Cell‐autonomous expression of membrane proteins in kidney thick ascending limb epithelium is vasopressin‐dependent

Abstract

The thick ascending limb (TAL) of the mammalian kidney exhibits functionally and structurally distinct patterns along its course from the border of the inner medulla to its transition to the distal convoluted tubule. While reabsorption of 25% of the filtered load is the principal task shared by all TAL cells, the transport of K+ and the divalent cations Ca++ and Mg++ is subject to axial and cellular heterogeneity and occurs along trans‐ or paracellular pathways. Differential expression of the renal outer medullary K+ channel (ROMK) has been associated with net K+ secretion or reabsorption, and claudin (Cldn) 10 and 16 with selective paracellular cation permeability. Earlier we had detected basolateral vasopressin (AVP) V2 receptor expression and substantial activating responsiveness of transport parameters along TAL upon adding AVP. Here we tested the hypothesis whether AVP modulates cell‐autonomous expression of membrane proteins involved in TAL epithelial ion transport.We analyzed perfusion‐fixed mouse and rat kidneys. Human kidney biopsies were studied for comparison. The effect of AVP was tested in AVP‐deficient Brattleboro rats using V2R‐specific desmopressin (dDAVP; 7 and 72 hours). NKCC2, ROMK, Kir4.1, Cldn‐10 and Cldn‐16 signals were analyzed using sensitive in situ hybridization, IHC, EM, and Western blot (WB).TAL morphological cell heterogeneity was obvious between kidney zones, but not at the cell‐to‐cell level within any zone. Morphologically heterogeneous “smooth” and “rough” patterns of neighboring TAL cells were not discovered, but instead, ROMK und Kir4.1 expression showed conspicuous heterogeneity in a mutually exclusive pattern in tubules of outer medulla and cortex of rodent and human kidneys. Most tight junctions of inner stripe TAL were Cldn‐10 positive. In its outermost layer, Cldn‐10‐ and ‐16 signals were co‐expressed; ROMK‐positive cells displayed Cldn‐10‐positive tight junctions between the adjacent ROMK‐positive cells whereas between ROMK‐negative cells, junctions were Cldn‐16‐positive. 72h dDAVP administration significantly increased the number of ROMK‐positive cells in outer medullary TAL. Kir4.1‐positive cells increased in parallel. Cldn‐10‐positive junctions increased at the expense of Cldn‐16‐positive junctions. WB showed 2.2‐fold increase for ROMK and 1.3‐fold for Kir4.1 (p<.05). Sensitive ISH revealed differential quantities of ROMK mRNA abundances in TAL, with increases in the number of cells expressing ROMK mRNA strongly upon dDAVP.These results demonstrate cell‐autonomous regulation of crucial TAL cell membrane proteins in response to activating stimuli in the absence of major cell morphological changes. Variable expression of ROMK, Kir4.1, and Cldn‐10 and ‐16 signals likely reflected vasopressindependent adaptive features in renal solute handling in the renal outer medulla.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.