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

Abstract

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-17β (E 2) regulation of forebrain (hypothalamus and telencephalon) tyrosine hydroxylase (TH) activity in the female catfish Heteropneustes fossilis in vitellogenic phase. E 2 produced biphasic effects on TH activity: low concentrations (10 −12–10 −5M) 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 E 2 (10 −9 M) increased TH activity significantly. However, the co-incubations with a high concentration of E 2 (10 −3 M) decreased it significantly. Pre-incubations of the enzyme preparations with cAMP (0.1 mM), followed by different concentrations of E 2 (10 −12, 10 −9, 10 −4, and 10 −3 M) produced concentration-dependent biphasic effects. The pre-incubations with a low concentration of E 2 (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 E 2 (10 −3 M) produced a significant decrease in TH activity. Co-incubations of high and low E 2, with or without cAMP, and PKA inhibitor (H-89) decreased TH activity significantly. The incubations with H-89 abolished the stimulatory effect of low E 2 or low E 2 + cAMP and intensified the inhibitory effect of high E 2 or high E 2 + cAMP combination. Co-incubations with PKC inhibitor (calphostin C) did not influence the stimulatory effect of low E 2 but lowered the stimulatory effect of low E 2 + cAMP treatment. Kinetic studies showed that the stimulatory effect of a low E 2 concentration was due to a decrease in apparent K m and an increase in apparent V max for both cofactor and substrate, and the inhibitory effect of a high E 2 concentration was due to reverse changes in the kinetics. The stimulatory effect of cAMP alone or in combination with low E 2 was related to decreased K m and increased V max for the cofactor. The inhibitory effect of PKA and PKC blockers, alone or in combination with E 2 and/or cAMP was due to increased K m and decreased V max of the enzyme for the cofactor. The present data suggest that E 2 modulates the short-term activation of brain TH activity differentially and may involve mainly the cAMP-PKA system.