Rise In Tyrosine Hydroxylase Activity Research Articles

20‐hydroxyecdysone and juvenile hormone influence tyrosine hydroxylase activity in Drosophila females under normal and heat stress conditions

The effects of exogenous 20-hydroxyecdysone (20E) and the juvenile hormone (JH) on the activity of the tyrosine hydroxylase (TH), the first and rate-limiting DA biosynthetic enzyme, has been studied in young females of wild type D. virilis and D. melanogaster under normal conditions and under heat stress (38 degrees C). Both 20E feeding of the flies and JH application led to a substantial rise in TH activity. A rise in JH and 20E levels was found not to prevent the response of TH to heat stress, but to change the intensity of its response to the stress exposure. Putative mechanisms of regulation of DA level by 20E and JH in Drosophila females are discussed.

Modification of the expression of adenosine 3′,5′-cyclic monophosphate-induced differentiated functions in neuroblastoma cells by beta-carotene and D-alpha-tocopheryl succinate.

The role of beta-carotene and vitamin E in modifying the effect of cell differentiating agent has not been studied. This study has investigated the effects of beta-carotene and d-alpha-tocopheryl succinate (alpha-TS) on adenosine 3',5'-cyclic monophosphate (cAMP) induced differentiated functions in murine neuroblastoma cells (NBP2) in culture. Prostaglandin E1 (PGE1), a stimulator of adenylate cyclase, and 4-(3-butoxy-4-methoxy-benzyl)-2-imidazolidinone (R020-1724), an inhibitor of cyclic nucleotide phosphodiesterase, were used to induce differentiation in NB cells. Both beta-carotene and alpha-TS markedly enhanced the level of morphologic differentiation (neurite formation) induced by both PGE1 and R020-1724. However, beta-carotene and alpha-TS by themselves were ineffective. These vitamins increased tyrosine hydroxylase (TH) activity. However, beta-carotene did not significantly affect PGE1- and R020-1724-stimulated rise in TH activity. alpha-TS at a higher concentration inhibited PGE1- and R020-1724-stimulated increase in TH activity. None of the above treatments affected basal choline acetyltransferase (ChAT) activity. beta-carotene and alpha-TS caused a transient increase in cAMP level, and they also enhanced the effect of PGE1 and R020-1724 on cAMP level in a transient manner. These results suggest that beta-carotene and alpha-TS modify the effects of cAMP stimulating agents on differentiation of NB cells in culture.

Expression and regulation of tyrosine hydroxylase in adult sensory neurons in culture: Effects of elevated potassium and nerve growth factor

To determine whether similar molecular mechanisms regulate the same proteins in diverse neuronal populations, the present study compared regulation of tyrosine hydroxylase (TOH) in placodal sensory and neural crest-derived sympathetic neurons in tissue culture. Long-term explant cultures of adult nodose and petrosal sensory ganglia (NPG) contained abundant TOH-immunoreactive neurons and exhibited TOH catalytic activity, as in vivo. After an initial decline during the first week of culture, enzyme activity was maintained at a stable plateau of 60% of zero time values for at least 3 weeks. However, exposure of 2-week-old cultures to depolarizing concentrations of potassium (K +; 40 mM) increased TOH activity approximately two-fold; total protein was unchanged, suggesting that the rise was due to increased TOH specific activity. Therefore, membrane depolarization in vitro appears to regulate this specific catecholaminergic (CA) trait in sensory, as in sympathetic CA cells. In sympathetic neurons, NGF regulates TOH activity throughout life. In marked contrast, TOH activity in adult NPG cultures was unchanged in the presence of 0, 10 or 100 units NGF/ml or in the presence of high concentrations of antiserum against the β-subunit of NGF. Adult sympathetic neurons, however, grown under identical conditions, exhibited a 5- to 10-fold rise in TOH activity in the presence of NGF. Thus, unlike sympathetics, CA metabolism in adult NPG neurons is not regulated by NGF in vitro; NGF is therefore unlikely to mediate target effects on CA metabolism in placodal sensory neurons in vivo. Our findings indicate that certain mechanisms of CA regulation are shared by placodal sensory and neural crest-derived sympathetic neurons, whereas others are not. Therefore, expression of the CA phenotype in peripheral neurons does not imply an obligatory repertoire of regulatory mechanisms.

Depolarizing stimuli increase tyrosine hydroxylase in the mouse locus coeruleus in culture

The influence of membrane depolarization on the development and regulation of brain noradrenergic neurons was studied in explant cultures of the mouse locus coeruleus (l.c.). Exposure to the depolarizing agents veratridine or elevated K + significantly increased the catalytic activity of tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine biosynthesis. The effects of veratridine were prevented by tetrodotoxin, suggesting that transmembrane Na + influx was necessary for the rise in TH. Morphometric analysis indicated that the rise in TH activity was not accompanied by altered TH-positive cell number or cell diameter. Rather, TH fluorescence intensity increased in each neuron, suggesting that depolarization increased TH per neuron. Immunoblot and densitometric analysis indicated that depolarization did, indeed, increase TH immunoreactive protein. Moreover, depolarization elevated enzyme activity in cultured neurons expressing the normal developmental increase in TH, as well as those in which plateau levels had already been attained. We conclude that depolarization and/or Na + influx regulates a critical transmitter macromolecule in brain neurons, as in the periphery, by altering enzyme molecule number.

Trans-synaptic increase in RNA coding for tyrosine hydroxylase in a rat sympathetic ganglion

To begin examining molecular mechanisms underlying trans-synaptic regulation, tyrosine hydroxylase (TH) and its messenger RNA (mRNA) were examined in the superior cervical sympathetic ganglion (SCG) of adult rats. Basal levels of TH mRNA were detectable in control ganglia by RNA dot hybridization, using the 32P nick translated PstI-KpnI restriction fragment of pTH.4 as a probe. Reserpine induced a 3-fold rise in TH activity per μg protein, and a simultaneous 3-fold increase in ganglion TH mRNA. As expected, ganglion decentralization (denervation) prevented the trans-synaptic induction of TH. In addition, decentralization prevented the increase in TH mRNA, suggesting that the increase in message was dependent on trans-synaptic stimulation. Northern blot analysis indicated that the cDNA (complementary DNA) probe hybridized to a single band of approximately 1900 nucleotides, which was the same size in all ganglia. Our observations indicate that induction of TH is associated with a trans-synaptic increase in mRNA coding for the enzyme. Consequently, trans-synaptic increases in impulse activity may induce TH by increasing neuronal levels of TH mRNA in the SCG.

Development of catecholaminergic phenotypic characters in the mouse locus coeruleus in vivo and in culture

While abundant studies have begun to elucidate ontogeny of the peripheral nervous system, molecular mechanisms underlying brain development remain obscure. To approach this problem, we initiated parallel in vivo and in vitro studies of the mouse locus coeruleus (l.c.), a brainstem noradrenergic nucleus. The catecholaminergic enzymes tyrosine hydroxylase (TH) and dopamine-β-hydroxylase (DBH) were used to monitor phenotypic expression and development. TH catalytic activity and immunocytochemical reactivity were initially detectable on gestational Day 13 (E-13) in vivo, and adult levels of activity were approximated by the third postnatal week. Immunotitration studies indicated that the developmental increase was due to accumulation of enzyme molecules and not enzyme activation. The in vivo developmental profile of DBH approximated that of TH. To begin defining regulatory mechanisms, explants of embryonic brainstem were placed in culture. Explantation on E-12, prior to expression of TH or DBH, resulted in the de novo appearance of these phenotypic characters after 4 days. Explantation on E-18, after the enzymes are already expressed, was followed by a striking sixfold rise in TH activity. Immunotitration studies revealed that the increase in TH activity in E-18 cultures was attributable to increased molecule number, reproducing the in vivo results. Moreover, the E-18 explants, cultured for 3 weeks, attained higher plateau levels of TH activity than E-12 cultures, and this difference was due to increased molecule number. Morphometric analysis indicated that 3-week E-12 cultures actually had more l.c. cells than E-18 cultures, indicating that differences in TH were not due to increased cells in the E-18 l.c. Finally, systematic study revealed that the development of TH activity in culture increased progressively from E-11 to E-12 to E-13, suggesting that critical regulatory events occur at this time. Our studies suggest that the l.c. is an excellent model for the study of brain development in vivo and in vitro. Initial phenotypic expression and dramatic development occur in culture in the absence of normal targets, normal afferent innervation and, presumably, normal humoural milieu.

Effects of MK-771, a novel TRH analog, on brain dopaminergic and serotonergic systems

The effect of acute treatment with MK-771, a thyrotropin-releasing hormone analog, has been investigated on the synthesis and release of dopamine and 5-hydroxytryptamine in rat brain. Single injection of MK-771 in doses of 10 and 15 mg/kg produced a rise in tyrosine hydroxylase activity as well as homovanillic acid level in striatum (a site of nigrostriatal dopamine system) and olfactory tubercles (a site of mesolimbic dopamine system). The dopamine levels remained unchanged (olfactory tubercles, hippocampus) or decreased (striatum, hypothalamus, pons-medulla) after MK-771 treatment. Acute MK-771 injection at the dose of 15 mg/kg increased mid-brain tryptophan hydroxylase activity and tryptophan levels by 26% and 55%, respectively. A consistent increase in the levels of 5-hydroxytryptamine and its metabolite 5-hydroxyindoleacetic acid was reported in several brain areas. These data suggest that MK-771 elicits a marked increase in the synthesis and turnover of brain dopamine and 5-hydroxytryptamine.

Lack of effect of chronically administered thyrotropin-releasing hormone (TRH) on regional rat brain tyrosine hydroxylase activity

Chronic treatment of adult male rats with TRH (1 or 10 mg/kg IP) for 5 or 9 days failed to alter the activity of tyrosine hydroxylase (TH), the enzyme regulating the rate-limiting step in catecholamine biosynthesis. In contrast, as previously described, chronic reserpine administration (0.5 mg/kg IP: 9 days) resulted in a significant rise in TH activity in midbrain, hypothalamus, pons-medulla and forebrain. These results suggest that the enhanced brain norepinphrine turnover reported to occur after treatment with TRH is not due to synthesis of new TH enzyme protein.

Mechanisms of tyrosine hydroxylase and dopamine beta-hydroxylase induction in organ cultures of rat sympathetic ganglia by potassium depolarization and cholinomimetics.

It was the aim of the present study to elucidate the mechanisms involved in specific tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) induction by potassium depolarization and cholinomimetics in rat superior cervical ganglia kept in organ culture. The effect of high (54 mM) potassium concentration on intact ganglia seems to result in a dual action: a) a specific induction of TH and DBH via release of acetylcholine from preganglionic cholinergic nerve terminals. b) a non-specific effect on terminal adrenergic neurons resulting in a general increase of protein synthesis as indicated by the increase in DOPA decarboxylase (DDC) and monoamine oxidase (MAO) activities. In decentralized superior cervical ganglia potassium depolarization failed to produce the specific TH and DBH induction although a small increase in DDC activity persisted. Carbamylcholine, acetylcholine and nicotine at concentrations of 10(-4) M elicited a selective induction of TH and DBH both in intact and decentralized ganglia via nicotinic receptor stimulation. Bethanechol, predominantly stimulating muscarinic receptors had no significant effect on TH activity. A 4 h pulse of 10(-4) M carbamylcholine produced optimal induction of DBH and TH 24 h and 48 h later respectively. Longer exposure to carbamylcholine resulted in a significantly smaller rise in TH activity.

ORGAN CULTURE OF THE RAT ADRENAL MEDULLA: A MODEL SYSTEM FOR THE STUDY OF TRANS‐SYNAPTIC ENZYME INDUCTION

Abstract—It was the aim of the present study to develop organ culture conditions for the rat adrenal medulla which are representative for the in vivo situation. This is a prerequisite for studying the complex processes involved in trans‐synaptic enzyme induction. The processes of trans‐synaptic enzyme induction initiated in vivo by injecting 5 mg/kg of reserpine 2 h prior to the removal of the adrenal medulla, continued in this culture system and final levels of tyrosine hydroxylase were comparable to those seen in vivo. That these culture conditions are representative for the in vivo induction is also supported by the fact that transection of the splanchnic fibres supplying the adrenal medulla or administration of actinomycin D prior to reserpine abolished the rise in tyrosine hydroxylase activity not only in vivo, but also in culture.The findings that high concentrations (0·29 mm) of corticosterone in the culture medium inhibited the increase in tyrosine hydroxylase activity caused by reserpine support the hypothesis that glucocorticoids act as modulatory agents in trans‐synaptic enzyme induction. This inhibition was exhibited only when corticosterone was added at the initiation of the culture period. If added 2 or 4 h after the beginning of the culture period there was little or no effect on the subsequent increase of tyrosine hydroxylase.

Action of 6-hydroxydopamine on lamb sympathetic ganglia, vas deferens and adrenal medulla: a combined histochemical, ultrastructural and biochemical comparison with the effects of reserpine.

1. The effects of a single dose of 6-hydroxydopamine (6-OHDA) compared with those of chronic reserpine treatment were studied in lamb sympathetic neurones and adrenal medulla by a combination of fluorescence histochemistry, electron microscopy and radiochemical assay.2. In sympathetic ganglia, 6-OHDA produced a rise in noradrenaline concentration within 24 h, and falls in tyrosine hydroxylase and monoamine oxidase activities, whereas reserpine caused a fall in noradrenaline, a rise in tyrosine hydroxylase activity and no change in monoamine oxidase activity. The fluorescence of intra- and postganglionic axons increased greatly within 24 h of 6-OHDA, and there was a corresponding accumulation of large dense-core vesicles within many axons whose neurotubules were disrupted. The changes were almost reversed after 3 weeks.3. In the vas deferens, the concentration of noradrenaline and tyrosine hydroxylase and monoamine oxidase activities had all fallen 24 h after 6-OHDA treatment and had started to recover 3 weeks later. In the adrenal medulla, 6-OHDA did not alter NA concentrations but increased tyrosine hydroxylase activity whereas reserpine depleted noradrenaline and increased tyrosine hydroxylase activity.4. The changes produced in sympathetic ganglia by 6-OHDA may be due both to a direct action on the axoplasmic transport of noradrenaline containing vesicles and indirectly to the reaction of the neurones to loss of the integrity of their axons.