Gastroesophageal reflux is common in children after successful repair of esophageal atresia (EA), and may be related to a congenital neuronal abnormality of the esophagus. This study employed a fetal rat model of adriamycin-induced EA to investigate whether the innervation of the esophagus is abnormal in EA. The fetal rats were divided into four groups: (1) normal controls; (2) a saline-injected controls; (3) adriamycin administered but without the development of EA; and (4) adriamycin-induced EA. The distal esophageal segments were immunostained with a general neural marker, protein gene product 9.5 (PGP). Immunoreactivity per cross-sectional area (/xsa) was measured with an image analyzer. The extent of the esophageal circumference encircled by PGP-stained nerve tissue was assessed. While there was no significant difference in PGP immunoreactivity/xsa between the groups, the near-complete ring of nerve tissue along the plane of the myenteric plexus was replaced by clusters of nerve tissue in the atretic group (normal vs EA, P = 0.001, Mann-Whitney U test). The abnormal distribution of nerve tissue in the atretic esophagus may be contributing factor in the esophageal dysmotility seen in EA.