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Pendrin is a Cl-/HCO3- exchanger expressed in the apical regions of renal intercalated cells. Following pendrin gene ablation, blood pressure falls, in part, from reduced renal NaCl absorption. We asked if pendrin is expressed in vascular tissue and if the lower blood pressure observed in pendrin null mice is accompanied by reduced vascular reactivity. Thus, the contractile responses to KCl and phenylephrine (PE) were examined in isometrically mounted thoracic aortas from wild-type and pendrin null mice. Although pendrin expression was not detected in the aorta, pendrin gene ablation changed contractile protein abundance and increased the maximal contractile response to PE when normalized to cross sectional area (CSA). However, the contractile sensitivity to this agent was unchanged. The increase in contractile force/cross sectional area observed in pendrin null mice was due to reduced cross sectional area of the aorta and not from increased contractile force per vessel. The pendrin-dependent increase in maximal contractile response was endothelium- and nitric oxide-independent and did not occur from changes in Ca2+ sensitivity or chronic changes in catecholamine production. However, application of 100 nM angiotensin II increased force/CSA more in aortas from pendrin null than from wild type mice. Moreover, angiotensin type 1 receptor inhibitor (candesartan) treatment in vivo eliminated the pendrin-dependent changes contractile protein abundance and changes in the contractile force/cross sectional area in response to PE. In conclusion, pendrin gene ablation increases aorta contractile force per cross sectional area in response to angiotensin II and PE due to stimulation of angiotensin type 1 receptor-dependent signaling. The angiotensin type 1 receptor-dependent increase in vascular reactivity may mitigate the fall in blood pressure observed with pendrin gene ablation.

Angiotensin II increases apical plasma membrane pendrin abundance and function. This study explored the role of the small GTPase Rac1 in the regulation of pendrin by angiotensin II. To do this, we generated intercalated cell (IC) Rac1 knockout mice and observed that IC Rac1 gene ablation reduced the relative abundance of pendrin in the apical region of intercalated cells in angiotensin II-treated mice but not vehicle-treated mice. Similarly, the Rac1 inhibitor EHT 1864 reduced apical pendrin abundance in angiotensin II-treated mice, through a mechanism that does not require aldosterone. This IC angiotensin II-Rac1 signaling cascade modulates pendrin subcellular distribution without significantly changing actin organization. However, NADPH oxidase inhibition with APX 115 reduced apical pendrin abundance in vivo in angiotensin II-treated mice. Moreover, superoxide dismutase mimetics reduced Cl- absorption in angiotensin II-treated cortical collecting ducts perfused in vitro. Since Rac1 is an NADPH subunit, Rac1 may modulate pendrin through NADPH oxidase-mediated reactive oxygen species production. Because pendrin gene ablation blunts the pressor response to angiotensin II, we asked if pendrin blunts the angiotensin II-induced increase in kidney superoxide. Although kidney superoxide was similar in vehicle-treated wild-type and pendrin knockout mice, it was lower in angiotensin II-treated pendrin-null kidneys than in wild-type kidneys. We conclude that angiotensin II acts through Rac1, independently of aldosterone, to increase apical pendrin abundance. Rac1 may stimulate pendrin, at least partly, through NADPH oxidase. This increase in pendrin abundance contributes to the increment in blood pressure and kidney superoxide content seen in angiotensin II-treated mice. NEW & NOTEWORTHY This study defines a new signaling mechanism by which angiotensin II modulates oxidative stress and blood pressure. [ABSTRACT FROM AUTHOR]

The urinary potassium (K+) excretion machinery is upregulated with increasing dietary K+, but the role of accompanying dietary anions remains inadequately characterized. Poorly absorbable anions, including HCO−3 , are thought to increase K+ secretion through a transepithelial voltage effect. Here, we tested if they also influence the K+ secretion machinery. Wild-type mice, aldosterone synthase (AS) knockout (KO) mice, or pendrin KO mice were randomized to control, high-KCl, or high-KHCO3 diets. The K+ secretory capacity was assessed in balance experiments. Protein abundance, modification, and localization of K+-secretory transporters were evaluated by Western blot analysis and confocal microscopy. Feeding the high-KHCO3 diet increased urinary K+ excretion and the transtubular K+ gradient significantly more than the high-KCl diet, coincident with more pronounced upregulation of epithelial Na+ channels (ENaC) and renal outer medullary K+ (ROMK) channels and apical localization in the distal nephron. Experiments in AS KO mice revealed that the enhanced effects of HCO−3 were aldosterone independent. The high-KHCO3 diet also uniquely increased the large-conductance Ca2+-activated K+ (BK) channel β4-subunit, stabilizing BKα on the apical membrane, the Cl−/HCO−3 exchanger, pendrin, and the apical KCl cotransporter (KCC3a), all of which are expressed specifically in pendrin-positive intercalated cells. Experiments in pendrin KO mice revealed that pendrin was required to increase K+ excretion with the high-KHCO3 diet. In summary, HCO−3 stimulates K+ excretion beyond a poorly absorbable anion effect, upregulating ENaC and ROMK in principal cells and BK, pendrin, and KCC3a in pendrin-positive intercalated cells. The adaptive mechanism prevents hyperkalemia and alkalosis with the consumption of alkaline ash-rich diets but may drive K+ wasting and hypokalemia in alkalosis. NEW & NOTEWORTHY Dietary anions profoundly impact K+ homeostasis. Here, we found that a K+-rich diet, containing HCO−3 as the counteranion, enhances the electrogenic K+ excretory machinery, epithelial Na+ channels, and renal outer medullary K+ channels, much more than a high-KCl diet. It also uniquely induces KCC3a and pendrin, in B-intercalated cells, providing an electroneutral KHCO3 secretion pathway. These findings reveal new K+ balance mechanisms that drive adaption to alkaline and K+-rich foods, which should guide new treatment strategies for K+ disorders. [ABSTRACT FROM AUTHOR]

The Cl-/HCO3- exchanger pendrin in the kidney maintains acid-base balance and intravascular volume. Pendrin is upregulated in models associated with high circulating aldosterone concentration, such as dietary NaCl restriction or an aldosterone infusion. However, it has not been established if pendrin is similarly regulated by aldosterone with a high-K+ diet because the effects of accompanying anions have not been considered. Here, we explored how pendrin is modulated by different dietary potassium salts. Wild-type (WT) and aldosterone synthase (AS) knockout (KO) mice were randomized to control, high-KHCO3, or high-KCl diets. Dietary KCl and KHCO3 loading increased aldosterone in WT mice to the same extent but had opposite effects on pendrin abundance. KHCO3 loading increased pendrin protein and transcript abundance. Conversely, high-KCl diet feeding caused pendrin to decrease within 8 h of switching from the high-KHCO3 diet, coincident with an increase in plasma Cl- and a decrease in HCO3-. In contrast, switching the high-KCl diet to the high-KHCO3 diet caused pendrin to increase in WT mice. Experiments in AS KO mice revealed that aldosterone is necessary to optimally upregulate pendrin protein in response to the high-KHCO3 diet but not to increase pendrin mRNA. We conclude that pendrin is differentially regulated by different dietary potassium salts and that its regulation is prioritized by the dietary anion, providing a mechanism to prevent metabolic alkalosis with high-K+ base diets and safeguard against hyperchloremic acidosis with consumption of high-KCl diets. NEW & NOTEWORTHY Regulation of the Cl-/HCO3- exchanger pendrin has been suggested to explain the aldosterone paradox. A high-K+ diet has been proposed to downregulate a pendrin-mediated K+-sparing NaCl reabsorption pathway to maximize urinary K+ excretion. Here, we challenged the hypothesis, revealing that the accompanying anion, not K+, drives pendrin expression. Pendrin is downregulated with a high-KCl diet, preventing acidosis, and upregulated with an alkaline-rich high-K+ diet, preventing metabolic alkalosis. Pendrin regulation is prioritized for acid-base balance. [ABSTRACT FROM AUTHOR]

Pendrin is expressed in the apical regions of type B and non-A, non-B intercalated cells, where it mediates [Cl.sup.-] absorption and HC[O.sup.-.sub.3] secretion through apical [Cl.sup.-]/HC[O.sup.-.sub.3] exchange. Since pendrin is a robust [I.sup.-] transporter, we asked whether pendrin is upregulated with dietary [I.sup.-] restriction and whether it modulates [I.sup.-] balance. Thus [I.sup.-] balance was determined in pendrin null and in wild-type mice. Pendrin abundance was evaluated with immunoblots, immunohistochemistry, and immunogold cytochemistry with morphometric analysis. While pendrin abundance was unchanged when dietary [I.sup.-] intake was varied over the physiological range, [I.sup.-] balance differed in pendrin null and in wild-type mice. Serum [I.sup.-] was lower, while [I.sup.-] excretion was higher in pendrin null relative to wild-type mice, consistent with a role of pendrin in renal [I.sup.-] absorption. Increased [H.sub.2]O intake enhanced differences between wild-type and pendrin null mice in [I.sup.-] balance, suggesting that [H.sub.2]O intake modulates pendrin abundance. Raising water intake from ~4 to ~11 ml/day increased the ratio of B cell apical plasma membrane to cytoplasm pendrin label by 75%, although circulating renin, aldosterone, and serum osmolality were unchanged. Further studies asked whether [H.sub.2]O intake modulates pendrin through the action of AVP. We observed that [H.sub.2]O intake modulated pendrin abundance even when circulating vasopressin levels were clamped. We conclude that [H.sub.2]O intake modulates pendrin abundance, although not likely through a direct, type 2 vasopressin receptor-dependent mechanism. As water intake rises, pendrin becomes increasingly critical in the maintenance of [Cl.sup.-] and [I.sup.-] balance. chloride; apical anion exchange; vasopressin; intercalated cells; vasopressin escape doi: 10.1152/ajprenal.90581.2008.

Pendrin (Slc26a4) localizes to type B and non-A, non-B intercalated cells in the distal convoluted tubule, the connecting tubule, and the cortical collecting duct (CCD), where it mediates apical [Cl.sup.-]/HC[O.sup.-.sub.3] exchange. The purpose of this study was to determine whether angiotensin II increases transepithelial net chloride transport, [J.sub.Cl] in mouse CCD through a pendrin-dependent mechanism. [J.sub.Cl] and transepithelial voltage, [V.sub.T], were measured in CCDs perfused in vitro from wild-type and Slc26a4 null mice ingesting a NaCl-replete diet or a NaCl-replete diet and furosemide. In CCDs from wild-type mice ingesting a NaClreplete diet, [V.sub.T] and [J.sub.Cl] were not different from zero either in the presence or absence of angiotensin II ([10.sup.-8] M) in the bath. Thus further experiments employed mice given the high-NaCl diet and furosemide to upregulate renal pendrin expression. CCDs from furosemide-treated wild-type mice had a lumen-negative [V.sub.T] and absorbed [Cl.sup.-]. With angiotensin II in the bath, [Cl.sup.-] absorption doubled although [V.sub.T] did not become more lumen negative. In contrast, in CCDs from furosemide-treated Slc26a4 null mice, [Cl.sup.-] secretion and a [V.sub.T] of ~0 were observed, neither of which changed with angiotensin II application. Inhibiting ENaC with benzamil abolished [V.sub.T] although [J.sub.Cl] fell only ~50%. Thus substantial [Cl.sup.-] absorption is observed in the absence of an electromotive force. Attenuating apical anion exchange with the peritubular application of the [H.sup.+]-ATPase inhibitor bafilomycin abolished benzamil-insensitive [Cl.sup.-] absorption. In conclusion, angiotensin II increases transcellular [Cl.sup.-] absorption in the CCD through a pendrin- and [H.sup.+]-ATPase-dependent process. transepithelial voltage; bafilomycin; [H.sup.+]-ATPase; knockout mice; Slc26a4; intercalated cell

Pendrin is an intercalated cell Cl−/HCO−3 exchanger thought to participate in K+-sparing NaCl absorption. However, its role in K+ homeostasis has not been clearly defined. We hypothesized that pendrin-null mice will develop hypokalemia with dietary K+ restriction. We further hypothesized that pendrin knockout (KO) mice mitigate urinary K+ loss by downregulating the epithelial Na+ channel (ENaC). Thus, we examined the role of ENaC in Na+ and K+ balance in pendrin KO and wild-type mice following dietary K+ restriction. To do so, we examined the relationship between Na+ and K+ balance and ENaC subunit abundance in K+-restricted pendrin-null and wild-type mice that were NaCl restricted or replete. Following a NaCl-replete, K+-restricted diet, K+ balance and serum K+ were similar in both groups. However, following a Na+, K+, and Cl−-deficient diet, pendrin KO mice developed hypokalemia from increased K+ excretion. The fall in serum K+ observed in K+-restricted pendrin KO mice was enhanced with ENaC stimulation but eliminated with ENaC inhibition. The fall in serum K+ observed in K+-restricted pendrin KO mice was enhanced with ENaC stimulation but eliminated with ENaC inhibition. However, reducing ENaC activity also reduced blood pressure and increased apparent intravascular volume contraction, since KO mice had lower serum Na+, higher blood urea nitrogen and hemoglobin, greater weight loss, greater metabolic alkalosis, and greater NaCl excretion. We conclude that dietary Na+ and K+ restriction induces hypokalemia in pendrin KO mice. Pendrin-null mice limit renal K+ loss by downregulating ENaC. However, this ENaC downregulation occurs at the expense of intravascular volume. NEW & NOTEWORTHY Pendrin is an apical Cl−/HCO−3 exchanger that provides renal K+-sparing NaCl absorption. The pendrin-null kidney has an inability to fully conserve K+ and limits renal K+ loss by downregulating the epithelial Na+ channel (ENaC). However, with Na+ restriction, the need to reduce ENaC for K+ balance conflicts with the need to stimulate ENaC for intravascular volume. Therefore, NaCl restriction stimulates ENaC less in pendrin-null mice than in wild-type mice, which mitigates their kaliuresis and hypokalemia but exacerbates volume contraction. [ABSTRACT FROM AUTHOR]

The present study explored whether the intercalated cell Cl-/HCO3- exchanger pendrin modulates epithelial Na+ channel (ENaC) function by changing channel open probability and/or channel density. To do so, we measured ENaC subunit subcellular distribution by immunohistochemistry, single channel recordings in split open cortical collecting ducts (CCDs), as well as transepithelial voltage and Na+ absorption in CCDs from aldosterone-treated wild-type and pendrin-null mice. Because pendrin gene ablation reduced 70-kDa more than 85-kDa γ-ENaC band density, we asked if pendrin gene ablation interferes with ENaC cleavage. We observed that ENaC-cleaving protease application (trypsin) increased the lumen-negative transepithelial voltage in pendrin-null mice but not in wild-type mice, which raised the possibility that pendrin gene ablation blunts ENaC cleavage, thereby reducing open probability. In mice harboring wild-type ENaC, pendrin gene ablation reduced ENaC-mediated Na(+) absorption by reducing channel open probability as well as by reducing channel density through changes in subunit total protein abundance and subcellular distribution. Further experiments used mice with blunted ENaC endocytosis and degradation (Liddle's syndrome) to explore the significance of pendrin-dependent changes in ENaC open probability. In mouse models of Liddle's syndrome, pendrin gene ablation did not change ENaC subunit total protein abundance, subcellular distribution, or channel density, but markedly reduced channel open probability. We conclude that in mice harboring wild-type ENaC, pendrin modulates ENaC function through changes in subunit abundance, subcellular distribution, and channel open probability. In a mouse model of Liddle's syndrome, however, pendrin gene ablation reduces channel activity mainly through changes in open probability. [ABSTRACT FROM AUTHOR]

Pendrin is expressed in the apical regions of type B and non-A, non-B intercalated cells, where it mediates Cl- absorption and HCO3- secretion through apical Cl-/HCO3- exchange. Since pendrin is a robustI- transporter, we asked whether pendrin is upregulated with dietaryI- restriction and whether it modulatesI- balance. ThusI- balance was determined in pendrin null and in wild-type mice. Pendrin abundance was evaluated with immunoblots, immunohistochemistry, and immunogold cytochemistry with morphometric analysis. While pendrin abundance was unchanged when dietaryI- intake was varied over the physiological range,I- balance differed in pendrin null and in wild-type mice. SerumI- was lower, whileI- excretion was higher in pendrin null relative to wild-type mice, consistent with a role of pendrin in renalI- absorption. Increased H2O intake enhanced differences between wildtype and pendrin null mice inI- balance, suggesting that H2O intake modulates pendrin abundance. Raising water intake from ∼4 to ∼11 ml/day increased the ratio of B cell apical plasma membrane to cytoplasm pendrin label by 75%, although circulating renin, aldosterone, and serum osmolality were unchanged. Further studies asked whether H2O intake modulates pendrin through the action of AVP. We observed that H2O intake modulated pendrin abundance even when circulating vasopressin levels were clamped. We conclude that H2O intake modulates pendrin abundance, although not likely through a direct, type 2 vasopressin receptor-dependent mechanism. As water intake rises, pendrin becomes increasingly critical in the maintenance of Cl- and I- balance. [ABSTRACT FROM AUTHOR]

Pendrin is an intercalated cell Cl - /[Formula: see text] exchanger thought to participate in K + -sparing NaCl absorption. However, its role in K + homeostasis has not been clearly defined. We hypothesized that pendrin-null mice will develop hypokalemia with dietary K + restriction. We further hypothesized that pendrin knockout (KO) mice mitigate urinary K + loss by downregulating the epithelial Na + channel (ENaC). Thus, we examined the role of ENaC in Na + and K + balance in pendrin KO and wild-type mice following dietary K + restriction. To do so, we examined the relationship between Na + and K + balance and ENaC subunit abundance in K + -restricted pendrin-null and wild-type mice that were NaCl restricted or replete. Following a NaCl-replete, K + -restricted diet, K + balance and serum K + were similar in both groups. However, following a Na + , K + , and Cl - -deficient diet, pendrin KO mice developed hypokalemia from increased K + excretion. The fall in serum K + observed in K + -restricted pendrin KO mice was enhanced with ENaC stimulation but eliminated with ENaC inhibition. The fall in serum K + observed in K + -restricted pendrin KO mice was enhanced with ENaC stimulation but eliminated with ENaC inhibition. However, reducing ENaC activity also reduced blood pressure and increased apparent intravascular volume contraction, since KO mice had lower serum Na + , higher blood urea nitrogen and hemoglobin, greater weight loss, greater metabolic alkalosis, and greater NaCl excretion. We conclude that dietary Na + and K + restriction induces hypokalemia in pendrin KO mice. Pendrin-null mice limit renal K + loss by downregulating ENaC. However, this ENaC downregulation occurs at the expense of intravascular volume. NEW & NOTEWORTHY Pendrin is an apical Cl - /[Formula: see text] exchanger that provides renal K + -sparing NaCl absorption. The pendrin-null kidney has an inability to fully conserve K + and limits renal K + loss by downregulating the epithelial Na + channel (ENaC). However, with Na + restriction, the need to reduce ENaC for K + balance conflicts with the need to stimulate ENaC for intravascular volume. Therefore, NaCl restriction stimulates ENaC less in pendrin-null mice than in wild-type mice, which mitigates their kaliuresis and hypokalemia but exacerbates volume contraction.

Pendrin (Slc26a4) is a Cl(-)/HCO3 (-) exchanger expressed in renal intercalated cells and mediates renal Cl(-) absorption. With pendrin gene ablation, blood pressure and vascular volume fall, which increases plasma renin concentration. However, serum aldosterone does not significantly increase in pendrin-null mice, suggesting that pendrin regulates adrenal zona glomerulosa aldosterone production. Therefore, we examined pendrin expression in the adrenal gland using PCR, immunoblots, and immunohistochemistry. Pendrin protein was detected in adrenal lysates from wild-type but not pendrin-null mice. However, immunohistochemistry and qPCR of microdissected adrenal zones showed that pendrin was expressed in the adrenal medulla, rather than in cortex. Within the adrenal medulla, pendrin localizes to both epinephrine- and norepinephrine-producing chromaffin cells. Therefore, we examined plasma catecholamine concentration and blood pressure in wild-type and pendrin-null mice under basal conditions and then after 5 and 20 min of immobilization stress. Under basal conditions, blood pressure was lower in the mutant than in the wild-type mice, although epinephrine and norepinephrine concentrations were similar. Catecholamine concentration and blood pressure increased markedly in both groups with stress. With 20 min of immobilization stress, epinephrine and norepinephrine concentrations increased more in pendrin-null than in wild-type mice, although stress produced a similar increase in blood pressure in both groups. We conclude that pendrin is expressed in the adrenal medulla, where it blunts stress-induced catecholamine release.