A rapid and efficient method for neuronal induction of the P19 embryonic carcinoma cell line.

ABSTRACT

BACKGROUND: P19 mouse embryonic carcinoma cells are conventionally induced to differentiate into neural cells by suspension culture in the presence of retinoic acid to form cell aggregates, followed by adhesion culture in a poly-l-lysine-coated dish. Drawbacks of this procedure include it taking more than 10 days to obtain mature neurons, and non-neuronal proliferating cells occupying the majority of the cell population with time. NEW METHOD: Here, we show a novel method for the rapid and efficient neurogenesis of P19 cells, without aggregate formation in a suspension culture. The new approach is based on an adherent serum-free culture in a laminin-coated dish in the presence of FGF8, a γ-secretase inhibitor, and cytosine arabinoside. RESULTS: The new method efficiently induced P19 cells to differentiate into neurons within 4 days, and subsequently into mature neurons that were responsive to several neurotransmitters, giving spontaneous neuronal network activity within 6 days. COMPARISON WITH EXISTING METHOD: The novel method accelerated neuritogenesis and enhanced population of neuron selectively compared to the conventional method. Proliferating non-neuronal cells were eliminated by adding cytosine arabinoside during neuronal maturation. CONCLUSIONS: The method is useful for studying neuronal differentiation or activities.