BINDING CHARACTERISTICS OF CILIARY NEUROTROPHIC FACTOR TO SYMPATHETICNEURONS AND NEURONAL CELL-LINES

Ciliary neurotrophic factor (CNTF) is a cytokine whose actions are lar gely restricted to the nervous system because of the predominant neuro nal distribution of its receptor, CNTFR alpha. In this study, we sough t to define the binding characteristics of CNTF to cultured sympatheti c neurons and cell lines of neuronal origin. We report that I-125-CNTF binds to cultured sympathetic neurons, MAH, PC12, and EW-1 cells via high and low affinity receptors that can be distinguished on the basis of their dissociation constants (K(D1 similar to)10(-12) M and K(D2)s imilar to 10(-9) M). Competition experiments showed that the IC50 for rat and human CNTF were, respectively, 65 pM and 5 an for sympathetic neurons and 75 pM. and 1.2 nM for EW-1 cells. Interestingly, leukemia inhibitory factor (LIF) did not compete for CNTF binding even at 100 n M concentration. The binding of I-125-CNTF to sympathetic neurons invo lved all three components of the CNTF receptor complex, namely CNTFR a lpha; LIFR, and gp130, as shown by cross-linking experiments. CNTF and LIF treatments down-regulated CNTF binding to sympathetic neurons and EW-1 cells, suggesting that heterologous ligands can regulate CNTF re ceptor levels, which may in turn modulate the efficacy of CNTF in vitr o and in vivo.