Novel method for studying myelination in vivo reveals that EDTA is a potent inhibitor of myelin protein and mRNA expression during development of the rat sciatic nerve

Abstract

To probe the effects of possible inhibitors or enhancers of in vivo myelination, we have modified a technique widely used in studies of the developing neuromuscular system that involves incorporation of test compounds into a silicon rubber solution, which solidifies on contact with air. U-shaped rubber implants are inserted around the sciatic nerve of 1-day-old rats and left in place for 24-48 h. Sections from the region of the nerve lying within the implant, with or without the test compound, are then immunolabeled, examined with in situ hybridization or electron microscopy. Application of EDTA (440 microg/implant) in this way strongly suppressed the levels of the myelin-associated molecules protein P0, myelin basic protein (MBP), and galactocerebroside (Galc). mRNA levels for P0 and the myelin-related transcription factor Krox-20 were also reduced, further supporting association of the EDTA-induced effect with the myelinating Schwann cells. In contrast, no obvious differences were observed in either neurofilament (NF) protein or glial fibrillary acidic protein (GFAP) expression, suggesting absence of influence on axons or nonmyelinating Schwann cells. Despite the severely altered molecular composition of myelin in the presence of EDTA, examination in the electron microscope did not reveal any apparent ultrastructural changes in the myelin sheaths or nerve development. This work introduces a novel method for studying nerve development and shows that EDTA, which chelates divalent cations such as Ca(2+) and Mg(2+), strongly and selectively reduces levels of molecules, which, on postnatal days 1-4, are expressed in myelinating cells at much higher levels than in cells not engaged in myelination.

Μοριακές Αλληλεπιδράσεις Στην Εγγύς Της Παρακομβικής Περιοχή Των Εμμύελων Νευρικών Ινών

“…(Εμβρυϊκή ημέρα 13) τα πρόδρομα κύτταρα Schwann ξεκινούν να εκφράζουν τρεις παράγοντες διαφοροποίησης: την myelin protein 0 (P0), η οποία και αποτελεί το 50% των συνολικών πρωτεϊνών που βρίσκονται στην μυελίνη του ΠΝΣ (Oguievetskaia et al, 2007), την growth associated protein 43 (GAP43) και την F-spondin (Corfas et al, 2004 Corfas et al, 2004;Jessen and Mirsky, 1999;Meintanis et al, 2004).…”