Adult sensory neurons differ chemically, morphologically, and functionally, but the factors that generate their diversity remain unclear. For example, neuropeptides are generally found in small neurons, whereas abundant neurofilament is common in large neurons. Neurons containing the neuropeptides calcitonin gene-related peptide (CGRP) or substance P were quantified using immunohistochemistry in rat lumbar dorsal root ganglion (DRG) at times before and after sensory neurons contact central and peripheral targets in vivo. No neurons in the newly formed DRG expressed neuropeptide or neuropeptide mRNA, but neuropeptides were detectable about the time that axons connect with peripheral targets. To determine the requirement for target in neuropeptide regulation, embryonic DRG neurons were isolated at times before central and peripheral connections had formed, placed in culture, and immunocytochemically assayed for CGRP and substance P. Cultured neurons expressed neuropeptides with a time course and in proportions similar to those in vivo. Thus, some neurons in the embryonic DRG seem to be intrinsically specified to later express CGRP and substance P. The percentage of CGRP-immunoreactive neurons was not changed by cell density, non-neuronal cells, neurotrophins in addition to nerve growth factor (NGF), or antibody inactivation of neurotrophin-3 in the presence of NGF. To test the role of extrinsic cues on CGRP expression, DRG neurons were co-cultured with potential target tissues. Coculture with a rat epidermal or smooth muscle cell line increased the proportion of CGRP-containing neurons, whereas primary skeletal muscle and 3T3 cells had no effects. Thus, multiple appropriate sensory neuron phenotypes arise in a regulated fashion in cultured neurons isolated before target connections have formed, and some candidate target tissues can modulate that intrinsic expression pattern. Key words: neuropeptides; substance P; calcitonin generelated peptide; neurofilament; nociceptive; sensory ganglion; target tissue; skin; neurotrophinA striking feature of the adult vertebrate sensory nervous system is the diversity of neuronal cell types that perform together to maintain homeostasis and convey information about the environment. During development, specific neurons must be generated in appropriate locations and connect discretely to both peripheral targets and the CNS for functional integrity. Thus, it is important to understand not only how the phenotype is generated but also how neurons match targets. This study examines the developmental regulation of sensory neurons containing calcitonin generelated peptide (CGRP) that predominantly contact visceral and cutaneous peripheral target end organs in vivo.Adult rat sensory neurons can express neuromodulatory neuropeptides, but the factors that regulate their initial expression remain unknown. Cutaneous afferents appear in rat proximal hindlimb on embryonic days 14 -15 (E14 -E15) and in skin of distal toes at E16 -E17 (Reynolds et al