Biomedical subjects

W H Dantzler

Publications and source records attributed to W H Dantzler.

At least 37 records · Page 2Linked to original sources

PubMed1998

Regulation of intracellular pH in proximal tubules of avian long-looped mammalian-type nephrons.

In nonperfused proximal tubules isolated from chicken long-looped mammalian-type nephrons, intracellular pH (pHi), measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, was approximately 7.3 under control conditions (HEPES-buffered medium with pH 7.4 at 37 degrees C) and was reduced to approximately 7.0 in response to NH4Cl pulse. The rate of recovery of pHi from this level to the resting level was 1) significantly reduced by the removal of Na+ from the bath, 2) significantly increased by the removal of Cl- from the bath, 3) unchanged by the removal of both Na+ and Cl- from the bath, 4) significantly reduced by the addition of either ethylisopropylamiloride or DIDS to the bath, 5) significantly increased by a high bath K+ concentration, and 6) unchanged by the addition of Ba2+ to the bath. These data suggest that both Na+-coupled and Cl--coupled basolateral acid-base fluxes are involved in determining the rate of recovery of pHi after acidification. The most likely ones to be important in regulating pHi are a Na+/H+ exchanger and a Na+-coupled Cl-/HCO-3 exchanger. In birds, long-looped mammalian-type nephrons resemble short-looped transitional nephrons but differ markedly from superficial loopless reptilian-type nephrons.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1998

Effect of alpha-ketoglutarate on organic anion transport in single rabbit renal proximal tubules.

The effect of exogenous alpha-ketoglutarate (alpha KG) and the peritubular Na(+)-dicarboxylate (Na-DC) cotransporter on organic anion/dicarboxylate (OA/DC) exchange in S2 segments of single, nonperfused rabbit proximal tubules was measured using 1 microM fluorescein (FL), a model OA, and epifluorescence microscopy. The effect of different transmembrane distributions of 10 microM alpha KG on peritubular FL uptake was measured at 37 degrees C using bicarbonate-buffered, nutrient-containing buffers, which are conditions similar to those found in vivo. Compared with FL uptake in the absence of exogenous alpha KG, preloading tubules with alpha KG (trans-configuration) or acute exposure to alpha KG (cis-configuration) increased FL uptake 62% and 54%, respectively, whereas a cis-trans-configuration of alpha KG increased FL uptake by 76%. The cis-stimulation of FL uptake by alpha KG was rapid, within 5-7 s. This stimulation was blocked 96% by simultaneous exposure to 2 mM Li+, indicating that stimulation of transport was secondary to the uptake of exogenous alpha KG. In the absence of exogenous alpha KG, selective inhibition of Na-DC cotransport using 2 mM Li+ or 1 mM methylsuccinate decreased FL uptake by 25% (effects that were reversible but not additive), suggesting that the Na-DC cotransporter recycles endogenous alpha KG that has left the cell in exchange for FL and that this activity supports approximately 25% of baseline activity of the OA/DC exchanger. With recycling of alpha KG accounting for approximately 25% of FL uptake and with accumulation of exogenous alpha KG accounting for another approximately 75% increase in FL uptake, Na-DC cotransport appears to directly support (25% + 75%)/175%, or approximately 57%, of total FL transport.

Animals↗

PubMed1998

Relation of cysteine conjugate nephrotoxicity to transport by the basolateral organic anion transport system in isolated S2 segments of rabbit proximal renal tubules.

We examined basolateral transport of the radiolabeled zwitterionic nephrotoxic cysteine S-conjugate, S-(1,2-dichlorovinyl)-L-cysteine (DCVC), inhibition of such transport and the effects of inhibition of transport on the toxicity produced by DCVC in isolated S2 segments of rabbit proximal tubules. High concentrations of unlabeled DCVC itself and an unlabeled nontoxic cysteine S-conjugate, S-(2-benzothiazole)-L-cysteine cis-inhibited the basolateral uptake of radiolabeled DCVC by approximately 80 to 85%. High concentrations of para-aminohippurate, the prototype substrate for the basolateral organic anion transport system, and probenecid, a well-known inhibitor of basolateral organic anion transport, cis-inhibited the basolateral uptake of radiolabeled DCVC by approximately 70%, whereas a high concentration of L-phenylalanine had little effect. High concentrations of S-(2-benzothiazole)-L-cysteine and para-aminohippurate in the bathing medium with DCVC inhibited the loss of 86Rb (used as a K+ surrogate to measure toxicity) from S2 segments produced by DCVC alone to approximately the same extent as they inhibited uptake of DCVC. Under the same circumstances, probenecid completely inhibited 86Rb loss. These data indicate that in rabbit proximal renal S2 tubules basolateral entry of DCVC can occur to a major extent via the organic anion transport pathway and that inhibition of such entry can reduce toxicity to approximately the same extent that entry is reduced. They also suggest that probenecid provides additional protection from DCVC toxicity.

Animals↗

PubMed1998

Detailed mapping of ochratoxin A reabsorption along the rat nephron in vivo: the nephrotoxin can be reabsorbed in all nephron segments by different mechanisms.

Ochratoxin A (OTA) is a widespread nephrotoxin excreted to a substantial degree via the kidney. Previously we showed that [3H]OTA can be reabsorbed along the rat nephron in vivo (Zingerle et al., 1997). In this study we investigated in detail the contribution of different nephron segments to [3H]OTA reabsorption and determined the possible mechanisms involved by microinfusion and microperfusion experiments. At pH 6 (approximately 94% of OTA neutral), OTA is reabsorbed in all nephron segments investigated. The estimated fractional reabsorptions (FR) at a tubular load of 20 fmol/min are: proximal convoluted tubule (PCT), 14.8%; proximal straight tubule (PST), 27.4%; ascending limb of Henle's loop (ALH), 13.6%; distal tubule (DT), 11.6%; collecting duct (CD), 24.6%; terminal CD, 22.0%. At pH 8 (approximately 10% of OTA neutral) FR are as follows: PCT, 0%; PST, 25.9%; ALH, 14.0%; DT, 3.2%; CD, 8.2%. Thus, OTA reabsorption in PST and ALH in pH-independent. Reabsorption in PST but not in DT or CD was inhibited by sulfobromophthalein, a substrate of the apical organic anion carrier. L-Phenylalanine did not reduce OTA reabsorption. After intravenous injection of unlabeled OTA, resulting in a plasma concentration of approximately 10(-5) mol/l, the FR of [3H]OTA during early proximal microinfusion was reduced slightly. From our results we conclude: 1) OTA can be reabsorbed in all nephron segments investigated. 2) Under physiological conditions the predominant sites of reabsorption are PST, ALH and terminal CD. 3) Reabsorption in PST and ALH is not pH-dependent. 4) pH-independent reabsorption in PST is mediated by the apical organic anion transporter (OAT-K1), whereas pH-dependent reabsorption in PCT is mediated by H(+)-dipeptide cotransporter(s). 5) Reabsorption also takes place during natural exposure, i.e., when OTA is present in plasma and renal tissue. 6) The high FR in ALH and CD explains, at least in part, the preferential impairment of postproximal functions and the accumulation in renal inner medulla and papila.

Absorption↗

PubMed1997

Ca2+ and Ba2+ effects on basolateral tetraethylammonium transport in isolated snake renal proximal tubules.

Previous work on snake renal tubules suggested that basolateral transport of tetraethylammonium (TEA) is symmetrical. To examine regulation of this transport step more closely, we determined the effects of (1) reductions in the extracellular Ca2+ concentration and basolateral Ca2+ entry, and (2) the presence of extracellular Ba2+ on TEA uptake and efflux across the basolateral membrane of isolated snake renal proximal tubules. Removal of extracellular Ca2+ reduced initial TEA uptake and enhanced TEA efflux. Blocking Ca2+ entry also reduced initial TEA uptake. Extracellular Ba2+ depolarized the basolateral membrane and reduced both TEA uptake and efflux. Inhibition of basolateral TEA uptake with a reduced membrane potential supports previous data indicating that uptake involves potential-driven facilitated diffusion. Inhibition of basolateral TEA efflux by Ba2+ even with a reduced membrane potential not only supports previously obtained data indicating that efflux is not influenced by the potential difference and that basolateral TEA transport is asymmetrical, but also suggests that TEA uptake and efflux may occur by separate pathways.

Animals↗

PubMed1997

Regulation of intracellular pH in avian renal proximal tubules.

In proximal tubules isolated from chicken transitional nephrons, intracellular pH (pHi), measured with the pH-sensitive fluorescent dye 2'.7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF), was approximately 7.3-7.4 under control conditions [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered medium with pH 7.4 at 39 degrees C] and was reduced to approximately 6.8 in response to NH4Cl pulse. The rate of recovery of pHi (dpHi/dt) from this acid level to the resting level and the resting pHi were 1) significantly reduced by the removal of Na+ from the bath, 2) significantly increased by the removal of Cl from the bath, and 3) unchanged by the removal of both Na+ and Cl from the bath. The addition of either amiloride or 4,4'-diisothiocyanostilbene-2,2'-disulfonate to the bath reduced dpHi/dt to about the same extent as the removal of Na+. These data suggest that both Na(+)-coupled and Cl-coupled acid-base fluxes at the basolateral membrane are involved in determining the resting pHi and the rate of recovery of pHi after acidification. The most likely possibilities appear to be a basolateral Na+/Hi exchanger, a basolateral Na(+)-coupled Cl/HCO3 exchanger, a basolateral Na(+)-HCO3(-)CO(3)2 cotransporter, and a basolateral Na(+)-independent Cl-/HCO3 exchanger.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1997

Effects of pH on basolateral tetraethylammonium transport in snake renal proximal tubules.

Previous work on snake renal proximal tubules suggested that pH might influence tetraethylammonium (TEA) transport across the basolateral membrane. To examine this more directly, we determined the effects of altering either extracellular pH (pHo) or intracellular pH (pHi) on TEA uptake and efflux across the basolateral membrane of isolated snake renal proximal tubules. We found no evidence for trans-stimulation of either TEA uptake or efflux by H+. Therefore, there was no evidence for a TEA/H+ exchanger. However, we found evidence for trans-inhibition of both TEA uptake and efflux as well as for cis-inhibition of TEA uptake by increasing H+ concentration. H+ concentration appeared to have some type of direct effect on basolateral transport independent of any effect on membrane potential. Moreover, there appeared to be an optimal intracellular H+ concentration for entry of TEA into the cells that corresponded to the one found at the physiological pHi of 7.1. There also appeared to be an optimal extracellular H+ concentration for efflux of TEA from the cells that corresponded to the one found at the physiological pHo of 7.4. The mechanism involved in this relationship is unknown, but the data support a concept derived from previous studies that TEA transport across the basolateral membrane is asymmetric.

Animals↗

PubMed1997

Regulation of intracellular pH in proximal tubules of avian loopless reptilian-type nephrons.

In proximal tubules isolated from chicken superficial loopless reptilian-type nephrons, intracellular pH (pHi), measured with pH-sensitive fluorescent dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein, was approximately 7.1-7.2 under control conditions (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered medium with pH 7.4 at 37 degrees C), and was reduced to approximately 6.9 in response to NH4Cl pulse. The rate of recovery of pHi (control value approximately equal to 5 x 10(-3) pH U/s) from this acid level was 1) significantly decreased by removal of Na+ or both Na+ and Cl- from the bath or addition of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (0.25 mM) to the bath, 2) significantly increased by high bath K+ (75 mM), and 3) unchanged by removal of Cl- alone from the bath or addition of ethylisopropylamiloride (1 mM) or Ba2+ (5 mM) to the bath. Resting pHi was 1) significantly decreased by Na+ or simultaneous Na+ and Cl- removal, 2) significantly increased by high K+, and 3) unchanged by Cl- removal alone or addition of Ba2+. The data do not fit the concept of pHi regulation by the most commonly suggested basolateral transporters (Na+/H+ exchanger, Na(+)-dependent and Na(+)-independent Cl-/HCO3- exchangers, or Na(+)-HCO3(-)-CO3(2-) cotransporter).

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1997

Taurine reabsorption by a carrier interacting with furosemide in short and long Henle's loops of rat nephrons.

Taurine is net reabsorbed in the proximal convolution by Cl- -stimulated Na+ symport specific for beta-amino acids but not in later nephron segments. However, large unidirectional taurine transport takes place there. To investigate unidirectional taurine reabsorption, we comicroinfused [3H]taurine and [14C]inulin into late proximal (LP), early distal (ED), and late distal (LD) tubule segments, as well as into the tips of long loops of Henle (LLH) of rats in vivo, and determined fractional reabsorption of [3H]taurine (FR) in the ipsilateral urine. FR (9 micromol/l taurine) was 80-93 (LP), 16 (ED), and 8% (LD). At 26 mmol/l taurine, FR decreased to 13 (LP) and 6% (ED). FR also decreased when Na+ or Cl- was absent or furosemide (5 x 10(-5) mol/l) was added. Bumetanide (5 x 10(-5) mol/l) had no effect, whereas aniline-2-sulfonic acid (ASA) inhibited. During LLH microinfusion, FR was 55% at 66 micromol/l and 17% at 228 micromol/l and was again inhibited by furosemide and ASA but not by bumetanide. [14C]taurine reabsorption from microperfused proximal convolutions was not influenced by furosemide. Chronic water diuresis did not affect taurine reabsorption in short Henle's loops. We conclude that taurine can enter cells of the distal nephron from the lumen by an Na+- and partly Cl- -dependent carrier with which C alpha,beta-substituted taurine (ASA) and C alpha,beta- and N-substituted beta-alanine (furosemide) directly interact. Thus proximal and distal taurine carriers seem to be different.

Absorption↗

PubMed1996

Changes in the countercurrent system in the renal papilla: diuresis increases pH and HCO3- gradients between collecting duct and vasa recta.

This study was designed to elucidate the acid-base balance local to the collecting duct urine (CD) and vasa recta blood (VR) in the rat renal papilla in diuresis. The pH changes were measured in both a furosemide-induced and a volume-load-induced diuresis, whereas the PCO2 (i.e., CO2 tension) and HCO3- concentration were measured only in a furosemide-induced diuresis. In an antidiuresis, the pH of the VR was more acidic than that of the systemic arterial blood (DeltapH = 0.44-0.73). Additionally, the pH of the ascending VR was significantly lower than that of the descending VR (DeltapH = 0.14-0. 16). In diuresis, the pH of the CD decreased (DeltapH = 0.81-0.97), while the pH of the descending and the ascending VR increased; however, the increase was only significant in the ascending VR (DeltapH = 0.23-0.30). Consequently, the significant difference in the pH gradient between the descending and the ascending VR was eliminated. The PCO2 values in the CD and the ascending VR were not different from those in antidiuresis, while the HCO3- concentration in the CD and the ascending VR, respectively, decreased and increased significantly. Thus, in diuresis, the decrease in the pH of the CD and the increase in the pH of the ascending VR result, respectively, from the decrease and the increase in the HCO3- concentration, with no changes in the PCO2 values.

Animals↗

PubMed1996

Comparative aspects of renal urate transport.

Urate is the major excretory end product of nitrogen metabolism in birds, most reptiles, and some amphibians and undergoes net secretion by the renal tubules. Secretion has been studied in isolated renal proximal tubules from reptiles and birds. Net secretion is influenced by passive backflux between the tubule cells that varies with perfusion rate. Transepithelial transport from bath to lumen involves uptake into the cells against an electrochemical gradient at the basolateral membrane and movement from the cells to the lumen down an electrochemical gradient. However, the apparent permeability of the basolateral membrane to urate is much higher than that of the luminal membrane. Transport into the cells at the basolateral membrane is dependent, in part, on the presence of filtrate (or an appropriate substitute) in the lumen. Because reptilian and avian nephrons filter intermittently, passive back-diffusion between the cells, high permeability of the basolateral membrane, and dependence of basolateral transport on luminal perfusion may prevent accumulation of urate in the cells or lumens of nonfiltering nephrons. In reptiles, basolateral transport is Na-independent but K-dependent and may involve countertransport for some unknown anion. In birds, this transport step, which is Na- and K-dependent, may occur by two separate systems, one involving counter-transport for an unknown anion and the other involving the general tertiary transport process for other organic anions such as PAH. It has not yet been possible to demonstrate that transport from the cells to the lumen down an electrochemical gradient is carrier-mediated in either reptiles or birds and some other process may be involved.

Animals↗

PubMed1996

Specificity of basolateral organic cation transport in snake renal proximal tubules.

We examined the specificity of basolateral organic cation transport in isolated snake (Thamnophis spp.) renal proximal tubules by determining the inhibitory effect of a series of n-tetraalkylammonium (n-TAA) compounds (n = 1-5) on the basolateral uptake of [3H]tetraethylammonium (TEA). The inhibitory potency increased with increasing alkyl chain length, with the apparent Michaelis constants for inhibition of TEA uptake ranging from 3.3 mM for tetramethylammonium (TMA) to 1.0 microM for tetrapentylammonium (TPeA). Thus the apparent affinity of the carrier for n-TAA compounds increases with their increasing hydrophobicity. Because previous data suggested that TEA transport across the basolateral membrane may be asymmetrical and that the exit step may be regulated differently from the entry step, we examined the kinetics of [3H]TEA efflux across the basolateral membrane, Efflux, like entry, occurred by a saturable process that could be described adequately by Michaelis-Menten kinetics. However, the concentration of TEA at one-half Jmax (Kt) for efflux (approximately 110 microM) was about six times the Kt for uptake (approximately 18 muM), indicating that the affinity of the carrier for TEA is greater in the uptake direction than in the efflux direction or that there are separate carriers with different affinities for uptake and efflux. In either case, this difference would favor movement of TEA taken up at the basolateral side across the cells and into the lumen over movement back into the peritubular fluid.

Animals↗

PubMed1996

Compartmentation of amino acids in the rat kidney.

Amino acid concentrations ([AA]) were determined in cortical, outer and inner medullary (OM and IM), and papillary tissue of rat kidney (Cti, mmol/kg wet wt), in plasma (Cpl), and in urine. In all regions, Cti values were highest for Tau, Gly, and Glu-, making up 54-65% of the total [AA]:27, 21, and 11 mmol/kg wet wt in cortex, OM, and IM and papilla, respectively. Cortical cell water [AA] values (CcH2O, mmol/kgH2O) were between 12.4 (Tau) and 0.09 (Orn+), representing cell water-to-plasma water ratios (CcH2O/CpH2O) between 134 (Asp-) and 0.9 (Thr and Cit). Short-term water diuresis did not change the total tissue [AA] throughout the kidney. Treatment of the tissue with Triton X-100 instead of sulfosalicylic acid (SSA) resulted in much higher [AA], except for Glu-, Glu-NH2, Tau, and exogenous L-homoarginine+ (hoArg+). When hoArg+ was infused (leading to a Cpl = 5.9 mmol/l), Cti of hoArg+ was similar throughout the kidney (13-22 mmol/kg wet wt). In the presence of hoArg+, CcH2O/CpH2O of Arg+ rose 13-fold. We conclude that 1) AA contribute 20% to cytosolic osmolality in renal cortex, 2) total [AA] decreases from cortex to papilla, 3) cellular uptake of Tau and anionic AA must be rheogenic, whereas cationic AA (except for Arg+ in cortex) are passively distributed, and 4) AA do not seem to contribute quantitatively to short-term medullary osmotic adaptation during diuresis.

Absorption↗

PubMed1996

Effect of alpha-KG in lumen on PAH transport by isolated perfused rabbit renal proximal tubules.

To determine whether dicarboxylate taken up at the luminal membrane could function in the p-aminohippurate (PAH) countertransport at the basolateral membrane, we examined the effect of adding alpha-ketoglutarate (alpha-KG) or glutarate (a nonmetabolized dicarboxylate that is countertransported for PAH at the basolateral membrane) to the luminal perfusate on net secretion of radiolabeled PAH in isolated perfused S2 segments of rabbit proximal tubules. Addition of 100 microM alpha-KG or glutarate to the luminal perfusate in tubules perfused and bathed with HEPES-buffered medium (in the absence of bicarbonate, glycine, lactate, malate, and citrate) produced a reversible twofold stimulation of net PAH transepithelial secretion. Addition of 4 mM LiCl (an inhibitor of Na-dicarboxylate transport that does not directly affect PAH transport) to the luminal perfusate along with alpha-KG eliminated stimulation of net PAH secretion. Addition of 100 microM or 1 mM alpha-KG or glutarate to the luminal perfusate in tubules perfused and bathed with bicarbonate-buffered medium containing glycine, lactate, malate, and citrate had no effect on net PAH transport from bath to lumen. These data indicate that alpha-KG (or glutarate) that enters the tubule cells via the luminal Na-dicarboxylate cotransporter can stimulate net PAH secretion, apparently via countertransport at the basolateral membrane, but only when tubules are not in an optimal metabolic state to produce intracellular alpha-KG.

Animals↗

PubMed1995

Relation of membrane potential to basolateral TEA transport in isolated snake proximal renal tubules.

We measured the effects of changes in bath K+ concentration ([K+]) on basolateral membrane potential difference (PD) and [3H]tetraethylammonium (TEA) transport in isolated snake (Thamnophis) proximal renal tubules (25 degrees C; pH 7.4). Increasing bath [K+] from 3 to 65 mM decreased PD from -60 mV (inside of cells negative) to -20 mV and 2-min uptake of [3H]TEA by approximately 25%, indicating that PD influences TEA entry into the cells. Uptake of [3H]TEA was inhibited similarly at both K+ concentrations by unlabeled TEA, indicating that uptake is carrier mediated. Kt (approximately 18 microM) for 2-min uptake of [3H]TEA in 3 mM K+ increased significantly in 65 mM K+, suggesting that the decrease in PD or the increase in [K+] alters the affinity of the transporter for TEA. The steady-state cell-to-bath ratio for [3H]TEA with 3 mM K+ (-60 mV PD) was approximately 16, significantly above the ratio of 10 predicted for passive distribution at electrochemical equilibrium. With 65 mM K+ (-20 mV PD) this ratio decreased to approximately 6, again significantly above the predicted ratio of 2. These data suggest that the PD can account for much, but not all, of the steady-state uptake of TEA. Efflux of [3H]TEA across the basolateral membrane was identical with either 3 or 65 mM K+ in the bath but was almost completely inhibited in either case by tetrapentylammonium, a potent inhibitor of TEA uptake. These data indicate that virtually all TEA transport across the basolateral membrane is carrier mediated and that transport out of the cells is unaffected by PD.

Animals↗

PubMed1995

Intracellular pH in snake renal proximal tubules.

Intracellular pH (pHi) was studied in isolated proximal renal tubules of garter snakes (Thamnophis spp.) with oil-filled lumens under control conditions [N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered medium with pH 7.4 at 25 degrees C] and in response to NH4Cl pulse. pHi was measured with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5,6-carboxyfluorescein (BCECF). Control resting pHi (7.1) and acidification in response to NH4Cl pulse (minimum pHi, 6.6) were essentially the same in snake tubules with oil-filled lumens or perfused lumens and in rabbit S2 proximal tubules with oil-filled lumens. Rate of recovery of pHi (dpHi/dt) from acid to resting level in snake tubules (2.5 x 10(-3) pH U/s was about one-third of that in rabbit tubules. Resting pHi and dpHi/dt from acid to resting level were Na+ dependent in the distal portion but not the proximal portion of snake proximal tubules. However, dpHi/dt was not influenced by amiloride or 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid in snake proximal tubules, suggesting that the effect of Na+ on dpHi/dt and resting pHi may involve membrane potential. This study also indicates that oil-filled lumens do not interfere with measurements of resting pHi and do permit evaluation of pHi regulation at the basolateral membrane without complications from transport at the luminal membrane.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1995

Interaction of alpha-KG with basolateral organic anion transporter in isolated rabbit renal S3 proximal tubules.

To understand the basolateral p-aminohippurate (PAH) transporter in the S3 segment of rabbit proximal tubules and its relationship to the transporter in the S2 segment, we measured the 30-s uptake and efflux of PAH across the basolateral membrane of single isolated S3 segments at 37 degrees C in bicarbonate-buffered media. Kinetic analysis of uptake data revealed a concentration of PAH at one-half Jmax of approximately 107 microM (same as in the S2 segment) but a Jmax of 600 fmol.min-1.nl-1 (one-tenth that of S2 segment). The coefficient for efflux across the basolateral membrane was also only one-sixth to one-tenth of that in the S2 segment. These data suggest that the basolateral PAH transporter is the same in both segments but that there are fewer transporters in the S3 than in the S2 segment. However, the apparent inhibitor constant values for cis-inhibition by probenecid (approximately 29 microM in S3, approximately 15 microM in S2) and by alpha-ketoglutarate (alpha-KG) in the presence of LiCl (approximately 40 microM in S3, approximately 160 microM in S2) suggest that the transporters may not be identical in the two segments. In bicarbonate-buffered medium, preloading the tubules with 100 microM alpha-KG did not trans-stimulate PAH uptake across the basolateral membrane, whereas preloading with 1.0 microM alpha-KG caused a significant stimulation of 43%. However, in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered medium, preloading the tubules with 100 microM alpha-KG caused a twofold increase in PAH uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Peritubular paraquat transport in isolated renal proximal tubules.

To better understand the characteristics of peritubular transport of organic cations (OCs), the uptake of the polyvalent OC dimethylbipyridinium (paraquat) and the structurally similar monovalent OC 1-methyl-4-phenylpyridinium (MPP+) was measured in suspensions of rabbit renal proximal tubules. Compared to the uptake of MPP+, the uptake of paraquat across the peritubular membrane was a low affinity, low capacity carrier-mediated process with a Jmax of 0.52 +/- 0.19 nmol.mg of protein.-1 min-1 and a Km of 162 +/- 25 microM. The uptake of MPP+ was a carrier-mediated process with a measured Jmax and Km of 1.8 +/- 0.09 nmol.mg of protein.-1min-1 and 28 +/- 8 microM, respectively. To determine whether paraquat is a substrate for the monovalent OC pathway, the effect of unlabeled MPP+ and tetraethylammonium (TEA) on paraquat uptake was examined. A 1 mM concentration of the monovalent OC MPP+ and TEA reduced the uptake of [14C]paraquat and [3H]MPP+ by approximately 30 and 90%, respectively, whereas 1 mM paraquat had no effect on [3H]MPP+ or [14C]TEA uptake. Thus, MPP+, but not paraquat, appears to interact with the monovalent OC transporter. On the other hand, the polyvalent OC substrates, including the polyamines putrescine and spermine, the herbicide diquat and the divalent hexamethonium bromidehydrate had no effect on either paraquat or MPP+ uptake. However, the synthetic polyamine methylglyoxal bis(guanyl-hydrazone)dihydrochloride (MGBG; 1 mM) reduced both paraquat and MPP+ uptake (by 60 and 90%, respectively). The ability of MGBG, unlike the other polyvalent substrates, to interact with paraquat transport may be related to structural similarities in the relative location of the two charged nitronium moieties in paraquat and MGBG.(ABSTRACT TRUNCATED AT 250 WORDS)

1-Methyl-4-phenylpyridinium↗