Abstract: SA-PO0484
ASIC2 Assembles with Epithelial Sodium Channel (ENaC) Subunits to Form Mechanosensitive Channels
Session Information
- Fluid, Electrolyte, and Acid-Base Disorders: Basic Research
November 08, 2025 | Location: Exhibit Hall, Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Fluid, Electrolytes, and Acid-Base Disorders
- 1101 Fluid, Electrolyte, and Acid-Base Disorders: Basic
Authors
- Shi, Shujie, University of Pittsburgh Department of Medicine, Pittsburgh, Pennsylvania, United States
- Whelan, Sarah Christine, University of Pittsburgh Department of Medicine, Pittsburgh, Pennsylvania, United States
- Szekely, Kennedy, University of Pittsburgh Department of Medicine, Pittsburgh, Pennsylvania, United States
- Drummond, Heather A., University of Mississippi Medical Center Holmes County, Jackson, Mississippi, United States
- Kleyman, Thomas R., University of Pittsburgh Department of Medicine, Pittsburgh, Pennsylvania, United States
Background
The pressure-induced constriction response within pre-glomerular vessels is an essential mechanism to protect kidney from high intraluminal pressure. This response is initiated by vascular smooth muscle (VSM) cells stretch which is sensed by mechanosensitive ion channel complexes. Previous studies suggest that ASIC2 and ENaC subunits are required for stretch-induced mechanoreceptor currents in renal VSM cells.
Methods
To examine whether ENaC β and γ subunits and ASIC2 are components of a mechanosensitive ion channel complex, we expressed wild type (WT) or a degenerin mutant mouse ASIC2a with or without ENaC β and γ subunits in Xenopus oocytes. Oocytes injected with αβγ ENaC were included as a control. Two-electrode voltage clamp was performed to test the channel’s response to flow-mediated shear stress by changing the perfusion rates (0.5 to 5 ml/min) with Na+ as the conducting ion.
Results
Under baseline conditions (0.5 ml/min), oocytes expressing WT ASIC2a and β/γENaC (2a/β/γ) displayed larger inward Na+ currents than ASIC2a alone. In oocytes expressing 2a/β/γ, not ASIC2a alone, baseline currents increased in response to the faster perfusion (5 ml/min), which were further activated by acidic pH, suggesting the hybrid (2a/β/γ) channel contains properties of both ASIC and ENaC. The inward Na+ currents carried by 2a/β/γ were insensitive to 10 μM amiloride, a concentration that blocks > 90% ENaC currents, but were inhibited when extracellular Na+ was replaced with NMDG+. We generated a gain-of-function mutation of ASIC2a by introducing a bulky residue at the degenerin site (G430V). ASIC2a G430V (2aG430V) had larger baseline Na+ currents and higher sensitivity to amiloride, an open pore blocker. Interestingly, we observed flow-mediated channel activation in oocytes expressing either 2aG430V alone or 2aG430V/β/γ. The higher perfusion rate (5 ml/min) elicited a two-fold increase in Na+ currents with 2aG430V, which was further increased to approximately four-fold when ENaC β/γ subunits are co-expressed with 2aG430V.
Conclusion
Collectively, our findings suggest that ASIC2a assembles with ENaC β/γ subunits to form mechanosensitive channels. Future studies will examine whether the presence of ENaC β/γ subunits in the channel complex alter ion permeability or sensitivity to amiloride.
Funding
- NIDDK Support