Human fetal brain cells in aggregate culture: a model system to study regulatory processes of the developing human...

Neuropeptide Y (NPY) is one of the most abundant neuropeptides in the brain and it has been implicated in a wide range of brain functions, including mentation. The aim of this study was to establish a culture system of human fetal brain cells expressing NPY in a regulated manner. The NPY production in response to forskolin and phorbol 12-myristate 13-acetate (PMA) was taken as a criterion for regulated expression of NPY. Aggregates were formed from dissociated cells derived from the cerebral hemispheres of human fetuses (12.5-19 weeks' gestation) by constant rotation and were maintained in serum-free medium. A 24 hr exposure to 10 microM forskolin + 20 nM PMA led to a 2-6-fold increase in NPY content of the cultures, most of which (80-90%) was secreted into the medium. The latter consisted of two substances differing in size: one corresponding in size to proNPY and the other to NPY. Thus, forskolin + PMA led to an increased production of NPY. Exposure to PMA alone led to an increase in NPY production comparable to that seen after forskolin + PMA and this effect of PMA was dose-dependent. In contrast, forskolin alone did not induce NPY production. Conditioned medium, derived from monolayer cultures enriched with human astrocytes, enhanced NPY production in response to forskolin+PMA in an age-dependent manner. The NPY production by aggregates derived from a 12.5 week-, 14-week- and 18-week-old fetus was enhanced 3-3.6-fold, 1.6-2-fold and 1.1-fold, respectively. Thus, expression of the NPY neurons in this culture system is a regulated process. The NPY production is enhanced markedly by activation of the protein kinase C pathway and by an astrocyte-derived soluble substance(s). Based on these results, we propose that this culture system can serve as a model for the study of regulatory processes of the human developing NPY neuron.