There are several lines of evidence suggesting that, in addition to neurotrophins, member(s) of glial cell line-derived neurotrophic factor (GDNF) family play important roles in the development of sympathetic neurons. However, the mechanism regulating the responsiveness of the neurons to GDNF family members is not known. Previously, we reported on the cooperative roles of bone morphogenetic protein-2 (BMP2) and retinoic acid (RA) in the enhancement of neurotrophin-3 (NT3) responsiveness in cultured sympathetic neurons dissociated from perinatal rat superior cervical ganglia (SCG). In the present study, we further examined the effects of BMP2 and RA on the regulation of the responsiveness of SCG neurons to GDNF family members. Consequently, we found that RA alone induced the responsiveness of SCG neurons specifically to GDNF by upregulating the ligand-specifying receptor for GDNF (GFR␣-1) at both the mRNA and protein levels. The expression levels of mRNAs for other ligand-specifying receptors for GDNF family (GFR␣-2 and GFR␣-3) were unaffected by RA. Although the upregulation of signal-transducing receptor Ret by the RA treatment was rather small, this treatment significantly increased the efficacy of tyrosine phosphorylation of Ret by GDNF. Experiments using synthetic retinoids suggested that RA acts through ␣-type of nuclear retinoic acid receptor to exert the induction of GDNF responsiveness. On the other hand, BMP2, which had no significant effect by itself on the GDNF responsiveness, promoted the action of RA to upregulate GFR␣-1 and enhance the GDNF responsiveness. These results indicate that RA and BMP2 play important roles in the induction of GDNF responsiveness, as well as NT3 responsiveness, of developing SCG neurons.

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