Ontogenesis of peptidylglycyl alpha-amidation activity in the mouse hypothalamus in vivo and in serum-free medium cultures. Relation with thyroliberin (TRH) accumulation and release in vitro.

The influence of peptidylglycine alpha-amidating monooxygenase (PAM) and of its co-factor, ascorbate, were studied in relation to thyroliberin (TRH) activity during mouse hypothalamus development. In vivo, PAM activity developed slowly at fetal stages, and exhibited a sharp rise around the 5th-8th postnatal day, the adult level being reached around day 15. The same developmental pattern was observed when studied in serum-free cultures initiated from fetal mouse hypothalamus. Using this in vitro model, we investigated the effects of ascorbate, a necessary co-factor of PAM, on TRH. Upon ascorbate supplementation of the culture medium, the TRH accumulation normally occurring in our cultures was further enhanced. The half maximum effect was attained with 20 microM, and the amplitude of the response to ascorbate was maximum around 9-13 days in vitro. Moreover, ascorbate increased to an even larger extent the amounts of TRH released upon chemical depolarization. These results are consistent with a direct role of ascorbate on PAM activity, but other more general effects on the maturation of the neuronal response to physiological stimuli cannot be excluded.