Bile acids are transported across the ileal enterocyte brush border membrane by the well characterized apical sodium-dependent bile acid transporter (Asbt) Slc10a2; however, the carrier(s) responsible for transporting bile acids across the ileocyte basolateral membrane into the portal circulation have not been fully identified. Transcriptional profiling of wild type and Slc10a2 null mice was employed to identify a new candidate basolateral bile acid carrier, the heteromeric organic solute transporter (Ost)␣-Ost␤. By Northern blot analysis, Ost␣ and Ost␤ mRNA was detected only in mouse kidney and intestine, mirroring the horizontal gradient of expression of Asbt in the gastrointestinal tract. Analysis of Ost␣ and Ost␤ protein expression by immunohistochemistry localized both subunits to the basolateral surface of the mouse ileal enterocyte. The transport properties of Ost␣-Ost␤ were analyzed in stably transfected Madin-Darby canine kidney cells. Coexpression of mouse Ost␣-Ost␤, but not the individual subunits, stimulated Na ؉ -independent bile acid uptake and the apical-to-basolateral transport of taurocholate. In contrast, basolateral-to-apical transport was not affected by Ost␣-Ost␤ expression. Co-expression of Ost␣ and Ost␤ was required to convert the Ost␣ subunit to a mature glycosylated endoglycosidase H-resistant form, suggesting that co-expression facilitates the trafficking of Ost␣ through the Golgi apparatus. Immunolocalization studies showed that co-expression was necessary for plasma membrane expression of both Ost␣ and Ost␤. These results demonstrate that the mouse Ost␣-Ost␤ heteromeric transporter is a basolateral bile acid carrier and may be responsible for bile acid efflux in ileum and other ASBT-expressing tissues.

The cellular and subcellular localization and mechanism of transport of the heteromeric organic solute transporter (OST) OST␣-OST␤ was examined in human and rodent epithelia. The two subunits of the transporter were expressed together in human small intestine, kidney, and liver, tissues that also express the apical sodium-dependent bile acid uptake transporter ASBT (

Toxic organic cations can damage nigrostriatal dopaminergic pathways as seen in most parkinsonian syndromes and in some cases of illicit drug exposure. Here, we show that the organic cation transporter 3 (Oct3) is expressed in nondopaminergic cells adjacent to both the soma and terminals of midbrain dopaminergic neurons. We hypothesized that Oct3 contributes to the dopaminergic damage by bidirectionally regulating the local bioavailability of toxic species. Consistent with this view, Oct3 deletion and pharmacological inhibition hampers the release of the toxic organic cation 1-methyl-4-phenylpyridinium from astrocytes and protects against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced dopaminergic neurodegeneration in mice. Furthermore, Oct3 deletion impairs the removal of the excess extracellular dopamine induced by methamphetamine and enhances striatal dopaminergic terminal damage caused by this psychostimulant. These results may have far-reaching implications for our understanding of the mechanism of cell death in a wide range of neurodegenerative diseases and may open new avenues for neuroprotective intervention.astrocytes ͉ Parkinson's disease ͉ extraneuronal monoamine transporter ͉ dopamine ͉ methamphetamine

Mutations in the mitochondrial encoded protein PTEN-induced putative kinase 1 (PINK1) cause autosomal recessive Parkinson disease (PD). In mammalian cells, mutant PINK1 has been reported to promote fission or inhibit fusion in mitochondria; however, the mechanism by which this process occurs remains elusive. Using an ecdysone-inducible expression system in mammalian dopaminergic neuronal cells, we report here that human mutant PINK1 (L347P and W437X) mediates an overall fission effect by increasing the ratio of mitochondrial fission over fusion proteins, leading to excessive dysfunctional fragmented mitochondria. Knocking down endogenous Pink1 produces similar effects. In contrast, overexpressing human wild type PINK1 produces a pro-fusion effect by increasing the ratio of mitochondrial fusion/fission proteins without resulting in functionally compromised mitochondria. Parkin knockdown blocks the imbalance in fission/fusion proteins. Furthermore, overexpressing parkin and ubiquitin increases degradation of the mitochondrial fission hFis1 protein, suggesting PINK1 and parkin maintain proper mitochondrial function and integrity via the fission/fusion machinery. Through genetic manipulations and treatment with the small molecule mitochondrial division inhibitor (mdivi-1), which inhibits DLP1/Drp1, both structural and functional mitochondrial defects induced by mutant PINK1 were attenuated, highlighting a potential novel therapeutic avenue for Parkinson disease.The discoveries of mutations in the mitochondrial protein PTEN-induced putative kinase 1 (PINK1) 2 as a cause of autosomal recessive PD (1-3) have fueled the longstanding interest in the role of mitochondrial dysfunction in PD. Currently about 50 PINK1 mutations have been reported (4), making it the second most common causative gene for autosomal recessive PD after parkin, an E3 ubiquitin ligase that has been shown to function downstream of PINK1 and to affect mitochondrial morphology (5-8). Although PINK1 mutations are spread throughout the gene, they are most commonly found in the region encoding the functional serine/threonine kinase domain at the C terminus, leading to loss of PINK1 kinase activity. Recently this kinase domain has been shown to face the cytoplasm (9), providing evidence of spatial proximity to allow this mitochondrial protein to directly interact with cytosolic parkin.Current consensus is that PINK1 is a protective protein. Supporting this role, PINK1 overexpression confers resistance to staurosporine, MPP ϩ , and rotenone toxicity in cultured cells (10, 11) as well as to MPTP-induced dopaminergic neuronal loss in mice (12). Conversely, reducing PINK1 levels by RNAi in cultured cells leads to enhanced cell death in the presence of MPP ϩ and rotenone (12, 13). Mitochondrial defects leading to degeneration of flight muscles and loss of dopaminergic neurons have also been reported in Pink1-deficient Drosophila (5, 14). In recent years, PINK1 has gained significant attention for its role in mitochondrial dynamics (fission, fusion, and migrati...

Mice deficient in the organic solute transporter (Ost)-α subunit of the heteromeric organic solute and steroid transporter, Ostα-Ostβ, were generated and were found to be viable and fertile but exhibited small intestinal hypertrophy and growth retardation. Bile acid pool size and serum levels were decreased by more than 60% in Ostα−/− mice, whereas fecal bile acid excretion was unchanged, suggesting a defect in intestinal bile acid absorption. In support of this hypothesis, when [3H]taurocholic acid or [3H]estrone 3-sulfate were administered into the ileal lumen, absorption was lower in Ostα−/− mice. Interestingly, serum cholesterol and triglyceride levels were also ∼15% lower in Ostα−/− mice, an effect that may be related to the impaired intestinal bile acid absorption. After intraperitoneal administration of [3H]estrone 3-sulfate or [3H]dehydroepiandrosterone sulfate, Ostα−/− mice had higher levels of radioactivity in their liver and urinary bladder and less in the duodenum, indicating altered hepatic, renal, and intestinal disposition. Loss of Ostα was associated with compensatory changes in the expression of several genes involved in bile acid homeostasis, including an increase in the multidrug resistance-associated protein 3, ( Mrp3)/ Abcc3, an alternate basolateral bile acid export pump, and a decrease in cholesterol 7α-hydroxylase, Cyp7a1, the rate-limiting enzyme in bile acid synthesis. The latter finding may be explained by increased ileal expression of fibroblast growth factor 15 ( Fgf15), a negative regulator of hepatic Cyp7a1 transcription. Overall, these findings provide direct support for the hypothesis that Ostα-Ostβ is a major basolateral transporter of bile acids and conjugated steroids in the intestine, kidney, and liver.