A series of 'in vivo' and 'in vitro' approaches were used to investigate the regulation of testis function by immune-derived factors in the goldfish. Administration of lipopolysaccharide (LPS) ' in vivo' lead to a significant decrease in plasma testosterone (T) levels of goldfish exhibiting a low gonadosomatic index (GSI) value. The murine cytokines, tumor necrosis factor [alpha] (TNF[alpha]) and interleukin-1[beta] (IL-1[beta]), LPS, and teleost macrophage conditioned media (MCM) inhibited human chorionic gonadotropin (hCG)-stimulated T production by goldfish testis pieces 'in vitro'. Basal T levels were affected more variably by the compounds. The predominant site of activity of TNF[alpha] in the gonadotropin-stimulated steroid biosynthetic pathway appears to be post-cAMP generation but prior to the conversion of pregnenolone to other sterol precursors. TNF- and IL-1-like peptides were immunologically detectable in LPS-stimulated RTS11 cell culture supernatants but the same mammalian antibodies failed to ablate the activity of the MCM on T levels 'in vitro' or detect TNF- and IL-1-like peptides in the tissues of male goldfish. Collectively, these studies provide support for the involvement of cytokines and possibly other immune-derived factors in the regulation of teleost testicular steroidogenesis.