All-trans retinoic acid stimulates and maintains neurite outgrowth in nerve growth factor-supported developing chick...

In explanted embryonic chick sympathetic neurons, all-trans retinoic acid (RA) as well as nerve growth factor (NGF) were found to be required for neuronal survival and neurite outgrowth at early stages of development (day 6.5-7) in agreement with previous work (Rodriguez-Tébar and Rohrer [1991] Development 112:813-820). The dependence of neurons on all-trans RA for survival diminished at later stages of development. However, all-trans RA was found to be needed at all stages of development in order to maximize neurite outgrowth. Further, removal of all-trans RA from the cultures led to a rapid degeneration of the formed neurites, demonstrating the essentiality of all-trans RA for both the development of neurites, and for the maintenance of existing neurites in cultured embryonic sympathetic neurons. The mechanism whereby all-trans RA exerts its effects on embryonic sympathetic neurons may involve activation of the nuclear retinoic acid and retinoid-X receptor (RAR and RXR) families. The results of Northern blot analyses and/or reverse transcriptase-polymerase chain reaction (RT-PCR) studies show that embryonic sympathetic ganglia express RAR beta, RAR gamma and RXR gamma mRNAs. RXR gamma mRNA is expressed at highest levels in immature neurons that are not yet responsive to NGF (day 6.5-7) and message levels decline with increasing developmental age. In contrast, RAR beta transcript levels are barely detectable at day 6.5-7, and increase approximately 4-fold in ganglia from embryos at day 8.5-9 and decline thereafter. RT-PCR studies show that RAR gamma mRNA is expressed both early (day 6.5-7) and late (day 15) in ganglionic development. Transcripts for the NGF receptors, p75NGFR and p140trk were also examined. The appearance of a single 2.7 kb p140trk transcript coincides with the time when RAR beta mRNA is maximally expressed, raising the possibility that NGF receptors may be targets of retinoid action. Evidence is also presented that all-trans RA may enhance neurite outgrowth by mechanisms other than simply inducing NGF-responsiveness of neurons.