The development of the nervous system is regulated by trophic signals that control cell proliferation, differentiation, and survival. Numb is an evolutionarily conserved protein identified by its ability to control cell fate in the nervous system of Drosophila. Mammals express four isoforms of Numb that differ in the length of a phosphotyrosine-binding (PTB) domain and a proline-rich region (PRR). Using PC12 cells stably expressing each of the human isoforms, we show that Numb regulates sensitivity of the cells to neurotrophic factor-induced differentiation and neurotrophic factor withdrawal-induced death in an isoform-specific manner. Numb isoforms containing a short PTB domain enhance the differentiation response to NGF and enhance apoptosis upon NGF withdrawal; Numb isoforms containing a long PTB domain exhibit the same sensitivity to NGF as vector-transfected cells. These effects of Numb were found to be independent of the length of the PRR. In undifferentiated conditions, the levels of full-length TrkA and of phosphorylated p44/p42 mitogen-activated protein kinase (MAPK) are increased in cells expressing Numb isoforms with a short PTB domain, indicating an up-regulation of NGF signaling pathways. Furthermore, we provide evidence that the mechanism whereby short PTB domain Numb isoforms sensitize cells to trophic factor deprivation-induced apoptosis involves elevations in intracellular calcium concentrations. Our results suggest that Numb sensitizes cells to neurotrophin responses in an isoform-specific manner, an effect that may play an important role in the development and plasticity of the nervous system.