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Tubular secretion of the organic cation, creatinine, limits its value as a marker of glomerular filtration rate (GFR) but the molecular determinants of this pathway are unclear. The organic anion transporters, OAT1 and OAT3, are expressed on the basolateral membrane of the proximal tubule and transport organic anions but also neutral compounds and cations. Here, we demonstrate specific uptake of creatinine into mouse mOat1- and mOat3-microinjected Xenopus laevis oocytes at a concentration of 10 μM (i.e., similar to physiological plasma levels), which was inhibited by both probenecid and cimetidine, prototypical competitive inhibitors of organic anion and cation transporters, respectively. Renal creatinine clearance was consistently greater than inulin clearance (as a measure of GFR) in wild-type (WT) mice but not in mice lacking OAT1 (Oat1-/-) and OAT3 (Oat3-/-). WT mice presented renal creatinine net secretion (0.23 ± 0.03 μg/min) which represented 45 ± 6% of total renal creatinine excretion. Mean values for renal creatinine net secretion and renal creatinine secretion fraction were not different from zero in Oat1-/- (-0.03 ± 0.10 μg/min; -3 ± 18%) and Oat3-/- (0.01 ± 0.06 μg/min; -6 ± 19%), with greater variability in Oat1-/-. Expression of OAT3 protein in the renal membranes of Oat1-/- mice was reduced to ~6% of WT levels, and that of OAT1 in Oat3-/- mice to ~60%, possibly as a consequence of the genes for Oat1 and Oat3 having adjacent chromosomal locations. Plasma creatinine concentrations of Oat3-/- were elevated in clearance studies under anesthesia but not following brief isoflurane anesthesia, indicating that the former condition enhanced the quantitative contribution of OAT3 for renal creatinine secretion. The results are consistent with a contribution of OAT3 and possibly OAT1 to renal creatinine secretion in mice. [ABSTRACT FROM AUTHOR]

Acute renal injury induces metabolic acidosis, but its specific effects on the collecting duct, the primary site for urinary ammonia secretion, the primary component of net acid excretion, are incompletely understood. We induced ischemia-reperfusion (I/R) acute renal injury in Sprague-Dawley rats by clamping the renal pedicles bilaterally for 30 mm followed by reperfusion for 6 h. Control rats underwent sham surgery without renal pedicle clamping. I/R injury decreased urinary ammonia excretion significantly but did not persistently alter urine volume, Na+, K+ or bicarbonate excretion. Histological examination demonstrated cellular damage in the outer and inner medullary collecting duct, as well as in the proximal tubule and the thick ascending limb of the loop of Henle. A subset of collecting duct cells were damaged and/or detached from the basement membrane; these cells were present predominantly in the outer medulla and were less frequent in the inner medulla. Immunohistochemistry identified that the damaged/detached cells were A-type intercalated cells, not principal cells. Both TdT-mediated dUTP nick-end labeling (TUNEL) staining and transmission electron microscopic examination demonstrated apoptosis but not necrosis. However, immunoreactivity for caspase-3 was observed in the proximal tubule, but not in collecting duct intercalated cells, suggesting that mechanism(s) of collecting duct intercalated cell apoptosis differ from those operative in the proximal tubule. We conclude that I/R injury decreases renal ammonia excretion and is associated with intercalated cell-specific detachment and apoptosis in the outer and inner medullary collecting duct. These effects likely contribute to the metabolic acidosis frequently observed in acute renal injury. [ABSTRACT FROM AUTHOR]

The flounder renal organic anion transporter (fOat) has substantial sequence homology to mammalian basolateral organic anion transporter orthologs (OAT1/Oat1 and OAT3/Oat3), suggesting that fOat may have functional properties of both mammalian forms. We therefore compared uptake of various substrates by rat Oat1 and Oat3 and human OAT1 and OAT3 with the fOat clone expressed in Xenopus oocytes. These data confirm that estrone sulfate is an excellent substrate for mammalian OAT3/Oat3 transporters but not for OAT1/Oat1 transporters. In contrast, 2,4-dichlorophenoxyacetic acid and adefovir are better transported by mammalian OAT1/Oat1 than by the OAT3/Oat3 clones. All three substrates were well transported by fOat-expressing Xenopus oocytes. fOat Km values were comparable to those obtained for mammalian OAT/Oat1/3 clones. We also characterized the ability of these substrates to inhibit uptake of the fluorescent substrate fluorescein in intact teleost proximal tubules isolated from the winter flounder (Pseudopleuronectes americanus) and killifish (Fundulus heteroclitus). The rank order of the IC50 values for inhibition of cellular fluorescein accumulation was similar to that for the Km values obtained in fOat-expressing oocytes, suggesting that fOat may be the primary teleost renal basolateral Oat. Assessment of the zebrafish (Danio rerio) genome indicated the presence of a single Oat (zfOat) with similarity to both mammalian OAT1/Oat1 and OAT3/Oat3. The puffer fish (Takifugu rubripes) also has an Oat (pfOat) similar to mammalian OAT1/Oat1 and OAT3/Oat3 members. Furthermore, phylogenetic analyses argue that the teleost Oat1/3-like genes diverged from a common ancestral gene in advance of the divergence of the mammalian OAT1/Oat1, OAT3/Oat3, and, possibly, Oat6 genes. [ABSTRACT FROM AUTHOR]