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Identification and localization of BK- subunits in the distal nephron of the mouse kidney

¹Department of Cellular and Integrative Physiology, University of Nebraska Medical Center, Nebraska Medical Center, Omaha, Nebraska; and ²Department of Physiology, University of Texas Health Science Center at San Antonio, San Antonio, Texas

Submitted 10 January 2007 ; accepted in final form 23 April 2007

Large-conductance, Ca²⁺-activated K⁺ channels (BK), comprised of pore-forming - and accessory -subunits, secrete K⁺ in the distal nephron under high-flow and high-K⁺ diet conditions. BK channels are detected by electrophysiology in many nephron segments; however, the accessory -subunit associated with these channels has not been determined. We performed RT-PCR, Western blotting, and immunohistochemical staining to determine whether BK-1 is localized to the connecting tubule's principal-like cells (CNT) or intercalated cells (ICs), and whether BK-2-4 are present in other distal nephron segments. RT-PCR and Western blots revealed that the mouse kidney expresses BK-1, BK-2, and BK-4. Available antibodies in conjunction with BK-1^–/– and BK-4^–/– mice allowed the specific localization of BK-1 and BK-4 in distal nephron segments. Immunohistochemical staining showed that BK-1 is localized in the CNT but not ICs of the connecting tubule. The localization of BK-4 was discerned using an anti-BK-4 antibody on wild-type tissue and anti-GFP on GFP-replaced BK-4 mouse (BK-4^–/–) tissue. Both antibodies (anti-BK-4 and anti-GFP) localized BK-4 to the thick ascending limb (TAL), distal convoluted tubule (DCT), and ICs of the distal nephron. It is concluded that BK-1 is narrowly confined to the apical membrane of CNTs in the mouse, whereas BK-4 is expressed in the TAL, DCT, and ICs.

connecting tubule; thick ascending limb; intercalated cells; mice; maxi K; BK_Ca

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