Characterizing novel aldosterone transport mechanisms in the murine collecting duct

Within the renal collecting duct (CD), Na + reabsorption and K + secretion is stimulated by the volume-regulating hormone aldosterone (ALDO) in response to hyperkalemia, hyponatremia, or hypovolemia. Due to its lipophilic nature, ALDO can diffuse across the plasma membrane to enter target cells, binding target receptors rapidly. Recent studies have shown corticosteroids may also be actively transported across the plasma membrane e.g. in the brain, adrenal glands and adipose tissue. Preliminary data from our group suggest that active transport mechanisms may also contribute to corticosteroid movement across CD cells. AIM: To evaluate the dynamics of ALDO distribution across murine CD cells: comparing apical (AP) vs basolateral (BL) application; distribution over 48h; and distribution in the absence of Na + /K + -ATPase activity. Murine CD cells (mCCD cl1 and primary cells) were cultured on filters for 9–10 days to establish a compartment model of polarized transporting epithelia. [ALDO] was measured across three compartments: AP, BL and cell lysate (CL). Equivalent-short circuit current ( I eq ) was quantified throughout experiments by epithelial volt-ohm meter. [ALDO] was determined via solid-liquid extraction and targeted LC-MS/MS. Distribution of ALDO following BL application (300nM, 3h) was unequal, with AP: 14.1±2.0%, BL: 85.3±1.8% and CL: 0.6±0.7% ( n =6). Treatment of the AP compartment (300nM, 3h) reversed the distribution, AP: 91.4±1.5%, BL: 8.3±1.4%, CL: 0.3±0.3% (n=6). This distribution is consistent with our previous findings applying 3nM ALDO or 150nM CORT (BL, 3h). I eq was stimulated in a similar manner when ALDO was applied to AP or BL bath. Application of ALDO to BL bath (300nM, 3-48h) demonstrated an unequal distribution at all time points, with AP increasing and BL decreasing 3-12h, then plateauing 12h-48h with AP ~30%, BL ~70%. CL remained at ~0.5% ( n =6). I eq was stimulated above control values at all time points. It is notable that in all experiments, at all times points, CL measured ~0.5-1.5nM. Pre-treating cells with ouabain (2mM, 30min) near-abolished I eq and ALDO treatment (300nM, 3h) did not alter I eq . The distribution of ALDO however, remained unchanged: AP: ~10.9%, BL: ~ 88.8%, and CL: ~0.4% ( n =6). Together these data suggest that an active transport mechanism may be maintaining cellular bioavailability of ALDO within CD cells independently of the electrochemical gradients maintained by the Na + /K + ATPase enabling transepithelial ion transport. Supported by the EASTBIO DTP. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.