Sex and estrous cycle-related differences in the amphetamine (AMT)-stimulated release (pg/mg/min) of catecholamines (CA) from rat striatal and mediobasal hypothalamus (MBH) fragments were measured in an in vitro perifusion system. In striatal tissue from intact males, AMT stimulated the release of both norepinephrine (NE) and dopamine (DA). The AMT-stimulated release of DA from striatal tissue obtained from intact females varied with the stage of the estrous cycle. This increase in DA release was lower in striatal tissue from proestrous females than from females in estrus (P less than 0.05) or diestrus 1 (P less than 0.01). The NE release stimulated by AMT was greater than basal release only on estrus and diestrus. 2. Following castration (CAST) or CAST plus 500 micrograms testosterone propionate (TP), daily for 4 days, striatal tissue fragments from male rats continued to release CA in response to AMT stimulation. In contrast, ovariectomy (OVX) severely attenuated the AMT-stimulated release of both CA. Treatment of OVX females with 5 micrograms estradiol benzoate (EB), daily for 4 days, or 1.2 mg progesterone (P) slightly increased the AMT-stimulated release of DA but not NE. Treatment of OVX females with 5 micrograms EB, daily for 4 days, plus 1.2 mg P completely restored the AMT-stimulated release of both CA. Interestingly, MBH fragments from intact or gonadectomized rats, with or without hormonal treatment, demonstrated a consistent AMT-stimulated release of DA regardless of the sex of the animal. The AMT-stimulated NE release from these MBH fragments was less consistent, but there was no significant differences between the groups. These results demonstrate that the AMT-stimulated release of DA from striatal tissue in vitro is sex, hormonal, and tissue dependent.