Collecting duct–specific gene inactivation of αENaC in the mouse kidney does not impair sodium and potassium balance

Abstract

Aldosterone controls the final sodium reabsorption and potassium secretion in the kidney by regulating the activity of the epithelial sodium channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN). ASDN consists of the last portion of the distal convoluted tubule (late DCT), the connecting tubule (CNT), and the collecting duct (CD) (i.e., the cortical CD [CCD] and the medullary CD [MCD]). It has been proposed that the control of sodium transport in the CCD is essential for achieving sodium and potassium balance. We have tested this hypothesis by inactivating the α subunit of ENaC in the CD but leaving ENaC expression in the late DCT and CNT intact. Under salt restriction or under aldosterone infusion, whole-cell voltage clamp of principal cells of CCD showed no detectable ENaC activity, whereas large amiloride-sensitive currents were observed in control littermates. The animals survive well and are able to maintain sodium and potassium balance, even when challenged by salt restriction, water deprivation, or potassium loading. We conclude that the expression of ENaC in the CD is not a prerequisite for achieving sodium and potassium balance in mice. This stresses the importance of more proximal nephron segments (late DCT/CNT) to achieve sodium and potassium balance. Isabelle Rubera, Johannes Loffing, and Edith Hummler contributed equally to this work. Conflict of interest:The authors have declared that no conflict of interest exists. Nonstandard abbreviations used: epithelial sodium channel (ENaC); aldosterone-sensitive distal nephron (ASDN); distal convoluted tubule (DCT); connecting tubule (CNT); collecting duct (CD); cortical CD (CCD); medullary CD (MCD); outer MCD (OMCD); inner MCD (IMCD); adrenalectomized (adx); glomerular filtration rate (GFR); pseudohypoaldosteronism type 1 (PHA-1); X-galactosidase (X-gal); aquaporin-2 (AQP2); sodium/calcium exchanger (NCX); calbindin D28K (CB); sodium thiazide-sensitive chloride cotransporter (NCC).

Cell models for studying renal physiology

“…These authors showed that mice lacking α-ENaC in the CCD maintain their Na + balance, suggesting that most of the ENaC-Na + absorption occurs in the early segments of the collecting duct system, i.e., the late DCT and connecting tubule (CNT) segments. Electrophysiological analyses of primary cultures of isolated DCTs dissected before their branching in the CCD (i.e., CNT) and isolated CCDs fully confirmed the pioneering study of Rubera et al [116]: Primary cultured DCT dissected from α-ENaC-deficient mice exhibited the same high electrical transepithelial resistance (R T ) and potential (P D ), and amiloride-sensitive I sc as DCT dissected from wild-type mice counterparts. In sharp contrast, CCD dissected from the kidneys of α-ENaC-deficient mice developed high R T and very low P D and did not respond to arginine vasopressin (Fig.…”

“…3 Electrophysiological properties of primary cultures of late distal and early collecting duct cells dissected from kidneys of transgenic mice harboring specific, αENaC-activating gene in the collecting duct. Isolated late DCT (including CNT) microdissected from the kidneys of Scnn 1a loxlox (loxlox) and Scnn 1a loxloxCre (loxloxCre) mice [116] and grown until confluency on collagencoated filters (14 days after seeding) were incubated in hormone-free, serum-free medium. The transepithelial electrical resistance (R T ), potential (P D ), and amiloride-sensitive short-circuit current (AmsI sc ) were then measured on cultured loxlox DCT or CCD and loxloxCre DCT or CCD.…”

“…Primary cultures of distinct segments of the renal tubule may also provide a complementary approach to in vivo analyses of transgenic mice harboring a conditional knock-out gene. Rubera et al generated transgenic mice expressing Cre recombinase placed under the Hoxb7 promoter to inactivate the α-ENaC subunit in the collecting duct [116]. These authors showed that mice lacking α-ENaC in the CCD maintain their Na + balance, suggesting that most of the ENaC-Na + absorption occurs in the early segments of the collecting duct system, i.e., the late DCT and connecting tubule (CNT) segments.…”

In Liddle Syndrome, Epithelial Sodium Channel Is Hyperactive Mainly in the Early Part of the Aldosterone-Sensitive Distal Nephron

“…These nephron segments have not yet been studied in Liddle mice but are thought to contribute substantially to ENaC-mediated sodium absorption in the kidney. [20][21][22] Previously, we reported that ENaC activity in DCT2/CNT is constitutively high and independent of aldosterone. This is in contrast to the aldosterone dependence of ENaC activity in CNT/CCD.…”

The epithelial sodium channel: from molecule to disease

“…Recently, a tissue-specific knockout of the a ENaC subunit in the CCD has been generated in the mouse (Rubera et al 2003). The absence of a ENaC protein in the CCD and OMCD, together with an undetectable ENaC activity in these nephron segments, has surprisingly little effect on Na + homeostasis.…”