Mice lacking the ␤1-subunit (gene, Kcnmb1; protein, BK-␤1) 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-␤1 is an ancillary subunit. However, the ␤1-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. Because of the correlation between certain forms of hypertension and renal defects, particularly in the distal nephron, it was determined whether the hypertension of Kcnmb1 ؊/؊ has a renal origin. We found that Kcnmb1 ؊/؊ are hypertensive, volume expanded, and have reduced urinary K and Na clearances. These conditions are exacerbated when the animals are fed a high K diet (5% K; HK). Supplementing HK-fed Kcnmb1 ؊/؊ with eplerenone (mineralocorticoid receptor antagonist) corrected the fluid imbalance and more than 70% of the hypertension. Finally, plasma [aldo] was elevated in Kcnmb1 ؊/؊ under basal conditions (control diet, 0.6% K) and increased significantly more than wild type when fed the HK diet. We conclude that the majority of the hypertension of Kcnmb1 ؊/؊ is due to aldosteronism, resulting from renal potassium retention and hyperkalemia.adrenal medulla ͉ BK ͉ eplerenone ͉ mineralcorticoid ͉ volume expansion

Estrogens exert a variety of effects in both reproductive and non-reproductive tissues. With the discovery of ERα splice variants, prior assumptions concerning tissue-specific estrogen signaling need to be re-evaluated. Accordingly, we sought to determine the expression of the classical estrogen receptors and ERα splice variants across reproductive and non-reproductive tissues of male and female mice. Western blotting revealed that the full-length ERα66 was mainly present in female reproductive tissues but was also found in non-reproductive tissues at lower levels. ERα46 was most highly expressed in the heart of both sexes. ERα36 was highly expressed in the kidneys and liver of female mice but not in the kidneys of males. ERβ was most abundant in non-reproductive tissues and in the ovaries. Because the kidney has been reported to be the most estrogenic non-reproductive organ, we sought to elucidate ER renal expression and localization. Immunofluorescence studies revealed ERα66 in the vasculature and the glomerulus. It was also found in the brush border of the proximal tubule and in the cortical collecting duct of female mice. ERα36 was evident in mesangial cells and tubular epithelial cells of both sexes, as well as podocytes of females but not males. ERβ was found primarily in the podocytes in female mice but was also present in the mesangial cells in both sexes. Within the renal cortex, ERα46 and ERα36 were mainly located in the membrane fraction although they were also present in the cytosolic fraction. Given the variability of expression patterns demonstrated herein, identification of the specific estrogen receptors expressed in a tissue is necessary for interpreting estrogenic effects. As this study revealed expression of the ERα splice variants at multiple sites within the kidney, further studies are warranted in order to elucidate the contribution of these receptors to renal estrogen responsiveness.

While there is evidence that sex hormones influence multiple systems involved in salt and water homeostasis, the question of whether sex hormones regulate aquaporin-2 (AQP2) and thus water handling by the collecting duct has been largely ignored. Accordingly, the present study investigated AQP2 expression, localization and renal water handling in intact and ovariectomized (OVX) female rats, with and without estradiol or progesterone replacement. OVX resulted in a significant increase in urine osmolality and increase in p256-AQP2 in the renal cortex at 7 days post-OVX, as well as induced body weight changes. Relative to OVX alone, estradiol repletion produced a significant increase in urine output, normalized urinary osmolality and reduced both total AQP2 (protein and mRNA) and p256-AQP2 expression, whereas progesterone repletion had little effect. Direct effects of estradiol on AQP2 mRNA and protein levels were further tested in vitro using the mpkCCD principal cell line. Estradiol treatment of mpkCCD cells reduced AQP2 at both the mRNA and protein level in the absence of deamino-8-d-AVP (dDAVP) and significantly blunted the dDAVP-induced increase in AQP2 at the protein level only. We determined that mpkCCD and native mouse collecting ducts express both estrogen receptor (ER)α and ERβ and that female mice lacking ERα displayed significant increases in AQP2 protein compared with wild-type littermates, implicating ERα in mediating the inhibitory effect of estradiol on AQP2 expression. These findings suggest that changes in estradiol levels, such as during menopause or following reproductive surgeries, may contribute to dysregulation of water homeostasis in women.

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)β1 subunit (gene: Kcnmb1), which has a role in K+ secretion in the CNT, is absent in the CCD in mice on a control diet. We hypothesized that BKα/β1 helps to maximize Na+ reabsorption during Na+ deficiency. With LNa+, the Na+ clearance of Kcnmb1-mutant mice ( Kcnmb1−/−) was 45% greater and the plasma Na+ concentration and osmolality were significantly reduced compared with wild-type mouse (WT) controls. On LNa+, Kcnmb1−/− exhibited exacerbated volume depletion (higher Hct and weight loss) compared with WT. LNa+, which did not affect the mean arterial blood pressure (MAP) of WT, significantly reduced MAP of Kcnmb1−/−. The plasma aldosterone concentration of Kcnmb1−/− on LNa+ was significantly elevated compared with Kcnmb1−/− on a control diet but was not different from WT on LNa+. Immunohistochemical staining revealed that BKα and BKβ1, which were absent in the principal cells (PCs) of the CCD, were localized on the basolateral membrane (BSM) of PCs of WT on LNa+. Moreover, BKα was absent from the BSM of PCs of Na+-deficient Kcnmb1−/−. We conclude that part of the mechanism to maximize Na+ reabsorption during Na+ deficiency is the placement of BKα/β1 channels in the BSM of CCD PCs.

We tested the hypothesis that oral NaHCO intake stimulates splenic anti-inflammatory pathways. Following oral NaHCO loading, macrophage polarization was shifted from predominantly M1 (inflammatory) to M2 (regulatory) phenotypes, and FOXP3CD4 T-lymphocytes increased in the spleen, blood, and kidneys of rats. Similar anti-inflammatory changes in macrophage polarization were observed in the blood of human subjects following NaHCO ingestion. Surprisingly, we found that gentle manipulation to visualize the spleen at midline during surgical laparotomy (sham splenectomy) was sufficient to abolish the response in rats and resulted in hypertrophy/hyperplasia of the capsular mesothelial cells. Thin collagenous connections lined by mesothelial cells were found to connect to the capsular mesothelium. Mesothelial cells in these connections stained positive for the pan-neuronal marker PGP9.5 and acetylcholine esterase and contained many ultrastructural elements, which visually resembled neuronal structures. Both disruption of the fragile mesothelial connections or transection of the vagal nerves resulted in the loss of capsular mesothelial acetylcholine esterase staining and reduced splenic mass. Our data indicate that oral NaHCO activates a splenic anti-inflammatory pathway and provides evidence that the signals that mediate this response are transmitted to the spleen via a novel neuronal-like function of mesothelial cells.