The prenatal development of growth hormone-releasing hormone (GHRH) gene expression was studied in rat hypothalamus using hybridization histochemistry. Two populations of neurons expressing the GHRH gene were found and they had a different pattern of development. Neurons in the arcuate nucleus first expressed GHRH on E16 and 4 days later an adult-like pattern was found. Few neurons along the laterodorsal aspect of the ventromedial hypothalamic nucleus expressed GHRH on E17, with a subsequent increase of expression over the next days. On the day before birth, adult-like patterns were seen. These results demonstrate the differential regulation of GHRH gene expression in the rat hypothalamus.