bluuebunny/crossref_metadata_2025 huggingface dataset:
Na(+)-Ca2+ exchange and Ca2+ depletion in rat proximal tubules
Authors: Dominguez J. H. , Mann C. , Rothrock J. K. , Bhati V.
Published: December 2017

The removal of external Ca2+ ([Ca2+]o) reduces cytosolic Ca2+ ([Ca2+]i) in rat proximal tubules. In this report the role of external Na+ ([Na+]o) on the changes of [Ca2+]i and Ca2+ efflux caused by withdrawal of [Ca2+]o is described in rat renal proximal tubules. In aequorin-loaded tubules [Ca2+]i decreased from 235 +/- 25 to 48 +/- 16 (n = 4, P = 0.017), and 45Ca2+ fractional efflux ratio (45Ca2+ FER) increased from 0.94 +/- 0.03 to 1.64 +/- 0.19 (n = 6, P = 0.021) when Ca2+ was withdrawn from the bathing media of Krebs buffer (KB). The fall of [Ca2+]i, as well as the activation of 45Ca2+ FER, was reversed when [Na+]o in Ca(2+)-free KB was lowered isosmotically from 150 to 15 mM. However, when tubules were superfused with only 5 mM [Na+]o before [Ca2+]o was removed, [Ca2+]i also declined, but 45Ca2+ FER did not increase. The Na(+)-Ca2+ exchange inhibitor dichlorobenzamil (DCB) added after [Ca2+]o was removed evoked responses similar to [Na+]o removal, although DCB also inhibited internal Ca2+ release. These results are congruous with stimulation of Na+ influx in exchange for [Ca2+]i in Ca(2+)-free KB. However, even though total tubular Na+ was higher in Ca(2+)-free KB after 10 min, the initial rate of 22Na+ influx was not different without or with [Ca2+]o.(ABSTRACT TRUNCATED AT 250 WORDS)
Cellular distribution of H(+)-ATPase following acute unilateral ureteral obstruction in rats
Authors: Purcell H. , Bastani B. , Harris K. P. , Hemken P. , Klahr S. , Gluck S.
Published: December 2017

Unilateral ureteral obstruction for 24 h produces an acidification defect in the rat kidney that closely resembles the human disorder. We examined the role of renal vacuolar H(+)-ATPase distribution and content in the generation of the postobstructive abnormality in distal hydrogen ion secretion. Rats were subjected to unilateral ureteral obstruction for 24 h, and the obstructed and contralateral kidneys were removed at 3 h, 5 days, and 10 days after release of the obstruction. The postobstructed and contralateral kidneys and kidneys from sham-operated rats were analyzed for intercalated cell number and subtype and for the cellular distribution of ATPase staining by means of a monoclonal antibody specific for the 31-kDa subunit of the vacuolar H(+)-ATPase. No change in the number or distribution of subtypes was detected in the cortex nor in the outer or inner stripe of the outer medulla. Immunoreactive H(+)-ATPase increased in both the cortex and medulla at 3 h after obstruction, and thereafter it declined to control values. The major morphological changes in H(+)-ATPase staining detected were an alteration in the intracellular distribution of the enzyme, which we refer to as discontinuity of (or “gaps” in) apical staining, and a decrease in the percent of intercalated cells showing a rim (or plasma membrane) staining pattern in the inner medulla. The changes observed may be a morphological representation of the physiological abnormalities underlying the postobstructive acidification defect.
Organic cation transport in human renal brush-border membrane vesicles
Authors: Ott R. J. , Hui A. C. , Yuan G. , Giacomini K. M.
Published: December 2017

The renal proximal tubule is responsible for the active elimination of organic cations. Studies in brush-border membrane vesicles (BBMV) suggest that active organic cation transport is mediated by an organic cation-proton antiporter. The goals of this study were to determine whether this transporter is expressed in human kidney and to elucidate its characteristics. Transport of the organic cations N1-[3H]methylnicotinamide (NMN) and [14C]tetraethylammonium (TEA) was determined by rapid filtration in BBMV of donor human kidneys. The uptake of NMN and TEA was driven against a concentration gradient by an outwardly directed proton gradient. NMN uptake was inhibited by the organic cations TEA, NMN, quinine, and cimetidine, but was not affected by p-aminohippuric acid, cephalexin, and polyamines. The electroneutral transport of NMN was characterized by a Km of 0.44 +/- 0.07 mM and a Vmax of 24.4 +/- 15.2 pmol.mg protein-1.s-1. The rate of proton efflux from BBMV increased in the presence of an inwardly directed TEA gradient. Preloading BBMV with NMN or TEA trans-stimulated the initial rate of uptake of TEA. Therefore the human proximal tubule expresses an organic cation-proton antiporter in the brush-border membrane. The antiporter is similar to that described in other mammalian species but shows some differences in its transport characteristics.
Abnormal intestinal regulation of calbindin-D9K and calmodulin by dietary calcium in genetic hypertension
Authors: Roullet C. M. , Roullet J. B. , Duchambon P. , Thomasset M. , Lacour B. , McCarron D. A. , Drueke T.
Published: December 2017

Using isolated duodenal cells from spontaneously hypertensive rats (SHR) and their normotensive controls, Wistar-Kyoto rats (WKY), we previously showed that cellular calcium flux was decreased in SHR and that increasing dietary calcium (from 1 to 2%) eliminated strain differences in Ca2+ fluxes. The present study was carried out to investigate the role of calbindin-D9K and calmodulin in the flux difference and dietary calcium effects. Calbindin-D9K and calmodulin were separated by sodium dodecyl sulfate (SDS) gel electrophoresis in duodenal protein extracts of SHR and WKY (12–14 and 24–26 wk old) fed either a 1 or 2% calcium diet and measured by a ligand blotting (45Ca) technique. Young SHR had a significantly lower calbindin-D9K (P less than 0.001) than did WKY on either diet. Calmodulin was significantly lower in young SHR than in WKY (P less than 0.002). There was no strain difference in calmodulin in older rats fed the normal calcium diet. Calbindin-D9K was significantly decreased by the high-calcium diet in both strains at both ages. There was a significant correlation between duodenal calbindin-D9K and plasma levels of calcitriol (r = +0.80, P less than 0.001) in WKY but not in SHR. Calmodulin was significantly decreased by dietary calcium in mature WKY (4.8 +/- 0.2 vs. 3.7 +/- 0.4 micrograms/mg cell protein, P less than 0.03), demonstrating a potential regulation by dietary calcium of this protein. Finally, there was a significant correlation between calbindin-D9K and calmodulin (r = 0.59, P less than 0.001) in WKY but not in SHR.(ABSTRACT TRUNCATED AT 250 WORDS)
Low-dose glucocorticoids maintain Na-H exchange in distal colon of adrenalectomized rats
Authors: Bastl C. P. , Bressler L. , Schulman G. , Mendez M. , Cragoe E. J.
Published: December 2017

With in vivo perfusion we demonstrated that physiological doses of glucocorticoids restore Na and Cl absorption in adrenalectomized rat colon. The absorption is spironolactone and amiloride resistant and is inhibited by the Na-H inhibitor, 5-(N-ethyl-N-isopropyl)amiloride (EIPA), suggesting that glucocorticoids modulate Na-H antiport. The present in vitro study examines pathways mediated by glucocorticoids in adrenalectomized rat distal colon and rectum. In vivo administration of 2.5 micrograms/100 g body wt dexamethasone did not alter serosal-to-mucosal flux or tissue electrical parameters but restored mucosal-to-serosal flux and net Na and Cl absorption within 2–3 h of administration to levels found in intact rat colon. Transport was not inhibited by 10(-5) M amiloride but was eliminated by 10(-5) M EIPA. After 26 h of dexamethasone, an amiloride-resistant short-circuit current was stimulated, accompanied by increased residual ion flux in rectum, but not distal colon, suggesting that a delayed or secondary effect of glucocorticoids is stimulation of electrogenic anion secretion. Thus adrenalectomy reduces net ion flux in distal colon by its effect on electroneutral mucosal-to-serosal NaCl flux. Small doses of glucocorticoids completely ameliorate this effect via stimulation of the Na-H antiport. Glucocorticoids maintain basal electroneutral NaCl absorption in distal rat colon.
Direct renal interstitial volume expansion causes exaggerated natriuresis in SHR
Authors: Khraibi A. A.
Published: December 2017

In Okamoto spontaneously hypertensive rats (SHR), elevated arterial blood pressure is not transmitted to the renal interstitium, and therefore pressure natriuretic and diuretic responses are attenuated. The objective of this study was to determine the effect of increasing renal interstitial hydrostatic pressure (RIHP) by direct renal interstitial volume expansion (DRIVE) on natriuresis and diuresis of SHR and Wistar-Kyoto rats (WKY). Unilateral nephrectomy and implantation of two polyethylene (PE) matrices were performed 3-4 wk before the acute experiment. Four groups of rats, two experimental and two time control, were used. A control clearance period was taken in all groups. In experimental groups and at the beginning and middle of the second period DRIVE was accomplished by bolus injection of a solution of 2.5% human albumin in saline directly into interstitium through one of the PE matrices. In time-control groups saline was infused in renal interstitium at the beginning of the second period. The second PE matrix was used to continuously measure RIHP in all groups. In experimental groups, DRIVE produced a significant increase in RIHP from 3.8 +/- 0.4 to 5.7 +/- 0.8 mmHg (P less than 0.05) in SHR and 4.3 +/- 0.4 to 7.1 +/- 0.5 mmHg (P less than 0.05) in WKY. In both groups the significant increase in RIHP was associated with significant increases in urinary sodium excretion (UNaV), fractional excretion of sodium (FENa), and urine flow rate (V).(ABSTRACT TRUNCATED AT 250 WORDS)
Axial heterogeneity of potassium transport across hamster thick ascending limb of Henle's loop
Authors: Tsuruoka S. , Koseki C. , Muto S. , Tabei K. , Imai M.
Published: December 2017

Functional significance of morphological heterogeneities along the thick ascending limb of Henle's loop of hamsters was explored by the in vitro microperfusion technique with special reference to K+ transport. The transmission electron microscopic study confirmed that there are two types of cells, with smooth surface (S-cell) and rough surface (R-cell), respectively, and that the former is abundant in the medullary thick ascending limb (MTAL), whereas the latter is in the cortical portion (CTAL). The electrophysiological study revealed that in both segments there are two cell populations, one having high basolateral and low apical membrane K+ conductances (HBC) and the other having low basolateral and high apical K+ conductances (LBC). Random cell puncture revealed that the ratios of HBC/LBC were 24/7 (77%/23%) in the MTAL and 7/22 (24%/76%) in the CTAL, suggesting that HBC corresponds to S-cell, whereas LBC corresponds to R-cell. Net K+ transport was determined in two segments by measuring K+ concentration in the collected and perfused fluid by ultramicroflame photometry. In all six tubules of MTAL, net K+ flux had a direction to reabsorption with a mean of 4.87 +/- 0.46 pmol.min-1.mm-1. In marked contrast, in all six tubules of CTAL, we observed K+ secretion with a mean of -3.81 +/- 0.49 pmol.min-1.mm-1. The transmural voltage was positive in both segments and was significantly higher in the CTAL (7.8 +/- 0.5 mV) than in the MTAL (2.5 +/- 0.2 mV). From these observations, we conclude that the S-cell corresponding to the HBC cell reabsorbs K+, whereas the R-cell corresponding to the LBC cell secrets K+.(ABSTRACT TRUNCATED AT 250 WORDS)
Ontogeny of cortical collecting duct sodium transport
Authors: Vehaskari V. M.
Published: December 2017

The postnatal maturation of Na transport in the rabbit cortical collecting duct (CCD) was investigated. CCD segments from rabbits of three different age groups (3-9 days, 10-15 days, and 22-27 days) were perfused in vitro. Lumen-to-bath 22Na fluxes were 38.5 +/- 4.7, 17.0 +/- 2.9, and 30.2 +/- 4.0 pmol.mm-1.min-1 in the three groups, respectively. The high flux in the youngest group was explained by a high passive flux (28.3 +/- 4.2 pmol.mm-1.min-1) determined in the presence of ouabain; the passive 22Na flux in the two other groups (7.1 +/- 2.6 and 3.1 +/- 2.4 pmol.mm-1.min-1) was not significantly different from previously reported adult values. Ouabain-sensitive 22Na flux, reflecting active transport, was low in the two younger groups (10.3 +/- 2.5 and 9.9 +/- 1.9 pmol.mm-1.min-1), but exhibited a rapid increase to 27.1 +/- 2.6 pmol.mm-1.min-1 by 22-27 days of age. In vivo glucocorticoid pretreatment did not affect the Na transport in any age group. Mineralocorticoid pretreatment for 2 days had no effect in the two younger groups, but increased lumen-to-bath 22Na flux from 30.2 +/- 4.0 to 51.1 +/- 4.3 pmol.mm-1.min-1 in the 22- to 27-day-old group. The findings demonstrate that the maturation of rabbit CCD Na transport occurs in two stages, with the first consisting of a decrease in passive permeability during the first 2 wk of life, followed by an increase in active transport and simultaneous development of mineralocorticoid responsiveness.
ATP and calcium modulation of nonselective cation channels in IMCD cells
Authors: Ono S. , Mougouris T. , DuBose T. D. , Sansom S. C.
Published: December 2017

The mIMCD-3 cell line, developed from simian virus 40 transformed mice, was grown on coverslips for single-channel analysis of the apical membrane. An amiloride-sensitive nonselective cation channel (NCATP) was demonstrated that occurred predominantly in excised patches. The selectivity sequence for NCATP was NH4+ = Na+ = K+ = Li+ = Rb+ > Cl-. The single-channel conductance was 24 pS and nonrectifying in 140 mM KCl or NaCl solutions. NCATP was not permeable to barium from the extracellular side. NCATP was not voltage gated but was activated spontaneously upon patch excision or after applying negative pressure (20-40 mmHg) to an excised patch in bath solutions containing 1 microM Ca2+ [mean number of open channels (NPl) = 1.45]. NCATP was not activated when excised into a bath solution in which Ca2+ was reduced to 100 nM. The open probability of NCATP was reduced by 68% when 2 mM ATP was added to the intracellular side of an excised patch but was unaffected when 0.1 mM 8-bromoguanosine 3',5'-cyclic monophosphate was added to the intracellular side. In cell-attached patches, NCATP was activated upon response to a hyposmotic (210 mosmol/kgH2O) bathing solution containing 0.5 mM Ca2+ (NPo = 0.35). These results show that the mIMCD-3 cell line contains a volume-sensitive nonselective cation channel that is modulated by ATP and calcium but not guanosine 3',5'-cyclic monophosphate. It is postulated that NCATP may act to initiate the volume regulatory response in IMCD cells.
Inhibition of locally produced nitric oxide resets tubuloglomerular feedback mechanism
Authors: Thorup C. , Persson A. E.
Published: December 2017

This study was designed to compare the effects of systemic and intratubular infusions of the nitric oxide (NO) synthase inhibitor N omega-nitro-L-arginine (L-NNA) on the tubuloglomerular feedback (TGF) mechanism in anesthetized rats. We recently showed that intravenous infusion of L-NNA led to increases in mean arterial blood pressure (Pa), proximal tubular stop-flow pressure (Psf), and enhanced TGF sensitivity and reactivity. To avoid major systemic effects, in this study TGF was studied after intratubular NO inhibition. Intratubular infusion of L-NNA (10(-3) M) yielded similar results as shown with intravenous infusion, without systemic effects. TGF sensitivity and reactivity were increased, indicated by decreased turning point (TP) from 19.8 +/- 1.0 to 15.2 +/- 0.7 nl/min and increased delta Psf from 10.0 +/- 0.8 to 23.9 +/- 1.9 mmHg (24.3 vs. 59.1%). L-NNA at a concentration of 10(-4) M showed significant changes in both TP (from 20.9 +/- 1.1 to 17.8 +/- 1.0 nl/min) and delta Psf (from 7.6 +/- 0.6 to 13.9 +/- 0.7 mmHg), whereas 10(-5) M only increased delta Psf (9.7 +/- 1.0 vs. 12.1 +/- 1.1 mmHg). However, at low tubular perfusion rates Psf was not influenced by L-NNA. The early proximal flow rate (EPFR) showed no change at low tubular perfusion rates with L-NNA. At maximal TGF activation (40 nl/min), delta EPFR was increased from 34% in control to 62%. Our results suggest that NO not only regulates glomerular capillary pressure but also decreases the sensitivity of the TGF mechanism.
NH+4 transport mediated by Na(+)-K(+)-ATPase in rat inner medullary collecting duct
Authors: Wall S. M. , Koger L. M.
Published: December 2017

The interaction of K+ and NH+4 on the basolateral membrane of cultured rat inner medullary collecting duct cells (IMCD) was explored using 86Rb+ as a K+ congener. Ouabain addition decreased Rb+ uptake to 22.0 +/- 2.6% of control. Possible ouabain-sensitive NH+4 transport was therefore explored. Replacement of N-methyl-D-glucamine chloride with NH4Cl or KCl decreased both total and ouabain-sensitive Rb+ uptake. This inhibition of Rb+ uptake by NH+4 was not due to changes in intracellular pH or differences in cell viability. We conclude that NH+4 competes with K+ for binding on the Na(+)-K(+)-adenosinetriphosphatase (ATPase). An inhibitory constant (Ki) for NH+4 of 11.0 +/- 1.6 and a Michaelis constant (Km) for K+ of 1.9 +/- 0.7 mM were measured. To extend these observations, NH+4 was tested as a substrate for ouabain-sensitive ATPase activity in native permeabilized IMCD cells. With 20 mM K+, activity was 202 +/- 73 nmol ATP hydrolyzed.min-1.mg protein-1, and with 20 mM NH+4, activity was 259 +/- 81 nmol ATP hydrolyzed.min-1.mg protein-1. Thus NH+4 substitution for K+ on the Na(+)-K(+)-ATPase is a likely mechanism for NH+4 uptake by the IMCD.
Effect of norepinephrine on cellular sodium transport in Ambystoma kidney proximal tubule
Authors: Abdulnour-Nakhoul S. , Khuri R. N. , Nakhoul N. L.
Published: December 2017

The effect of norepinephrine (NE) on mechanisms of cellular Na+ transport in the isolated, perfused proximal tubule of Ambystoma tigrinum was examined. Single-barreled voltage and ion-selective microelectrodes were used to determine basolateral (V1), luminal (V2), and transepithelial (V3) membrane potentials and intracellular Na+ activity (alpha Nai). In CO2/HCO3- control solution, addition of NE (10(-6) M) to the bath caused depolarizations of V1, V2, and V3 are decreased alpha Nai. These effects were mimicked by isoproterenol and inhibited by propranolol. Addition of NE in the absence of luminal Na+ and substrates did not cause any changes in V1, V2, V3, or alpha Nai. NE did not affect the changes in membrane potential difference (PD) or alpha Nai caused by removal and readdition of luminal substrates and/or Na+. To study the effect of NE on Na-K-adenosinetriphosphatase (Na-K-ATPase), the pump was inhibited by external K+ removal and then reactivated by readdition of 12 mM K+ to the bath in the presence and absence of NE. Reactivation of the pump caused hyperpolarization of membrane PDs, and alpha Nai recovered monotonically in 3-5 min. The peak hyperpolarizations of V1 and V2 (approximately 1 min) were significantly larger in the presence of NE. During the first 3 min, and also at the same alpha Nai, the rate of decrease of alpha Nai was significantly faster in the presence of NE. In conclusion, these results show a direct effect of NE on cell membrane PDs and alpha Nai in the kidney proximal tubule. Most likely, beta-receptors are involved in mediating the action of NE. Neither Na/H exchange nor Na-substrate cotransport at the luminal membrane are affected by NE. On the other hand, NE activates Na-K-ATPase.
Anti-AP-1 activity of all-trans retinoic acid in glomerular mesangial cells
Authors: Simonson M. S.
Published: December 2017

Functional antagonism between retinoic acid (RA) receptors and activator protein-1 (AP-1) transcription factors might regulate expression of genes involved in the response to injury in the kidney. We designed experiments to analyze the mechanisms by which RA inhibits AP-1-directed transcriptional responses in glomerular mesangial cells. RA inhibited serum-stimulated mesangial cell proliferation as assessed by measurements of [3H]thymidine uptake and cell number. In transient transfection assays with a chloramphenicol acetyltransferase reporter, RA completely blocked transcription directed by an AP-1 cis-element in cells stimulated by serum. AP-1 DNA binding was analyzed in electrophoretic gel mobility shift assays using nuclear extracts from control or RA-pretreated cells stimulated with serum. RA did not abolish AP-1 DNA binding activity under the conditions of this assay. The apparent equilibrium dissociation constant, maximal density of binding, and association rate for the AP-1-DNA interaction were similar in serum-stimulated cells or RA-pretreated cells stimulated with serum. RA repressed serum-stimulated induction of the immediate early genes c-fos and c-jun, whose protein products dimerize to form AP-1. Repression was relatively selective for c-fos/c-jun; induction of other immediate early transcription factors (junB, c-myc, and egr-1) was not downregulated by RA. That repression of c-fos by RA might contribute to anti-AP-1 activity was suggested by experiments with an antisense c-fos expression vector, which demonstrated that c-fos induction was required for serum-stimulated AP-1 activity. Together, these data demonstrate that RA antagonizes AP-1-directed transcription without inhibiting AP-1 DNA-binding in mesangial cells. Selective repression of c-fos and c-jun might contribute to the anti-AP-1 activity of RA.
Autoregulation of afferent arteriolar blood flow in juxtamedullary nephrons
Authors: Takenaka T. , Harrison-Bernard L. M. , Inscho E. W. , Carmines P. K. , Navar L. G.
Published: December 2017

Utilizing the in vitro blood-perfused juxtamedullary nephron preparation, we examined the effects of alterations in renal arterial pressure on afferent arteriolar blood flow. With video microscopy and cross-correlation techniques, arteriolar inside diameters and centerline erythrocyte velocity were measured to estimate single afferent arteriolar blood flow. In response to random changes in perfusion pressure, afferent arteriolar diameter (n = 8) varied inversely (-0.53 +/- 0.02%/mmHg), and erythrocyte velocity was directly related (1.4 +/- 0.1%/mmHg). Above 95 mmHg, the slope of the relationship between perfusion pressure and afferent arteriolar blood flow did not differ from zero (0.081 +/- 0.053%/mmHg), suggesting efficient autoregulation. When the tubuloglomerular feedback pathway was interrupted by the addition of furosemide (n = 9) or papillectomy (n = 7), there was attenuation of pressure-induced afferent arteriolar constriction, with impairment in blood flow autoregulation (0.60 +/- 0.05%/mmHg). Superfusion with diltiazem abolished autoregulatory responses in afferent arteriolar diameter and blood flow (1.5 +/- 0.2%/mmHg). These data demonstrate the autoregulation of blood flow of individual afferent arterioles in juxtamedullary nephrons and suggest that both tubuloglomerular feedback-dependent and -independent mechanisms are required for autoregulatory responses.
Proteinuria and impaired glomerular permselectivity in uninephrectomized fawn-hooded rats
Authors: Oliver J. D. , Simons J. L. , Troy J. L. , Provoost A. P. , Brenner B. M. , Deen W. M.
Published: December 2017

Previous studies of glomerular permselectivity have indicated that both size selectivity and charge selectivity changes play a role in the pathogenesis of proteinuria. In this study, we measured Ficoll sieving coefficients, hemodynamic parameters, and urinary protein excretion rates in the FHH strain of fawn-hooded rats. These animals spontaneously develop systemic and glomerular hypertension, proteinuria, and focal and segmental glomerulosclerosis at a relatively young age. Three groups of FHH rats were studied: two-kidney controls (2K), untreated uninephrectomized rats (CON-NX), and uninephrectomized rats treated with the angiotensin I converting enzyme inhibitor enalapril (ENA-NX). CON-NX rats had higher glomerular transcapillary pressures (delta P) and higher urinary excretion rates of both total protein (UpV) and albumin (UaV) than did 2K rats, whereas treatment with enalapril prevented both glomerular hypertension and the increased proteinuria. Ficoll sieving coefficients were significantly higher in both groups of NX rats compared with 2K rats only for Stokes-Einstein radii (rs) < or = 46 A. Fits of sieving data to pore models showed a small increase in the number of large, nonselective pores in NX, which was not prevented by enalapril treatment. Total clearances of Ficoll with rs = 36 A (the size of albumin) in CON-NX and ENA-NX groups were unchanged compared with 2K animals. In contrast, UaV in CON-NX rats was more than six times that of 2K and ENA-NX rats. Across groups, UpV, UaV, and the ratio (UaV)/(UpV) all correlated strongly with delta P.(ABSTRACT TRUNCATED AT 250 WORDS)
The microtubule network of renal epithelial cells is disrupted by ischemia and reperfusion
Authors: Abbate M. , Bonventre J. V. , Brown D.
Published: December 2017

Ischemia results in alterations in the integrity of the plasma membrane of renal epithelial cells, changes in cell polarity, and initiation of cell division. Because microtubules are implicated in these processes, we examined the effects of ischemia and reperfusion on the microtubular cytoskeleton in rat kidney. Major alterations in the microtubule network of S3 proximal tubules were detected after 40 min of ischemia followed by 1 h of reperfusion. There was fragmentation of microtubules and considerably less intense staining with antitubulin antibodies than with normal kidneys. Some thick ascending limbs of Henle close to medullary vascular bundles showed a variable loss of tubulin staining. After 24 and 48 h of reperfusion, tubulin labeling was again present in most proximal tubule cells in contact with the basement membrane but was not detectable in exfoliated cells. Numerous mitotic figures were present in kidneys 48 h after reperfusion. Kidneys subjected to 40 min of ischemia without reperfusion and contralateral kidneys studied after 1 h of reperfusion showed only mild microtubular disruption. Because of the established role of microtubules in the generation and maintenance of epithelial cell polarity, their loss may contribute to structural changes that occur after ischemia and reperfusion.
Localization of angiotensin II type 1 receptor subtype mRNA in rat kidney
Authors: Healy D. P. , Ye M. Q. , Troyanovskaya M.
Published: December 2017

The physiological effects of angiotensin II (ANG II) on the kidney are mediated primarily by the ANG II type 1 (AT1) receptor. Two highly similar AT1 receptor subtypes have been identified in the rat by molecular cloning techniques, namely AT1A and AT1B. The intrarenal localization of the AT1A and AT1B receptor subtypes has not been studied by hybridization methods with subtype-specific receptor probes. Using radiolabeled probes from the 3' noncoding region of the AT1A and AT1B cDNAs, we localized AT1 mRNA in rat kidney by in situ hybridization. Specificity of the 3' noncoding region probes was tested by Northern blot and solution hybridization methods. AT1A mRNA levels were highest in the liver, kidney, and adrenal. In contrast, AT1B mRNA levels were highest in the adrenal and pituitary and low in kidney. Autoradiographic localization of 125I-[Sar1,Ile8]ANG II binding indicated that the highest levels of AT1 receptors were found in glomeruli and vascular elements. In situ hybridization with a nonselective AT1 receptor riboprobe indicated that the highest levels of AT1 mRNA were in the outer medullary vasa recta and cortical glomeruli with additional diffuse labeling of the cortex and outer medulla, consistent with labeling of tubular elements. In contrast, in situ hybridization with the AT1 subtype selective probes revealed that AT1A receptor mRNA was primarily localized to the vasa recta and diffusely to the outer stripe of the outer medulla and the renal cortex.(ABSTRACT TRUNCATED AT 250 WORDS)
Acute inflammation is the harbinger of glomerulosclerosis in anti-glomerular basement membrane nephritis
Authors: Scandrett A. L. , Kissane J. , Lefkowith J. B.
Published: December 2017

The degree of glomerular inflammation and injury during immune-mediated glomerulonephritis is felt to be critical to the eventual development of glomerulosclerosis, although the relative contributions of acute and chronic inflammation are uncertain. By grading the initial dose of antibody in accelerated anti-glomerular basement membrane nephritis, we observed that only animals with the most substantial acute inflammation (in terms of neutrophil influx and acute proteinuria) developed sustained proteinuria followed by an increase in serum creatinine and evidence of severe glomerulosclerosis. Chronic inflammation (i.e., glomerular macrophage influx and evidence of glomerular cell proliferation), in contrast, was evident without the development of glomerulosclerosis. Decreasing the degree of acute inflammation during severe nephritis by complement depletion diminished both the initial and sustained proteinuria and the influx of neutrophils, prevented the terminal increase in serum creatinine, and attenuated the evolution of glomerulosclerosis. Complement depletion, however, did not affect peak proteinuria, macrophage influx, or glomerular cell proliferation. Regression analysis of the entire data set demonstrated that acute (day 1) proteinuria was predictive of the eventual histopathological index, more so than chronic (day 7) proteinuria. To recapitulate, glomerulosclerosis following antiglomerular basement membrane nephritis appears to be substantially dependent on the degree of acute inflammatory injury.
Dopamine and protein phosphatase activity in renal proximal tubules
Authors: Slobodyansky E. , Aoki Y. , Gaznabi A. K. , Aviles D. H. , Fildes R. D. , Jose P. A.
Published: December 2017

In the brain, dopamine, via protein kinase A (PKA) activation of dopamine- and cAMP-regulated phosphoprotein (DARPP-32), inhibits protein phosphatase 1 (PP1) activity and keeps Na(+)-K(+)-adenosinetriphosphatase (ATPase) in its phosphorylated inactive state. In the present study, we examined the relationship among dopamine, PP1, and Na(+)-K(+)-ATPase activities in renal proximal tubules. PP1 activity in proximal tubules was not decreased by dopamine (5 x 10(-9)-10(-4) M), fenoldopam (5 x 10(-6) M), or norepinephrine (5 x 10(-7) M). In contrast, in the medullary thick ascending limb of Henle and in the brain striatum, PP1 activity was decreased by fenoldopam (5 x 10(-6) M). We also showed that the ability of dopamine (10(-6) M) to inhibit Na(+)-K(+)-ATPase activity in proximal tubules (assessed by ouabain-sensitive 86Rb uptake) occurred in the absence or presence of a sodium clamp with 5 microM monensin. Thus the inhibitory effect of dopamine on Na(+)-K(+)-ATPase activity in proximal tubules is not regulated by PP1 activity. Tautomycin and okadaic acid by themselves, at concentrations that inhibited PP1 activity, had no effect on Na(+)-K(+)-ATPase activity in proximal tubules. The ability of a dopamine D1 agonist, fenoldopam, to inhibit PP1 activity in brain striatum and in medullary thick ascending limb, but not in proximal tubules, suggests differential organ and nephron segment regulation of PP activity.
Pathways involved in PTH-induced rise in cytosolic Ca2+ concentration of rat renal proximal tubule
Authors: Tanaka H. , Smogorzewski M. , Koss M. , Massry S. G.
Published: December 2017

Parathyroid hormone (PTH) raises cytosolic Ca2+ concentration ([Ca2+]i) in isolated or cultured renal proximal tubule cells. The pathways through which this action is mediated are not fully delineated. This study explored these pathways utilizing fura 2. [Ca2+]i of freshly prepared renal proximal tubular cells increased from 150 +/- 3.6 to 281 +/- 9.0 nM after the exposure to 10(-7) M angiotensin II, which served as a positive control. Both PTH-(1-84) and PTH-(1-34) produced a dose-dependent rise in [Ca2+]i. The effects of both moieties were similar up to 10(-7) M, but with higher doses the rise in [Ca2+]i with PTH-(1-84) was greater (P < 0.01) than with PTH-(1-34). This effect of the hormone occurred in the presence or absence of calcium in the media, but the rise in [Ca2+]i was significantly greater in the presence of calcium. The PTH-induced rise in [Ca2+]i was markedly inhibited by PTH antagonist [Nle8,18,Tyr34]bPTH-(7-34)-NH2 (bPTH is bovine PTH), verapamil, or nifedipine. 12-O-tetradecanoylphorbol-13-acetate (TPA), an activator of protein kinase C, increased [Ca2+]i of cells, but its effect was less than PTH. Staurosporine abolished the TPA effect and partially inhibited that of PTH. A G protein activator raised [Ca2+]i, whereas a G protein inhibitor and pertussis toxin partially blocked the effect of PTH. Sodium or chloride channel blockers or sodium-free media did not modify the effect of PTH.(ABSTRACT TRUNCATED AT 250 WORDS)
Myosin heavy chain turnover during cardiac mass changes by glucocorticoids
Authors: Czerwinski S. M. , Kurowski T. T. , McKee E. E. , Zak R. , Hickson R. C.
Published: December 2017

One aim of this investigation was to determine whether the cardiac enlargement observed with glucocorticoid treatment is temporary or remains a permanent adaptation if steroid treatment is prolonged. A second aim was to study whether myosin heavy chain (MHC) synthesis rates are coordinated with the cardiac mass responses. Female rats received either a vehicle (1% aqueous carboxymethyl cellulose in saline) or hydrocortisone 21-acetate for 1, 3, 7, 11, and 15 days. Peak cardiac enlargement (10–15%) was observed after 7 days of hormone treatment in two separate series of experiments. The enlargement was maintained through 11 days of steroid injections but by 15 days had declined toward control levels. MHC synthesis measurements were performed by constant infusion of [3H]leucine. Leucine specific activities were similar among precursor pools (intracellular, extracellular, and leucyl-tRNA) and did not vary with steroid treatments. Fractional synthesis rates of ventricular MHC (%/day) did not change during the period of increase in ventricular mass but were reduced to 56–59% of controls (-11/19.5) at 7 and 11 days of treatment, when ventricular mass increases were highest. MHC breakdown (%/day) was reduced to approximately 60% (-11.5/18.7) of controls at 7 and 11 days. Changes in total protein synthesis, which was measured in isolated perfused hearts, were similar to the MHC responses and indicated that the alterations in MHC synthesis are synchronized with the hormonal effects on total protein metabolism. These results demonstrate that peak cardiac enlargement is not maintained with long-term glucocorticoid treatment.(ABSTRACT TRUNCATED AT 250 WORDS)
Effects of high-intensity cycle exercise on sympathoadrenal-medullary response patterns
Authors: Kraemer W. J. , Patton J. F. , Knuttgen H. G. , Hannan C. J. , Kettler T. , Gordon S. E. , Dziados J. E. , Fry A. C. , Frykman P. N. , Harman E. A.
Published: December 2017

Plasma proenkephalin peptide F immunoreactivity and catecholamines were examined on separate days in nine healthy males before and after maximal exercise to exhaustion at four intensities [36, 55, 73, and 100% of maximal leg power (MLP)] by use of a computerized cycle ergometer. The mean duration of 36, 55, 73, and 100% MLP was 3.31, 0.781, 0.270, and 0.1 min, respectively. All intensities were greater than those eliciting peak O2 uptake for the individual subjects. Blood samples were obtained before, immediately after exercise, and 5 and 15 min after exercise. Significant (P less than 0.05) increases in plasma peptide F immunoreactivity (i.e., from mean resting value of 0.18 to 0.43 pmol/ml) were observed immediately after exercise at 36% MLP. Significant increases in plasma epinephrine were observed immediately after exercise at 36% MLP (i.e., from mean resting value of 2.22 to 3.11 pmol/ml) and 55% MLP (i.e., from mean resting value of 1.67 to 2.98 pmol/ml) and 15 min after exercise at 100% MLP (i.e., from mean resting value of 1.92 to 3.88 pmol/ml). Significant increases for plasma norepinephrine were observed immediately after exercise (36, 55, 73, and 100% MLP), 5 min after exercise (36, 55, and 73% MLP), and 15 min after exercise (36% MLP). Increases in whole blood lactate were observed at all points after exercise for 36, 55, and 73% MLP and 5 min after exercise for 100% MLP. These data show that brief high-intensity exercise results in differential response patterns of catecholamines and proenkephalin peptide F immunoreactivity.
Differential response of rat limb bones to strenuous exercise
Authors: Li K. C. , Zernicke R. F. , Barnard R. J. , Li A. F.
Published: December 2017

We examined the influence of a strenuous exercise regimen on tibial and metatarsal bones to show not only how the geometric, histological, and mechanical properties of immature bone respond to strenuous exercise but also how long bones within the same limb may respond differentially to exercise. Female Sprague-Dawley rats (8 wk old) were divided randomly into two groups: a sedentary control (n = 15) and an exercised group (n = 15). The exercise intensity was 80-90% of maximum oxygen capacity 5 days/wk for 10 wk. Mechanical properties of tibia and second metatarsus (MT) were determined with three-point bending, and contralateral bones were used for geometric and histological analyses. Length and middiaphyseal cross-sectional geometry of the exercised tibiae were significantly less than controls, but material properties were not different. The exercised tibiae had significantly lower structural properties (e.g., loads at the proportional limit and maximum and energy at failure load). The middiaphyseal dorsal cortex of exercised MT was significantly thicker than controls, but tensile stress at the proportional limit and elastic modulus of exercised MT were significantly less than controls. The average number of osteons and osteocytes per unit area of the tibial middiaphysis was significantly greater in the exercised group--especially in lateral and posterior cortices. The number of osteons and osteocytes per unit area in the MT, however, was significantly less in the exercised group. The differential effects of strenuous exercise on tibia and MT suggest that local loading and bone-specific responses have important roles in modulating the response of immature bone to strenuous exercise.
Positive-pressure ventilation alters blood-to-brain and blood-to-CSF transport in neonatal pigs
Authors: Mirro R. , Leffler C. W. , Armstead W. M. , Busija D. W.
Published: December 2017

These experiments examine the transfer of sucrose, urea, sodium, and albumin from blood to brain in new-born pigs exposed to an increase in ventilation pressure. We also studied the movement of urea and sodium from blood to cerebrospinal fluid (CSF). By use of a standard time-cycled pressure-limited infant respirator, mean airway pressure (Paw) was increased from approximately 3 to 17 cmH2O. Urea and albumin transfer into the brain were unchanged with increased Paw. Sodium transport decreased significantly in all brain regions, while sucrose transfer was increased in the cerebrum [transfer constant (Kin) = 3.5 +/- 0.04 vs. 9.9 +/- 1.0 cm3.g-1.s-1.10(6)] at the increased Paw. Transport of urea nd sodium from blood to CSF decreased to half of control values with increased Paw. Thus, in newborn pigs, increasing Paw selectively alters blood-to-brain transport. In addition, movement of tracers from blood to CSF was severely restricted, possibly by a decrease in CSF production. It appears likely that the increased cerebral venous pressure causes the observed changes in tracer transport. Such altered blood-to-brain transport could adversely affect neuronal function.
Localization and in vitro actions of atrial natriuretic peptide in the cat carotid body
Authors: Wang Z. Z. , He L. , Stensaas L. J. , Dinger B. G. , Fidone S. J.
Published: December 2017

Previous studies of atrial natriuretic peptide (ANP) have indicated that its release from the heart and from discrete areas of the central nervous system evokes coordinated physiological and behavioral adjustments that mitigate the adverse hypertensive effects of volume overload and/or acute increases in sodium intake. Because the reflex activity initiated by arterial chemoreceptors of the carotid body directly contributes to the integrated regulation of systemic blood pressure, we have investigated the possibility that ANP has a significant role in the chemosensory process as well. Our immunocytochemical studies show that ANP-like immunoreactivity is present in the preneural chemosensitive type I cells in the cat carotid body. Furthermore we found that the biologically active ANP fragment atriopeptin III is a potent inhibitor of carotid sinus nerve activity evoked by hypoxia. Our findings suggest that circulating and/or endogenous ANP may modulate carotid body function as part of a coordinated response to changes in systemic volume and solute balance.
Restraining hamsters alters their breathing pattern
Authors: Sweeney T. D. , Leith D. E. , Brain J. D.
Published: December 2017

Does the restraint required for head or nose-only exposure of rodents to inhaled aerosols or gases alter their breathing pattern? And does prior exercise training, which may increase muscle strength, affect this response to restraint? To answer those questions, we measured breathing pattern in 11 adult male hamsters while they were either 1) free to move in small cages or 2) closely restrained in head-out cones. The measurements were repeated after hamsters spent 6 wk either sedentary in standard cages or in cages with exercise wheels. Hamsters were placed in a plethysmograph to measure respiratory frequency (f) and tidal volume (VT). Their product is minute volume (V). When restrained, f and V were 1.9 and 1.7 times, respectively, greater than when hamsters were free, but VT did not change. After 6 wk, the sedentary group responded differently to restraint; f increased 3-fold, VT decreased by one-half, and V increased 1.6-fold. Exercised hamsters increased f 2.3-fold and decreased VT by one-third; V increased by 1.5-fold. In inhalation studies, changes in breathing pattern would significantly influence the amount of material inhaled, the fraction retained, and thus the amount and distribution of material deposited in the lungs.
Effect of amiloride on diaphragmatic contractility: evidence of a role for Na(+)-Ca2+ exchange
Authors: Zavecz J. H. , Anderson W. M. , Adams B.
Published: December 2017

Extracellular Ca2+ has been shown to be important for the normal function of the diaphragm. In this study we have examined the potential importance of Na(+)-Ca2+ exchange as a mechanism for Ca2+ influx during the contractile process by studying the effect of inhibition or stimulation of Na(+)-Ca2+ exchange. Blockade of Na(+)-Ca2+ exchange with amiloride attenuated the twitch response, altered the force-frequency response curve, and enhanced the development of fatigue. The effect of amiloride could be partially reversed by increasing the extracellular Ca2+ concentration. The ability of amiloride to decrease force was associated with decreased Ca2+ uptake by the diaphragm. Enhancing intracellular Na(+)-extracellular Ca2+ exchange by inhibiting the Na(+)-K+ pump significantly decreased the rate of the development of muscle fatigue (89%). The maximal inhibition of diaphragmatic force produced by the amiloride analogue benzamil, which possesses 10-fold greater selectivity for Na(+)-Ca2+ exchange, was not significantly different from that produced by amiloride (76.2 +/- 1.1%), with a concentration that decreased maximum force by 50% equal to 46 microM compared with 460 microM for amiloride. Both agents slowed the maximal rate of relaxation up to 90%. Benzamil elevated resting tension during continuous stimulation of the diaphragm at 0.15 Hz. The results suggest that Na(+)-Ca2+ exchange may have a role in the normal function of the diaphragm.
Stimulation of glucose transport in skeletal muscle by hypoxia
Authors: Cartee G. D. , Douen A. G. , Ramlal T. , Klip A. , Holloszy J. O.
Published: December 2017

Hypoxia caused a progressive cytochalasin B-inhibitable increase in the rate of 3-O-methylglucose transport in rat epitrochlearis muscles to a level approximately six-fold above basal. Muscle ATP concentration was well maintained during hypoxia, and increased glucose transport activity was still present after 15 min of reoxygenation despite repletion of phosphocreatine. However, the increase in glucose transport activity completely reversed during a 180-min-long recovery in oxygenated medium. In perfused rat hindlimb muscles, hypoxia caused an increase in glucose transporters in the plasma membrane, suggesting that glucose transporter translocation plays a role in the stimulation of glucose transport by hypoxia. The maximal effects of hypoxia and insulin on glucose transport activity were additive, whereas the effects of exercise and hypoxia were not, providing evidence suggesting that hypoxia and exercise stimulate glucose transport by the same mechanism. Caffeine, at a concentration too low to cause muscle contraction or an increase in glucose transport by itself, markedly potentiated the effect of a submaximal hypoxic stimulus on sugar transport. Dantrolene significantly inhibited the hypoxia-induced increase in 3-O-methylglucose transport. These effects of caffeine and dantrolene suggest that Ca2+ plays a role in the stimulation of glucose transport by hypoxia.
Polymyxin B inhibits contraction-stimulated glucose uptake in rat skeletal muscle
Authors: Young J. C. , Kurowski T. G. , Maurice A. M. , Nesher R. , Ruderman N. B.
Published: December 2017

Glucose transport in muscle is activated by contractile activity, an effect that persists in the postexercise state. Polymyxin B, a cyclic decapeptide antibiotic, inhibits the stimulation of glucose uptake in isolated muscle by contractile activity but also decreases tension development in electrically stimulated muscle. The purpose of this study was to determine whether polymyxin B also inhibits contraction-stimulated glucose uptake after in vivo administration of the drug and to examine the relationship between the effects of polymyxin B on tension development and its effects on contraction-stimulated glucose uptake. When polymyxin B was administered to rats in vivo, glucose uptake in muscle after electrical stimulation was decreased, despite the same amount of tension developed as in control rats, indicating an effect of polymyxin B on glucose transport independent of tension development. Our results also indicate that the postexercise increase in glucose uptake is a function of the tension developed by prior contractions. When muscles were perfused with medium containing polymyxin B, this relationship was disrupted. These results provide evidence that polymyxin B causes a decrease in muscle glucose uptake independent of its effects on tension development. The extent to which its effects on glucose uptake are also the result of a diminution in contractile force is uncertain.