Estrogen regulation of in vitro brain tyrosine hydroxylase activity in the catfish Heteropneustes fossilis: interactions with cAMP-protein kinase A and protein kinase C systems in enzyme activation.

In the present in vitro study, interactions of both cAMP-protein kinase A (PKA) and protein kinase C (PKC) systems were investigated in the estradiol-17beta (E2) regulation of forebrain (hypothalamus and telencephalon) tyrosine hydroxylase (TH) activity in the female catfish Heteropneustes fossilis in vitellogenic phase. E2 produced biphasic effects on TH activity: low concentrations (10(-12)-10(-5) M) stimulated, and high concentrations (10(-3)-10(-4) M) inhibited enzyme activity (Tukey's test, P<0.05). Co-incubations of the enzyme preparations with cAMP (1.0 mM), IBMX (1.5 mM) or theophylline (1.5 mM) and a low concentration of E2 (10(-9) M) increased TH activity significantly. However, the co-incubations with a high concentration of E2 (10(-3) M) decreased it significantly. Pre-incubations of the enzyme preparations with cAMP (0.1 mM), followed by different concentrations of E2 (10(-12), 10(-9), 10(-4), and 10(-3) M) produced concentration-dependent biphasic effects. The pre-incubations with a low concentration of E2 (10(-9) M), followed by different concentrations of cAMP (0.05-1.0 mM) produced a significant concentration-dependent stimulation of TH activity and that with a high concentration of E2 (10(-3) M) produced a significant decrease in TH activity. Co-incubations of high and low E2, with or without cAMP, and PKA inhibitor (H-89) decreased TH activity significantly. The incubations with H-89 abolished the stimulatory effect of low E2 or low E2+cAMP and intensified the inhibitory effect of high E2 or high E2+cAMP combination. Co-incubations with PKC inhibitor (calphostin C) did not influence the stimulatory effect of low E2 but lowered the stimulatory effect of low E2+cAMP treatment. Kinetic studies showed that the stimulatory effect of a low E2 concentration was due to a decrease in apparent Km and an increase in apparent Vmax for both cofactor and substrate, and the inhibitory effect of a high E2 concentration was due to reverse changes in the kinetics. The stimulatory effect of cAMP alone or in combination with low E2 was related to decreased Km and increased Vmax for the cofactor. The inhibitory effect of PKA and PKC blockers, alone or in combination with E2 and/or cAMP was due to increased Km and decreased Vmax of the enzyme for the cofactor. The present data suggest that E2 modulates the short-term activation of brain TH activity differentially and may involve mainly the cAMP-PKA system.