Altered expression of type II sodium/phosphate cotransporter in polycystic kidney disease.
(73/939)
Renal phosphate (Pi) absorption is mediated via the type II sodium/Pi cotransporter (NaPi-2) in the brush border membrane (BBM) of proximal tubules. Simultaneous detection of NaPi-2 mRNA by in situ hybridization and of NaPi-2 immunoreactivity by immunohistochemistry was performed to investigate the distribution of the cotransporter in healthy control rats and during progression of autosomal dominant polycystic kidney disease (ADPKD). The purpose of the study was to disclose a relation between proximal tubular cell differentiation and NaPi-2 expression. In controls, NaPi-2 expression was present in the entire proximal tubule. In the Han:SPRD (cy/+) model for ADPKD, the proximal nephron is primarily affected by the cystic changes. Epithelial proliferation and impaired epithelial-matrix interaction result in a loss of cell differentiation that eventually leads to cystic enlargement of the nephron. Normal expression of NaPi-2 in this model was found only in tubules with intact BBM. Loss of BBM and cellular interdigitation were paralleled by the loss of NaPi-2 in situ hybridization and immunoreactive signals. These changes were moderate and focal in 2-mo-old rats and generalized all over the cortex after 8 mo. Advanced renal damage in the older PKD group was associated with mild phosphaturia, which suggests functional insufficiency of tubular NaPi-2 reabsorption. These data show how proliferative changes and loss of tubular epithelial differentiation in ADPKD may prevent functional expression of the NaPi-2 system in the proximal tubule in a rapidly progressive manner. NaPi-2 in proximal tubule BBM is suggested to play an important role in impaired tubular absorption of Pi in renal disease. (+info)
Dynamic interaction between myogenic and TGF mechanisms in afferent arteriolar blood flow autoregulation.
(74/939)
The dynamic activity of afferent arteriolar diameter (AAD) and blood flow (AABF) responses to a rapid step increase in renal arterial pressure (100-148 mmHg) was examined in the kidneys of normal Sprague-Dawley rats (n = 11) before [tubuloglomerular feedback (TGF)-intact] and after interruption of distal tubular flow (TGF-independent). Utilizing the in vitro blood-perfused juxtamedullary nephron preparation, fluctuations in AAD and erythrocyte velocity were sampled by using analog-to-digital computerized conversion, video microscopy, image shearing, and fast-frame, slow-frame techniques. These assessments enabled dynamic characterization of the autonomous actions and collective interactions between the myogenic and TGF mechanisms at the level of the afferent arteriole. The TGF-intact and TGF-independent systems exhibited common initial (0-24 vs. 0-13 s, respectively) response slope kinetics (-0.53 vs. -0.47% DeltaAAD/s; respectively) yet different maximum vasoconstrictive magnitude (-11.28 +/- 0.1 vs. -7. 02 +/- 0.9% DeltaAAD; P < 0.05, respectively). The initial AABF responses similarly exhibited similar kinetics but differing magnitudes. In contrast, during the sustained pressure input (13-97 s), the maximum vasoconstrictor magnitude (-7.02 +/- 0.9% DeltaAAD) and kinetics (-0.01% DeltaAAD/s) of the TGF-independent system were markedly blunted whereas the TGF-intact system exhibited continued vasoconstriction with slower kinetics (-0.20% DeltaAAD/s) until a steady-state plateau was reached (-25.9 +/- 0.4% DeltaAAD). Thus the TGF mechanism plays a role in both direct mediation of vasoconstriction and in modulation of the myogenic response. (+info)
Localization of inward rectifier potassium channel Kir7.1 in the basolateral membrane of distal nephron and collecting duct.
(75/939)
Inward rectifier potassium channels (Kir) play an important role in the K(+) secretion from the kidney. Recently, a new subfamily of Kir, Kir7.1, has been cloned and shown to be present in the kidney as well as in the brain, choroid plexus, thyroid, and intestine. Its cellular and subcellular localization was examined along the renal tubule. Western blot from the kidney cortex showed a single band for Kir7.1 at 52 kD, which was also observed in microdissected segments from the thick ascending limb of Henle, distal convoluted tubule (DCT), connecting tubule, and cortical and medullary collecting ducts. Kir7.1 immunoreactivity was detected predominantly in the DCT, connecting tubule, and cortical collecting duct, with lesser expression in the thick ascending limb of Henle and in the medullary collecting duct. Kir7.1 was detected by electron microscopic immunocytochemistry on the basolateral membrane of the DCT and the principal cells of cortical collecting duct, but neither type A nor type B intercalated cells were stained. The message levels and immunoreactivity were decreased under low-K diet and reversed by low-K diet supplemented with 4% KCl. By the double-labeling immunogold method, both Kir7.1 and Na(+), K(+)-ATPase were independently located on the basolateral membrane. In conclusion, the novel Kir7.1 potassium channel is located predominantly in the basolateral membrane of the distal nephron and collecting duct where it could function together with Na(+), K(+)-ATPase and contribute to cell ion homeostasis and tubular K(+) secretion. (+info)
Mouse beta(6) integrin sequence, pattern of expression, and role in kidney development.
(76/939)
Integrins mediate cell-cell and cell-extracellular matrix interactions and play key roles in development. beta(6) integrin expression has been demonstrated in human fetal kidney at a higher level than in the adult, making beta(6) integrin a marker of interest for the study of development of the nephron. The aims of this study were to determine the cDNA sequence for the mouse beta(6) integrin and to characterize beta(6) integrin expression in the developing mouse kidney. Two embryonic mouse kidney cDNA libraries were screened, and the coding region was sequenced. The mouse beta(6) nucleotide coding region sequence shows 82% nucleotide identity to the human sequence. The putative amino acid sequence has 89.5% identity to human beta(6) integrin and contains many conserved domains. By reverse transcription-PCR, beta(6) integrin mRNA expression is very low at 11 d of gestation in the mouse, increases dramatically by E14 and E17 (20-fold, normalized for increases in ss actin), and plateaus by 2 wk of age. beta(6) integrin expression is induced 15- to 20-fold after 5 d in metanephric explant culture. Reverse transcription-PCR of adult rat microdissected nephron segments demonstrates ss(6) integrin mRNA expression in proximal tubule, cortical thick ascending limb, distal nephron segments (inner and outer medullary collecting ducts), and macula densa-containing segments. Lectin-peroxidase and in situ colocalization studies demonstrated expression of ss(6) integrin mRNA in developing proximal tubules and thick ascending limb. Culture of mouse metanephric kidneys with antisense oligonucleotides to beta(6) integrin resulted in inhibition of ureteric bud branching and complete lack of mesenchyme condensation. These studies demonstrate a high homology between the human and mouse beta(6) integrin sequence, a different pattern of expression in the developing mouse kidney compared with the primate kidney, and abnormal metanephric development in culture in the absence of beta(6) integrin. These findings suggest an important role for beta(6) integrin in normal development of the mouse kidney. (+info)
The alpha(1G)-subunit of a voltage-dependent Ca(2+) channel is localized in rat distal nephron and collecting duct.
(77/939)
The molecular type and localization of calcium channels along the nephron are not well understood. In the present study, we assessed the distribution of the recently identified alpha(1G)-subunit encoding a voltage-dependent calcium channel with T-type characteristics. Using a RNase protection assay, alpha(1G)-mRNA levels in kidney regions were determined as inner medulla >> outer medulla congruent with cortex. RT-PCR analysis of microdissected rat nephron segments revealed alpha(1G) expression in the distal convoluted tubule (DCT), in the connecting tubule and cortical collecting duct (CT+CCD), and inner medullary collecting duct (IMCD). alpha(1G) mRNA was expressed in the IMCD cell line mIMCD-3. Single- and double-labeling immunohistochemistry and confocal laser microscopy on semithin paraffin sections of rat kidneys by using an anti-alpha(1G) antibody demonstrated a distinct labeling at the apical plasma membrane domains of DCT cells, CT principal cells, and IMCD principal cells. (+info)
Na(+)-K(+)-ATPase-mediated basolateral rubidium uptake in the maturing rabbit cortical collecting duct.
(78/939)
Within the renal cortical collecting duct (CCD), transepithelial Na(+) absorption and K(+) secretion are linked to basolateral Na(+)-K(+)-ATPase activity. Our purpose was to examine the developmental changes in basolateral Na(+)-K(+)-ATPase-mediated (86)rubidium (Rb) uptake, its inhibitor sensitivity and relationship to pump hydrolytic activity and Na(+) transport. Multiple CCDs ( approximately 6 mm) from maturing rabbits were affixed to coverslips, preincubated at 37 degrees C for 10 min (+/-1-2.5 mM ouabain or 10 or 100 micro M Schering-28080, an inhibitor of H(+)-K(+)-ATPase), and then transferred to prewarmed incubation solution containing tracer amounts of (86)Rb (+/-inhibitors). After 1 min at 37 degrees C, tubular samples were rinsed and permeabilized and isotope counts were measured to calculate basolateral Rb uptake. Ouabain-inhibitable Rb uptake, an index of basolateral Na(+)-K(+) pump activity, increased approximately 3-fold during the 1st 8 wk of postnatal life (P < 0.03). The approximately 2-fold increase in absolute rate of Rb uptake between 1 and 6 wk (2.64 +/- 0.45 to 5.02 +/- 0.32 pmol. min(-1). mm(-1)) did not reach statistical significance. The rate of basolateral Rb uptake increased further after the 6th wk of life to 7.29 +/- 0.53 pmol. min(-1). mm(-1) in adult animals (P < 0.03 vs. 6 wk). Schering-28080 failed to inhibit Rb uptake, implying that functional H(+)-K(+)-ATPase is absent at the basolateral membrane. Na(+)-K(+)-ATPase hydrolytic activity, determined by using a microassay that measured inorganic phosphate release from [gamma-(32)P]ATP under maximum velocity (V(max)) conditions, also increased in the differentiating CCD (from 316.2 +/- 44.4 pmol. h(-1). mm(-1) at 2 wk to 555.9 +/- 105.1 at 4 wk to 789.7 +/- 145.0 at 6 wk; r = 1.0 by linear regression analysis; P < 0.005). The parallel approximately 2.5-fold increases in Na(+)-K(+)-ATPase activity and ouabain-sensitive Rb uptake between 2- and 6-wk postnatal age suggest that the developmental increase in basolateral transport capacity is due predominantly to an increase in enzyme abundance. The signals mediating the developmental increase in Na(+)-K(+)-ATPase activity in the CCD remain to be defined. (+info)
Dynamics of glomerular ultrafiltration in the rat. VIII. Effects of hematocrit.
(79/939)
This study was undertaken in an effort to examine the effects of selective variations in systemic hematocrit on the preglomerular, glomerular, and postglomerular micocirculation in the rat. By isovolemic exchange transfusions, systemic hematocrit (control 51 ml/100 ml) was either reduced (21 ml/100 ml, N equal 7 rats) or elevated (62 ml/100 ml, N equal 7). Single nephron glomerular filtration rate varied inversely and filtration fraction varied directly with the changes in hematocrit. The fall in filtration fraction with decreased hematocrit was due to a decline in the measured glomerular transcapillary hydraulic pressure difference and to a marked increased in the initial glomerular plasma flow rate. Afferent (RA)and efferent (RE) arteriolar resistance declined with the fall in hematocrit; RA fell proportionately more than did RE. The rise in filtration fraction with the elevation in hematocrit was due to a marked increase in in part due to a relatively greater rise in RE than in RA. These findings provide an attractive explanation for the general tendency for filtration fraction to vary directly with hematocrit in anemic and polycythemic states in man. (+info)
Differential renal distribution of NHERF isoforms and their colocalization with NHE3, ezrin, and ROMK.
(80/939)
Na(+)/H(+) exchanger regulatory factor (NHERF) and NHERF2 are PDZ motif proteins that mediate the inhibitory effect of cAMP on Na(+)/H(+) exchanger 3 (NHE3) by facilitating the formation of a multiprotein signaling complex. With the use of antibodies specific for NHERF and NHERF2, immunocytochemical analysis of rat kidney was undertaken to determine the nephron distribution of both proteins and their colocalization with other transporters and with ezrin. NHERF was most abundant in apical membrane of proximal tubule cells, where it colocalized with ezrin and NHE3. NHERF2 was detected in the glomerulus and in other renal vascular structures. In addition, NHERF2 was strongly expressed in collecting duct principal cells, where it colocalized with ROMK. These results indicate a striking difference in the nephron distribution of NHERF and NHERF2 and suggests NHERF is most likely to be the relevant biological regulator of NHE3 in the proximal tubule, while NHERF2 may interact with ROMK or other targets in the collecting duct. The finding that NHERF isoforms occur in different cell types suggests that NHERF and NHERF2 may subserve different functions in the kidney. (+info)