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.
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.
Meintanis
Stathis
S
Department of Biochemistry, Laboratory of Cellular and Molecular Neurobiology, Hellenic Pasteur Institute, 115 21 Athens, Greece.
Thomaidou
Dimitra
D
Jessen
Kristjan R
KR
Mirsky
Rhona
R
Matsas
Rebecca
R
D000818:Animals; D000831:Animals, Newborn; D002614:Chelating Agents / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000235:genetics; D004195:Disease Models, Animal; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D004337:Drug Carriers / Q000008:administration & dosage*; D051767:Early Growth Response Protein 2; D004492:Edetic Acid / Q000494:pharmacology*; D005699:Galactosylceramides / Q000235:genetics; D006131:Growth Inhibitors / Q000494:pharmacology; D020007:Implants, Experimental; D046529:Microscopy, Electron, Transmission; D004676:Myelin Basic Protein / Q000235:genetics; D018993:Myelin P0 Protein / Q000235:genetics; D009185:Myelin Proteins / Q000235:genetics*; D009186:Myelin Sheath / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D009416:Nerve Regeneration / Q000187:drug effects / Q000235:genetics; D019635:Neurosurgical Procedures / Q000295:instrumentation / Q000379:methods; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D012583:Schwann Cells / Q000187:drug effects / Q000378:metabolism; D012584:Sciatic Nerve / Q000187:drug effects* / Q000254:growth & development / Q000648:ultrastructure; D012826:Silicone Elastomers / Q000008:administration & dosage*; D014157:Transcription Factors / Q000235:genetics
D002614:Chelating Agents; D004268:DNA-Binding Proteins; D004337:Drug Carriers; D051767:Early Growth Response Protein 2; C497328:Egr2 protein, rat; D005699:Galactosylceramides; D006131:Growth Inhibitors; D004676:Myelin Basic Protein; D018993:Myelin P0 Protein; D009185:Myelin Proteins; D012333:RNA, Messenger; D012826:Silicone Elastomers; D014157:Transcription Factors; C002454:galactocerebroside; D004492:Edetic Acid