Neurite penetration into collagen gels requires Ca2+-dependent metalloproteinase activity.
Cultured sympathetic neurons from neonatal rats release a Ca2+-dependent metalloproteinase capable of degrading native and denatured type I collagen. A large fraction of the metalloproteinase activity is released from growth cones of growing neurites, which suggests that this proteinase may be involved in neurite penetration into collagen-rich extracellular matrices during development. In the present study, we investigated the possibility that neurite penetration into 3-dimensional collagen gels requires the Ca2+-dependent metalloproteinase. Sympathetic neurons were plated on top of collagen gels and the neuronal metalloproteinase activity was inhibited with the collagenase inhibitor HSCH2CH[CH2CH(CH3)2]-CO-Phe-Ala-NH2 (HS-LFA). HS-LFA inhibited the Ca2+-dependent metalloproteinase activity, penetration of neurites into collagen gels and release of 3H from 3H-collagen gels in a dose-dependent manner over very similar concentration ranges (EC50 = 150-200 nM). Highly significant correlations existed between neurite penetration into collagen gels and metalloproteinase activity (r = 0.99), and metalloproteinase activity and release of 3H from 3H-collagen gels (r = 0.99). Although HS-LFA inhibited the growth of neurites within collagen gels, it had no effect on neurite outgrowth on collagen films. Thus, the neuronal Ca2+-dependent metalloproteinase appears to be required for neurite penetration into and growth within 3-dimensional collagen matrices, but not for growth along a 2-dimensional collagen substrate.
Pittman
R N
RN
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia.
Williams
A G
AG
D000818:Animals; D001345:Autoradiography; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003094:Collagen; D004591:Electrophoresis, Polyacrylamide Gel; D005728:Ganglia, Sympathetic / Q000166:cytology / Q000201:enzymology / Q000254:growth & development*; D008666:Metalloendopeptidases / Q000378:metabolism*; D009474:Neurons / Q000166:cytology* / Q000201:enzymology; D051381:Rats
D003094:Collagen; D008666:Metalloendopeptidases; D002118:Calcium