEA at 0.1 to 10 mM also inhibits osmotic water flux through human AQP1 channels expressed in Xenopus oocytes, decreasing the net swelling rate in hypotonic saline by 30–40% as compared to AQP1-expressing oocytes not treated with TEA . This blocking effect on osmotic water flux was demonstrated to involve AQP1 channels specifically by using site-directed mutagenesis (tyrosine 186 to phenylalanine) to generate a Y186F AQP1 channel that is insensitive to block by TEA, but retains sensitivity to block by mercury. The blocking effect of mercury in AQP1 channels is dependent on a neighboring residue, cysteine 189 . TEA offers attractive advantages over mercury as a reversible and less toxic blocker for AQP1 channels in experimental analyses of water channel function; however, the relevance of TEA as a blocker for AQP1 channels outside the Xenopus expression system has not been examined previously.
Differences in properties (including pharmacological sensitivities) have been observed for ion channels expressed in oocytes as compared to those in native tissues. For example, the Torpedo acetylcholine receptor and the Shaker K+ channel protein differ in the fractions of protein glycosylated, the composition of the oligosaccharide chains, and the degree of protein maturation when expressed in Xenopus oocytes as compared with channels in native tissue or other expression systems [7, 8]. Differences in glycosylation patterns can influence the binding of external blocking agents. The presence of tissue-specific targeting signals not recognized in Xenopus oocytes may lead to protein degradation . These discrepancies raised the question of whether AQP1 channels in native tissues are sensitive to TEA, as they are in Xenopus oocytes. Data presented here show that TEA is effective as a blocker of AQP1 channels expressed in a mammalian renal cell line and in native renal epithelial membrane.
AQP1 channels are complex solute conductors. They are constitutively permeable to water, and also function as regulated non-selective cation channels  when gated by intracellular cyclic GMP [11, 12]. Even though only a small proportion of the AQP1 channels that are present in the membrane appear to be available for cGMP-activation of the ionic conductance (1/56,000), model-based calculations nonetheless suggest that this contribution could be meaningful in a physiological setting . Unlike transporters which move substantial amounts of substrate by design, ion channels function to change membrane potential by the net movement of relatively tiny amounts of charge (~10 picomoles per cm2 membrane to generate a change of 100 mV) , hence the low proportion of active AQP1 ion channels is consistent with a functional goal of gated ionic signaling that is distinct from that of massive constitutive osmotic water permeability.
The role of the AQP1 channel in mediating both water and ion fluxes raises interesting questions about whether the pathways are shared or structurally distinct. Expression of AQP1 channels in bilayers has shown that the water permeability but not the ionic conductance is blocked by p-chloromethylbenzenesulfonate, indicating that the two fluxes may involve distinct pathways . Work presented here provides an additional line of evidence that the pathways for water and ions in AQP1 can be distinguished pharmacologically, using the blocker TEA as a probe. In combination with data from three-dimensional imaging analyses , these results support the suggestion that the AQP1 channel may contain separate parallel pathways for ion and water fluxes. These findings have potential significance in understanding the complex role for AQP1 in processes of salt and water movement across cell membranes.
Results and Discussion
TEA inhibition of water flux in AQP1-expressing MDCK cells
TEA inhibition of water flux in renal tubule
Lack of TEA block of ion channel conductance
Results presented here show that TEA serves as an effective, though partial, blocker of the osmotic water permeability mediated by human AQP1 channels expressed in a heterologous mammalian cell line and by native AQP1 channels in isolated rat renal descending thin limbs of Henle's loops. Before the characterization of TEA, mercurial compounds were among the only known blockers of AQP1, affecting both the constitutive water flux and the regulated ion conductance [6, 10]. For AQP1 channels expressed in Xenopus oocytes, the block of osmotic water flux is 5% in 0.01 mM TEA, 21% in 1 mM TEA, and 36% in 10 mM TEA, as referenced to the osmotic water flux in control AQP1-expressing oocytes . Data presented here show that the level of block by TEA is comparable or greater for AQP1 channels expressed in mammalian cells. 1 mM TEA produces a 34% block of osmotic water flux in MDCK cells, and 10 mM TEA produces a 50% block in renal descending thin limbs. The effect of TEA on water permeability in cells and tissues that express high levels of AQP1 channels is likely to be significant, given the substantial amount of water that can be moved across AQP1-expressing epithelia (for example, see data on descending thin limbs in Fig. 3). The idea that TEA might be a candidate for a lead compound for the development of drugs with possible clinical applications  is supported by the present data that show the effectiveness of TEA in two different mammalian models of AQP1-mediated osmotic water permeability. While the usefulness of TEA in vivo as a research tool may be limited by toxicity and effects on other channels at higher concentrations, it offers a degree of reversibility and selectivity not seen with mercury, an agent which acts by covalent modification of all exposed cysteine residues [21, 22].
The regulated ion channel function of AQP1 is not blocked by external TEA. An intriguing question remains regarding the location of the permeation pathway for cations in AQP1 channels. Analysis of the crystal structure of GlpF suggests that potential ion binding sites may be located in the central pore, at the four-fold axis of symmetry in the tetrameric association of subunits that form the channel . Interestingly, this arrangement of a tetramer of subunits to form a central ion pore is a hallmark of the family of voltage-gated ion channels , which rely on a tetrameric organization to create a central pore for ions that is lined by sequences contributed from all four subunits. In contrast, the four individual pores for water or glycerol, located within each subunit of AQP1 or GlpF, appear to be lined with hydrophobic residues expected to preclude ion permeability [15, 24, 25]. Importantly, TEA distinguishes between the two pathways in AQP1, supporting the idea that the ion and water pores are physically distinct although they co-exist in the same channel complex.
The AQP1-mediated ion conductance may contribute measurably to the function of tissues in which AQP1 is abundantly expressed, such as in the mammalian renal proximal tubule . Further work is needed to assess the additional levels of regulatory control that may constrain the availability of the AQP1 proteins to serve as ion channels gated by cGMP, within a background of constitutive water channel activity. The maintenance of an ionic pore, as well as a conserved cGMP-binding domain in the AQP1 carboxy tail for regulating activation , suggests that the ion channel function is likely to be essential for physiological regulatory processes that have yet to be fully appreciated.
Materials and Methods
Madin-Darby Canine Kidney (MDCK) cells were obtained as a gift from Dr. R. Lynch, University of Arizona, and used between passages 13–14. Cells were grown and maintained in Dulbecco's Modified Eagle Medium (Life Technologies) that was supplemented with 10% fetal bovine serum (Hyclone), 170 mM glutamine and penicillin/streptomycin (100 μg/ml, Life Technologies) in humidified air containing 5% carbon dioxide at 37°C.
The adenovirus (AV) backbone for the aquaporin-1 (AQP1) sense and antisense AV constructs was a replication-deficient "first-generation" AV with deletions of the E1 and E3 genes . This "empty" AV contains the cytomegalovirus (CMV) promoter and bovine growth hormone polyadenylation (bHG) site separated by a polylinker that was used to clone AQP1 DNA as described previously . A recombinant AQP1 virus was constructed using a plasmid containing the coding sequence for AQP1, pCHIPev . pCHIPev was digested and the AQP1 insert was subcloned into the shuttle vector pSKAC, creating pSKAC/AQP1. pSKAC contains map units 0.0–1.3 of the AV which includes the left terminal repeat of AV, a CMV promoter, an AMV translation enhancer and a polylinker region. DNA fragments containing AQP1 DNA were liberated from pSKAC after restriction and ligated into adenovirus as described previously . Human embryonic kidney cells (293 cells) were transfected with ligation mixture and individual viruses were isolated from cell lysates by two consecutive rounds of plaque purification using an agar overlay as described previously . Individual viruses were amplified in 293 cells and purified over cesium step gradient. Individual AV DNA titers were determined by three different methods: 1) plaque titration on 293 cells, 2) immunofluorescence microscopy of AV protein expression (anti-penton group antigen, clone 143, Biodesign, Kennebunk, ME) in 293 cells infected with serial dilutions of AV and 3) absorbance at 260 nm (pfu/ml = A260 × dilution × 1010).
Osmotic water flux assay
Water permeability was measured as the net fluid movement driven by an osmotic gradient across intact monolayers of adenovirus-infected MDCK cells, by methods similar to those reported previously [27, 29]. Cells were seeded onto Transwell filters (Costar, 1 cm2, 0.4 μm pore size) at a density of 1.5 × 105 cells per well. Cells were maintained in humidified air containing 5% CO2 for two weeks to allow for cell-cell junctions to mature. Cells were infected at the apical surface with adenovirus containing AQP1 cDNA or with empty adenovirus at a multiplicity of infection (MOI) of 10. Five days post-infection, all medium was removed carefully and completely from both upper (apical) and lower (basolateral) chambers. Exactly one milliliter of fresh prewarmed isotonic medium (~300 mOsM) was added to the lower chamber and exactly 175 μl of hypertonic medium (~450 mOsM) was added to the upper chamber at time zero. In experimental groups, TEA chloride was present at a 1 mM concentration in both the upper and lower chambers. After four hours of incubation at 37°C, all of the medium from each of the upper chambers was removed carefully and volume was measured using an analytical balance.
Sodium dodecyl sulfate (SDS) solubilized whole cell lysates containing 5% β-mercaptoethanol were electrophoresed into 12% polyacrylamide gels containing 0.1 % SDS. Fractionated proteins were blotted onto nitrocellulose using the Transblot system as per manufacturer instructions (Biorad, Hercules, CA). The blots were preincubated for 30 minutes at 22°C in Tris-buffered saline containing 5% nonfat powdered milk and 0.2% Tween-20 (TBS-T), and were then probed with affinity purified anti-AQP1 IgG (1:5000) for 2 hr at 22°C. The blots were washed (3 × 15 min) in TBS-T and were incubated for 2 hours with horseradish peroxidase-conjugated secondary antibodies (goat anti-rabbit, 1:5000, Pierce). The blots were washed (3 × 15 min) in TBS-T and specific labeling was visualized following enhanced chemiluminescence (Pierce, Rockford, IL) and exposure (10 seconds) to ECL-Hyperfilm (Amersham, Arlington Heights, IL). Immunoblots were digitized using the UVP gel documentation system and densitometry was performed using LabWorks software (Upland, CA).
Isolated perfused rat inner medullary thin limbs
Animals and dissection of tubules for immunocytochemistry and for in vitro fluid movement measurements
Young male Munich-Wistar rats (Harlan Sprague-Dawley, Indianapolis, In), 90 g average body weight, were maintained on Teklad Mouse/Rat Diet No. 7001 with free access to water. Dissection of inner medullary thin limbs of Henle's loop from fresh rat renal tissue for immunocytochemistry and for in vitro microperfusion was performed as described in detail previously . We used descending thin limb (DTL) and ascending thin limb (ATL) segments from the top 70% of the inner medulla above the pre-bend level. Descending and ascending thin limbs were identified by cell type using an inverted microscope with Nomarski differential interference contrast (DIC) optics at 400× magnification [17, 18].
Rabbit antibody recognizing the carboxy tail domain of AQP1 (from W.D. Stamer and J.W. Regan, University of Arizona) was used for immunocytochemical confirmation of protein expression in isolated thin limbs of Henle as described previously . Tubules were positioned onto a glass microscope slide covered with a layer of Cell-Tak® adhesive (Becton Dickinson, Bedford, MA). Tubules were then fixed in 4% paraformaldehyde for 10 min and permeabilized with 100% methanol for 2 min at -20°C. Primary antibody or non-immune rabbit serum (diluted 1:500) was applied overnight at 4°C., followed by incubation with biotinylated goat anti-rabbit antibody (diluted 1:100, 60 min), and incubation with streptavidin conjugated to Cy5 (60 min). Digital fluorescent images were obtained with a Leica-TCS confocal microscope.
Perfusion and bathing solution for in vitro microperfusions
The Ringer solution used for perfusing and bathing the tubules was that initially used by Chou and Knepper  in perfusion of thin limbs from chinchilla kidneys and modified by us. It contained (in mM): 130 NaCl, 20 HEPES, 5 NaHCO3, 2.5 K2HPO4, 2 CaCl2, 1.2 MgSO4, 5.5 glucose, and 5 urea. The osmolality was about 290 mOsm/kg H2O, the pH was adjusted to 7.4 while the solution was gassed with 95 O2/5% CO2.
Perfusion of tubules in vitro
We perfused the dissected tubules in vitro by a technique essentially the same as that originally described by Burg et al.  and modified for use in our laboratory [17, 32, 33]. The perfusion rate was about 15–20 nl/min. The temperature of the bathing chamber was maintained at 37°C during perfusion and the bath was covered with a layer of light paraffin oil. To check for leaks and to measure net fluid movement across the epithelium with an imposed osmotic gradient, [14C]dextran (MW ~70,000) was added to the perfusate.
Determination of net water movement and water permeability of thin limbs perfused in vitro
There is no net transepithelial water movement with solutions of identical osmolalities in the bath and lumen. For this study, however, we determined the occurrence of net water absorption and water permeability when we imposed an osmotic gradient from bath to lumen (see below). Net water absorption, Jv (nl min-1 mm tubule length-1), was determined in each collection period with [14C]dextran in the perfusate using the following relationship: Jv= (Vi - Vo)/L. In this equation, Vi(initial perfusion rate, nl/min) is calculated by dividing the cpm of 14C in the collected fluid by the cpm/nl of 14C in the initial perfusate and by the time of the collection period; Vo (collection rate, nl/min) is measured directly from the collection; and L (mm) is the length of the tubule perfused, measured with an ocular micrometer. For purposes of determining the water permeability of DTL and ATL, we imposed a 100 mOsmol/kg H2O osmotic gradient from bath to lumen with sucrose and determined Jv as just described. Water permeability (Pf, μm s-1) was then determined from the equation Pf = Jv/(As Vw δCosmol) where As = luminal surface area (π DL), Vw = partial molar volume of water (18 ml/M), δCosmol = transepithelial osmolality gradient . When the effect of TEA on water movement was examined, TEA bromide (10 mM) was added to the bathing medium and the NaCl concentration was reduced by 10 mM to maintain the osmolality constant. Collection periods were 5 min in length and each tubule served as its own control for measuring net water flux and water permeability in the presence and absence of TEA. [14C]dextran (sp act 1.08 mCi/g) was obtained from American Radiolabeled Chemicals, St. Louis, MO., USA. Results are summarized as means ± SE. The n value is the number of tubules. Each tubule came from a different animal.
Adult female Xenopus laevis were anesthetized with tricaine methane sulfonate (MS-222, Sigma Chemical Co). Stage V-VI oocytes were removed by partial laparotomy and defolliculated by collagenase treatment . Cloned human Aquaporin1 DNA was provided by Dr. P. Agre , linearized with XbaI and used as a template for in vitro synthesis of cRNA with T3 RNA polymerase. Prepared oocytes were injected with 50 nl of sterile water (control oocytes) or 50 nl of sterile water containing AQP1 cRNA (~0.2 to 0.5 ng/nl) and were incubated for 2–5 days at 18°C in culture medium (ND96: 96 mM NaCl, 2 mM KCl, 1.8 mM CaCl2, 1 mM MgCl2, 5 mM Hepes, 2.5 mM pyruvic acid, 100 U/ml penicillin, and 100 μg/ml streptomycin, pH 7.6) to allow protein expression.
Two-electrode voltage clamp recordings were performed at room temperature with electrodes (0.5–3 MΩ) filled with 3 M KCl. Data were recorded with a GeneClamp 500 (Axon Instruments, Foster City, CA), filtered at 2 kHz, digitized at 50 to 2000 μs and analyzed with pClamp software (Axon Instruments). Control recording saline for two-electrode voltage clamp contained (in mM): 100 NaCl, 5 MgCl2, and 5 HEPES, pH 7.3. TEA saline was made with (in mM) 10 TEA.Cl, 90 mM NaCl, 5 MgCl2, and 5 HEPES, pH 7.3. Activation of the ionic current was induced by preincubation of AQP1-expressing oocytes for 10–30 minutes in 10 mM 8Br-cGMP (Sigma) in high K+ saline, containing (in mM): 30 KCl, 80 K gluconate, 5 MgCl2, 5 HEPES, pH 7.3, or by addition of 2–4 mM sodium nitroprusside (SNP), a stimulator of guanylate cyclase and cGMP production [34, 35]. Control oocytes treated in the same conditions showed no induction of an ionic current. Higher concentrations of SNP (>10 mM) affected ionic permeability in some batches of control oocytes and were not used. Current voltage relationships were assessed in Na+ control saline and in TEA saline to assess potential blocking effects of TEA.
All chemicals were purchased from standard sources except as specified above and were of the highest purity available.
List of abbreviations
Net osmotic water absorption rate
Madin-Darby Canine Kidney cell
Osmotic water permeability
Sodium nitroprusside (nitric oxide donor)
Tetraethylammonium (chloride salt)
We thank Dr. John Regan for antibodies to AQP1, Amy Marble and Dr. Kathryn Bolles for technical assistance, Dr. Heddwen Brooks for helpful discussions, Neil Atodaria and Eileen Ryan for technical assistance with permeability assays using MDCK cells, and Dr. Ron Lynch for the gift of MDCK cells (University of Arizona). Funding was provided by NIH R01 GM59986 (AJY), American Health Assistance Foundation #G2001-026 (WDS), and NIH R01 DK16294 (WHD) and Research to Prevent Blindness Foundation.
- Armstrong CM: Interaction of tetraethylammonium ion derivatives with the potassium channels of giant axons. J Gen Physiol. 1971, 58: 413-437. 10.1085/jgp.58.4.413.PubMed CentralView ArticlePubMedGoogle Scholar
- Latorre R, Vergara C, Hidalgo C: Reconstitution in planar lipid bilayers of a Ca2+-dependent K+ channel from transverse tubule membranes isolated from rabbit skeletal muscle. Proc Natl Acad Sci U S A. 1982, 79: 805-809.PubMed CentralView ArticlePubMedGoogle Scholar
- Lang DG, Ritchie AK: Tetraethylammonium blockade of apamin-sensitive and insensitive Ca2+-activated K+ channels in a pituitary cell line. J Physiol. 1990, 425: 117-132.PubMed CentralView ArticlePubMedGoogle Scholar
- Bissada NK, Welch LT, Finkbeiner AE: Uropharmacology: VII. Ganglionic stimulating and blocking agents. Urology. 1978, 11: 425-431. 10.1016/0090-4295(78)90252-2.View ArticlePubMedGoogle Scholar
- Brooks HL, Regan JW, Yool AJ: Inhibition of aquaporin-1 water permeability by tetraethylammonium: involvement of the loop E pore region. Mol Pharmacol. 2000, 57: 1021-1026.PubMedGoogle Scholar
- Preston GM, Jung JS, Guggino WB, Agre P: The mercury-sensitive residue at cysteine 189 in the CHIP28 water channel. J Biol Chem. 1993, 268: 17-20.PubMedGoogle Scholar
- Buller AL, White MM: Altered patterns of N-linked glycosylation of the Torpedo acetylcholine receptor expressed in Xenopus oocytes. J Membr Biol. 1990, 115: 179-189.View ArticlePubMedGoogle Scholar
- Santacruz-Toloza L, Huang Y, John SA, Papazian DM: Glycosylation of shaker potassium channel protein in insect cell culture and in Xenopus oocytes. Biochemistry. 1994, 33: 5607-5613.View ArticlePubMedGoogle Scholar
- Chen PX, Mathews PM, Good PJ, Rossier BC, Geering K: Unusual degradation of alpha-beta complexes in Xenopus oocytes by beta-subunits of Xenopus gastric H-K-ATPase. Am J Physiol. 1998, 275: C139-145.PubMedGoogle Scholar
- Yool AJ, Stamer WD, Regan JW: Forskolin stimulation of water and cation permeability in aquaporin 1 water channels. Science. 1996, 273: 1216-1218.View ArticlePubMedGoogle Scholar
- Anthony TL, Brooks HL, Boassa D, Leonov S, Yanochko GM, Regan JW, Yool AJ: Cloned human aquaporin-1 is a cyclic GMP-gated ion channel. Mol Pharmacol. 2000, 57: 576-588.PubMedGoogle Scholar
- Saparov SM, Kozono D, Rothe U, Agre P, Pohl P: Water and ion permeation of aquaporin-1 in planar lipid bilayers. Major differences in structural determinants and stoichiometry. J Biol Chem. 2001, 276: 31515-31520. 10.1074/jbc.M104267200.View ArticlePubMedGoogle Scholar
- Yool AJ, Weinstein AM: New roles for old holes: Ion channel function in aquaporin-1. News Physiological Sciences. 2001,Google Scholar
- Hille B: Ion Channels of Excitable Membranes. Edited by: Sunderland MA. 2001, Sinauer Associates Inc., 3Google Scholar
- Fu D, Libson A, Miercke LJ, Weitzman C, Nollert P, Krucinski J, Stroud RM: Structure of a glycerol-conducting channel and the basis for its selectivity. Science. 2000, 290: 481-486. 10.1016/S0009-2614(98)00538-7.View ArticlePubMedGoogle Scholar
- Kovbasnjuk O, Leader J, Weinstein A, Spring A: Water does not flow across the tight junctions of MDCK cell epithelium. Proc. Natl. Acad. Sci. USA. 1998, 95: 6526-6530. 10.1073/pnas.95.11.6526.PubMed CentralView ArticlePubMedGoogle Scholar
- Brokl OH, Dantzler WH: Amino acid fluxes in rat thin limb segments of Henle's loop during in vitro microperfusion. Am J Physiol. 1999, 277: F204-210.PubMedGoogle Scholar
- Chou CL, Knepper MA: In vitro perfusion of chinchilla thin limb segments: segmentation and osmotic water permeability. Am J Physiol. 1992, 263: F417-426.PubMedGoogle Scholar
- Nielsen S, Pallone T, Smith B, Christensen E, Agre P, Maunsbach A: Aquaporin-1 water channels in short and long loop descending thin limbs and in descending vasa recta in rat kidney. Am. J. Physiol. 1995, 268: F1023-F1037.PubMedGoogle Scholar
- Nielsen S, Smith B, Christensen E, Knepper M, Agre P: Chip28 waterchannels are localized in consitutively water-permeable segments of thenephron. J. Cell Biol. 1993, 120: 371-383. 10.1083/jcb.120.2.371.View ArticlePubMedGoogle Scholar
- Marston AW, Wright HT: A method for covalent insertion of mercury into the cysteine disulfide bridges of proteins. J Biochem Biophys Methods. 1984, 9: 307-314. 10.1016/0165-022X(84)90014-9.View ArticlePubMedGoogle Scholar
- Pallone TL, Kishore BK, Nielsen S, Agre P, Knepper MA: Evidence that aquaporin-1 mediates NaCl-induced water flux across descending vasa recta. Am J Physiol. 1997, 272: F587-596.PubMedGoogle Scholar
- Jan LY, Jan YN: Tracing the roots of ion channels. Cell. 1992, 69: 715-718. 10.1016/0092-8674(92)90280-P.View ArticlePubMedGoogle Scholar
- Murata K, Mitsuoka K, Hirai T, Walz T, Agre P, Heymann JB, Engel A, Fujiyoshi Y: Structural determinants of water permeation through aquaporin-1. Nature. 2000, 407: 599-605. 10.1038/35036519.View ArticlePubMedGoogle Scholar
- Ren G, Reddy VS, Cheng A, Melnyk P, Mitra AK: Visualization of a water-selective pore by electron crystallography in vitreous ice. Proc Natl Acad Sci U S A. 2001, 98: 1398-1403. 10.1073/pnas.041489198.PubMed CentralView ArticlePubMedGoogle Scholar
- Drazner MH, Peppel KC, Dyer S, Grant AO, Koch WJ, Lefkowitz RJ: Potentiation of beta-adrenergic signaling by adenoviral-mediated gene transfer in adult rabbit ventricular myocytes. J Clin Invest. 1997, 99: 288-296.PubMed CentralView ArticlePubMedGoogle Scholar
- Stamer WD, Peppel K, O'Donnell ME, Roberts BC, Wu F, Epstein DL: Expression of aquaporin-1 in human trabecular meshwork cells: role in resting cell volume. Invest Ophthalmol Vis Sci. 2001, 42: 1803-1811.PubMedGoogle Scholar
- Preston GM, Agre P: Isolation of the cDNA for erythrocyte integral membrane protein of 28 kilodaltons: member of an ancient channel family. Proc Natl Acad Sci U S A. 1991, 88: 11110-11114.PubMed CentralView ArticlePubMedGoogle Scholar
- Delporte C, Hoque AT, Kulakusky JA, Braddon VR, Goldsmith CM, Wellner RB, Baum BJ: Relationship between adenovirus-mediated aquaporin 1 expression and fluid movement across epithelial cells. Biochem Biophys Res Commun. 1998, 246: 584-588. 10.1006/bbrc.1998.8668.View ArticlePubMedGoogle Scholar
- Pannabecker TL, Dahlmann A, Brokl OH, Dantzler WH: Mixed descending- and ascending-type thin limbs of Henle's loop in mammalian renal inner medulla. Am JPhysiol Renal Physiol. 2000, 278: F202-208.Google Scholar
- Burg M, Grantham J, Abramow M, Orloff J: Preparation and study of fragments of single rabbit nephrons. Am J Physiol. 1966, 210: 1293-1298.PubMedGoogle Scholar
- Dantzler WH: Characteristics of urate transport by isolated perfused snake proximal renal tubules. Am J Physiol. 1973, 224: 445-453.PubMedGoogle Scholar
- Dantzler WH: PAH transport by snake proximal renal tubules: differences from urate transport. Am J Physiol. 1974, 226: 634-641.PubMedGoogle Scholar
- Omerovic A, Leonard JP, Kelso SR: Effects of nitroprusside and redox reagents on NMDA receptors expressed in Xenopus oocytes. Brain Res Mol Brain Res. 1994, 22: 89-96. 10.1016/0169-328X(94)90035-3.View ArticlePubMedGoogle Scholar
- Sawada M, Ichinose M, Stefano GB: Nitric oxide inhibits the dopamine-induced K+ current via guanylate cyclase in Aplysia neurons. J Neurosci Res. 1997, 50: 450-456. 10.1002/(SICI)1097-4547(19971101)50:3<450::AID-JNR11>3.0.CO;2-A.View ArticlePubMedGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article: verbatim copying and redistribution of this article are permitted in all media for any purpose, provided this notice is preserved along with the article's original URL.