Acute inactivation of tryptophan hydroxylase by amphetamine analogs involves the oxidation of sulfhydryl sites

doi:10.1016/0922-4106(89)90048-5

European Journal of Pharmacology: Molecular Pharmacology

Volume 172, Issue 1, 7 March 1989, Pages 93-97

https://doi.org/10.1016/0922-4106(89)90048-5 Get rights and content

Abstract

The activity of rat hippocampal tryptophan hydroxylase was reduced from 30–60% 3 h after the administration of a 10–15 mg/kg dose of either fenfluramine, methamphetamine or 3,4-methylenedioxymethamphetamine (MDMA). Tryptophan hydroxylase inactivated by these drug treatments could be reconstituted by a prolonged anaerobic incubation in the presence of 5 mM dithiothreitol and 50 μM Fe²⁺. Drug-inactivated enzyme obtained from rats killed 18 h after multiple doses of either D(+)- or L(−)-MDMA could not be similarly restored. These observations suggest that the rapid decrease in central tryptophan hydroxylase activity induced by amphetamine analogs results from the reversible oxidation of a sulfhydryl site(s) within the enzyme molecule, whereas the irreversible decrease in enzymatic activity measured 18 h after multiple-dose MDMA treatment may reflect serotonergic toxicity.

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Cited by (63)

Tryptophan hydroxylase and serotonin synthesis regulation
2020, Handbook of Behavioral Neuroscience
Citation Excerpt :
This return of TPH2 activity was not due to new protein synthesis because the recovery of TPH2 activity was not prevented by the protein synthesis inhibitor cyclohexamide (Stone et al., 1989a). These investigators concluded that MDMA and related drugs inhibit TPH2 by oxidizing key cysteine sulfhydryl groups in the enzyme and, in a time dependent manner, this recovery becomes irreversible and probably represents another process (Stone et al., 1989a, 1989b). In a more recent study, recombinant TPH2 was tested in vitro for its response to oxidation by hydrogen peroxide and diamide, and the results indicated that TPH2 activity was inhibited by both compounds, leading to the formation of high molecular weight aggregates in a DTT-reversible manner (Kuhn, Sykes, Geddes, Jaunarajs, & Bishop, 2011).
Tryptophan hydroxylase (TPH) is the rate-limiting enzyme in the biosynthesis of the neurotransmitter serotonin (5-HT). Increases or decreases in TPH catalytic activity can therefore lead to corresponding changes in the neuronal content of 5-HT. TPH requires a reduced pteridine cofactor, molecular oxygen, and nonheme iron to hydroxylate its substrate l-tryptophan. Alterations in the tissue content of any of these four factors could change TPH activity. Posttranslational modifications of TPH, including phosphorylation and cysteine oxidation, cause significant changes in its catalytic activity. The discovery of a brain-specific isoform of TPH, termed TPH2 to distinguish this enzyme from the peripheral form TPH1, opened new avenues of research that allowed in-depth studies of TPH molecular function, solutions of 3D crystal structures, creation of TPH1 and TPH2 knockout mouse lines, and the successful search for specific inhibitors of TPH isoforms some of which are entering clinical trials for the treatment of disorders linked to excess peripheral 5-HT.
Effects of long-term exposure of 3,4-methylenedioxymethamphetamine (MDMA; "ecstasy") on neuronal transmitter transport, brain immuno-regulatory systems and progression of experimental periodontitis in rats
2014, Neurochemistry International
Citation Excerpt :
Single or repeated injections of MDMA can cause long-lasting changes in neurochemical and histological markers of 5-HTergic function in rat (Schmidt et al., 1986, 1987; Aguirre et al., 1998; Barrionuevo et al., 2000; Schenk et al., 2011; Do and Schenk, 2013) and human brain (McCann et al., 2005). The neurotoxicity is characterized by a decline in the activity of tryptophan hydroxylase (Stone et al., 1989), a decrease in the brain tissue levels of 5-HT (Aguirre et al., 1995; Schenk et al., 2011; Do and Schenk, 2013), reduced number of SERT + fibres in rats (Lopez-Rodriguez et al., 2014), and a lower density of [3H]paroxetine-labeled 5-HT transporters (SERT) in several regions of the rat brain (Battaglia et al., 1987; Hervías et al., 2000). The latter has also been confirmed by nuclear imaging studies (PET or SPECT) in humans displaying a lower density of brain SERT sites and a reduction in their function in MDMA users (McCann et al., 1998, 2005; Ricaurte et al., 2000; Buchert et al., 2003, 2004; Erritzoe et al., 2011).
The present study was designed to investigate the effects of long-term exposure (4 weeks) to the widely used narcotic drug and putative neurotoxicant 3,4-methylenedioxymetamphetamine (MDMA; “ecstasy”) on neuronal transmitter transport and progression of experimental periodontitis in male Wistar rats. The rats were exposed to MDMA (10 mg/kg/day i.p.) or saline five days a week for four consecutive weeks. Exposure to MDMA induced a significant reduction in the synaptosomal reuptake of serotonin, while the uptake of dopamine was significantly increased 24 h after the last injection of MDMA. In contrast, the synaptosomal uptake of noradrenaline and the vesicular uptake through the vesicular monoamine transporter 2 were not affected. In the experiments of periodontitis development, ligature-induced periodontitis was induced three days prior to MDMA administration. Compared to controls, MDMA-treated rats developed significantly more periodontitis. In conclusion, our results show that long-term exposure to MDMA affects the serotonergic and dopaminergic transport systems in the rat brain and increased the susceptibility to the psychosomatic ailment periodontitis following disturbances of brain immune-regulatory systems. These results are interesting with respect to recent research showing that changes in neurotransmitter signalling may alter the reactivity of brain-controlled immunoregulatory systems controlling pathogenic microorganisms colonizing mucosal surfaces.
Prior methamphetamine self-administration attenuates serotonergic deficits induced by subsequent high-dose methamphetamine administrations
2012, Drug and Alcohol Dependence
Citation Excerpt :
This suggestion is based, at least in part, on findings by Bowyer et al. (2008) that METH-induced 5HT depletions within the striatum occurred only in hyperthermic mice maintained in a warm environment (Bowyer et al., 2008). METH-induced hyperthermia promotes the formation of oxygen radicals and reduces tryptophan hydroxylase activity (Fleckenstein et al., 1997; see also Stone et al., 1989), which may contribute to oxidative damage (Tata and Yamamoto, 2007; Yamamoto et al., 2010). Additionally, increased temperature has been shown to increase human SERT uptake of [3 H]DA in HEK-293 cells, which may further promote oxidative damage (Saldaña and Barker, 2004).
Pre-clinical studies indicate that high-dose, non-contingent methamphetamine (METH) administration both rapidly and persistently decreases serotonergic neuronal function. Despite research indicating the hippocampus plays an important role in METH abuse and is affected by METH use, effects of METH self-administration on hippocampal serotonergic neurons are not well understood, and were thus an important focus of the current study. Because humans often administer METH in a binge-like pattern, effects of prior METH self-administration on a subsequent “binge-like” METH treatment were also examined.
Rats were treated as described above, and sacrificed 1 or 8 d after self-administration or 1 h or 7 d after the final binge METH or saline exposure. Hippocampal serotonin (5-hydroxytryptamine; 5HT) content and transporter (SERT) function were assessed.
METH self-administration per se had no persistent effect on hippocampal 5HT content or SERT function. However, this treatment attenuated the persistent, but not acute, hippocampal serotonergic deficits caused by a subsequent repeated, high-dose, non-continent METH treatment administered 1 d the last self-administration session. No attenuation in persistent deficits were seen when the high-dose administration of METH occurred 15 d after the last self-administration session.
The present findings demonstrate that METH self-administration alters serotonergic neurons so as to engender “tolerance” to the persistent serotonergic deficits caused by a subsequent METH exposure. However, this “tolerance” does not persist. These data provide a foundation to investigate complex questions including how the response of serotonergic neurons to METH may contribute to contingent-related disorders such as dependence and relapse.
Tryptophan Hydroxylase and Serotonin Synthesis Regulation
2010, Handbook of Behavioral Neuroscience
Citation Excerpt :
A similar protocol was used for ‘activation’ of the brain enzyme. Activation (or recovery of lost activity) with the brain TPH required a prolonged incubation, 20–24h, under similar anaerobic conditions (Hamon et al., 1978; Kuhn et al., 1980b; Stone et al., 1989a). These features of TPH activation (or restoration) suggested a homology between TPH1 and TPH2, but at the same time we noted a great difference in the rate of the process, presumably due to their respective iron-binding affinities, which differed by more than two orders of magnitude.
Potential determinants of existing tryptophan hydroxylase (TPH) activity in vivo are (1) a supply of tryptophan via the tryptophan transporter, (2) de novo synthesis of tetrahydrobiopterin and efficient regeneration of this coenzyme, and (3) iron metabolism. TPH is an iron protein. Iron binding is relatively strong in TPH2, while it is very weak in TPH1. Owing to the poorly liganded iron of TPH, various complexities arise. Irreversible inactivation of TPH and neural deterioration seem to be caused by an H₂O₂ reaction with non-liganded or poorly liganded Fe²⁺. Within the cell, the amount of ferrous iron available to TPH is below saturation. The metabolism of iron may control serotonin production by attenuating actual TPH activity in the cell interior. Phosphorylation of TPH was studied in terms of its functional aspects with the use of brain TPH, while the identification of phosphorylation sites was mainly conducted on the basis of the amino acid sequence of TPH1. Protein turnover is cell-type specific in terms of its machinery, and its regulation is one of the major post-translational modifications of a protein. TPH degradation is likely linked to protein phosphorylation. Functionally, TPH, the rate-limiting enzyme in serotonin biosynthesis, is involved in neural development, which likely affects animal behaviors in the adult stages. Future studies will define the sensitive period during development when TPH activity is required for the fine-tuning of neuronal organization that is responsible for each behavior in the adult.
Methamphetamine toxicity and messengers of death
2009, Brain Research Reviews
Methamphetamine (METH) is an illicit psychostimulant that is widely abused in the world. Several lines of evidence suggest that chronic METH abuse leads to neurodegenerative changes in the human brain. These include damage to dopamine and serotonin axons, loss of gray matter accompanied by hypertrophy of the white matter and microgliosis in different brain areas. In the present review, we summarize data on the animal models of METH neurotoxicity which include degeneration of monoaminergic terminals and neuronal apoptosis. In addition, we discuss molecular and cellular bases of METH-induced neuropathologies. The accumulated evidence indicates that multiple events, including oxidative stress, excitotoxicity, hyperthermia, neuroinflammatory responses, mitochondrial dysfunction, and endoplasmic reticulum stress converge to mediate METH-induced terminal degeneration and neuronal apoptosis. When taken together, these findings suggest that pharmacological strategies geared towards the prevention and treatment of the deleterious effects of this drug will need to attack the various pathways that form the substrates of METH toxicity.
Distinctive iron requirement of tryptophan 5-monooxygenase: TPH1 requires dissociable ferrous iron
2005, Biochemical and Biophysical Research Communications
A peripheral type of tryptophan 5-monooxygenase (EC 1.14.16.4), TPH1, is very unstable in vitro, but the inactivation was reversible and full reactivation occurs upon anaerobic incubation with a high concentration of dithiothreitol (DTT, 15 mM). In this study, distinctive iron requirement of TPH1 was revealed through analysis of the enzyme’s inactivation and activation by DTT. For this purpose, all the glasswares, plastics, Sephadex G-25 gels, and reagents including protein solutions had been treated with metal chelators, and apo-TPH was prepared by treatment with EDTA. Apo-TPH thus prepared exclusively required free Fe²⁺ for its catalytic activity; 10⁻⁸ M was enough under the strict absence of Fe³⁺ but 10⁻¹² M was too low. No other metal ions including Fe³⁺ were effective. It appeared that Fe³⁺ bound to the enzyme with a higher affinity than Fe²⁺, resulting in the inactivation. Ascorbate, a non-thiol reducing agent, did not substitute DTT in the activation of TPH1, but enhanced the Fe²⁺-dependent activity of apo-TPH as effectively as DTT. Thus, the DTT-activation was essentially substituted by preparation of apo-TPH by the EDTA treatment and the assay of apo-TPH in the presence of Fe²⁺ and ascorbate. The activation of TPH1 by incubation with DTT was accompanied by exposure of 9 sulfhydryls out of the total 10 cysteine residues, but the cleavage of disulfide bonds seemed not to be crucial, even if it occurred. The effect of DTT was substituted by some other sulfhydryls whose structure was analogous to that of commonly used metal chelators. Based on these observations, the following dual roles of DTT are proposed: (1) in the activation of TPH, DTT removes inappropriate bound iron (Fe³⁺) as a chelator, keeping Fe³⁺ away from the enzyme’s binding site which needs to bind Fe²⁺ for the catalytic activity, and (2) in both the activation and reaction processes, DTT prevents oxidation of Fe²⁺ to Fe³⁺ as a reducing agent.

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Short communicationAcute inactivation of tryptophan hydroxylase by amphetamine analogs involves the oxidation of sulfhydryl sites

Abstract

European J. Pharmacol.

Life Sci.

European J. Pharmacol.

J. Biol. Chem.

Brain Res.

Short communication
Acute inactivation of tryptophan hydroxylase by amphetamine analogs involves the oxidation of sulfhydryl sites