An immunomodulatory role for serotonin (5-HT) has been demonstrated in mammals and evidence for a similar role for 5-HT has recently emerged in fish. However, as limited studies are available, discrepancies often exist regarding the role of 5-HT in the teleost immune response. Therefore, studies were undertaken to help clarify this relationship. Lymphocyte proliferation and extracellular superoxide (O2.-) production were examined in cells from bluegill sunfish injected with either 5-HTP (the immediate precursor to 5-HT) or pCPA (an inhibitor of the rate-limiting enzyme in 5-HT synthesis), or, in vitro following exposure of immune cells to either 5-HT, the 5-HT(1A) receptor agonist, 8-OH-DPAT, or the receptor antagonist, NAN-190. Exposure of fish to 5-HTP increased whole brain 5-HT levels, while pCPA exposure decreased whole brain and splenic 5-HT. In vivo exposure of fish to pCPA depressed T- and B-lymphocyte proliferation; exposure to 5-HTP failed to alter either immune endpoint. In vitro exposure of bluegill splenocytes to 5-HT or 8-OH-DPAT inhibited lymphoproliferation; treatment with NAN-190 had no effect on immune function. Results suggest a link in bluegill between immune function and the serotonergic system. The disparity observed following in vivo- and in vitro-induced serotonergic alterations indicates the complexity of this neuro-immune relationship and emphasizes the need for further studies in this regard.