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Am J Physiol Cell Physiol
2022 Oct 01;3234:C1061-C1069. doi: 10.1152/ajpcell.00303.2022.
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Following Ussing's legacy: from amphibian models to mammalian kidney and brain.
Blazer-Yost BL
.
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Professor Hans H. Ussing (1911-2000) was one of the founding members of the field of epithelial cell biology. He is most famous for the electrophysiological technique that he developed to measure electrogenic ion flux across epithelial tissues. Ussing-style electrophysiology has been applied to multiple tissues and has informed fields as diverse as amphibian biology and medicine. In the latter, this technique has contributed to a basic understanding of maladies such as hypertension, polycystic kidney disease, cystic fibrosis, and diarrheal diseases to mention but a few. In addition to this valuable contribution to biological methods, Prof. Ussing also provided strong evidence for the concept of active transport several years before the elucidation of Na+K+ATPase. In addition, he provided cell biologists with the important concept of polarized epithelia with specific and different transporters found in the apical and basolateral membranes, thus providing these cells with the ability to conduct directional, active and passive transepithelial transport. My studies have used Ussing chamber electrophysiology to study the toad urinary bladder, an amphibian cell line, renal cell lines, and, most recently, choroid plexus cell lines. This technique has formed the basis of our in vitro mechanistic studies that are used in an iterative manner with animal models to better understand disease progress and treatment. I was honored to be invited to deliver the 2022 Hans Ussing Lecture sponsored by the Epithelial Transport Group of the American Physiological Society. This manuscript is a version of the material presented in that lecture.
Figure 2.
Current voltage clamp and double chamber Ussing chambers: current voltage clamp made by Gus Kelley and Gary Huber in the group of Professor Malcolm Cox at the University of Pennsylvania, Renal Electrolyte Section. A: the clamp shown was made to measure both short-circuit current and transepithelial resistance depending on the placement of the electrical connections. B: the picture shows a double chamber (made for toad urinary bladder) wherein the tissue to be measured was placed over both chambers. When the two halves were assembled, the rubber gaskets separated the tissue and provided two electrically isolated pieces. The white plugs on the sides had small ports for the insertion of agar bridges to connected to electrodes that interfaced with the clamp apparatus in A.
Figure 4.
Ussing-style electrophysiological experiments of TRPV4 agonist and antagonist in choroid plexus cell lines. The graphs represent a compilation of several studies. Electrophysiology graphs are displayed as ISC (left, a measure of net electrogenic ion flux) and conductance (right, a measure of barrier permeability). GSK1016790A, a TRPV4 agonist, was added to all cultures at time zero using a concentration that was determined optimal for each cell line.A: electrophysiological responses in the porcine choroid plexus-Reims cell line. The black trace represents the agonist-only treatment. In the cultures shown by the pick traces, a TRPV4 antagonist, RN1734 was added 10 min before the addition of the agonist. The antagonist prevents the TRPV4-mediated increase in ion flux and conductance. In the teal trace, RN 1734 added bilaterally 10 min after the addition of agonist. The antagonist immediately reverses both the current and conductance.B: electrophysiological responses in a human choroid plexus cell line, HIBCCP. The black trace represents the agonist only treatment. In the cultures shown by the pick traces, a TRPV4 antagonist, RN1734 was added 5 min before the addition of the agonist. The antagonist prevents the TRPV4-mediated increase in ion flux and conductance. In the teal trace, RN 1734 added bilaterally at time t = 5 min. The antagonist immediately reverses both the current and conductance. All traces represent means ± SE for the stated “n” of technical replicates. *P < 0.05 considered significant between condition and GSK control measured by multiple t tests grouped analysis and indicated by color. GSK, GSK1016790A; HIBCPP, human choroid plexus papilloma; Isc, short circuit current; TRPV4, transient receptor potential vanilloid 4.
Figure 1. A: original diagram of the Ussing chamber set-up and the figure legend from Ussing and Zerahn’s manuscript in 1951 (2). The diagram is reprinted with permission from John Wiley and Sons. B: Ussing chamber set-up as applied to cells cultured on permeable supports. Figure created with BioRender.com by Alexandra Hochstetler. TEER, transepithelial electrical resistance.
