Nesting male demoiselles Chromis dispilus were captured and blood sampled under water by divers, and plasma levels of reproductive hormones correlated with behavioural status at the time of capture. Plasma gonadotropin-II (dGtH-II) was measured using a heterologous assay based on carp GtH-II standards and an antibody to the ߭subunit of cGtH-II. Plasma samples diluted parallel to standards in the assay, and there was a significant transient increase in dGtH-II in response to treatment with gonadotropin releasing hormone analogue (GnRHa). Male demoiselles had elevated plasma levels of dGtH-II during periods of reproductive display, but low levels of dGtH-II during periods of brooding, irrespective of egg stage. Plasma testosterone levels followed a similar pattern whereas plasma 11-ketotestosterone and 17,20߭dihydroxy-4-pregnen-3-one levels tended to remain elevated during the early phases of egg brooding. The results are consistent with earlier studies showing that reproductive hormone levels in demoiselles are behaviourally mediated, and support the view that behavioural alterations of reproductive endocrine status in fishes are mediated through the action of GnRH.