A novel mouse model for an inducible gene-modification in the renal thick ascending limb

Abstract

The thick ascending limb (TAL) is critical for the renal control of fluid and ion homeostasis. The function of the TAL depends on the activity of the bumetanide-sensitive Na-K-2Cl co-transporter NKCC2, which is highly abundant in the luminal membrane of TAL cells. The TAL function is regulated by various hormonal and non-hormonal factors. However, many of the underlying signal transduction pathways remain elusive. Here, we describe the characterization of a novel gene-modified mouse model for an inducible and specific Cre/Lox-mediated gene modification in the TAL. In these mice, a tamoxifen-dependent Cre (CreERT2) was inserted into the 3’ UTR of the Slc12a1 gene, which encodes NKCC2 (Slc12a1-CreERT2). Although this gene-modification strategy reduced endogenous NKCC2 expression at the mRNA and protein level, the lowered NKCC2 abundance was not associated with an altered urinary fluid and ion excretion. Likewise, the renal response to loop-diuretics or water restriction was similar in wildtype (wt) and in Slc12a1-CreERT2wt/tg mice. Immunohistochemistry on kidneys from Slc12a1-CreERT2 mice revealed strong Cre expression exclusively in TAL cells but not in any other nephron portion. Cross-breeding of these mice with the mT/mG reporter mouse line showed a very low recombination rate (0.22%) at baseline, but a complete recombination (100%) after repeated tamoxifen administration. The achieved recombination encompassed the entire TAL and included also the macula densa. Thus, the new Slc12a1-CreERT2wt/tg mouse line allows an inducible and very efficient gene-targeting in the TAL and hence promises to be a powerful tool to advance our understanding of the regulation of TAL function. The research of Johannes Loffing is supported by the Swiss National Science Foundation (310030_143929/1) and the Swiss National Centre for Competence in Research “Kidney.CH” This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.