Tryptophan hydroxylase

The first step in the synthesis of 5-HT is hydroxylation of the essential amino acid, tryptophan, by the enzyme tryptophan hydroxylase (Fig. 9.4). This enzyme has several features in common with tyrosine hydroxylase, which converts tyrosine to /-DOPA in... 

Not a great deal is known about factors that actually activate tryptophan hydroxylase. In particular, the relative contribution of tryptophan supply versus factors that specifically modify enzyme activity under normal dietary conditions is unknown. However, removal of end-product inhibition of tryptophan hydroxylase has been firmly ruled out. Also, it has been established that this enzyme is activated by electrical stimulation of brain slices, even in the absence of any change in tryptophan concentration, and so other mechanisms are clearly involved. 

So far, it has been established from in vitro studies that the enzyme undergoes phosphorylation, a process that changes the conformation of the enzyme protein and leads to an increase in its activity. This involves Ca +/calmodulin-dependent protein kinase II and cAMP-dependent protein kinase which suggests a role for both intracellular Ca + and enzyme phosphorylation in the activation of tryptophan hydroxylase. Indeed, enzyme purified from brain tissue innervated by rostrally projecting 5-HT neurons, that have been stimulated previously in vivo, has a higher activity than that derived from unstimulated tissue but this increase rests on the presence of Ca + in the incubation medium. Also, when incubated under conditions which are appropriate for phosphorylation, the of tryptophan hydroxylase for its co-factor and substrate is reduced whereas its Fmax is increased unless the enzyme is purified from neurons that have been stimulated in vivo, suggesting that the neuronal depolarisation in vivo has already caused phosphorylation of the enzyme. This is supported by evidence that the enzyme activation caused by neuronal depolarisation is blocked by a Ca +/calmodulin protein kinase inhibitor. However, whereas depolarisation... 

The apparent reliance of enzyme activation on phosphorylation and intracellular Ca + gives a clue as to how the rate of 5-HT synthesis might be coupled to its impulse-evoked release. Certainly, the impulse-induced increase in intracellular Ca +, and/or activation of the G protein-coupled receptors that govern synthesis of cAMP, could modify the activity of tryptophan hydroxylase. Indeed, this could explain why activation of either somal 5-HTia autoreceptors in the Raphe nuclei (which depress the firing rate of 5-HT neurons) or terminal 5-HTib autoreceptors (which depress 5-HT release) can reduce the production of cAMP and attenuate 5-HT synthesis. 

Siuciak, JA, Clark, MS, Rind, HB, Whittemore, SR and Russo, AF (1998) BDNF induction of tryptophan hydroxylase mRNA levels in the rat brain. J. Neurosci. Res. 52 149-158. Sprague, JE, Everman, SL and Nichols, DE (1998) An integrated hypothesis for the serotonergic axonal loss induced by 3,4-methylenedioxymethamphetamine. Neuro toxicology 19 427-A42. Stock, MJ (1997) Sibutramine a review of the pharmacology of a novel anti-obesity agent. Int. J. Obesity 21 (Suppl 1) S25-S29. 

Figure 13.7 Synthesis and structure of the trace amines phenylethylamine, /)-tyramine and tryptamine. These are all formed by decarboxylation rather than hydroxylation of the precursors of the established monoamine neurotransmitters, dopamine and 5-HT. (1) Decarboxylation by aromatic L-amino acid decarboxylase (2) phenylaline hydroxylase (3) tyrosine hydroxylase (4) tryptophan hydroxylase...

QUESTION Did you measure tryptophan hydroxylase or just the 5-HT/5-HIAA depletion ... 

ANSWER We used it in the 20 mg/kg twice a day for a 4-day regimen with MDMA, and then corrected for molecular weight and used an equimolar dose of MBDB, sacrificed the animals 2 weeks later, and then measured. We used basically HPLC and used serotonin and 5-HIAA from one hemisphere and then measured tritiated pyroxetine from the other hemisphere. And we got something like 60 percent depletion of serotonin, and the pyroxetine binding site decreased by about 60 percent. With MBDB it was decreased by about 40 percent. It was a clear and significant decrease, but not quite to the extent that we had. But we have not looked at tryptophan hydroxylase. 

It is important to stress that these three eriteria must be met before neurotoxicity can be established. Similar effeets upon 5-HT levels, reuptake sites, and morphology must also be observed before it ean be concluded that 5-HT neurotoxicity has occurred. In this regard, multiple doses of METH have been shown to produee long-lasting reduetions in tryptophan hydroxylase activity (Hotchkiss et al. 1979) as well as 5-HT content and uptake sites (Rieaurte et al. 1980) in the rat brain. 

When administered in doses higher than 12 mg/kg/day, depletions of 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) last up to 6 months after cessation of drug treatment (Harvey and McMaster 1975 Harvey et al. 1977 Clineschmidt et al. 1978 Steranka and Sanders-Bush 1979 Schuster et al. 1986 Kleven et al. 1988). Other long-lasting effects of fenfluramine include a decrease in 5-HT uptake sites (Schuster et al. 1986) and tryptophan hydroxylase activity (Steranka and Sanders-Bush 1979). 

Hotchkiss, A.J. Morgan, M.E. and Gibb, J.W. The long-term effects of multiple doses of methamphetamine on neostriatal tryptophan hydroxylase, tyrosine hydroxylase, choline acetyltransferase and glutamate decarboxylase activities. Life Sci 25 1373-1378. 1979. 

Schmidt, C.J., and Taylor, V.L. Depression of rat brain tryptophan hydroxylase activity following the acute administration of methylenedioxymethamphetamine. Biochem Pharmacol 36 4095-4 102, 1987. 

Taylor, D., and Ho, B.T. Effect of short- and long-term treatment with cocaine on rat brain tryptophan hydroxylase. Res Commun Chem Pathol Pharmacol 15 805-808, 1976. 

QUESTION You have dopamine reuptake blockade, you have the SCH 23390 bloeking, speeifieally blocking this tryptophan hydroxylase effect. 

I have no doubt that the bearing on these data is that you have shown the pharmacological effects on tyrosine and tryptophan hydroxylase activity. I am not sure that I can equate those with effects on actual neurotoxicity. 

Bakhit, C., and Gibb, J.W. Methamphetamine-induced depression of tryptophan hydroxylase Recovery following acute treatment. Eur J Pharmacol 76 229-233, 1981. 

This similarity between MDMA and PCA is also observed in vivo in that PCA produces both an acute and long-term depletion of 5-HT (Fuller et al. 1975 Steranka et al. 1977). Like PCA, the acute decrease in 5-HT concentrations produced by MDMA is associated with a decrease in the activity of the rate-limiting enzyme for 5-HT synthesis, tryptophan hydroxylase (TPH). The timecourse of this change in cortical enzyme activity is also shown in figure 1. More detailed analysis of this acute effect of MDMA and kinetic analysis of TPH activity reveals that the decrease in enzyme activity actually precedes the decline in transmitter levels and is due to a reduction in the activity of the enzyme (Schmidt and Taylor 1987 Schmidt and Taylor 1988). As shown for the cortex in figure 3, the decrease in 5-HT... 

Knapp, S. Mandell, A.J. and Geyer, M.A. Effects of amphetamines on regional tryptophan hydroxylase aetivity and synaptosomal conversion of tryptophan to 5-hydroxytryptamine in rat brain. J Pharmacol Exp Ther 189 676-689, 1974. 

Ross, S.B., and Froden, O. On the mechanism of the acute decrease of rat brain tryptophan hydroxylase activity by 4-chloroamphetamine. Neurosci Lett 5 215-220, 1977. 

Typically, neurotoxic effects of drugs on monoamine neurons have been assessed from reductions in brain levels of monoamines and their metabolites, decreases in the maximal activity of synthetic enzymes activity, and decreases in the active uptake carrier. In the present study, the traditional markers described above have been used, including the measurement of the content of monoamines and their metabolites in brain at several different timepoints following drug administration. Since reports in the literature have documented that MDMA and MDA can inhibit the activity of tryptophan hydroxylase (TPH), the rate-limiting enzyme in serotonin synthesis (Stone et al. 1986 Stone et al. 1987). it is unclear whether MDMA-induced reductions in the content of serotonin and its metabolite 5-hydroxyin-doleacetic acid (5-HlAA) may be due to suppressed neurotransmission in otherwise structurally intact serotonin neurons or may represent the eonsequenee of the destruction of serotonin neurons and terminals. 

Sanders-Bush, E. Bushing, J.A. and Sulser, F. Long-term effects of p-chloroamphetamine on tryptophan hydroxylase activity and on levels of 5-hydroxytryptamine and 5-hydroxyindole aeetie acid in brain. 

A second approach might be to measure dopamine and serotonin along with their metabolites and other specific neuronal constituents such as tyrosine hydroxylase and tryptophan hydroxylase or uptake carrier sites in brain tissue obtained at autopsy. Accumulating data in this way might be a slow and tedious process, and drug dosing history might be uncertain and variable nonetheless, the approach deserves consideration. 

A fourth approach to evaluating the intactness of dopamine and/or serotonin neurons in human subjects who have taken one of the amphetamine analogs might be to use a probe for labeling a constituent of those neurons in position emission tomography scanning studies. A label for the serotonin or dopamine uptake carrier, or a label for tryptophan hydroxylase or tyrosine... 

Yoshida, K. et al. (2002). Monoamine oxidase a gene polymorphism, tryptophan hydroxylase gene polymorphism and antidepressant response to fluvoxamine in Japanese patients with major depressive disorder. Prog. Neuropsychopharmacol. Biol. Psychiatry, 26, 1279-83. 

Boularand, S., Darmon, M.C., Ganem, Y., Launay, J.M. Mallet J. (1990). Complete coding sequence of human tryptophan hydroxylase. Nucleic Acids Res. 18(14), 4257. 

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