Substance P (SP) is present in the anterior pituitary gland (AP), and its concentration there is regulated by the hormonal status of the animal. The observation that SP is releasable from hemipituitaries in a K(+)-stimulated, Ca(2+)-dependent manner and the demonstration of SP-binding sites in the AP have led to the suggestion that SP participates in a paracrine or autocrine manner in the regulation of AP function. Contradictory reports of the effects of SP on the secretion of AP hormones, particularly LH, led us to address the question of whether SP can act directly on the AP to effect LH secretion. We found that SP (100 nM) can stimulate LH release (300-400% of control values) in short term cultures of AP cells and that this effect varies as a function of the age and sex of the animal. There was no significant effect of SP on the release of LH from AP cells of male and female prepubertal rats (20-30 days). During the peripubertal period (30-35 days), a sharp increase in the response to SP occurred in both sexes. This responsiveness was dose dependent and persisted at all ages studied in AP cells from the female rat. In contrast, the responsiveness of AP cells from male rats that developed during the peripubertal period diminished during maturation and was absent after 60 days of age. When adult female rats were exposed to androgens for 6 weeks in vivo and tested for the ability of SP to stimulate the LH secretion, the response was significantly diminished. These studies support the speculation that SP has a functional role in the secretion of LH.