Neurotransmission in the hippocampus is modulated variously through presynaptic metabotropic glutamate receptors (mGluRs). To establish the precise localization of presynaptic mGluRs in the rat hippocampus, we used subtype-specific antibodies for eight mGluRs (mGluR1-mGluR8) for immunohistochemistry combined with lesioning of the three major hippocampal pathways: the perforant path, mossy fiber, and Schaffer collateral. Immunoreactivity for group II (mGluR2) and group III (mGluR4a, mGluR7a, mGluR7b, and mGluR8) mGluRs was predominantly localized to presynaptic elements, whereas that for group I mGluRs (mGluR1 and mGluR5) was localized to postsynaptic elements. The medial perforant path was strongly immunoreactive for mGluR2 and mGluR7a throughout the hippocampus, and the lateral perforant path was prominently immunoreactive for mGluR8 in the dentate gyrus and CA3 area. The mossy fiber was labeled for mGluR2, mGluR7a, and mGluR7b, whereas the Schaffer collateral was labeled only for mGluR7a. Electron microscopy further revealed the spatial segregation of group II and group III mGluRs within presynaptic elements. Immunolabeling for the group III receptors was predominantly observed in presynaptic active zones of asymmetrical and symmetrical synapses, whereas that for the group II receptor (mGluR2) was found in preterminal rather than terminal portions of axons. Target cell-specific segregation of receptors, first reported for mGluR7a , was also apparent for the other group III mGluRs, suggesting that transmitter release is differentially regulated by 2-amino-4-phosphonobutyrate-sensitive mGluRs in individual synapses on single axons according to the identity of postsynaptic neurons. Key words: metabotropic glutamate receptor; hippocampus; perforant path; mossy fiber; Schaffer collateral; axon terminal; preterminal; immunohistochemistry; lesionMetabotropic glutamate receptors (mGluRs) have various modulatory f unctions on neuronal excitability, transmitter release, and synaptic plasticity in the C NS (Pin and Duvoisin, 1995). These f unctions have been studied most extensively in the hippocampus because of its roles in learning and memory and of its architecture, which is compartmentalized well with the three major excitatory pathways: the perforant path, mossy fiber, and Schaffer collateral. The mGluRs consist of at least eight subtypes that are classified into three groups (Nakanishi and Masu, 1994;Pin and Duvoisin, 1995). Group I mGluRs (mGluR1/mGluR5) are selectively activated by 3,5-dihydroxyphenylglycine (DHPG) (Schoepp et al., 1994) and coupled to inositol phospholipid hydrolysis. On the other hand, group II mGluRs (mGluR2/ mGluR3) and group III mGluRs (mGluR4/mGluR6/mGluR7/ mGluR8), which are linked to inhibition of the cAM P cascade in receptor-transfected cell lines, are selectively activated by 2-(2,3-dicarboxycyclopropyl)glycine (DCG-IV) (Hayashi et al., 1993) and 2-amino-4-phosphonobutyrate (L-AP4), respectively.Excitability of hippocampal neurons is modulated directly through group I mGluRs (Davies et...

We have isolated a novel liver-specific organic anion transporter, LST-1, that is expressed exclusively in the human, rat, and mouse liver. LST-1 is a new gene family located between the organic anion transporter family and prostaglandin transporter. LST-1 transports taurocholate (K m ‫؍‬ 13.6 M) in a sodium-independent manner. LST-1 also shows broad substrate specificity. It transports conjugated steroids (dehydroepiandrosterone sulfate, estradiol-17␤-glucuronide, and estrone-3-sulfate), eicosanoids (prostaglandin E 2 , thromboxane B 2 , leukotriene C 4 , leukotriene E 4 ), and thyroid hormones (thyroxine, K m ‫؍‬ 3.0 M and triiodothyronine, K m ‫؍‬ 2.7 M), reflecting hepatic multispecificity.LST-1 is probably the most important transporter in human liver for clearance of bile acids and organic anions because hepatic levels of another organic anion transporter, OATP, is very low. This is also the first report of the human molecule that transports thyroid hormones.One of the major function of the liver is the removal of various endogenous and exogenous compounds from the circulation (1, 2). This clearance process involves basolateral membrane transport systems that mediate the hepatocellular uptake of bile acids, organic anions, and organic cations (3, 4). One well studied class of substrates are the bile acids. The uptake of taurocholate is mainly mediated by the Na ϩ /taurocholate cotransporting polypeptide (ntcp) in a Na ϩ -dependent manner (5). The uptake of other bile acids (e.g. cholate) occurs predominantly via a Na ϩ -independent mechanism (2, 4). Some amount of taurocholate is also transported by the Na ϩ -independent mechanism. This Na ϩ -independent carrier system further shows a broad substrate specificity transporting conjugated steroids, cardiac glycosides, and other xenobiotics (4).Initially, the organic anion transporter (oatp) 1 family (oatp1, oatp2, oatp3) was considered to represent the Na ϩ -independent transporting mechanisms in the liver (6 -8). Subsequently, a human cDNA, termed OATP, was isolated (9). However, significant differences were found between human OATP and rat oatp family. First, although the substrate specificities were qualitatively similar, significant differences were found between human OATP-and rat oatp family-mediated initial uptake rates and apparent K m values (10, 11). Second, Northern blot analysis of the human OATP showed considerably high expression in the brain, a pattern that is different from any of the oatp family members. These findings strongly suggest the existence of a different group of organic anion transporters in human liver.Here we report the isolation of a novel human organic anion transporter, termed LST-1, which is expressed exclusively in the liver. When expressed in Xenopus oocytes, many of the functional characteristics of LST-1 were identical to the multispecific transporting mechanisms of human liver. These results suggest that LST-1 is the predominant clearance mechanism of several endogenous and exogenous substrates in human liver. MATERIALS ...

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Two complementary DNAs for the organic anion transporter subtypes oatp2 and oatp3, which transport thyroid hormones as well as taurocholate, were isolated from a rat retina cDNA library. The sequence of oatp2 is identical to that recently reported (Noé , B., Hagenbuch, B., Stieger, B., and Meier, P. J. (1997) Proc. Natl. Acad. Sci. U. S. A. 94, 10346 -10350), whereas the sequence of oatp3 is novel. oatp3 consists of 670 amino acid residues and exhibits a structural architecture common to the organic anion transporter family, possessing the 12 putative membrane-spanning segments. Oocytes injected with oatp2 and oatp3 cRNAs showed taurocholate uptake in a saturable manner. The oatp2 and oatp3 cRNAinjected oocytes also showed significant uptake of both thyroxine and triiodothyronine. Northern blot and in situ analyses showed that the oatp2 mRNA was widely expressed in neuronal cells of the central nervous system, especially in the hippocampus, cerebellum, and choroid plexus as well as in the retina and liver. The oatp3 mRNA was highly expressed in the kidney and moderately abundant in the retina. This suggests that oatp2 and oatp3 are multifunctional transporters involved in the transport of thyroid hormones in the brain, retina, liver, and kidney.A homeostatic system controls the fluid environment in the brain and keeps its chemical composition relatively constant compared with that of plasma. One mechanism is the bloodbrain barrier, which selectively transports chemical substances via capillary endothelial cells (1). A second essential component is the choroid plexus (blood-cerebrospinal fluid barrier), which secretes or takes up specific chemical substances (2). Although the presence of specific transporting mechanisms has long been postulated, little is known about their molecular identity. Recent molecular biological studies revealed the organic anion transporter family: the Na ϩ -independent organic anion-transporting polypeptide oatp1 from rat liver, which transports bile acid, bromosulfophthalein (BSP), 1 and conjugated and unconjugated steroid hormones (3, 4); the kidney-specific transporter OAT-K1, which transports methotrexate in the basolateral membrane of renal tubules (5); and the prostaglandin transporter (6). Moreover, physiological studies have suggested the presence of other members of the organic anion transporter family (7). Noé et al. (8) have recently reported that a new organic anion transporter subtype (oatp2) is present in rat brain and liver and that the oatp2-expressed oocytes transported cardiac glycoside as well as taurocholate. However, the endogenous substrate of oatp2 and the regional distribution in the brain have not been revealed.It has been suggested that thyroid hormones are transported into the brain via the blood-brain barrier (9) or via the choroid plexus (10). To reveal this mechanism, we focused on the retina. In the retina, the retinal pigment epithelium is the unique source of transthyretin synthesis, and it serves to transport thyroxine (T4) across the blood-retina barr...

Digoxin, which is one of the most commonly prescribed drugs for the treatment of heart failure, is mainly eliminated from the circulation by the kidney. P-glycoprotein is well characterized as a digoxin pump at the apical membrane of the nephron. However, little is known about the transport mechanism at the basolateral membrane. We have isolated an organic anion transporter (OATP4C1) from human kidney. Human OATP4C1 is the first member of the organic anion transporting polypeptide (OATP) family expressed in human kidney. The isolated cDNA encodes a polypeptide of 724 aa with 12 transmembrane domains. The genomic organization consists of 13 exons located on chromosome 5q21. Its rat counterpart, Oatp4c1, is also isolated from rat kidney. Human OATP4C1 transports cardiac glycosides (digoxin, K m ‫؍‬ 7.8 M and ouabain, K m ‫؍‬ 0.38 M), thyroid hormone (triiodothyronine, Km ‫؍‬ 5.9 M and thyroxine), cAMP, and methotrexate in a sodiumindependent manner. Rat Oatp4c1 also transports digoxin (K m ‫؍‬ 8.0 M) and triiodothyronine (Km ‫؍‬ 1.9 M). Immunohistochemical analysis reveals that rat Oatp4c1 protein is localized at the basolateral membrane of the proximal tubule cell in the kidney. These data suggest that human OATP4C1͞rat Oatp4c1 might be a first step of the transport pathway of digoxin and various compounds into urine in the kidney.