Functional Characterization of Rat Brain-specific Organic Anion Transporter (Oatp14) at the Blood-Brain Barrier

Oatp14/blood-brain barrier-specific anion transporter 1 (Slc21a14) is a novel member of the organic anion transporting polypeptide (Oatp/OATP) family. Northern blot analysis revealed predominant expression of Oatp14 in the brain, and Western blot analysis revealed its expression in the brain capillary and choroid plexus. Immunohistochemical staining indicated that Oatp14 is expressed in the border of the brain capillary endothelial cells. When expressed in human embryonic kidney 293 cells, Oatp14 transports thyroxine (T4; prothyroid hormone) (Km = 0.18 μm), as well as amphipathic organic anions such as 17β estradiol-d-17β-glucuronide (Km = 10 μm), cerivastatin (Km = 1.3 μm), and troglitazone sulfate (Km = 0.76 μm). The uptake of triiodothyronine (T3), an active form produced from T4, was significantly greater in Oatp14-expressed cells than in vector-transfected cells, but the transport activity for T3 was ∼6-fold lower that for T4. The efflux of T4, preloaded into the cells, from Oatp14-expressed cells was more rapid than that from vector-transfected cells (0.032 versus 0.006 min–1). Therefore, Oatp14 can mediate a bidirectional transport of T4. Sulfobromophthalein, taurocholate, and estrone sulfate were potent inhibitors for Oatp14, whereas digoxin, p-aminohippurate, or leukotriene C4, or organic cations such as tetraetheylammonium or cimetidine had no effect. The expression levels of Oatp14 mRNA and protein were up- and down-regulated under hypo- and hyperthyroid conditions, respectively. Therefore, it may be speculated that Oatp14 plays a role in maintaining the concentration of T4 and, ultimately, T3 in the brain by transporting T4 from the circulating blood to the brain.

Brain capillary endothelial cells are characterized by tightly sealed cellular junctions (tight junctions) and the paucity of fenestra and pinocytotic vesicles, which prevent free exchange between brain and blood (1,2). Therefore, the uptake of nutrients by the brain occurs through the brain capillary endothelial cells via specific transport systems (3)(4)(5)(6)(7). Metabolic enzymes and efflux transporters expressed in the brain capillaries facilitate the elimination of endogenous wastes and xenobiotics from the brain, and restrict their brain accumulation (3)(4)(5)(6)(7). Because of these characteristics, the brain capillaries are referred to as the blood-brain barrier (BBB). 1 The organic anion transporting polypeptides (Oatps in rodents and OATPs in human) belong to the growing gene family of organic anion/prostaglandin transporters that can mediate sodium-independent membrane transport of numerous endogenous and xenobiotic amphipathic compounds (8,9). Fourteen members of the Oatp/OATP gene family have been identified in rodents and humans, and they are classified within the gene superfamily of solute carriers as the Slc21a/SLC21A gene family (Human Gene Nomenclature Committee DataBase) (8,9). Several members of the Oatp/OATP family have been identified in the brain (Oatp1-3 and moat1 in rodents and OATP-A in human) (10 -14). Especially, in the BBB, rat Oatp2 and human OATP-A have been shown to be expressed in the plasma membrane of the brain capillary endothelial cells (15,16). Involvement of rat Oatp2 in the uptake and efflux transport of its substrates was investigated in vivo (17,18). The uptake of [D-penicillamine 2,5 ]-enkephalin (DPDPE) from the blood to the brain was determined by the brain perfusion technique in the presence and absence of Oatp2 inhibitors (17). The brain uptake of DPDPE was increased in Mdr1a (P-glycoprotein) gene knockout mice, and the uptake in Mdr1a knockout mice was inhibited by the substrates and inhibitors of rat Oatp2 such as digoxin and 17␤ estradiol-D-17␤-glucuronide (E 2 17␤G). Vice versa, when E 2 17␤G was microinjected into the cerebral cortex, the subsequent elimination of E 2 17␤G from the brain was carrier-mediated (18), and the elimination of E 2 17␤G was completely inhibited by co-administration of taurocholate and probenecid, whereas digoxin had only a partial effect (18). Partial inhibition by digoxin suggested that additional efflux transport system(s) for E 2 17␤G, which is taurocholate-and probenecidsensitive, is involved in the brain capillary.
Li et al. (19) recently identified BBB-specific anion trans-porter 1 (BSAT1) using gene microarray techniques by comparing the gene expression profile of cDNA from the brain capillary with that from the liver and kidney. BSAT1 cDNA consisted of 2148 bp that encoded a 716-amino acid residues protein with 12 putative membrane-spanning domains. BSAT1 was highly enriched in the brain capillary compared with brain homogenate, liver, and kidney. Comparison of the cDNA sequences of BSAT1 revealed that it is the 14th member of the Oatp/OATP family (Oatp14). Although the localization at the BBB and the substrates of this isoform remain unknown, BBBspecific expression prompted us to hypothesize that Oatp14 accounts for the efflux of organic anions including E 2 17␤G via the BBB, together with Oatp2. The purpose of the present study is to characterize the substrate specificity and spectrum of inhibitors of Oatp14, as well as its tissue distribution and localization. Through this study, we found out that thyroxine (T 4 ) is a good substrate of Oatp14, and the expression level of Oatp14 in the BBB is affected by plasma thyroid conditions. The results of the present study suggest that Oatp14 plays an important role in regulating the concentration of T 4 in the central nervous system and in brain development. Unlabeled pravastatin, troglitazone, and its conjugated metabolites were kindly donated from Sankyo, unlabeled cerivastatin was from Bayer AG, and unlabeled E3040 glucuronide and E3040 sulfate were from Eisai. All other chemicals were commercially available, of reagent grade, and were used without any purification.
Capillary Isolation-Rat brain capillaries were isolated using a modification of the procedure of Boado et al. (22). All steps in the isolation procedure were carried out at 4°C in pregassed (95% O 2 -5% CO 2 ) solutions. Briefly, pieces of gray matter were gently homogenized in three volumes (v/w) of an artificial extracellular fluid buffer and, after addition of dextran (final concentration 15%), the homogenate was centrifuged at low speed. The resulting pellet was resuspended in Buffer B (103 mM NaCl, 25 mM NaHCO 3 , 10 mM D-glucose, 4.7 mM KCl, 2.5 mM CaCl 2 , 1.2 mM MgSO 4 , 1.2 mM K 2 HPO 4 , and 15 mM HEPES, 1 mM sodium pyruvate, 0.5% (w/v) bovine serum albumin, pH 7.4) and then filtered through a 200-m nylon mesh. The filtrate was passed over a column of glass beads, and after washing with Buffer B, the capillaries adhering to the beads were collected by gentle agitation.
Northern Blot Analysis-A commercially available hybridization blot containing poly(A) ϩ RNA from various rat tissues (rat multi-tissue Northern blot; Clontech) was used for the Northern blot analysis. A fragment (position numbers 1-807) from Oatp14 was used as a probe, and its nucleotide sequence showed less than 60% identity with other members of the Oatp family. The master blot filter was hybridized with the 32 P-labeled probe at 68°C according to manufacturer's instructions. The filter was washed finally under high stringency conditions (0.1ϫ SSC (1ϫ SSC ϭ 0.15 M NaCl and 0.015 M sodium citrate)) and 0.1% SDS at 65°C and then exposed to Fuji imaging plates (Fuji Photo Film, Kanagawa, Japan) for 3 h at room temperature and examined using an imaging analyzer (BAS 2000; Fuji Photo Film).
Western Blot Analysis-Antiserum against Oatp14 was raised in rabbits against a synthetic peptide consisting of the 17 carboxyl-terminal amino acids of Oatp14. Antiserum was purified by affinity column chromatography using the antigen and used for subsequent analyses. Choroid plexus, brain homogenate, and isolated brain capillary samples were diluted with Loading Buffer (BioLabs, Hertfordshire, United Kingdom). They were then boiled for 3 min and loaded onto an 8.5% SDS-polyacrylamide electrophoresis gel with a 3.75% stacking gel. Proteins were electroblotted onto a polyvinylidene difluoride membrane (Pall Filtran, Karlstein, Germany) using a blotter (Trans-blot; Bio-Rad) at 15 V for 1 h. The membrane was blocked with TBS-T (Tris-buffered  1 and 4, 50 g), brain homogenate (lanes 2 and 5, 50 g), and isolated brain capillary (lanes 3 and 6, 50 g) were separated by SDS-PAGE (10% separating gel). Oatp14 was detected by anti-Oatp14 polyclonal antibody.
FIG. 2. Immunohistochemical staining of Oatp14 for brain slices. Frozen sections of rat brain were used for immunohistochemical detection with peroxidase to probe for Oatp14 with a polyclonal antibody. The lined and dotted arrows represent luminal and abluminal sides of brain capillary endothelial cells, respectively. Positive labeling was only found in the border of brain capillary endothelial cells. saline containing 0.05% Tween 20) and 5% skimmed milk for 1 h at room temperature. After washing with TBS-T, the membrane was incubated with the antibodies (dilution 1:1000). The membrane was allowed to bind a horseradish peroxidase-labeled anti-rabbit IgG antibody (Amersham Biosciences) diluted 1:5000 in TBS-T for 1 h at room temperature followed by washing with TBS-T.
Immunohistochemical Study-Frozen sections from male Sprague-Dawley rats were prepared for the immunohistochemical study after fixing in acetone (Ϫ20°C). The brain slices adhered to the glass cover slips were washed with PBS and fixed for 10 min on ice in acetone. After washing with PBS, the capillaries were permearized in 0.2% (v/v) Triton X-100 in PBS and incubated with peroxidase blocking reagent (DAKO, Carpinteria, CA) for 10 min at room temperature to block nonspecific peroxidase. Slices were incubated with anti-Oatp14 antibody (1:100) for 60 min at room temperature, washed three times with PBS, and subsequently incubated for 60 min at room temperature with the horseradish peroxidase-labeled anti-rabbit secondary antibody (Envisionϩ system; DAKO). The immune reaction was visualized using diaminobenzidine and then nuclei were stained with hematoxylin (DAKO). The specificity of the antibody reaction was verified by negative controls, which were incubated with polyclonal antibody that had been blocked with the antigenic peptide.
Cloning of Rat Oatp14 cDNA-Based on the nucleotide sequence reported by Li et al. (19) (GenBank TM accession number NM 053441), the following primers were designed to isolate Oatp14 cDNA encoding a full open reading frame of Oatp14: forward primer, 5Ј-ggaattccgccaccatggacacttcatccaaaga-3Ј and reverse primer, 5Ј-ggattccttaaagtcgggtctccttgc-3Ј. PCR was performed using cDNA prepared from rat brain as template according to the following protocol: 96°C for 1 min, 55°C for 1 min, and 72°C for 2 min; 50 cycles. PCR products were subcloned to pGEM-T Easy Vector (Promega, Madison, WI) and sequenced. The nucleotide sequence of rat Oatp14 cDNA was identical as being that of BSAT1 except for one base change (A175G) resulting in a change of amino acid (T59A). However, it was confirmed that this change was not because of an error accumulated during PCR by sequencing the RT-PCR products directly.
Stable Expression of Oatp14 cDNA in HEK293 Cells-The Oatp14-cDNA was subcloned into the pcDNA3.1(ϩ) (Invitrogen) and introduced into HEK293 cells by lipofection with FuGENE 6 (Roche Diagnostics) according to the manufacturer's protocol and were selected by culturing them in the presence of G418 sulfate (800 g/ml; Invitrogen). HEK293 cells were grown in minimum essential medium (Invitrogen) supplemented with 10% fetal bovine serum, penicillin (100 units/ ml), streptomycin (100 g/ml), and G418 sulfate (400 g/ml) at 37°C with 5% CO 2 and 95% humidity. Cells were incubated for 24 h before starting the experiments with culture medium supplemented with sodium butyrate (5 mM).
Transport Study-Uptake was initiated by adding the radiolabeled ligands to the incubating buffer in the presence and absence of inhibitors after cells had been washed three times and preincubated with Krebs-Henseleit buffer (142 mM NaCl, 23.8 mM NaHCO 3 , 4.83 mM KCl, 0.96 mM KH 2 PO 4 , 1.20 mM MgSO 4 , 12.5 mM HEPES, 5 mM glucose, and 1.53 mM CaCl 2 , adjusted to pH 7.4) at 37°C for 15 min. For the efflux study, cells were preincubated with [ 125 I]T 4 at 37°C for 15 min and washed three times with ice-cold Krebs-Henseleit buffer, followed by incubation in the absence of [ 125 I]T 4 with Krebs-Henseleit buffer at 37°C. The uptake and efflux were terminated at designed times by adding ice-cold Krebs-Henseleit buffer. The radioactivity associated with the cells and medium specimens was determined in a liquid scintillation counter. The remaining aliquots of cell lysates were used to determine protein concentrations by the method of Lowry (23) with bovine serum albumin as a standard. Ligand uptake is given as the cell-to-medium concentration ratio determined as the amount of ligand associated with the cells divided by the medium concentration. Specific uptake was obtained by subtracting the uptake by vector-transfected cells from that by Oatp14-expressed cells.
Kinetic Analyses-Kinetic parameters were obtained from the follow- where v is the uptake rate of the substrate (pmol/min/mg protein), S is the substrate concentration in the medium (M), K m is the Michaelis-Menten constant (M), and V max is the maximum uptake rate (pmol/min/mg protein). To obtain the kinetic parameters, the equation was fitted to the initial uptake velocity. The experimental data were fitted to the equation by nonlinear regression analysis with weighting as the reciprocal of the observed values, and the Damping Gauss Newton Method algorithm was used for fitting. Inhibition constants (K i ) for Oatp14-mediated transport were calculated assuming competitive inhibition.
Production of Hyperthyroid and Hypothyroid Conditions-Male Sprague-Dawley rats, weighing 200 -220 g, were purchased from Japan SLC (Shizuoka, Japan). Rats had free access to food and water at all times during the study. Production of hyperthyroid and hypothyroid conditions involved a modification of the procedure of Burmeister et al. (24). Hypothyroidism was induced by the addition of 0.05% methimazole (MMI), an inhibitor for thyroid hormone synthesis in the thyroid gland, to the drinking water or thyroidectomy. Hypothyroidism was assessed clinically by failure to gain weight at the expected rate and could be observed within 2 weeks of the beginning MMI treatment and within 1 week after thyroidectomy. Hyperthyrodism was produced by giving L-T3 (50 g/100 g body weight, subcutaneously, daily) 4 days before capillary isolation.

RESULTS
Tissue Distribution of Oatp14 -The expression of Oatp14 mRNA in rat tissues was investigated by Northern blot analysis (Fig. 1A). A band was detected at 2.6 kbp, predominantly in the brain. No hybridization signals were detected in mRNA isolated from other tissues, including the heart, spleen, lung, liver, skeletal muscle, kidney, and testis.
Immunoblot and Immunohistochemical Staining of Oatp14 -The expression of Oatp14 in the choroid plexus, brain homog-enate, and brain capillary was examined by Western blot analysis (Fig. 1B). Immunoreactive protein was detected at ϳ90 kDa in the choroid plexus, brain homogenate, and brain capillary. These bands were abolished when preabsorbed polyclonal antibody for Oatp14 was used, suggesting that the positive bands were specific for the antigen peptide.
To investigate the localization of Oatp14 in brain capillary endothelial cells, immunohistochemical staining was carried out using anti-Oatp14 polyclonal antibody (Fig. 2). Positive signals for anti-Oatp14 polyclonal antibody were detected in brain capillary endothelial cells. The signals were detected along the plasma membrane of brain capillary endothelial cells. The signal was abolished by preincubating the polyclonal antibody of Oatp14 with antigen (data not shown).
Transport Properties of Oatp14 - Fig. 3 shows the time profiles of the uptake of

(D) by Oatp14-expressed HEK293 cells and vector-transfected HEK293 cells. Their uptake by
Oatp14-expressed cells is markedly greater than that by vector-transfected cells. This Oatp14-mediated uptake showed saturation kinetics and followed the Michaelis-Menten equation (Fig. 3, E-H). The kinetic parameters for the uptake by Oatp14 were determined by nonlinear regression analysis and summarized in Table I. The uptake of various organic anions by Oatp14 was investigated, and the results are summarized in Table II (Table II). Although the triiodothyronine (T 3 ) uptake by Oatp14-expressed cells was significantly greater than that by vector-transfected cells, the Oatp14-mediated uptake for T 3 was ϳ6-fold smaller than that of T 4 and reverse T 3 by Oatp14 (Table II) (Table II).
To investigate whether Oatp14 can mediate bidirectional transport, cells were preloaded with [ 125 I]T 4 for 15 min followed by incubation in the absence of [ 125 I]T 4 . The radioactivity associated with cell specimens was rapidly reduced in Oatp14expressed HEK293 cells compared with that in vector-trans- fected cells, and the elimination rate constants were 0.032 Ϯ 0.002 and 0.006 Ϯ 0.001 min Ϫ1 , respectively (Fig. 4).
Effects of Hyperthyroid and Hypothyroid Conditions on the Expression of Oatp14 in the Brain Capillary-The effects of hyper-and hypothyroid conditions on the expression of Oatp14 in the brain capillary were investigated by RT-PCR and Western blotting (Fig. 7, A and B). RT-PCR and Western blotting analyses revealed that the expression levels of Oatp14 mRNA and protein were up-and down-regulated under hypothyroid and hyperthyroid conditions, respectively.

DISCUSSION
In the present study, we reported the substrate specificity of Oatp14, as well as its tissue distribution and localization in the brain. Oatp14 is expressed in the brain capillary and choroid plexus. It mediated the uptake of T 3 , T 4 , and reverse T 3 , as well as organic anions such as E 2 17␤G, cerivastatin, and TRO-S, suggesting its involvement in the membrane transport of these ligands in the brain capillary.
T 3 and its prohormone, T 4 , are produced in the thyroid gland and released into the blood. T 3 plays an essential role in brain development via binding to specific nuclear receptors (thyroid hormone receptor) (25). Deficiency of thyroid hormones particularly during fetal and neonatal period in the brain causes mental retardation and cretinism (26,27). T 3 is supplied to the brain and peripheral tissues as T 4 from which T 3 is enzymatically produced by type 2 iodothyronine deiodinase (D2) (25). Therefore, the brain uptake process of T 4 from the circulating blood is the first step in all subsequent reactions of thyroid hormone in the brain. Whether there is a specific transport mechanism(s) for T 4 in brain capillary endothelial cells remains controversial. The brain uptake of T 4 was saturable in dogs (28) but not in mice (29). Analysis of the transport and molecular properties of Oatp14 should help us resolve this.
Transfection of Oatp14 cDNA into HEK293 cells resulted in a marked increase in the uptake of T 4 , as well as reverse T 3 , an inactive metabolite of T 4 produced by type 3 iodothyronine deiodinase. Although the uptake of T 3 by Oatp14-expressed cells was significantly greater than that by vector-transfected cells (Table II), T 3 was extensively taken up by vector-transfected cells (Table II). Whether the uptake in vector-transfected cells is ascribed to specific transport system(s) for T 3 or passive diffusion remains unknown. The transport activity for T 3 exhibited by Oatp14, obtained by subtraction of the uptake by vector-transfected cells from that by Oatp14-expressed cells, was ϳ6-fold lower than that for T 4 and reverse T 3 by Oatp14 (Table II), although the chemical structures of T 3 and reverse T 3 are quite similar. The K i value of T 3 for Oatp14 was 25-fold greater than that of T 4 (Table III), and this may result in apparent low transport activity for T 3 by Oatp14. Oatp14 can also mediate bidirectional transport, because the efflux of T 4 is facilitated in Oatp14-expressed cells (Fig. 4), and it is possible that Oatp14 is involved in both the uptake and efflux of its ligands through the brain capillary (i.e. BBB).
Involvement of Oatp14 in maintaining homeostasis of T 4 in the brain was also supported by the change in expression levels of Oatp14 in the brain capillary under hypo-and hyperthyroid conditions (Fig. 7). The expression of Oatp14 in the brain capillary changed as if Oatp14 was responsible for maintaining the concentration of T 4 in the central nervous system: up-and down-regulated under hypothyroid and hyperthyroid conditions, respectively (Fig. 7). This pattern is similar to that observed in D2 expression (25). Increased D2 expression increases the conversion of T 4 to T 3 to compensate for the decrease in the local brain concentration of T 4 and vice versa. Therefore, we hypothesize that Oatp14 is involved in the uptake of T 4 through the brain capillaries.
In addition to Oatp14, Oatp2, the other isoform of rat Oatp family, is also the candidate transporter for T 3 and T 4 uptake by the brain from the circulating blood in rodents. The uptake of both T 3 and T 4 was significantly increased in Oatp2-cRNA injected oocytes with similar K m values (ϳ5-7 M) (12). Oatp2 has been identified both in the luminal and abluminal mem-brane of brain capillary endothelial cells (15). It is possible that Oatp14 and Oatp2 serve high and low affinity sites for T 4 in the brain capillary, because the K m values of Oatp2 were ϳ30-fold greater than that of Oatp14 (12). Following uptake from the circulating blood into endothelial cells, T 4 has to cross the abluminal membrane to reach the brain interstitial space and brain parenchymal cells. Whether this process is carrier-mediated remains unknown. Bidirectional nature of Oatp2-mediated transport has been reported in Oatp2-cRNA-injected oocytes (30). Oatp14 and Oatp2 are candidate transporters involved in the abluminal secretion of thyroid hormones. Further studies are necessary to identify the exact localization of Oatp14 in the brain capillary and to evaluate its contribution to the total brain uptake of T 4 into the brain. Pardridge et al. (31) demonstrated that the brain uptake of T 3 was saturable and inhibited by T 4 using carotid arterial bolus injection technique of Oldendorf, and Oatp2 may account for the brain uptake of T 3 in the brain capillary.
Oatp14 was detected in the choroid plexus by Western blot analysis (Fig. 1). The choroid plexus is located in the lateral, third and fourth ventricles, and the interface between the cerebrospinal fluid and the circulating blood acting as a barrier to protect the central nervous system, in conjunction with the BBB (32,33). The brain distribution of T 3 and T 4 after intracerebroventricular administration is limited to ependymal cells and circumventricular organs and, thus, transport via the choroid plexus could account for the brain distribution near the ventricles (34). As speculated in the case of brain capillary endothelial cells, it is possible that Oatp14 acts as an uptake system to supply T 4 to ependymal cells and circumventricular organs in the choroid plexus.
In addition to thyroids, Oatp14 accepts certain types of amphipathic organic anions, such as E 2 17␤G, cerivastatin, and TRO-S, as substrates although their transport activity was markedly lower than that of T 4 , except TRO-S (see Fig. 3 and Table II). Because Oatp14 can mediate the bidirectional trans-  7. Effect of hyperthyroid and hypothyroid conditions on the expression of Oatp14 on the BBB. A, RT-PCR analysis. mRNA samples were prepared from isolated brain capillary from control rats (lanes 1, 3, and 5), hypothyroid rats (MMI-treated or thyroidectomized rats; lanes 2 and 4, respectively) and hyperthyroid rats (T 3 -treated rats; lane 6). PCR products stained with ethidium bromide were visualized under UV light. G3PDH, glucose-3-phosphate dehydrogenase. B, Western blotting. Brain capillaries (50 g/lane) isolated from control rats (lanes 1, 3, and 5), hypothyroid rats (MMI-treated or thyroidectomized rats; lanes 2 and 4, respectively), and hyperthyroid rats (T 3treated rats; lane 6) were separated by SDS-PAGE (10% separating gel). Oatp14 was detected by anti-Oatp14 polyclonal antibody. port, it is possible that Oatp14 is involved in the efflux of organic anions such as E 2 17␤G from the brain when it is microinjected into the cerebral cortex and possibly in the efflux of excess T 4 and reverse T 3 from the brain. The spectrum of inhibitors of Oatp14 was consistent with the transporter hypothesized based on in vivo studies (18), but further investigations will be required to confirm this speculation.
Whether the results obtained using cDNA from rodents can be applied to the human situation is an important issue. Human OATP-F, an isoform in which Oatp14 exhibits high homology (84% in amino acid level), has a similar substrate specificity to Oatp14 (35). Northern blot analysis demonstrated abundant expression of OATP-F in the brain and testis and, to a lesser extent, heart, but the localization in the brain remains unidentified. In terms of substrate specificity and homology, OATP-F is supposed to be the human ortholog of Oatp14, and it may be suggested that OATP-F is also involved in the uptake of T 4 from the circulating blood into the central nervous system though the brain capillary and choroid plexus. In view of the importance of supplying T 4 to the brain during development, it is possible that functional loss of the OATP-F gene may be associated with a thyroid hormone-related neuronal disorder characterized by resistance to thyroid hormone treatment.
In conclusion, we have characterized Oatp14 in terms of its substrate specificity and localization in the brain and demonstrated that Oatp14 accepts T 4 , as well as organic anions, including certain glucuronide and sulfate conjugates. Oatp14 is localized on the plasma membrane of brain capillary endothelial cells and involved in the uptake of T 4 from the blood to the central nervous system. Oatp14 is one of the mechanisms for maintaining homeostasis of T 4 and, ultimately, T 3 in the brain.