Aromatase activity is abundant in limbic and nonlimbic neural structures in zebra finches, especially in the telencephalon near neural circuits that control singing behavior. Also, male songbirds can have high estrogen levels in blood. Because it is difficult to detect aromatase activity in other male tissues, we have postulated that the brain itself is the source of the estrogen present in the blood of males. Previously, we developed methods to measure estrogen synthesis in vivo by injecting [3H]androgen into the systemic circulation or directly into tissues and then determining the quantity of [3H]estrogen entering or leaving the brain. Our results support our hypothesis that the brain is the primary site of estrogen synthesis in males of this species. Here, we confirm that [3H]estrogen emanates from the brain after systemic [3H]androgen injection by showing that the presumptive estrogen in jugular plasma is significantly reduced by treatment with an aromatase inhibitor. In females, but not males, estrogens can be found in large amounts in carotid plasma after systemic androgen injection, presumably due to ovarian estrogen synthesis. However, carotid estrogen levels are variable in females, so it is difficult to determine if the female brain contributes estrogen to blood as is seen in males. The data presented here suggest that the adrenal of males is not an important site of aromatization, because [3H]estrogens are undetected in the adrenals of castrated or sham-castrated males after adrenal injections of [3H] androgen. Nevertheless, at least 2.8- to 17.3-fold more [3H]estrogen was present in the jugular than in the carotid of castrated males after systemic injection of [3H]androgen. Because androgen levels were in the physiological range, we conclude that the brain routinely enriches the estrogen content of blood in normal adult males of this species.