Epithelial Na+ channels are activated by laminar shear stress.
The degenerin/epithelial Na+ channel (ENaC) superfamily is a group of structurally related ion channels that are involved in diverse biological processes, including responses to mechanical stimuli. In renal cortical collecting ducts, changes in rates of perfusion affect Na+ reabsorption through an amiloride-sensitive pathway, suggesting that ENaC may be a mechanosensitive channel. In this study, we examined whether ENaC expressed in oocytes is regulated by laminar shear stress (LSS). A 1.8-mm (internal diameter) perfusion pipette was placed within 0.5-1.0 mm of the oocyte to provide laminar flow across the oocyte surface. LSS induced a dose-dependent and reversible increase in benzamil-sensitive whole cell Na+ currents in oocytes expressing alphabetagamma ENaC. The half-time for activation by LSS was approximately 5 s. Repetitive stimulation by LSS of oocytes expressing ENaC was associated with a reduction in the response to LSS. Oocytes expressing alphabetaS518Kgamma, a pore region mutant with a high open probability, were insensitive to LSS. We demonstrated previously that channels with a Cys residue introduced at position alphaSer-580 had a low open probability, but, following modification by [2-(trimethylammonium)ethyl]methanethiosulfonate bromide (MTSET), channels exhibited a high open probability. Oocytes expressing alphaS580Cbetagamma ENaC respond to LSS similar to wild type; however, covalent modification by MTSET largely eliminated the response to LSS. Our results suggest that shear stress activates ENaC by modifying the gating properties of the channel.
Carattino
Marcelo D
MD
Renal-Electrolyte Division, Departments of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
Sheng
Shaohu
S
Kleyman
Thomas R
TR
D000584:Amiloride / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D004848:Epithelium / Q000378:metabolism; D005260:Female; D066298:In Vitro Techniques; D015640:Ion Channel Gating; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D011994:Recombinant Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D015222:Sodium Channels / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D013314:Stress, Mechanical; D014982:Xenopus laevis
D011994:Recombinant Proteins; D015222:Sodium Channels; C013407:benzamil; D000584:Amiloride