We have examined the ability of Schwann cell-conditioned medium (SCCM) to promote neurite growth from embryonic retina and tectum in explant culture. Both retinal and tectal explants adhered to polylysine substrates, but neurite outgrowth from the explants was minimal when grown in the presence of non-conditioned serum-free defined medium. The addition of SCCM resulted in a significant extension of neurites from both types of explants. The neurite outgrowth from the retinal explants was mainly radial and appeared to be in fascicles of small diameter. Neurite outgrowth from the tectal explants stimulated by the SCCM tended to be in larger fascicles and in a more convoluted pattern than that seen from the retinal explants. After serial passaging of the SCCM to remove any substrate-adsorbable factors all neurite-promoting activity was lost. Neurite growth from 100% of the explants was seen on the tissue culture plates that had been preincubated with the SCCM. SCCM was analyzed by immunoblot to determine whether it contained laminin, a potent promoter of neurite outgrowth. Samples of media were electrophoresed on a sodium dodecyl sulphate (SDS)-polyacrylamide gel alongside purified laminin, transferred to nitrocellulose and stained with an antibody to laminin. Purified laminin ran in two bands at 400 kDa and 200 kDa. The conditioned media had a laminin positive band at 200 kDa. Antibody to laminin, when added to the conditioned media, resulted in a loss of the neurite promoting activity of the SCCM. These results suggest that the conditioned media contained laminin or a laminin-like molecule. From these observations it is evident that Schwann cells in vitro produce a substrate-adsorbable neurite-promoting factor into the tissue culture medium that is effective on developing retina and tectum.
Bibliographical noteFunding Information:
technicaal ssistanceK,a thy Ensrud for maintaining the tissuec ulturef acilityand Gerald ~edgewicfko r the photographwico rk. This project was supported by NIH Grant EY05432 (L.K.M.), a grant from the Minnesota Medical Foundatio(nL .K.M.) and a grant fromR e-searchto PrevenBt lindnessI,n c.,to theD epartment of Ophthalmology.
- Glial cell
- Superior colliculus
- Tissue culture
- Visual system