Organ culture of the bovine subcommissural organ: evidence for synthesis and release of the secretory material.
The subcommissural organ (SCO) is a brain circumventricular organ formed by ependymal and hypendymal secretory cells. It secretes glycoproteins into the cerebrospinal fluid of the third ventricle where they condense into a thread-like structure known as Reissner's fiber (RF). The present study was designed to investigate whether or not the bovine SCO continues to synthesize and release glycoproteins after a long-term culture. Cultured explants of SCO survive for several months. The content of the secretory granules present in the cultured ependymocytes displayed immunoreactive and lectin-binding properties similar to those of the core glycosylated glycoproteins found in the bovine SCO. The explants actively incorporated (35)S-cysteine. In the cultured ependymocytes, the pattern of distribution of the radioactive label and that of the immunoreactive secretory material was similar, thus indicating that this material has been synthesized during culture. At the ultrastructural level, the cultured tissue exhibited a high degree of differentiation comparable to that of the bovine SCO in situ. A striking finding was the observation of similar results when cerebrospinal fluid was used as a culture medium. The addition of antibodies against RF-glycoproteins into the culture medium allowed visualization, by means of different immunocytochemistry protocols, deposits of extracellular immunoreactive secretory material on the free surface of the cultured ependymocytes, indicating that release of secretory glycoproteins into the culture medium does occur. Primary culture of dispersed SCO ependymocytes, obtained either from fresh or organ cultured bovine SCO, showed that these cells release RF-glycoproteins that aggregate in the vicinity of each cell. The present investigation has shown that: (1) two types of secretory ependymocytes become evident in the cultured SCO; (2) under culture conditions, the SCO cells increase their secretory activity; (3) explants of bovine SCO synthesize RF-glycoproteins and release them to the culture medium; (4) after release these proteins aggregate but do not form a RF; (5) a pulse of anti-RF antibodies into the culture medium blocks the secretion of RF-glycoproteins for several days.
Schöbitz
K
K
Instituto de Histología y Patología, Facultad de Medicina, Universidad Austral de Chile, Valdivia, Chile. kschoebi@uach.cl
Gonzalez
C
C
Peruzzo
B
B
Yulis
C R
CR
Rodríguez
E M
EM
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D002555:Cerebrospinal Fluid; D003470:Culture Media; D016895:Culture Media, Serum-Free; D004805:Ependyma / Q000166:cytology; D007150:Immunohistochemistry; D008855:Microscopy, Electron, Scanning; D009924:Organ Culture Techniques / Q000379:methods; D013351:Subcommissural Organ / Q000254:growth & development* / Q000378:metabolism* / Q000648:ultrastructure; D013997:Time Factors
D003470:Culture Media; D016895:Culture Media, Serum-Free