C-type natriuretic peptides (CNPs) are produced by endothelium and inhibit vascular smooth muscle cell (VSMC) proliferation. However, endothelial damage stimulates only transient VSMC proliferation in arteries. Here we report that a new source of CNP develops in rat carotid neointima 14 days after balloon angioplasty, when VSMC replication is subsiding despite continued absence of endothelium. CNP was detected immunohistochemically in neointimal but not medial VSMC. No other natriuretic peptides were detected immunohistochemically. CNP immunoreactivity (0.036 +/- 0.010 fmol/mg wet wt) was found in damaged arteries by radioimmunoassay, but none was detected in normal media. Reverse-phase chromatography suggested that this immunoreactivity consisted of CNP(1-53) and perhaps CNP(1-22). CNP transcript was identified by reverse transcription and polymerase chain reaction in carotid segments that had been stripped of endothelium but only once neointima had formed. Moreover, neointima expressed the NPR-C type of natriuretic peptide receptor at the same time as it synthesized CNP. Thus neointima develops an autocrine system for CNP that could regulate neointimal growth. Furthermore, the findings establish novel phenotypic differences between medial and neointimal VSMC in vivo.