Electrical stimulation increases phosphorylation of tyrosine hydroxylase in superior cervical ganglion of rat.

Abstract

Electrical stimulation of the superior cervical ganglion of the rat increased the phosphorylation of tyrosine hydroxylase (tyrosine 3-monooxygenase, EC 1.14.16.2) in this tissue. Ganglia were incubated with [32pJp1 for 90 min and were then electrically stimulated via the preganglionic nerve. Tyrosine hydroxylase was isolated from homogenates of the ganglia by immunoprecipitation followed by polyacrylamide gel electrophoresis. 32P-labeled tyrosine hydroxylase was visualized by radioautography, and the incorporation of 32p into the enzyme was quantitated by densitometry of the radioautograms. Stimulation of ganglia at 20 Hz for 5 min increased the incorporation of 32p into tyrosine hydroxylase to a level 5-fold that found in unstimulated control ganglia. The increase in phosphorylation of tyrosine hydroxylase was dependent on the duration and frequency of stimulation. Preganglionic stimulation did not increase the phosphorylation of tyrosine hydroxylase in a medium that contained low Ca2+ and high Mg2 . Increases in phosphorylation were reversible; within 30 min after the cessation of stimulation, the incorporation of 32p into tyrosine hydroxylase decreased to the level found in unstimulated ganglia. The nicotinic antagonist hexamethonium reduced the increase in 32p incorporation into tyrosine hydroxylase by about 50%, while the muscarinic antagonist atropine had no effect. Thus, preganglionic stimulation appeared to increase the phosphorylation of tyrosine hydroxylase in part by a nicotinic mechanism and in part by a noncholinergic mechanism. Antidromic stimulation of ganglia also increased the phosphorylation of tyrosine hydroxylase. Two-dimensional gel electrophoresis revealed that electrical stimulation also increased the incorporation of 32p into at least six other phosphoproteins in the ganglion.Electrical stimulation of sympathetic nerves increases the rate of catecholamine biosynthesis in these nerves (1). This acceleration of catecholamine synthesis was initially thought to be due to the release of catecholamines from the nerve terminals, resulting in a decrease in intraneuronal catecholamine levels and a relief of the feedback inhibition of tyrosine hydroxylase (tyrosine 3-monooxygenase, EC 1.14.16.2) by catecholamines (2). It is now clear, however, that stimulation of both peripheral and central adrenergic nerves causes a stable activation of tyrosine hydroxylase and a change in the kinetic properties of the enzyme (most commonly, a decrease in the apparent Km for pteridine cofactor and an increase in the Ki for catecholamines) (3-6). There is evidence that the activation of tyrosine hydroxylase may be mediated by phosphorylation of the enzyme. Tyrosine hydroxylase is a phosphoprotein (7) and can be activated in vitro by phosphorylation (8-12). Moreover, phosphorylation of tyrosine hydroxylase is increased in situ in adrenal chromaffin cells by stimuli that increase the rate of catecholamine synthesis in these cells (13,14).Preganglionic stimulation increases tyrosine hydroxylase activity in the...

Protein Phosphorylation and Neuronal Function

“…Subsequently, treatment of intact catecholaminergic cells with either cyclic AMP analogues or secretagogues was shown both to phosphorylate and to activate tyrosine hydroxylase in association with an acceleration of catecholamine biosynthesis (Meligeni et ai., 1982;Haycock et al, 1982~). To date, stimulation-dependent phosphorylation of tyrosine hydroxylase has been demonstrated in a number of intact tissues, including vas deferens, adrenal medulla, PC-12 cells, superior cervical ganglion, and intact and ''leaky'' adrenal chromaffin cells Meligeni et al, 1981;Cahill and Perlman, 1984;Yanagihara et al, 1984;Niggli et al, 1984). Taken together, the data strongly suggested that cyclic AMP-dependent phosphorylation of tyrosine hydroxylase is an important process in regulating catecholamine biosynthesis.…”

Dopamine Autoreceptors Modulate the Phosphorylation of Tyrosine Hydroxylase in Rat Striatal Slices

“…52, 1517-1522 (1989). (Lee et al, 1985;McTigue et al, 1985;Tachikawa et al, 1987), superior cervical ganglia (Cahill and Perlman, 1984), and CNS fragments or synaptosomes (Porter, 1986;Haycock, 1987). The phosphorylation of TH is consistently associated with an increase in enzyme activity (for review, see Goldstein and Greene, 1987).…”

Predicted Amino Acid Sequence of Bovine Tyrosine Hydroxylase and Its Similarity to Tyrosine Hydroxylases from Other Species

“…Tyrosine hydroxylase [TH; tyrosine 3-monooxygenase; L-tyrosine tetrahydr0pteridine:oxygen oxidoreductase (3-hydroxylating); EC 1.14.16.21 is the rate-limiting enzyme in the biosynthesis of catecholamines (Levitt et al, 1965;Molinoff and Axelrod, 1971;Kaufman and Kaufman, 1985). Besides the regulation of TH activity by posttranslational phosphorylation (Cahill and Perlman, 1984;Campbell et al, 1986;Cremins et al, 1986; for review, see Kaufman and Kaufman, 1989, TH synthesis is regulated by neuronal activity (transsynaptic induction) (Thoenen et al, 1969(Thoenen et al, , 1979Joh et al, 1973;Thoenen, 1975;Zigmond, 1985), nerve growth factor (Thoenen et al, 1971(Thoenen et al, , 1979(Thoenen et al, , 1985Acheson et al, 1984), glucocorticoids (Otten and Thoenen, 1975;Thoenen and Otten, 1978;Thoenen et al, 1979), laminin , and cell-cell contact (Acheson and Thoenen, 1983;Saadat and Thoenen, 1986;Saadat et al, 1987).…”