Model of neurotransmitter specification by transcription factors, activity and target-derived factors. A, Early in development, when neurons have not yet established contact with their targets, spontaneous calcium spike activity and transcription factors work together to specify their transmitter expression profile. B, After neurons contact their targets, target-derived factors, including both retrograde factors (yellow dots) and diffusible factors (red dots), suppress expression of non-cholinergic transmitters, block spontaneous calcium activity and disrupt evoked activity-dependent transmitter specification. These factors work in combination with transcription factors to further refine and lock inthe transmitter expression phenotype.