IN VIVO ELECTROPHYSIOLOGICAL MATURATION OF NEURONS DERIVED FROM A MULTIPOTENT PRECURSOR (EMBRYONAL CARCINOMA) CELL LINE

Abstract The multipotent embryonal carcinoma (EC) P19 cell line differentiates into neurons, glia and smooth muscle following exposure to retinoic acid (RA). RA-induced differentiation is irreversible and the neurons that develop are abundant, post-mitotic, and survive for prolonged periods in culture or when grafted into the CNS of adult rats. Striatal slices containing grafted P19 cells were studied with intracellular recording and labelling techniques to examine the development of electrophysiological and morphological properties of P19-derived neurons over a period of 6 to 120 days after grafting into ibotenic acid lesioned striatum. Cells from 1-week-old grafts had a range of immature electrophysiological characteristics including unstable resting membrane potentials (RMP's) and very high membrane input resistances (Rin's). Many were not able to produce action potentials (AP's). In contrast, the majority of cells recorded from 2- and 3-week-old grafts had stable RMP's, moderate Rin's, and were able to produce regenerative AP's. In grafts over 4 weeks of age, the majority of P19-derived neurons had mature neuronal electrophysiological characteristics including RMP's of −60 mV, Rin's of 100–300 M Ω, and overshooting AP's. Morphologically, P19 derived neurons increase in soma size from 12–15 μ in diameter in 7–14-day-old grafts, to 25–35 μ in diameter in grafts 50–120 days old. Developing neurons exhibited a variety of morphotypes with increasingly complex processes and lengths of process extension. Our results demonstrate a developmental progression of the electrophysiology of P19-derived neurons, culminating in mature characteristics closely resembling those of adult rodent hippocampal or cortical pyramidal neurons. The ability to easily alter these cells genetically provides a powerful model for addressing issues specific to neuronal development.