• The Korean Journal of Physiology and Pharmacology
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• v.4 no.4
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• pp.311-318
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• 2000

We cultured the rabbit inner medullary collecting duct (IMCD) cells as monolayers on collagen-coated membrane filters, and investigated distribution of the P2Y receptors by analyzing nucleotide-induced short circuit current $(I_{sc})$ responses. Exposure to different nucleotides of either the apical or basolateral surface of cell monolayers stimulated $I_{sc}.$ Dose-response relationship and cross-desensitization studies suggested that at least 3 distinct P2Y receptors are expressed asymmetrically on the apical and basolateral membranes. A $P2Y_2-like$ receptor, which responds to UTP and ATP, is expressed on both the apical and basolateral membranes. In addition, a uracil nucleotide receptor, which responds to UDP and UTP, but not ATP, is expressed predominantly on the apical membrane. In contrast, a $P2Y_1-like$ receptor, which responds to ADP and 2-methylthio-ATP, is expressed predominantly on the basolateral membrane. These nucleotides stimulated intracellular cAMP production with an asymmetrical profile, which was comparable to that in the stimulation of $I_{sc}.$ Our results suggest that the adenine and uracil nucleotides can interact with different P2Y nucleotide receptors that are expressed asymmetrically on the apical and basolateral membranes of the rabbit IMCD cells, and that both cAMP- and $Ca^{2+}-dependent$ signaling mechanisms underlie the stimulation of $I_{sc}$.

• The Korean Journal of Physiology
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• v.30 no.2
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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.

• The Korean Journal of Physiology
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• v.21 no.2
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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.

• Journal of Food Hygiene and Safety
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• v.8 no.4
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• pp.189-193
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• 1993

Cisplatin is useful for various cancers including advanced testicular and ovarian cancers. However, clinical use of cisplatin has been limited due to its dose-related neplrotoxicity. Transport studies across the membrane vesicles were performed to study the cisplatin nephrotoxicity. In these experiments, after cisplatin was administered to adult male New Zealand White rabbits, basolateral membrane (BLM) vesicles were prepared from the renal cortex. Para-aminohippurate (PAH) uptakes through BLM vesicles were measured to examine the interactioln of cisplatin on the transports of the substrates. As results of the uptake experiments using the vesicle systems, cisplatin had little effects on PAH transport through BLM vesicle. In conclusion, cisplatin did not cause the damage of basolateral membranes.

• The Korean Journal of Physiology
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• v.30 no.2
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• pp.279-288
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• 1996

Chronic exposure to cadmium impairs various renal tubular functions, including organic acid (anion) secretion. To investigate the mechanism of cadmium-induced alterations in the organic anion transport system, kinetics of p-aminohippurate (PAH) uptake was studied in renal cortical basolateral membrane vesicles (BLMV) isolated from cadmium-intoxicated rats (adult male Sprague-Dawley). Cadmium intoxication was induced by subcutaneous injections of $CdCl_{2}$ (2 mg Cd/kg per day) for 3 weeks. The renal plasma membrane vesicles were prepared by Percoll gradient centrifugation. The vesicular uptake of $^{14}C$-PAH was determined by rapid filtration technique using Millipore filter. Cadmium intoxication resulted in a marked attenuation of $Na^{+}$-dependent, ${\alpha}$-ketoglutarate (${\alpha}$KG)-driven PAH uptake with no changes in $Na^{+}$ and ${\alpha}$KG-independent transport component. Kinetic analysis indicated that Vmax, but not Km, of the $Na^{+}$-dependent, ${\alpha}$KG-driven component was reduced. A similar reduction of $Na^{+}$-dependent, ${\alpha}$KG-driven PAH uptake was observed in normal membrane vesicles directly exposed to inorganic cadmium in vitro, and this was accompanied by an inhibition of both $Na^{+}$-dependent ${\alpha}$KG uptake and ${\alpha}$KG-PAH exchange activity. These results indicate that during chronic exposure to cadmium, free cadmium ions liberated in the proximal tubular cytoplasm directly interact with the basolateral membrane and impair the active transport capacity for organic anions, most likely due to an inhibition of both $Na^{+}$-dicarboxylate cotransporter and dicarboxylate-organic anion antiporter activities.

• The Korean Journal of Physiology
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• v.25 no.2
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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.

• Toxicological Research
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• v.8 no.1
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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.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.3
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• pp.190-196
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• 2019

Tight junctions are constituents of the blood-epididymis barrier that play roles in regulating the unidirectional transcellular transport of ions, water, and solutes to maintain optimal conditions for sperm maturation and storage. Claudin 1 (Cldn1) and 4 (Cldn4) are known as tight junction proteins and are expressed in the basolateral membranes as well as tight junctions in the epididymis of rodents. Here, we examined the expression and localization of Cldn1 and 4 to determine the function of these proteins in the pig epididymis. Cldn1 was highly expressed in the basolateral membrane of epithelial cells in the caput and corpus regions of the epididymis. In the cauda region, however, Cldn1 labeling was significantly decreased in the basolateral membrane of epithelial cells. In contrast, labeling indicated that Cldn4 was expressed in the basolateral membrane in the cauda region of the epididymis and was present at punctate reactive sites in the caput and corpus regions. However, in no region of the epididymis did we detect colocalization of Cldn1 and 4 with labeled ZO-1, the distribution of which is restricted to the tight junctions. Our results indicate that Cldn1 and 4 were region-specifically expressed in the pig epididymis but not present in the tight junctions of epididymal epithelium. In addition, reciprocal regulation in specific regions of the epididymis between Cldn1 and 4 may play an important role in generating an optimal luminal environment for sperm maturation and storage in the pig epididymis.

• The Korean Journal of Physiology
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• v.22 no.2
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• pp.281-293
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• 1988

The effect of pH on the rate of PAH uptake was studied in rabbit renal basolateral membrane vesicles (BLMV) and brush border membrane vesicles (BBMV). In the absence of Na in incubation medium, a decrease in external $pH(pH_0)$ led to an increase in probenecid-sensitive PAH uptake by BLMV. In the presence of Na, the probenecid-sensitive PAH uptake was unaltered when the $pH_0$ decreased from 8.0 to 6.0 but further decrease in $pH_0$ to 5.5 increased significantly the uptake. The probenecid-sensitive PAH uptake was not affected by an alteration in pH per se in the absence of a pH gradient with or without the presence of Na. However, the presence of Na stimulated the probenecid-sensitive PAH uptake in all pH ranges tested over that measured in the absence of Na. A similar pattern of pH dependence on the PAH uptake was observed in BBMV but the presence of Na did not alter the probenecid-sensitive PAH uptake in the presence and absence of a pH gradient. Kinetic analysis for BLMV showed that Na or pH gradient increased Vmax of the probenecid-sensitive PAH uptake without a change in Km value. These results suggest that PAH is transported by $OH^-/PAH$ exchange process in the luminal membrane, but the pH dependence in the BLMV is not unequivocally consistent with an anion exchange process. The PAH transport is dependent on Na in BLMV but not in BBMV.

• The Korean Journal of Physiology
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• v.22 no.2
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• pp.307-318
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• 1988

Properties of succinate transport were examined in basolaterat membrane vesicles (BLMV) isolated from rabbit renal cortex. An inwardly directed $Na^+$ gradient stimulated succinate uptake and led to a transient overshoot. $K^+,{\;}Li^+,{\;}Rb^+$ and choline could not substitute for $Na^+$ in the uptake process. The dependence of the initial uptake rate of succinate on $Na^+$ concentration exhibited sigmoidal kinetics, indicating interaction of more than one $Na^+$ with transporter Hill coefficient for $Na^+$ was calculated to be 2.0. The $Na^+-dependent$ succinate uptake was electrogenic, resulting in the transfer of positive charge across the membrane. The succinate uptake into BLMV showed a pH optimum at external pH $7.5{\sim}8.0$, whereas succinate uptake into brush border membrane vesicles (BBMV) did not depend on external pH. Kinetic analysis showed that a Na-dependent succinate uptake in BLMV occurred via a single transport system, with an apparent Km of $15.5{\pm}0.94{\;}{\mu}M$ and Vmax of $16.22{\pm}0.25{\;}nmole/mg{\;}protein/min$. Succinate uptake was strongly inhibited by $4{\sim}5$ carbon dicarboxylates, whereas monocarboxylates and other organic anions showed a little or no effect. The succinate transport system preferred dicarboxylates in trans-configuration (furmarate) over cis-dicarboxylates (maleate). Succinate uptake was inhibited by the anion transport inhibitors DIDS, SITS and furosemide, and $Na^+-coupled$ transport inhibitor harmaline. These results indicate the existence of a $Na^+-dependent$ succinate transport system in BLMV that may be shared by the other Krebs cycle intemediates. This transport system seems to be very similar to the luminal transport system for dicarboxylates.