• The Korean Journal of Physiology and Pharmacology
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• 제1권4호
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• pp.403-411
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• 1997

To elucidate the mechanism of gentamicin induced renal dysfunction, renal functions and activities of various proximal tubular transport systems were studied in gentamicin-treated rats (Fisher 344). Gentamicin nephrotoxicity was induced by injecting gentamicin sulfate subcutaneously at a dose of 100 $mg/kg{\cdot}day$ for 7 days. The gentamicin injection resulted in a marked polyuria, hyposthenuria, proteinuria, glycosuria, aminoaciduria, phosphaturia, natriuresis, and kaliuresis, characteristics of aminoglycoside nephropathy. Such renal functional changes occurred in the face of reduced GFR, thus tubular transport functions appeared to be impaired. The polyuria and hyposthenuria were partly associated with a mild osmotic diuresis, but mostly attributed to a reduction in free water reabsorption. In renal cortical brush-border membrane vesicles isolated from gentamicin-treated rats, the $Na^+$ gradient dependent transport of glucose, alanine, phosphate and succinate was significantly attenuated with no changes in $Na^+-independent$ transport and the membrane permeability to $Na^+$. These results indicate that gentamicin treatment induces a defect in free water reabsorption in the distal nephron and impairs various $Na^+-cotransport$ systems in the proximal tubular brush-border membranes, leading to polyuria, hyposthenuria, and increased urinary excretion of $Na^+$ and other solutes.

• The Korean Journal of Physiology and Pharmacology
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• 제3권5호
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• pp.513-519
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• 1999

Direct exposure of renal tubular brush-border membranes (BBM) to free cadmium (Cd) causes a reduction in phosphate (Pi) transport capacity. Biochemical mechanism of this reduction was investigated in the present study. Renal proximal tubular brush-border membrane vesicles (BBMV) were isolated from rabbit kidney outer cortex by Mg precipitation method. Vesicles were exposed to $50{\sim}200\;{\mu}M\;CdCl_2$ for 30 min, then the phosphate transporter activity was determined. The range of Cd concentration employed in this study was comparable to that of the unbound Cd documented in renal cortical tissues of Cd-exposed animals at the time of onset of renal dysfunction. The rate of sodium-dependent phosphate transport $(Na^+-Pi\;cotransport)$ by BBMV was determined by $^{32}P-Iabeled$ inorganic phosphate uptake, and the number of $Na^+-Pi$ cotransporters in the BBM was assessed by Pi-protectable $^{14}C-labeled$ phosphonoformic acid $([^{14}C]PFA)$ binding. The exposure of BBMV to Cd decreased the $Na^+-Pi$ cotransport activity in proportion to the Cd concentration in the preincubation medium, but it showed no apparent effect on the Pi-protectable PFA binding. These results indicate that an interaction of renal BBM with free Cd induces a reduction in $Na^+-Pi$ cotransport activity without altering the carrier density in the membrane. This, in turn, suggest that the suppression of phosphate transport capacity $(V_{max})$ observed in Cd-treated renal BBM is due to a reduction in $Na^+-Pi$ translocation by existing carriers, possibly by Cd-induced fall in membrane fluidity.

• 한국식품위생안전성학회지
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• 제8권4호
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• pp.195-204
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• 1993

Basolateral and brush border membrane (BLM and BBM) vesicles of renal proximal tubules were prepared from adult male New Zealand White rabbits to evaluate as a potential model for assessment of nephrotoxicity. PAH uptakes using BLMV, glucose and leucine uptakes using BBMV were measured in the rabbits treated cephaloridine. In addition, urinalysis and histopathological studies were performed to investigate the correlationship with membrane vesicle uptakes. The results were as follows: (1) the activite of Na+, K+ -ATPase was enriched 12.3-fold in vasolateral memvrane vesicles (BLMV) and the specific activity of alkaline phosphatase in purified brush border memvrane vesicles (BBMV) was enriched 10.1-fold compared with each of microsomal homogenate. (2) In the uptake experiments, cephaloridine decreased initial and probenecid-sensitive PAH uptakes in BLMV. (3) Cephaloridine tested decreased initial and phlorizin-sensitive glucose uptakes in BBMV. (4) Cephaloridine tested decreased initial and Na+-dependent leucine uptakes in BBMV. (5) Cephaloridine tested significantly increased the urinary excretion of glucose and activity of ${\gamma}$-GTP. (6) Cephaloridine tested caused moderate necrotic changes in renal tubular cells and formation of urinary cast in the lumina of Henle's loop and collecting tubules besides the swelling of renal tubules.

• The Korean Journal of Physiology
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• 제25권2호
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• pp.179-188
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• 1991

Effect of a sulfhydryl reagent, p-chloromercuribenzoic acid (PCMB), on the transport of succinate was studied in brush border (BBMV) and basolateral (BLMV) membrane vesicles isolated from rabbit renal cortex. PCMB induced an irreversible inhibition of the $Na^+-dependent$ succinate uptake in a dose-dependent manner with $IC_{50}$ of 55 and $65\;{\mu}M$ in BBMV and BLMV, respectively. The inhibitory effect of PCMB was prevented by a pretreatment of vesicles with dithiothreitol. PCMB did not increase $Na^+$ permeability at concentrations inhibiting succinate uptake. The PCMB inhibition of succinate uptake was due to a change in Vmax, but not in Km. When membrane vesicles were pretreated with PCMB in the presence of unlabelled succinate, the inhibitory effect was significantly reduced. In both BBMV and BLMV, succinate uptake was inhibited by various sulfhydryl reagents with the inhibitory potency of following order: $HgCl_2$>DTNB>PCMBS>PCMB. These results suggest that sulfhydryl groups are essential for dicarboxylate transport and that they may be located at or near substrate binding sites of the transporters in renal brush border and basolateral membranes.

• The Korean Journal of Physiology and Pharmacology
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• 제3권2호
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• pp.191-198
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• 1999

This study was undertaken to examine the effect of ethanol on $Na^+-dependent$ transport systems (glucose, phosphate, and dicarboxylate) in renal brush-border membrane vesicles (BBMV). Ethanol inhibited $Na^+-dependent$ uptakes of glucose, phosphate, and succinate in a dose-dependent manner, but not the uptakes of $Na^+-dependent.$ The $H^+/TEA$ antiport was reduced by 8% ethanol. Kinetic analysis showed that ethanol caused a decrease in $V_{max}$ of three transport systems, leaving $K_m$ values unchanged. Ethanol decreased phlorizin binding, which was closely correlated with the decrease in $V_{max}$ of $Na^+-glucose$ uptake. These results indicate that ethanol inhibits $Na^+-dependent$ uptakes of glucose, phosphate, and dicaboxylate and that the reduction in $V_{max}$ of $Na^+-glucose$ uptake is caused by a decrease in the number of active carrier proteins in the membrane.

• Toxicological Research
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• 제8권1호
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• pp.95-107
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• 1992

Basolateral and brush border membrance (BLM and BBM) vesicles of renal proximal tubules were prepared from adult male New Zealand White rabbits to develop an experimental for assessment of nephrotoxicity. PAH uptakes using BLMV, and glucose and leucine uptakes using BBMV were measured in the rabbits treated uranyl acetate. In addition, urinalysis and histopathological studies were performed to investigate the correlationship with the membrance vesicle uptakes.

• The Korean Journal of Physiology
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• 제21권2호
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• pp.273-282
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• 1987

가토 신피질에서 분리한 brush border membrane (BBM)과 basolateral membrane vesicle (BLM)에서 유기 음이온인 p-aminohippuric acid (PAH)와 유기 양이온인 tetraethylammonium (TEA)의 이동에 대한 동력학적 분석을 하였다. BLM에서 PAH에 대한 Km과 Vmax값은 각각 $0.34{\pm}0.02\;mM$과 $0.22{\pm}0.07\;nmol/mg\;protein/20s$였으며 BBM에서 각 값은 $8.46{\pm}0.57\;mM$과 $4.43{\pm}0.40\;nmol/mg\;protein/20s$였다. BLM에서 용액내 Na의 제거는 PAH에 대한 Km 값에는 영향없이 Vmax 값을 변화시켰다. BBM에서 TEA이동에 대한 Km과 Vmax 값은 각각 $0.55{\pm}0.15\;mM$과 $1.04{\pm}0.23\;nmol/mg\;protein/20s$였으며 BLM에서 각 값은 $0.46{\pm}0.04\;mM$과 $0.61{\pm}0.06\;nmol/mg\;protein/20s$였다. BLM에서 측정한 유기 이온들의 이동에 대한 Km 값이 신절편이나 분리된 tubule에서 보고된 값과 일치함을 보였으며 이러한 결과는 신세뇨관 세포막을 통한 유기 이온들의 이동 특성이 membrane vesicle을 분리하는 과정에서 변하지 않았음을 가르킨다.

• 한국영양학회:학술대회논문집
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• 한국영양학회 1995년도 추계학술대회 초록
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• pp.39-39
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• 1995

• Toxicological Research
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• 제11권1호
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• pp.103-108
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• 1995

Rat renal proximal tubule suspension was prepared from adult male Sprague Dawley rat (250-300g) by mechanical (non-enzymatical) method and evaluated as a pontential model for mechanistic studies and early screening of nephrotoxicity, using anionic antibiotics (cephaloridine). Cephaloridine (CPL) produced an increase in LDH release into media. This release results from decrease a proximal tubule cell viability and subsequently increase the permeability of cell viability and subsequently increase the permeability of cell membrane. Since loss of intracellular potassium and ATP into media is the sign of disruption of cell membrane, especially basolateral membrane (BLM), CPL induced proximal tubule cell compromise also appear be associated with BLM, maybe $Na^+-K^+$ ATPase. Also seen was significant depression in brush border membrane (BBM) ALP activity and no significantly increase in BBM GGT activities. The inhibition of typical anion, PAH accumulation (especially, CPL 5 mM) and cation, TEA (especially, 4hours incubation) were seen dose dependently. This is because of CPL accumulation in renal proximal tubule and increase of cytotoxicity.

• The Korean Journal of Physiology
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• 제30권2호
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• pp.249-256
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• 1996

The effect of probenecid on the transport of tetraethylammonium (TEA) was studied in renal cortical slices and isolated membrane vesicles to investigate the interaction of organic anion with the organic cation transport system in proximal tubule. Probenecid reversibly inhibited TEA uptake by renal cortical slices in a dose-dependent manner over the concentration range of 1 and 5 mM. The efflux of TEA was not affected by the presence of 3 mM probenecid. Kinetic analysis indicated that probenecid decreased Vmax without significant change in Km. Probenecid inhibited significantly tissue oxygen consumption at concentrations of 3 and 5 mM. However, probenecid did not significantly reduce TEA uptake in brush border and basolateral membrane vesicles prepared from renal cortex even at a concentration as high as 10 mM. These results indicate that probenecid reduces TEA uptake in cortical slices by inhibiting tissue metabolism rather than by an interaction with the organic ration transporter.