Leptin is a key mediator in the maintenance of neuroendocrine homeostasis. The aim of this study was to determine the changes in serum leptin, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), nitric oxide (NO) levels in patients with hyperprolactinemia. The study consists of 16 consecutive patients with high prolactin (PRL) levels (group I) and a control group of 11 normoprolactinemic patients (group II). Pituitary tumor tissues of patients in groups I and II were analyzed for immunohistochemical (IHC) expression of prolactin and leptin. Group I has significantly higher levels of leptin than group II (P < 0.001). There is a strong correlation between PRL and leptin concentrations in group I. However, there were no statistically significant differences for NO, TNF-alpha, IL-6 between the two groups. IHC staining showed that there was strong immunoreactivity for leptin protein in PRL-secreting pituitary adenomas. Double immunostaining of adenoma tissues with PRL and leptin showed that the adenoma cells expressed both. These findings together are suggestive that leptin co-secretion from a prolactinoma may be the cause of increased serum leptin concentration, independently from the peripheral action of prolactin.