EPENDYMAL AND CHOROIDAL CELLS IN CULTURE - CHARACTERIZATION AND FUNCTIONAL-DIFFERENTIATION

During the past 10 years, our teams developed long-term primary cultur es of ependymal cells derived from ventricular malls of telencephalon and hypothalamus or choroidal cells (modified ependymal cells) derived from plexuses dissected out of fetal or newborn mouse or fat brains. Cultures were established in serum-supplemented or chemically defined media after seeding on serum-, fibronectin-, or collagen-laminin-coate d plastic dishes or semipermeable inserts. To identify and characteriz e cell types growing in our cultures, we used morphological features p rovided by phase contrast, scanning, and transmission electron microsc opy. We used antibodies against intermediate filament proteins (viment in, glial fibrillary acidic protein, cytokeratin, desmin, neurofilamen t proteins), actin, myosin, ciliary rootlets, laminin, and fibronectin in single or double immunostaining, and monoclonal antibodies against epitopes of ependymal or endothelial cells, to recognize ventricular wall cell type with immunological criteria. Ciliated or nonciliated ep endymal cells in telencephalic cultures, tanycytes and ciliated and no nciliated ependymal cells in hypothalamic cultures always exceeded 75% of the cultured cells under the conditions used. These cells were cha racterized by their cell shape and epithelial organization, by their a pical differentiations observed by scanning and transmission electron microscopy, and by specific markers (e.g., glial fibrillary acidic pro tein, ciliary rootlet proteins, DARPP 32) detected by immunofluorescen ce. All these cultured ependymal cell types-remarkably resembled in vi vo ependymocytes in terms of molecular markers and ultrastructural fea tures. Choroidal cells were also maintained for several weeks in cultu re, and abundantly expressed markers were detected in both choroidal t issue and culture (Na+-K+-dependent ATPase, DARPP 32, G proteins, ANP receptors). In this review, the culture models we developed (defined i n terms of biological material, media, substrates, duration, and subcu lturing) are also compared with those developed by other investigators during the last 10 years. Focusing on morphological and functional ap proaches, we have shown that these culture models were suitable to inv estigate and provide new insights on (1) the gap junctional communicat ion of ependymal, choroidal, and astroglial cells in long-term primary cultures by freeze-fracture or dye transfer of Lucifer Yellow CR afte r intracellular microinjection; (2) some ionic channels; (3) the hormo ne receptors to tri-iodothyronine or atrial natriuretic peptides; (4) the regulatory effect of tri-iodothyronine on glutamine synthetase exp ression; (5) the endocytosis and transcytosis of proteins; and (6) the morphogenetic effects of galactosyl-ceramide. Vie also discuss new in sights provided by recent results reported on in vitro ependymal and c horoidal expressions of neuropeptide-processing enzymes and neurosecre tory proteins or choroidal expression of transferrin regulated through , serotoninergic activation. (C) 1998 Wiley-Liss, Inc.