Abstract
The Ca2+-activated, maxi-K (BK) K+ channel, with low Ca2+-binding affinity, is expressed in the distal tubule of the nephron and contributes to flow-dependent K+ secretion. In the present study we demonstrate that the Ca2+-activated, SK3 (KCa2.3) K+ channel, with high Ca2+-binding affinity, is also expressed in the mouse kidney (RT-PCR, immunoblots). Immunohistochemical evaluations using tubule specific markers demonstrate significant expression of SK3 in the distal tubule and the entire collecting duct system, including the connecting tubule (CNT) and cortical collecting duct (CCD). In CNT and CCD, main sites for K+ secretion, the highest levels of expression were along the apical (luminal) cell membranes, including for both principal cells (PCs) and intercalated cells (ICs), posturing the channel for Ca2+-dependent K+ secretion. Fluorescent assessment of cell membrane potential in native, split-opened CCD, demonstrated that selective activation of the Ca2+-permeable TRPV4 channel, thereby inducing Ca2+ influx and elevating intracellular Ca2+ levels, activated both the SK3 channel and the BK channel leading to hyperpolarization of the cell membrane. The hyperpolarization response was decreased to a similar extent by either inhibition of SK3 channel with the selective SK antagonist, apamin, or by inhibition of the BK channel with the selective antagonist, iberiotoxin (IbTX). Addition of both inhibitors produced a further depolarization, indicating cooperative effects of the two channels on Vm. It is concluded that SK3 is functionally expressed in the distal nephron and collecting ducts where induction of TRPV4-mediated Ca2+ influx, leading to elevated intracellular Ca2+ levels, activates this high Ca2+-affinity K+ channel. Further, with sites of expression localized to the apical cell membrane, especially in the CNT and CCD, SK3 is poised to be a key pathway for Ca2+-dependent regulation of membrane potential and K+ secretion.
Highlights
Calcium-activated potassium channels, KCa, are a small group of potassium channels that are widely expressed in numerous tissues ranging from neurons to vascular endothelial cells [1,2,3,4,5]
We found that SK3 is expressed in the mouse kidney with immunohistochemical staining showing apparent strong expression in the thick ascending limb, the distal convoluted tubule, and the entire collecting duct systems, including the connecting tubule (CNT) and the cortical collecting duct (CCD)
Initial studies included co-staining for Aquaporin 2 (AQP2), a marker of principal cells (PCs) cells within the collecting duct system
Summary
Calcium-activated potassium channels, KCa, are a small group of potassium channels that are widely expressed in numerous tissues ranging from neurons to vascular endothelial cells [1,2,3,4,5]. As with other K+ channels, the KCa channels can play a major role in regulating the plasma membrane electrical potential difference, Vm. As with other K+ channels, the KCa channels can play a major role in regulating the plasma membrane electrical potential difference, Vm Their classical regulation by intracellular Ca2+, [Ca2+]i, leads to a highly dynamic coupling between Vm and [Ca2+]i which appears to underlie their central role in a wide array of functions ranging from neuronal excitability [6,7], to modulation of vascular smooth muscle tone [8,9], to cell volume regulation [10,11].
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