Flow‐induced shear stress reduces K content of MDCK cells by a Ca and TEA‐dependent K efflux mechanism

Abstract

High tubular flow induces K+ secretion by large conductance, Ca2+‐activated K+ channels (BKCa) in mammalian connecting tubules (CNTs) and cortical collecting ducts (CCDs). Although expressed in CNTs and PCs, BKCa are also highly expressed in intercalated cells (IC), which may not have the capacity to sustain K+ secretion because of minimal Na‐K‐ATPase. We used MDCK cells, a cell line with properties consistent with both PCs and ICs, in a parallel plate flow chamber (PPFC) to support the hypothesis that high flow activates BKCa in ICs to reduce IC size thereby increasing tubule internal diameter and decreasing resistance to flow. Using immunocytochemistry and RT‐PCR, we found that MDCKs contain BK‐α, BK‐β1, BK‐β2 and BK‐β4 subunits. MDCKs were exposed to static (control) or 10 dynes/cm2 shear stress in a PPFC at 37ºC for 6 hrs. Perfusate was similar to mammalian CNT luminal fluid. After incubation, K+content in MDCKs was measured by flame photometry and normalized by DNA. With high flow, intracellular K+ decreased by 40% compared to static conditions (4.2 vs. 7.1 μg K+/μg DNA, N=4). This decrease was attenuated by removal of luminal Ca2+ with EGTA (10.5 vs. 9.2, N=3) or addition of 5 mM TEA (11.0 vs. 11.6, N=2), a blocker of BKCa. These data show that MDCKs contain the BKCa molecular components and suggest that the elevated [Ca2+]i due to shear stress may result in activation of BKCa, which reduces cell K+ content and volume.