Shear stress-induced volume decrease in C11-MDCK cells by BK-alpha/beta4.
Large-conductance, calcium-activated potassium channels (BK) are expressed in principal cells (PC) and intercalated cells (IC) in mammalian nephrons as BK-alpha/beta1 and BK-alpha/beta4, respectively. IC, which protrude into the lumens of tubules, express substantially more BK than PC despite lacking sufficient Na-K-ATPase to support K secretion. We previously showed in mice that IC exhibit size reduction when experiencing high distal flows induced by a high-K diet. We therefore tested the hypothesis that BK-alpha/beta4 are regulators of IC volume via a shear stress (tau)-induced, calcium-dependent mechanism, resulting in a reduction in intracellular K content. We determined by Western blot and immunocytochemical analysis that C11-Madin-Darby canine kidney cells contained a predominance of BK-alpha/beta4. To determine the role of BK-alpha/beta4 in tau-induced volume reduction, we exposed C11 cells to tau and measured K efflux by flame photometry and cell volume by calcein staining, which changes inversely to cell volume. With 10 dynes/cm(2), calcein intensity significantly increased 39% and monovalent cationic content decreased significantly by 37% compared with static conditions. Furthermore, the shear-induced K loss from C11 was abolished by the reduction of extracellular calcium, addition of 5 mM TEA, or BK-beta4 small interfering (si) RNA, but not by addition of nontarget siRNA. These results show that BK-alpha/beta4 plays a role in shear-induced K loss from IC, suggesting that BK-alpha/beta4 regulate IC volume during high-flow conditions. Furthermore, these results support the use of C11 cells as in vitro models for studying BK-related functions in IC of the kidney.
Holtzclaw
J David
JD
Department of Cellular and Integrative Physiology, University of Nebraska Medical Center, Omaha, 68198-5850, USA.
Liu
Liping
L
Grimm
P Richard
PR
Sansom
Steven C
SC
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D048429:Cell Size; D004285:Dogs; D007668:Kidney / Q000166:cytology* / Q000378:metabolism*; D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits / Q000378:metabolism*; D051038:Large-Conductance Calcium-Activated Potassium Channel beta Subunits / Q000378:metabolism*; D023421:Models, Animal; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000494:pharmacology; D013314:Stress, Mechanical; D019789:Tetraethylammonium / Q000494:pharmacology
D051037:Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; D051038:Large-Conductance Calcium-Activated Potassium Channel beta Subunits; D026902:Potassium Channel Blockers; D034741:RNA, Small Interfering; D019789:Tetraethylammonium; D011188:Potassium; D002118:Calcium