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R K Packer

Publications and source records attributed to R K Packer.

16 recordsLinked to original sources
PubMed2006

Long-term regulation of proximal tubule acid-base transporter abundance by angiotensin II.

In the proximal tubule, angiotensin II (Ang-II) regulates HCO(-)(3) reabsorption and H+ secretion by binding the type 1 Ang-II (AT1) receptor, stimulating Na(+)/HCO(-)(3) cotransport and Na(+)/H(+) exchange. Studies were carried out to determine if long-term changes in Ang-II receptor occupation alter the abundance of the basolateral Na(+)/HCO(-)(3) cotransporter (NBC1) or the apical membrane type 3 Na(+)/H(+) exchanger (NHE3). In the first set of experiments, rats eating a low-sodium diet were infused with the AT1 blocker, candesartan, or vehicle. In the second, lisinopril-infused rats were infused with either Ang II or vehicle. Transporter abundances were determined in whole kidney homogenates (WKH) and in brush border membrane (BBM) preparations by semiquantitative immunoblotting. Tissue distribution of transporters was assessed by immunocytochemistry. Blockade of the AT1 receptor by candesartan caused decreased abundance of NBC1 in WKH (59 +/- 9% of control; P<0.05) and Ang-II infusion increased abundance (130 +/- 7% of control; P<0.05). Changes in NBC1 in response to candesartan were confirmed immunohistochemically. Neither candesartan nor Ang II infusion affected the abundance of NHE3 in WKH or cortical homogenates. Candesartan decreased type 2 sodium-phosphate cotransporter abundance in both WKH (52 +/- 7% of control; P<0.05) and BBM (32 +/- 7% of control; P<0.05). Serum bicarbonate was decreased by candesartan and increased by Ang-II. Candesartan also decreased urinary ammonium excretion (P<0.05). The long-term effects of Ang-II in the proximal tubule may be mediated in part by regulation of NBC1 abundance, modifying bicarbonate reabsorption.

Angiotensin II↗
PubMed2000

Long-term regulation of renal Na-dependent cotransporters and ENaC: response to altered acid-base intake.

Increased systemic acid intake is associated with an increase in apical Na/H exchange in the renal proximal tubule mediated by the type 3 Na/H exchanger (NHE3). Because NHE3 mediates both proton secretion and Na absorption, increased NHE3 activity could inappropriately perturb Na balance unless there are compensatory changes in Na handling. In this study, we use semiquantitative immunoblotting of rat kidneys to investigate whether acid loading is associated with compensatory decreases in the abundance of renal tubule Na transporters other than NHE3. Long-term (i.e., 7-day) acid loading with NH(4)Cl produced large decreases in the abundances of the thiazide-sensitive Na-Cl cotransporter (TSC/NCC) of the distal convoluted tubule and both the beta- and gamma-subunits of the amiloride-sensitive epithelial Na channel (ENaC) of the collecting duct. In addition, the renal cortical abundance of the proximal type 2 Na-dependent phosphate transporter (NaPi-2) was markedly decreased. In contrast, abundances of the bumetanide-sensitive Na-K-2Cl cotransporter of the thick ascending limb and the alpha-subunit of ENaC were unchanged. A similar profile of changes was seen with short-term (16-h) acid loading. Long-term (7-day) base loading with NaHCO(3) resulted in the opposite pattern of response with marked increases in the abundances of the beta- and gamma-subunits of ENaC and NaPi-2. These adaptations may play critical roles in the maintenance in Na balance when changes in acid-base balance occur.

Acid-Base Equilibrium↗
PubMed1999

Vasopressin increases Na-K-2Cl cotransporter expression in thick ascending limb of Henle's loop.

To investigate whether the enhancement of thick ascending limb (TAL) NaCl transport in response to long-term increases in circulating vasopressin concentration is associated with increased expression levels of the apical Na-K-2Cl cotransporter in the rat TAL, we have carried out immunoblotting and immunofluorescence studies using affinity-purified, peptide-directed antibodies. Semiquantitative immunoblotting studies demonstrated a marked increase (193% of controls) in Na-K-2Cl cotransporter band density in response to restriction of water intake to 15 ml/day for 7 days. In contrast, the expression levels of two other apical proteins of the TAL (the type 3 Na/H exchanger and Tamm-Horsfall protein) were unchanged in the outer medulla. A 7-day subcutaneous infusion of the V2 receptor-selective vasopressin analog, 1-desamino-[8-D-arginine]vasopressin (DDAVP), to Brattleboro rats also markedly increased Na-K-2Cl cotransporter expression in the outer medulla (183% of controls). Immunofluorescence localization in outer medullary tissue sections confirmed the increase in Na-K-2Cl cotransporter expression in response to DDAVP. We conclude that vasopressin strongly upregulates the expression of the Na-K-2Cl cotransporter of the TAL and that it is likely to play an important role in the long-term regulation of the countercurrent multiplication system.

Animals↗
PubMed1999

Regulation of thick ascending limb ion transporter abundance in response to altered acid/base intake.

Changes in ammonium excretion with acid/base perturbations are dependent on changes in medullary ammonium accumulation mediated by active NH4+ absorption by the medullary thick ascending limb. To investigate whether alterations in the abundance of medullary thick ascending limb ion transporters, namely the apical Na+/K+(NH4+)/2Cl- -cotransporter (BSC-1), the apical Na+/H+ -exchanger (NHE3), and the Na+/K+ -ATPase alpha1-subunit, may be responsible in part for altered medullary ammonium accumulation, semiquantitative immunoblotting studies were performed using homogenates from the inner stripe of the rat renal outer medulla. After 7 d of NH4Cl (7.2 mmol/220 g body wt per d) loading (associated with increased medullary ammonium accumulation), neither BSC-1 nor Na+/K+ -ATPase protein expression was altered, but NHE3 protein abundance was significantly increased. On the other hand, both BSC-1 and Na+/K+ -ATPase protein abundance was increased significantly in rats fed NaHCO3 (7.2 mmol/220 g body wt per d) for 7 d. Rats fed a high-NaCl diet (7.7 mEq Na+/220 g body wt per d) for 5 d also showed marked increases in both BSC-1 and Na+/K+ -ATPase expression. The expression level of NHE3 protein did not change with either NaHCO3 or high NaCl intake. None of these three transporters showed a significant difference in abundance between the groups fed equimolar (7.2 mmol/220 g body wt per d for 7 d) NaHCO3 or NaCl. It is concluded that outer medullary BSC-1 and Na+/K+ -ATPase alpha1-subunit protein abundance is increased by chronic Na+ loading but not by acid/base perturbations and that outer medullary NHE3 protein abundance is increased by chronic NH4Cl loading.

Acid-Base Equilibrium↗
PubMed1997

Inhibition of IMCD 11 beta-hydroxysteroid dehydrogenase type 2 by low pH and acute acid loading.

Mineralocorticoid receptors in the inner medullary collecting duct (IMCD) are protected from glucocorticoid binding by an enzyme, 11 beta-hydroxysteroid dehydrogenase type 2 (11 beta-HSD2). To study the role of 11 beta-HSD2 in acid-base homeostasis, 11 beta-HSD2 activity was measured in rat IMCD-enriched cell suspensions. Homogenates of cell suspensions were incubated in buffers ranging in pH from 6.00 to 8.15 in the presence of 1 microCi of 3H-corticosterone (CS) and 400 microM NAD+. Enzyme activity was expressed as the amount of 3H-CS converted to 3H-11-dehydrocorticosterone (DHCS). IMCD 11 beta-HSD2 activity at pH 6.5 was 49% of activity at pH 7.5; 22.5 versus 11.0 fmol/microgram of protein per h. Experiments also were performed on intact cell suspensions at pH 7.5 and 6.5. There was a 42% inhibition in the IMCD cell suspension conversion rate of 3H-CS to 3H-11-DHCS at pH 6.5; 13.1 versus 7.6 fmol/microgram per h (P < 0.005). In cell suspensions at pH 7.5, 1-day acid loading caused a 26% inhibition in conversion rate, 13.2 versus 9.9 fmol/microgram per h (P < 0.05), when compared with controls. These results suggest that during acute metabolic acidosis, IMCD 11 beta-HSD2 is inhibited and may allow access to the mineralocorticoid receptors by glucocorticoids.

11-beta-Hydroxysteroid Dehydrogenases↗
PubMed1995

Urinary organic anion excretion in response to dietary acid and base loading.

Animals eating a base-loaded or base-forming diet excrete urine containing large amounts of organic anions (OA). Although citrate is the only OA previously identified as being excreted in appreciable amounts during base loading, citrate excretion accounts for only part of total OA excretion. The objectives of this study were to identify other OA excreted by rats and to see how their excretion changed in response to moderate (8 micro Eq/g per day) and heavy (30 micro Eq/g per day) loads of NaHCO3 and NH4Cl. Urinary OA were identified by high-performance liquid chromatography and were measured by enzymatic techniques as well. It was found that, in addition to citrate, significant quantities of alpha-ketoglutarate (alpha-KG) were excreted by base-loaded rats and that the excretion of citrate, alpha-KG, and succinate increased with base loading and decreased with acid loading. Citrate plus alpha-KG excretion rates were, respectively, two-thirds and one-third the rate of HCO3- excretion in rats given moderate and heavy base loads. The excretion of creatinine, glutamine, and hippurate showed no clear pattern in response to acid or base loading. It was concluded that, especially in animals experiencing moderate base loads, increases in the excretion of citrate and alpha-KG represent a much more significant component of base excretion than has been recognized previously.

Acid-Base Equilibrium↗
PubMed1992

Time course of renal glutamate dehydrogenase induction during NH4Cl loading in rats.

To study mechanisms involved in renal glutamate dehydrogenase (GDH) regulation in response to systemic acid loading, we have measured blood pH, ammonium excretion, renal GDH mRNA levels, and GDH activity in rats. Acid intake (0.28 M NH4Cl in drinking water for 3 days) increased GDH mRNA levels in the renal cortex, but had no effect in the outer stripe of the outer medulla, inner stripe of the outer medulla, or the inner medulla. Rats were subjected to a step change in acid intake by alkali loading for 3 days (7.2 meq NaHCO3 per day in food slurry) and shifting to acid loading for up to 7 days (7.2 meq NH4Cl in food slurry). Ammonium excretion rose rapidly, increasing by 14-fold in the first 24-h period and 38-fold in the second 24-h period. Cortical GDH mRNA levels were increased relative to alkali-loaded values by 3.7-fold in 24 h, 4.3-fold in 4 days, but only 2.2-fold in 7 days. GDH activity was unchanged after 24 h of acid intake, but was significantly increased after 48 h. We concluded the following: 1) GDH mRNA is present in all regions of the kidney, but levels increase in response to acid loading only in the renal cortex; 2) GDH mRNA levels increase within 1 day after the initiation of acid loading, but the associated increase in functional enzyme activity takes 2 or more days; and 3) the large increases in ammonium excretion that occur in the first day after initiation of acid loading are not dependent on increased GDH activity.

Ammonium Chloride↗
PubMed1991

Role of countercurrent multiplication in renal ammonium handling: regulation of medullary ammonium accumulation.

Ammonium (NH3 plus NH4+), produced predominantly in the proximal tubule, is transferred to the final urine by a process involving countercurrent multiplication of ammonium which generates an ammonium concentration gradient in the renal medulla. It was hypothesized that if urinary ammonium excretion rates are controlled in part by the medullary ammonium gradient, changes in hydration and acid-base state should cause changes in the medullary ammonium gradient consistent with expected changes in urinary ammonium concentrations. To test that hypothesis, rats were subjected to water diuresis, water deprivation, water deprivation plus furosemide, and dietary acid and base loads and corticomedullary ammonium gradients in their kidneys were then measured. Sections were cut along the corticomedullary axis to yield slices of cortex, outer stripe of outer medulla, inner stripe of outer medulla, and three levels of the inner medulla. The total ammonia content of homogenized slices was measured by either a membrane ammonia electrode or an enzymatic technique. Kidneys from water-deprived animals showed a distinct ammonium gradient along the corticomedullary axis, with the highest contents found at the tip of the papilla. The gradient was attenuated by water diuresis and abolished by furosemide. Acid loading enhanced the gradient, and base loading abolished it. These results indicate that the corticomedullary ammonium gradient is regulated in response to changes in hydration and acid-base state.

Acid-Base Equilibrium↗
PubMed1990

Apical acidification by rabbit papillary surface epithelium.

The rabbit papillary surface epithelium (PSE) is a simple cuboidal epithelium that covers the outer surface of the renal papilla and has an apical surface that faces the urinary space. We studied acid-base transport in this epithelium by dissecting it from the papilla, mounting it in a modified Ussing chamber, and following pH changes in the apical bathing solution. The experiments demonstrated that the PSE is capable of acidifying the apical solution at a substantial rate. The acidification rate was similar with 100% nitrogen and 100% oxygen (with and without 10 microM antimycin A), ruling out a dependence on oxidative metabolism. Addition of 1 mM iodoacetate decreased apical acidification by 55%, suggesting a dependence on glycolysis. The net rate of lactate secretion was only 17% of the total acid secretion rate, indicating that apical acidification was not directly caused by secretion of lactic acid alone. Removal of sodium or potassium from the apical solutions or the addition of 1 mM N-ethylmaleimide failed to eliminate the apical acidification. Although the rate of PSE apical acidification is comparable to that of the rabbit outer medullary collecting duct (on a unit surface area basis), its contribution to urinary net acid excretion is likely to be small, owing to the small relative surface area of the PSE. However, by altering the pH of urine locally within the pelvic recesses, the PSE has the potential of modifying the formation of renal stones within the pelvic recesses.

Acids↗
PubMed1989

Role of organic anions in renal response to dietary acid and base loads.

Bicarbonate is formed when organic anions are oxidized systemically. Therefore, changes in organic anion excretion can affect systemic acid-base balance. To assess the role of organic anions in urinary acid-base excretion, we measured urinary excretion in control rats, NaHCO3-loaded rats, and NH4Cl-loaded rats. Total organic anions were measured by the titration method of Van Slyke. As expected, NaHCO3 loading increased urine pH and decreased net acid excretion (NH4+ + titratable acid - HCO3-), whereas NH4Cl loading had the opposite effect. Organic anion excretion was increased in response to NaHCO3 loading and decreased in response to NH4Cl loading. We quantified the overall effect of organic ion plus inorganic buffer ion excretion on acid-base balance. The amounts of organic anions excreted by all animals in this study were greater than the amounts of NH4+, HCO3-, or titratable acidity excreted. In addition, in response to acid and alkali loading, changes in urinary organic anion excretion were 40-50% as large as changes in net acid excretion. We conclude that, in rats, regulation of organic anion excretion can contribute importantly to the overall renal response to acid-base disturbances.

Acid-Base Equilibrium↗
PubMed1989

Chloride and osmotic water permeabilities of isolated rabbit renal papillary surface epithelium.

The papillary surface epithelium (PSE) covers the mammalian renal papilla. It has been proposed that water or solute fluxes across the PSE might result in changes in urine or medullary interstitial fluid composition. To study osmotic water and chloride permeabilities, the PSE was dissected from underlying medullary tissue and mounted in a small Ussing chamber. Osmotic water permeability was low (14.2 +/- 3.0 microns/s) and was unaffected by 100 nM vasopressin added to the basolateral surface. In contrast, the PSE showed a substantial chloride permeability of 3.1 +/- 0.4 x 10(-5) cm/s that decreased reversibly to 2.2 +/- 0.3 x 10(-5) cm/s (P less than 0.01) with vasopressin. Vasopressin also reversibly increased the transepithelial resistance of the PSE from 87 +/- 9 to 106 +/- 13 omega.cm2 (P less than 0.02). Apical bumetanide (10(-6) M) had no significant effect on PSE chloride permeability. The apparent Na-Cl permeability ratio (0.75 +/- 0.01) calculated from dilution potential measurements was not affected significantly by vasopressin or apical bumetanide. We conclude that it is unlikely that physiologically significant osmotic water fluxes occur across the papillary surface epithelium. However, the NaCl permeability is sufficiently high that physiologically significant transepithelial NaCl fluxes could occur under conditions associated with reflux of urine backward from the papillary tip.

Animals↗