Publisher Summary This chapter review the effects of two different factors derived from muscle which enhance cholinergic properties of neurons in vitro. In the absence of muscle extract the development of choline acetyltransferase (CAT) in tissue culture follows a biphasic time course in which there is an initial decrease in CAT activity followed by a period of steadily increasing activity. The addition of muscle extract to the cultures prevents the initial decline and stimulates the subsequent development of CAT activity. A factor (cholinergic development factor, CDF) in the muscle extract which produces such effects has been purified and partially characterized. While CDF was being purified from rat skeletal muscle, attempts were made to purify neurotrophic activity from autopsied human skeletal muscle. Chick ciliary neurons were used to assay the enhancement of acetylcholine synthesis, CAT. Although neurotrophic factors may exhibit a range of specific behaviors in vitro such as neuron survival or enhancement of cholinergic activity, for complete validation such factors must be shown to be capable of influencing neuronal survival in vivo. An in vivo effect could eliminate a trivial explanation of the in vitro phenomena, i.e. that purified extracts may provide essential components missing or perturbed in the tissue culture environment, or that they act in a manner normally inoperative in vivo.