Nerve growth-promoting factor produced in culture media conditioned by specific CNS tissues of the snail Helisoma

Medium conditioned by tissue from the CNS of the snail, Helisoma, is capable of promoting neurite outgrowth in isolated neurons from adult central ganglia. The conditioning factor(s) (CF), contained in conditioned medium (CM), is produced only by central ganglionic rings and buccal ganglia and not by other tissues, including hemolymph. CF requires a minimum of 24 h to be produced or released into the medium. At 12 h growth-promoting activity was not detectable. CF binds tightly to the polylysine substratum and its activity is not mimicked by addition of various sera, NGF or fibronectin. CF activity is abolished by chymotrypsin, trypsin or heating to 100 °C, but is stable to DNase and RNase treatment. The percentage of cells exhibiting neurite outgrowth is approximately linear with the amount of neural tissue used to condition the medium up to 2 ganglionic rings/ml. Addition of more ganglia fails to stimulate a greater response. This apparent plateau of CM activity appears to be a function of production and/or release of CF, rather than a saturation effect on plated cells, since dose-response curves for dilutions of CM are approximately linear regardless of the number of ganglia used for conditioning. In addition, anisomycin inhibits 35% of CF appearance under conditions of over 90% protein synthesis inhibition in the ganglia used to produce the CM. Under these conditions anisomycin has no apparent effect on the maintenance of electrical excitability. The inhibitor data suggest that 65% of CF is derived from a pre-existing storage pool and that the remainder is synthesized during the 72 h conditioning period.