1) TrkA, in the absence of p75, is fully capable of mediating 125I-NGF endocytosis; 2) internalized 125I-NGF is slowly and inefficiently degraded; 3) following internalization, 125I-NGF is retroendocytosed; and 4) the ability of 125I-NGF to remain receptor-associated during acidic conditions may provide a mechanism for its retroendocytosis via recycling TrkA vesicles.

Nerve growth factor (NGF) treatment of Chinese hamster ovary ®broblast (CHO) cells exogenously expressing 2.5610 5 TrkA receptors (CHO/TrkA) results in inhibition of serum and insulin-like growth factor-I (IGF-I) stimulated cell proliferation in a dosedependent manner. Furthermore, NGF does not stimulate [ 3 H]thymidine incorporation and inhibits IGF-I mediated DNA synthesis in CHO/TrkA cells. NGF and IGF-I induce extracellular-signal regulated kinase 1 (ERK1) and ERK2 activation, but NGF is able to stimulate a higher and more sustained activation of these enzymes compared with IGF-I. Cotreatment with NGF and IGF-I yields an ERK1/2 activity pro®le similar to that of NGF treatment alone. While pretreatment with mitogen activated protein kinase kinase (MKK) inhibitor PD98059 (30 mM) results in 100% inhibition of IGF-I stimulated MAPK phosphorylation (IC 50 51 mM), NGF mediated MAPK phosphorylation is only decreased by 50% (IC 50 =3 mM). NGF, but not IGF-I, stimulates tyrosine phosphorylation and activation of PLC-g1 which can be inhibited in a dose-dependent manner by phosphoinositide-speci®c phospholipase C (PI-PLC) inhibitor U73122 (IC 50 =4 mM). Pretreatment with U73122 (IC 50 =7 mM) results in an 87% inhibition of NGF mediated MAPK phosphorylation, while cotreatment with PD98059 and U73122 results in 97% inhibition. U73122 pretreatment has no eect on NGF stimulated Akt activation. NGF, but not IGF-I, stimulates the tyrosine phosphorylation of Suc1-associated neurotrophic factor-induced tyrosine phosphorylation target (SNT-1)/®broblast growth factor receptor substrate 2 (FRS2) which can be completely prevented by pretreatment with 10 mM U73122. Finally, inhibition of PI-PLC results in NGF's ability to stimulate DNA synthesis in the absence and presence of IGF-I.

1) TrkA, in the absence of p75, is fully capable of mediating 125I-NGF endocytosis; 2) internalized 125I-NGF is slowly and inefficiently degraded; 3) following internalization, 125I-NGF is retroendocytosed; and 4) the ability of 125I-NGF to remain receptor-associated during acidic conditions may provide a mechanism for its retroendocytosis via recycling TrkA vesicles.