Renal water reabsorption: A physiologic retrospective in a molecular era

Renal water reabsorption: A physiologic retrospective in a molecular era. The cloning and sequencing of the aquaporin water channels has been an enormous advance in the biomedical sciences, as recognized by the award of the Nobel Prize to Peter Agre last year. Among many other examples, expression of aquaporin proteins in Xenopus oocytes and other heterologous expression systems has confirmed two important models of renal function: the increase in the water permeability of the collecting duct by antidiuretic hormone (ADH), and the mechanism of near isosmotic volume reabsorption by the proximal tubule. These mechanisms were the subjects of intensive investigation by numerous investigators, including Thomas E. Andreoli, who is being honored by this symposium, and who developed many of the key concepts in these areas. His early work with artificial lipid bilayer membranes and the pore-forming antibiotic amphotericin provided the rigorous foundation in experimental and conceptual modeling techniques that he later applied to physiologic and pathophysiologic mechanisms in the kidney, which are summarized in this retrospective. Dr. Andreoli and his colleagues proposed a water channel mechanism for the action of ADH, which has been confirmed by the cloning and heterologous expression of aquaporin-2. They also proposed that volume reabsorption by the proximal tubule depended on a very high hydraulic conductivity and the development of luminal hypotonicity produced by active solute reabsorption. This model has also been confirmed in mice in which aquaporin-1 expression is knocked out, resulting in a low proximal tubule water permeability that exaggerates the development of luminal hypotonicity

cAMP Increases Density of ENaC Subunits in the Apical Membrane of MDCK Cells in Direct Proportion to Amiloride-sensitive Na+ Transport

Antidiuretic hormone and/or cAMP increase Na+ transport in the rat renal collecting duct and similar epithelia, including Madin-Darby canine kidney (MDCK) cell monolayers grown in culture. This study was undertaken to determine if that increment in Na+ transport could be explained quantitatively by an increased density of ENaC Na+ channels in the apical membrane. MDCK cells with no endogenous ENaC expression were retrovirally transfected with rat α-, β-, and γENaC subunits, each of which were labeled with the FLAG epitope in their extracellular loop as described previously (Firsov, D., L. Schild, I. Gautschi, A.-M. Mérillat, E. Schneeberger, and B.C. Rossier. 1996. Proc. Natl. Acad. Sci. USA. 93:15370–15375). The density of ENaC subunits was quantified by specific binding of 125I-labeled anti-FLAG antibody (M2) to the apical membrane, which was found to be a saturable function of M2 concentration with half-maximal binding at 4–8 nM. Transepithelial Na+ transport was measured as the amiloride-sensitive short-circuit current (AS-Isc) across MDCK cells grown on permeable supports. Specific M2 binding was positively correlated with AS-Isc measured in the same experiments. Stimulation with cAMP (20 μM 8-p-chlorothio-cAMP plus 200 μM IBMX) significantly increased AS-Isc from 11.2 ± 1.3 to 18.1 ± 1.3 μA/cm2. M2 binding (at 1.7 nM M2) increased in direct proportion to AS-Isc from 0.62 ± 0.13 to 1.16 ± 0.18 fmol/cm2. Based on the concentration dependence of M2 binding, the quantity of Na+ channels per unit of AS-Isc was calculated to be the same in the presence and absence of cAMP, 0.23 ± 0.04 and 0.21 ±0.05 fmol/μA, respectively. These values would be consistent with a single channel conductance of ∼5 pS (typically reported for ENaC channels) only if the open probability is <0.02, i.e., less than one-tenth of the typical value. We interpret the proportional increases in binding and AS-Isc to indicate that the increased density of ENaC subunits in the apical membrane can account completely for the Isc increase produced by cAMP

From the Editors: A Guide for Peer Review in the Field of Exercise Science

International Journal of Exercise Science 11(1): 1112-1119, 2018. Since its inception, the mission of the International Journal of Exercise Science (IJES) has been to engage student researchers, to provide an outlet for peer-review and possible publication of their work, and to grant an opportunity for them to gain experience as peer-reviewers. The Editors of IJES take pride in providing these opportunities for student involvement, and we are constantly seeking new and innovative ways to enhance students’ professional development. As our readership has expanded across the globe and our scope has broadened to cross many Kinesiology related disciplines, we believe it is timely to revisit the purpose of peer-review, give advice on best practices, and provide a template for reviews. Presenting these guiding principles should simplify and streamline both the review and the revision processes for students and professionals alike

Transport of amino acids in ehrlich ascites cells: Competitive stimulation

1. 1. The term `competitive stimulation' refers to an experimental finding in which the uptake of one amino acid is increased in the presence of another amino acid when both are initially present extracellularly. The uptake of both amino acids of such pairs was investigated with use of double-label techniques.2. 2. For all amino acid pairs examined which showed competitive stimulation, the uptake of one amino acid was increased whereas that of the other was decreased. Furthermore, the decrease in uptake of the latter was considerably greater than the increase in uptake of the former.3. 3. In time studies it was found that the distribution ratio of the amino acid whose uptake was stimulated was increased at all incubation times up to 30 min. The amino acid whose uptake was inhibited showed not only a markedly decreased uptake but also a significantly altered curve of time dependent of uptake.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/33287/1/0000679.pd

Na+ and K+ electrochemical potential gradients and the transport of [alpha]-aminoisobutyric acid in Ehrlich ascites tumor cells

1. 1. The uptake of [alpha]-aminoisobutyric acid (AIB) was measured in Ehrlich ascites tumor cells which were treated by osmotic shock and cold incubation to elevate cell Na+ and lower K+. These cells were suspended in media in which Na+ was partially replaced by choline or K+, thereby reversing the Na+ concentration gradient or both the Na+ and K+ concentration gradients.2. 2. There was an inward transport of AIB against its concentration gradient when the Na+ concentration gradient was reversed and also when both the Na+ and K+ concentration gradients were reversed.3. 3. In other experiments, the steady-state AIB and ion distribution ratios were measured, and the energy expenditure needed to maintain the AIB concentration gradient was calculated and compared with the energy expenditure calculated to be available from the ion electrochemical potential gradients in two model carrier systems.4. 4. A model in which there was a 1:1 cotransport of Na+ and AIB and no associated K+ movement was found to be inadequate to explain the entire AIB accumulation.5. 5. A model in which the carrier operated electro-neutrally, returning one K+ for each Na+ and AIB brought in, comes closer to satisfying the data from the studies of the steady state. However, there are a few discrepancies and the hypothesis does require a 100% efficiency of coupling between the ion fluxes and the uptake of AIB to explain most of the data from the steady-state studies.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/32865/1/0000243.pd