In Heteropneustes fossilis, ovariectomy inhibited in vivo brain (hypothalamus-pituitary, telencephalon and medulla oblongata) tyrosine hydroxylase (TH) activity with significant effects in weeks 2, 3, 4 and 5 of the gonadal resting phase and in weeks 3, 4 and 5 of the prespawning phase (P<0.05, Tukey's test). Oestradiol-17beta (OE(2)) replacement in 3-week ovariectomised fish produced biphasic responses in both seasons; the low dosages of 0.05 and 0.5 micro g/g body weight (BW) elevated TH activity, whereas the high dosages of 1.0 and 2.0 micro g/g BW decreased it. The magnitude of the inhibition was higher in the resting phase than in the prespawning phase. The inhibitory effect of ovariectomy may be produced by elevating the apparent K(m) values (decreased affinity) of the enzyme for both L-tyrosine (substrate) and dimethyltetrahydropteridine (cofactor) and consequently decreasing the V(max). Significant changes (P<0.05) in both these parameters were noticed but showed minor differences with regard to the length of ovariectomy, season or brain regions. The biphasic effects of OE(2) replacement on TH activity seemed to be produced by differential effects on apparent K(m) and V(max). The stimulatory effect of the low dosages of OE(2) coincides with a decrease in the apparent K(m) values (increased affinity) for both substrate and cofactor and an increase in the V(max) of the enzyme. The inhibitory effect of the high dosages of OE(2) correlated with an increase in the apparent K(m) values (decreased affinity) for both substrate and cofactor, and a decrease in the V(max) compared with the lower dosage groups. The results strongly suggested that OE(2) can modulate brain catecholaminergic activity at the level of tyrosine hydroxylation which, in turn, may alter gonadotrophin secretion. OE(2) may elicit biphasic effects by differentially altering the enzyme affinity towards the substrate and cofactor.