Site-specific regulation of the epithelial sodium channel (ENaC) and the renal outer medullary K+ channel (ROMK) in the aldosterone-sensitive distal nephron (ASDN)

• Applicant: Professor Dr. Christoph Korbmacher
• Subject Area: Anatomy and Physiology ; Nephrology
• Term: from 2017 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 391676485

Pathophysiological disturbances of Na+ or K+ homeostasis result in potentially life threatening disorders. This justifies the need to investigate cellular and molecular mechanisms involved in renal Na+ and K+ handling. The aldosterone-sensitive distal nephron (ASDN) comprises the late distal convoluted tubule (DCT2), connecting tubule (CNT), cortical collecting duct (CCD) and its distal parts. In the ASDN the epithelial sodium channel (ENaC) and the renal outer medullary K channel (ROMK) are critically important for the final adjustment of tubular Na+ absorption and K+ secretion, respectively. Aldosterone stimulates ENaC through the mineralocorticoid receptor (MR). This promotes tubular Na+ absorption and enhances the electrical driving force for ROMK mediated renal K+ secretion. Recent evidence supports the idea that ENaC and ROMK are expressed and regulated in a site-specific manner. This has important implications for Na+ homeostasis and blood pressure control as well as for K+ homeostasis. This renewal proposal is a logical continuation of our current project with a focus on patch-clamp studies in microdissected murine renal tubules. In addition, we will use a differentiated mouse CCD cell line (mCCDcl1) and native mouse distal colon in a comparative approach to reveal common and tissue-specific regulatory mechanisms of ENaC mediated Na+ transport. Based on published findings and preliminary experiments, we have two key objectives: (1) Identify common and distinct mechanisms of ENaC regulation in DCT2/CNT versus CNT/CCD; (2) Explore site-specific mechanisms of ROMK regulation in the distal nephron. Whole-cell and outside-out patch-clamp recordings will be performed in the transitional zones between DCT2 and early CNT (DCT2/CNT) and between late CNT and early cortical collecting duct (CNT/CCD). In combination with transgenic mouse models this provides a powerful approach to address unresolved physiological questions regarding the site-specific regulation of ENaC and ROMK in native tubules. In particular, we will use inducible nephron-specific MR deficient (MR KO) mice, a nephron specific glucocorticoid receptor (GR) deficient mouse model and angiotensin II type 1A (AT1A) receptor deficient (Agtr1A-/-) mice to explore the differential effects of MR, GR and AT1A deficiency on ENaC and ROMK activity in DCT2/CNT versus CNT/CCD. Recent evidence suggest that mTORC2 plays a role in ENaC regulation in response to changes in dietary K+ intake and local [K+]. To explore the role of mTORC2 in site-specific ENaC and ROMK regulation, we will use a novel mouse model (TRKO mice) with doxycycline-inducible tubular specific mTORC2 deficiency. With the experiments detailed in our project proposal, we expect to obtain new findings regarding the mechanisms involved in the site-specific regulation and function of ENaC and ROMK. This will enhance our understanding of the physiological interplay of these channels in Na+ and K+ homeostasis.

• DFG Programme: Research Grants