Potassium-sparing effects of furosemide in mice on high-potassium diets.

In individuals on a regular "Western" diet, furosemide induces a kaliuresis and reduction in plasma K concentration by inhibiting Na reabsorption in the thick ascending limb of Henle's loop, enhancing delivery of Na to the aldosterone-sensitive distal nephron. In the aldosterone-sensitive distal nephron, the increased Na delivery stimulates K wasting due to an exaggerated exchange of epithelial Na channel-mediated Na reabsorption of secreted K. The effects of furosemide are different in mice fed a high-K, alkaline (HK) diet: the large-conductance Ca-activated K (BK) channel, in conjunction with the BK β-subunit (BK-α/β), mediates K secretion from intercalated cells (IC) of the connecting tubule and collecting ducts.

Furosemide reduces BK-αβ4-mediated K secretion in mice on an alkaline high-K diet.

Special high-K diets have cardioprotective effects and are often warranted in conjunction with diuretics such as furosemide for treating hypertension. However, it is not understood how a high-K diet (HK) influences the actions of diuretics on renal K handling. Furosemide acidifies the urine by increasing acid secretion via the Na-H exchanger 3 (NHE3) in TAL and vacuolar H-ATPase (V-ATPase) in the distal nephron.

Net K secretion in the thick ascending limb of mice on a low-Na, high-K diet.

Because of its cardio-protective effects, a low-Na, high-K diet (LNaHK) is often warranted in conjunction with diuretics to treat hypertensive patients. However, it is necessary to understand the renal handling of such diets in order to choose the best diuretic. Wild-type (WT) or Renal Outer Medullary K channel (ROMK) knockout mice (KO) were given a regular (CTRL), LNaHK, or high-K diet (HK) for 4-7 days.

Maintaining K balance on the low-Na, high-K diet.

A low-Na, high-K diet (LNaHK) is considered a healthier alternative to the "Western" high-Na diet. Because the mechanism for K secretion involves Na reabsorptive exchange for secreted K in the distal nephron, it is not understood how K is eliminated with such low Na intake. Animals on a LNaHK diet produce an alkaline load, high urinary flows, and markedly elevated plasma ANG II and aldosterone levels to maintain their K balance.

Physiological role of NBCe2 in the regulation of electrolyte transport in the distal nephron.

The electrogenic Na(+)-HCO3 (-) cotransporter 2 (NBCe2) is a newly discovered protein in the distal nephron. Our understanding is minimal regarding its physiological role in renal electrolyte transport. In this mini-review, we summarize the potential function of NBCe2 in the regulation of blood pressure, acid-base, and K(+) and Ca(2+) transport in the distal nephron.

Deficient acid handling with distal RTA in the NBCe2 knockout mouse.

In many circumstances, the pathogenesis of distal renal tubular acidosis (dRTA) is not understood. In the present study, we report that a mouse model lacking the electrogenic Na(+)-HCO3 (-) cotransporter [NBCe2/Slc4a5; NBCe2 knockout (KO) mice] developed dRTA after an oral acid challenge. NBCe2 expression was identified in the connecting tubule (CNT) of wild-type mice, and its expression was significantly increased after acid loading.

Increased Epithelial Sodium Channel Activity Contributes to Hypertension Caused by Na+-HCO3- Cotransporter Electrogenic 2 Deficiency.

The gene SLC4A5 encodes the Na(+)-HCO3 (-) cotransporter electrogenic 2, which is located in the distal nephron. Genetically deleting Na(+)-HCO3 (-) cotransporter electrogenic 2 (knockout) causes Na(+)-retention and hypertension, a phenotype that is diminished with alkali loading. We performed experiments with acid-loaded mice and determined whether overactive epithelial Na(+) channels (ENaC) or the Na(+)-Cl(-) cotransporter causes the Na(+) retention and hypertension in knockout.

Low Na, high K diet and the role of aldosterone in BK-mediated K excretion.

A low Na, high K diet (LNaHK) is associated with a low rate of cardiovascular (CV) disease in many societies. Part of the benefit of LNaHK relies on its diuretic effects; however, the role of aldosterone (aldo) in the diuresis is not understood. LNaHK mice exhibit an increase in renal K secretion that is dependent on the large, Ca-activated K channel, (BK-α with accessory BK-β4; BK-α/β4).

Interacting influence of diuretics and diet on BK channel-regulated K homeostasis.

Large conductance, Ca-activated K channels (BK) are abundantly located in cells of vasculature, glomerulus, and distal nephron, where they are involved in maintaining blood volume, blood pressure, and K homeostasis. In mesangial cells and smooth muscle cells of vessels, the BK-α pore associates with BK-β1 subunits and regulates contraction in a Ca-mediated feedback manner. The BK-β1 also resides in connecting tubule cells of the nephron.

Relation between BK-α/β4-mediated potassium secretion and ENaC-mediated sodium reabsorption.

The large-conductance, calcium-activated BK-α/β4 potassium channel, localized to the intercalated cells of the distal nephron, mediates potassium secretion during high-potassium, alkaline diets. Here we determine whether BK-α/β4-mediated potassium transport is dependent on epithelial sodium channel (ENaC)-mediated sodium reabsorption. We maximized sodium-potassium exchange in the distal nephron by feeding mice a low-sodium, high-potassium diet.

Regulation of BK-α expression in the distal nephron by aldosterone and urine pH.

In the distal nephron, the large-conductance Ca-activated K (BK) channel, comprised of a pore-forming-α (BK-α) and the BK-β4 subunit, promotes K excretion when mice are maintained on a high-K alkaline diet (HK-alk). We examined whether BK-β4 and the acid-base status regulate apical membrane expression of BK-α in the cortical (CCD) and medullary collecting ducts (MCD) using immunohistochemical analysis (IHC) and Western blot. With the use of IHC, BK-α of mice on acontrol diet localized mostly cytoplasmically in intercalated cells (IC) of the CCD and in the perinuclear region of both principle cells (PC) and IC of the MCD.

Flow-sensitive K+-coupled ATP secretion modulates activity of the epithelial Na+ channel in the distal nephron.

The epithelial Na(+) channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN) is under tonic inhibition by a local purinergic signaling system responding to changes in dietary sodium intake. Normal BK(Ca) channel function is required for flow-sensitive ATP secretion in the ASDN. We tested here whether ATP secreted through connexin channels in a coupled manner with K(+) efflux through BK(Ca) channels is required for inhibitory purinergic regulation of ENaC in response to increases in sodium intake.

Bicarbonate promotes BK-α/β4-mediated K excretion in the renal distal nephron.

Ca-activated K channels (BK), which are stimulated by high distal nephron flow, are utilized during high-K conditions to remove excess K. Because BK predominantly reside with BK-β4 in acid/base-transporting intercalated cells (IC), we determined whether BK-β4 knockout mice (β4KO) exhibit deficient K excretion when consuming a high-K alkaline diet (HK-alk) vs. high-K chloride diet (HK-Cl).

Role of BK channels in hypertension and potassium secretion.

Purpose Of Review: This review summarizes recent studies of hypertension associated with a defect in renal K excretion due to genetic deletions of various components of the large, Ca-activated K channel (BK), and describes new evidence and theories regarding K secretory roles of BK in intercalated cells.

Recent Findings: Isolated perfused tubule methods have revealed the importance of BK in flow-induced K secretion. Subsequently, mice with genetically deleted BK subunits revealed the complexities of BK-mediated K secretion.

Coupled ATP and potassium efflux from intercalated cells.

Increased flow in the distal nephron induces K secretion through the large-conductance, calcium-activated K channel (BK), which is primarily expressed in intercalated cells (IC). Since flow also increases ATP release from IC, we hypothesized that purinergic signaling has a role in shear stress (τ; 10 dynes/cm(2)) -induced, BK-dependent, K efflux. We found that 10 μM ATP led to increased IC Ca concentration, which was significantly reduced in the presence of the P(2) receptor blocker suramin or calcium-free buffer.

BK channels and a new form of hypertension.

Large, Ca-activated K channels (BK) are comprised of an α pore (BKα) and one of four β subunits (BKβ1-4). When the gene for BKβ1 is knocked out (BKβ1-KO), the result is increased myogenic tone of vascular smooth muscle and hypertension. We reexamined whether the hypertension is entirely due to increased vascular tone, because most monogenic forms of hypertension have renal origins and BKβ1 resides in renal connecting tubule (CNT) cells.

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.

Intercalated cell BK-alpha/beta4 channels modulate sodium and potassium handling during potassium adaptation.

The large-conductance, calcium-activated potassium (BK) channels help eliminate potassium in mammals consuming potassium-rich diets. In the distal nephron, principal cells contain BK-alpha/beta1 channels and intercalated cells contain BK-alpha/beta4 channels. We studied whether BK-beta4-deficient mice (Kcnmb4(-/-)) have altered renal sodium and potassium clearances compared with wild-type mice when fed a regular or potassium-rich diet for ten days.

Biomechanical strain causes maladaptive gene regulation, contributing to Alport glomerular disease.

Patients with Alport's syndrome develop a number of pro-inflammatory cytokine and matrix metalloproteinase (MMP) abnormalities that contribute to progressive renal failure. Changes in the composition and structure of the glomerular basement membranes likely alter the biomechanics of cell adhesion and signaling in these patients. To test if enhanced strain on the capillary tuft due to these structural changes contributes to altered gene regulation, we subjected cultured podocytes to cyclic biomechanical strain.

Hypertension of Kcnmb1-/- is linked to deficient K secretion and aldosteronism.

Mice lacking the beta1-subunit (gene, Kcnmb1; protein, BK-beta1) of the large Ca-activated K channel (BK) are hypertensive. This phenotype is thought to result from diminished BK currents in vascular smooth muscle where BK-beta1 is an ancillary subunit. However, the beta1-subunit is also expressed in the renal connecting tubule (CNT), a segment of the aldosterone-sensitive distal nephron, where it associates with BK and facilitates K secretion.

Role of BKbeta1 in Na+ reabsorption by cortical collecting ducts of Na+-deprived mice.

On a low-Na(+) diet (LNa(+)), urinary Na(+) loss is prevented by aldosterone-induced Na(+) reabsorption through epithelial Na(+) channels (ENaC) in the connecting tubules (CNT) and cortical collecting ducts (CCD). However, the mechanism whereby K(+) loss is minimized and Na(+) reabsorption is maximized in the face of a reduced lumen-to-bath Na(+) gradient is not fully understood. The large-conductance calcium-activated potassium channel (BK)beta1 subunit (gene: Kcnmb1), which has a role in K(+) secretion in the CNT, is absent in the CCD in mice on a control diet.

Insulin increases the activity of mesangial BK channels through MAPK signaling.

Glomerular hyperfiltration and mesangial expansion have been described in mouse models of a hyperinsulinemic early stage of type 2 diabetes mellitus (DM). Large-conductance Ca(2+)-activated K(+) channels (BK) have been linked to relaxation of human mesangial cells (MC) and may contribute to MC expansion and hyperfiltration. We hypothesized that high insulin levels increase BK activity in MC by increasing the number and/or open probability (P(o)) of BK in the plasma membrane.

Association of VASP with TRPC4 in PKG-mediated inhibition of the store-operated calcium response in mesangial cells.

We tested the hypotheses that the NO-cGMP-PKG pathway mediates inhibition of the store-operated cation channel (SOC) in human glomerular mesangial cells (HMC) and that TRPC4, a molecular component of SOC in HMC, is associated with PKG-phosphorylated vasodilator-stimulated phosphoprotein (VASP). Using fura 2 ratiometry, we measured intracellular Ca(2+) concentration [Ca(2+)](i) to determine whether sodium nitroprusside (SNP), an NO donor, and 8-Br-cGMP affected SOC-TRPC4 via PKG. We found that the SOC response in HMC was attenuated in the presence of 100 microM SNP, an NO donor, or 100 microM 8-Br-cGMP.

BK channels in the kidney.

Purpose Of Review: Large, BK (calcium-activated potassium) channels are now regarded as relevant players in many aspects of renal physiology, including potassium secretion. This review will highlight recent discoveries regarding the function and localization of BK in the kidney.

Recent Findings: Patch clamp electrophysiology has revealed BK in cultured podocytes, glomerular mesangial cells, and in several tubule segments including principal cells (connecting tubules/principal cells), and intercalated cells of connecting tubules and cortical collecting ducts.

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

Large-conductance, Ca(2+)-activated K(+) channels (BK), comprised of pore-forming alpha- and accessory beta-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 beta-subunit associated with these channels has not been determined. We performed RT-PCR, Western blotting, and immunohistochemical staining to determine whether BK-beta1 is localized to the connecting tubule's principal-like cells (CNT) or intercalated cells (ICs), and whether BK-beta2-4 are present in other distal nephron segments.