CHARACTERIZATION OF A CNS CELL-LINE, CAD, IN WHICH MORPHOLOGICAL-DIFFERENTIATION IS INITIATED BY SERUM DEPRIVATION

A CNS catecholaminergic cell line, Cath.a, was established by targeted oncogenesis in transgenic mice. Cath.a cells express neuronal propert ies but lack neuronal morphology. Here, we describe a variant of Cath. a, called CAD (Cath.a-differentiated), in which reversible morphologic al differentiation can be initiated by removal of serum or exogenously added protein from the medium. In serum- or protein-free media, CAD c ells stop proliferating and extend long processes. Differentiated CAD cells can be maintained without serum or protein for at least 6 weeks. CAD cells are distinct from Cath.a cells; most significant, the origi nal immortalizing oncogene, SV40 T antigen, was spontaneously lost. By immunostaining or immunoblotting, we show that CAD cells express neur on-specific proteins, such as class III beta-tubulin, GAP-43, SNAP-25, and synaptotagmin, but not GFAP. Ultrastructurally, processes from di fferentiated CAD cells have abundant parallel microtubules and interme diate filaments, and bear varicosities that contain both large dense-c ore vesicles/granules (120-160 nm) and smaller clear vesicles (60-80 n m). Additionally, CAD cells express enzymatically active tyrosine hydr oxylase and accumulate L-DOPA. CAD cells exhibit biochemical and morph ological characteristics of primary neurons and provide an unique tool for studying neuronal differentiation.