Organic anion-transporting polypeptide 1a4-mediated heterogeneous distribution of sulforhodamine-101 in rat hepatic lobules.

It has been known that organic anion-transporting polypeptides (Oatps) involve hepatic transports several organic anionic compounds and drugs. This study aimed to investigate sulforhodamine-101 (SR-101) distribution in the rat liver, determine the molecules responsible for the distribution, and delineate the manner of distribution. After intravenous SR-101 administration, its distribution in frozen rat hepatic sections was examined. SR-101-derived signals were detected in regions around the hepatic central vein (CV), where immunohistochemistry (IHC) indicated high Oatp1a4 expression. The signals decreased with treatment by digoxin, a specific substrate for Oatp1a4. In vitro studies using isolated rat hepatocytes and rat Oatp1a4-expressing Xenopus laevis oocytes have suggested that SR-101 is an Oatp1a4 substrate and is taken up into rat hepatocytes mainly via Oatp1a4. Therefore, results suggested SR-101 zonation because of Oatp1a4 involvement and that Oatp1a4 function is dominant in the region around the hepatic CV in rat hepatic lobules.

Involvement of carrier-mediated transport in the retinal uptake of clonidine at the inner blood-retinal barrier.

In the present study, the blood-to-retina transport across the inner BRB was investigated for clonidine, a compound which is expected to exhibit a neuroprotective effect for the treatment of severe retinal diseases. In the in vivo study, the integration plot analysis for [(3)H]clonidine exhibited an apparent influx permeability clearance of 457 µL/(min·g retina) in the retina. The in vivo inhibition study suggests that the blood-to-retina transport of clonidine at the BRB is organic cation-sensitive since clonidine, pyrilamine, and propranolol, at a concentration of 40 mM, significantly reduced the retinal uptake index (RUI) of [(3)H]clonidine, and an inhibitory effect on the RUI was also exhibited by verapamil, at a concentration of 3 mM. The in vitro study with TR-iBRB2 cells, an in vitro model cell line of the inner BRB, suggests that carrier-mediated transport is involved in the blood-to-retina transport of clonidine at the inner BRB since the results obtained demonstrated time-, temperature-, pH-, and concentration-dependent [(3)H]clonidine uptake, with a Km of 286 µM. In the in vitro inhibition study, the [(3)H]clonidine uptake was significantly reduced by several organic cations, such as clonidine, verapamil, pyrilamine, and propranolol, and was competitively inhibited by 200 µM verapamil, in spite of slight or no significant alteration being produced with organic anions. Furthermore, the typical substrates and inhibitors of well-known organic cation transporters had no significant effect on the uptake of [(3)H]clonidine to suggest the involvement of novel transporter molecules in the transport of clonidine across the inner BRB. These results suggest that the blood-to-retina transport of clonidine across the inner BRB involves a carrier-mediated transport manner, suggesting the contribution of a novel organic cation transporter expressed by the retinal capillary endothelial cells.

Involvement of the carrier-mediated process in the retina-to-blood transport of spermine at the inner blood-retinal barrier.

The elimination of spermine, the end product of cellular polyamine, from the retina to the blood across the blood-retinal barrier (BRB) was investigated. The in vivo microdialysis study revealed that the elimination of [(3)H]spermine from vitreous humor after vitreous bolus injection was in a biexponential manner. The rate constant for the elimination of [(3)H]spermine during the terminal phase was estimated to be 1.67-fold greater than that of [(14)C]d-mannitol, a bulk flow marker, and the difference in the terminal elimination rate constant between [(3)H]spermine and [(14)C]d-mannitol was reduced in the presence of 50 mM spermine, suggesting a retina-to-blood transport system for [(3)H]spermine across the BRB. The retina-to-blood transport of [(3)H]spermine was also supported by a study of the retinal uptake index (RUI). The in vitro transport study with TR-iBRB2 cells, a model cell line of the inner BRB, revealed time-, concentration- and temperature-dependent transport of [(3)H]spermine, suggesting the involvement of carrier-mediated processes in spermine transport across the inner BRB. The in vitro study also suggested that the transport of spermine at the inner BRB is pH-, membrane potential- and Cl(-)-sensitive and Na(+)-insensitive, and these functional properties of spermine transport suggest only a minor contribution of spermine transporters, such as CCC9 (SLC12A8), the expression of which was suggested at the inner BRB. In the inhibition study, [(3)H]spermine transport was markedly inhibited by putrescine, spermidine, spermine and agmatine while substrates of a well-characterized organic cation transporter (OCTs/SLC22A) and a cationic amino acid transporter (CATs/SLC7A) had no effect, suggesting the involvement of unknown transporters in spermine elimination from the retina.