Protein tyrosine kinase (PTK) activity associated with synaptosomal membrane glycoprotein (SMGP) fractions of rat brain was examined. The synthetic substrate poly(Glu,-Tyr) was phosphorylated by SMGP in the presence of Mg2+ and Mn2+, whose stimulatory effects were additive. In contrast, endogenous tyrosine phosphorylation in SMGPs was strictly dependent on Mn2+. Anti-phosphotyrosine antibodies (PY20) immunoprecipitated two polypeptides in SMGPs of Mr 170K and 60K. Upon preincubation with IGF-I, 97/90K polypeptides were phosphorylated, corresponding to the IGF-I receptor beta-subunits, and were immunoprecipitated with both PY20 and anti-IGF-I-receptor antibodies. Immunoblot analysis using anti-src antibody revealed that there was src protein associated with the glycoprotein fractions of solubilized synaptosomal membranes. Additional experiments revealed that the 60K tyrosinephosphorylated polypeptide present in the PY20 precipitates was indeed pp60'-"". This was confirmed by subjecting the PY20 immunoprecipitates to immunoblotting using anti-src antibodies. In addition, src protein was directly immunoprecipitated by anti-src antibodies from the SMGP preparations. Hence, IGF-I receptors and glycoprotein-associated PTKs including pp60'~"'" may play an important role in synaptic transmembrane signalling, plasticity, and neuronal survival.