Increase In Tryptophan Hydroxylase Activity Research Articles

Acoustic stimulation in vivo and corticotropin-releasing factor in vitro increase tryptophan hydroxylase activity in the rat caudal dorsal raphe nucleus

Exposure of rats to unpredictable loud sound pulses increases activity of the rate-limiting enzyme for serotonin synthesis, tryptophan hydroxylase (TPH), in the median raphe nucleus (MnR) and a mesolimbocortical serotonergic system. Corticotropin-releasing factor (CRF)-induced activation of a subset of serotonergic neurons in the caudal dorsal raphe nucleus (DR) may underlie stress-related increases in TPH activity in the MnR and a mesolimbocortical serotonergic system. An in vivo acoustic stimulation paradigm and an in vitro brain slice preparation were designed to test the hypothesis that stress-related stimuli and CRF receptor activation have convergent actions on TPH activity in the caudal DR (DRC). We measured 5-hydroxytryptophan (5-HTP) accumulation as an index of TPH activity following inhibition of aromatic amino acid decarboxylase (using NSD-1015). To examine effects of acoustic stimulation on TPH activity, male Wistar rats, pretreated with NSD-1015, were exposed to a 30 min sham, predictable or unpredictable acoustic stimulation paradigm; brains were frozen and microdissected for analyses of tissue 5-HTP concentrations in subregions of the DR. To examine the effect of CRF receptor activation on TPH activity, freshly prepared brain slices were exposed to CRF (0–2000 nM) for 10 min in the presence of NSD-1015, then frozen and microdissected for analysis of tissue 5-HTP concentrations. Increases in TPH activity in the DRC, but not other subregions, were observed in both paradigms. These findings are consistent with the hypothesis that stress-related increases in TPH activity are mediated via effects of CRF or CRF-related neuropeptides on a mesolimbocortical serotonergic system originating in the DRC.

Anti-inflammatory drugs ameliorate opposite enzymatic changes in ileal 5-hydroxytryptamine metabolism in the delayed phase after cisplatin administration to rats

The effects of anti-inflammatory drugs on ileal 5-hydroxytryptamine (5-HT) metabolic dynamics at 72 h after a single administration of cisplatin were investigated in rats. Cisplatin 5 mg/kg i.p. caused pathological changes, with an inflammatory response occurring 72 h after its administration. The inflammatory response was associated with the induction of cyclooxygenase-2, but not cyclooxygenase-1, in the ileal mucosa at 72 h after the cisplatin administration. Daily treatment with meloxicam 3 mg/kg s.c. ameliorated the cisplatin-induced mucosal damage, whereas dexamethasone 1 mg/kg s.c. did not. Cisplatin administration also caused a significant increase in cyclooxygenase-2 mRNA expression at 72 h after administration, which was blunted by dexamethasone, but not by meloxicam. Cisplatin increased the content of 5-HT and its metabolite, 5-hydroxyindoleacetic acid (5-HIAA), but had no effect on 5-HT turnover (5-HIAA/5-HT ratio). Meloxicam and dexamethasone did not significantly decrease 5-HT and 5-HIAA content. Cisplatin significantly decreased monoamine oxidase activity but increased tryptophan hydroxylase (TPH) activity and TPH 1 mRNA expression in ileal tissue. Meloxicam and dexamethasone significantly restored the decreased monoamine oxidase activity and inhibited the cisplatin-induced increase in tryptophan hydroxylase activity toward the control levels. These drugs also decreased the cisplatin-induced increase in TPH 1 mRNA expression. Neither cisplatin nor the anti-inflammatory drugs had significant effect on mRNA expression of the serotonin re-uptake transporter. These results suggest that the inflammatory response associated with cyclooxygenase-2 induction is involved in the opposite change in ileal tryptophan hydroxylase and monoamine oxidase activities in the delayed phase after single administration of cisplatin to rats.

Regional serotonin metabolism in the brain of transgenic mice lacking monoamine oxidase A.

The effect of a lack of the gene encoding monoamine oxidase A (MAO A) in transgenic Tg8 mice on the activity of tryptophan hydroxylase (TPH), the rate-limiting enzyme in serotonin (5-HT) biosynthesis, and on the levels of 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) in the midbrain, hypothalamus, hippocampus, striatum, amygdala, and frontal cortex was studied. It was shown that mice with a genetic MAO A knockout differed from mice of the initial C3H/HeJ strain in having a higher level of 5-HT and a lower level of its metabolite, 5-HIAA, in all brain regions but the frontal cortex, where the changes were insignificant. Although the 5-HIAA/5-HT ratio in various brain regions differed considerably, the decrease of the 5-HT oxidative deamination index in Tg8 mice was similar in different brain regions (to 41-45% of control values), with the exception of the frontal cortex, where the decrease of the 5-HIAA/5-HT was somewhat smaller (to 54%). The presence of the remaining 45% +/- 1.9% of the control ratio value indicates rather effective oxidative deamination of 5-HT in MAO A knockout mice and explains the lack of severe behavioral and pathological consequences in MAO A genetic deficiency. An increase of TPH activity in mice lacking MAO A was found in the frontal cortex, hippocampus, and amygdala. No significant changes were found in the striatum, hypothalamus, and midbrain. The data show an effect of the MAO A gene mutation on TPH and indicate a uniform decrease of 5-HT catabolism in different brain regions except for the frontal cortex, which is somewhat more resistant to the lack of MAO A than other brain structures.

Changes in serotonin dynamics in the gastrointestinal tract of colon-26 tumour-bearing mice: effects of cisplatin treatment.

Severe nausea and vomiting are common side effects of anti-cancer chemotherapy. 5-HT3 receptor antagonists have been used for the treatment of these gastrointestinal symptoms. The purpose of this study was to examine whether specific changes in serotonin dynamics occurred in the gastrointestinal tract in mice in which Colon-26 adenocarcinoma cells were injected s.c., especially after treatment with cisplatin. The serotonin content of the small intestine of mice inoculated s.c. with Colon-26 adenocarcinoma increased significantly 2 weeks after the inoculation of the tumor cells; this was associated with an increase in tryptophan hydroxylase activity and the number of enterochromaffin cells as compared with control mice. Intravenous injection of cisplatin significantly reduced the serotonin content in the small intestine of Colon-26 tumour-bearing mice but not in control mice. The spontaneous release of serotonin from isolated intestine was not different between Colon-26 tumour-bearing and control mice; however, pretreatment of mice with cisplatin induced two fold increases in serotonin release from duodenum, jejunum and ileum in Colon-26 tumour-bearing mice but not in control mice. These results indicate that a region-specific increase in the number of enterochromaffin cells is observed in the intestine of Colon-26 tumour-bearing mice, associated with an increase in the serotonin content and tryptophan hydroxylase activity. Cisplatin treatment induced the release of serotonin from affected enterochromaffin cells in the gastrointestinal tract, which may be related to the occurrence of nausea in clinical use.

Sound stress activation of tryptophan hydroxylase blocked by hypophysectomy and intracranial RU 38486

The rapidly reversible increase in cortical or midbrain tryptophan hydroxylase activity observed ex vivo after exposure of rats to 1-h sound stress was blocked by hypophysectomy, but not sham hypophysectomy, and restored by dexamethasone administration to the hypophysectomized animals (500 μg/day i.p. for 3 days). The response to sound stress was also lost with deafferentation of the hypothalamus. These results indicate that hypothalamic control of adrenal glucocorticoid is required for the serotoenergic response to sound stress. The glucocorticoid antagonist, RU 38486, given intracerebroventricularly (200 μg/day for 4–5 days) or bilaterally, into the region of the central nucleus of the amygdala (100 μg 15 min before stress), blocked the sound stress-induced increase in tryptophan hydroxylase activity. In contrast, the antimineralocorticoid, RU 26752, was without effect. The block obtained with RU 38486 suggests that glucocorticoid is required by the neurons that relay the effects of sound stress to the rostrally projecting serotonergic neurons.

Intracranial Dehydroepiandrosterone Blocks the Activation of Tryptophan Hydroxylase in Response to Acute Sound Stress

Bilateral infusion of dehydroepiandrosterone (DHEA) given intracerebroventricularly blocked the sound stress-induced increase in tryptophan hydroxylase activity observed ex vivo in midbrain and cortex but had no effect on the level of tryptophan hydroxylase activity from sham-stressed rats. DHEA (20 μg total dose) given bilaterally into the region of the central nucleus of the amygdala, 30 min prior to 1 h sound stress, also blocked the increase in enzyme activity in a dose-dependent manner. The DHEA treatment did not alter the activation of the enzyme seen in vitro in the presence of phosphorylating conditions. The effect of DHEA was steroid specific in that other sex steroids, such as estrogen, androgens, or progesterone, were without any effect. Coadministration, 20 μg each, of the potent glucocorticoid agonist, RU 28362, with DHEA 30 min prior to 1 h sound stress completely blocked the DHEA suppressive effect on sound stress-induced increases in tryptophan hydroxylase activity. The results obtained suggest that DHEA blocks this increase in tryptophan hydroxylase activity by antagonizing the effects of glucocorticoid.

Involvement of brain tryptophan hydroxylase in the mechanism of hibernation

Marked changes were revealed in the activity of the key enzyme in serotonin biosynthesis, tryptophan hydroxylase (TPH), during entry into hibernation, hibernation, and arousal in ground squirrels ( Citellus erythrogenys). An increase in TPH activity was found in the midbrain, hippocampus, and striatum during the prehibernation period in euthermic ground squirrels. A further increase in TPH activity was observed during the entry into hibernation. Significant elevation was found not only in potential TPH activity measured at the incubation temperature of 37°C but also at incubation temperature of 7°C, approximating the body temperature in hibernation. V max in the midbrain of hibernating animals was about 50% higher than in active ones without significant changes in K m. Thus, brain TPH maintains functionality during torpidity and is activated before the entry into hibernation. The results support the idea that brain serotonin is crucially involved in the transition to the maintenance of the hibernation state.

Acute effects of reserpine on tryptophan hydroxylase activity and mRNA in rat brain

The present study examined changes in tryptophan hydroxylase (TPH) activity in the rat brainstem 1–4 days after i.p. reserpine administration. TPH enzyme activity was significantly decreased day 1 posttreatment and followed by a small but significant increase on days 2–4. Maximal TPH activity on day 4 was elevated only 42% above control level. This small increase in TPH activity was not preceded by any change of in situ hybridization signal of TPH message on day 1. In contrast, adrenal tyrosine hydroxylase (TH) activity in these same reserpinized animals was 2.5-fold higher than control on 4 day posttreatment. In addition, TH message was correspondingly increased day 1 posttreatment. Taken together, these results indicate a trend of small increase in TPH activity and no alteration in message after reserpine treatment unlike catecholaminergic systems.

Activity of tryptophan hydroxylase in brain of hereditary predisposed to catalepsy rats

The activity of the rate-limiting enzyme of serotonin biosynthesis, tryptophan hydroxylase (TPH), was studied in the brain of rats bred for 20 generations for predisposition to catalepsy (an excessive freezing). Increased TPH activity was found in the striatum but not in the hippocampus and midbrain of cataleptic rats compared with Wistar ones. K m for the enzyme from the striatum of cataleptics was twice as low as that in control rats, although no difference in their V max was found. The increase in TPH activity in the striatum of cataleptics was nonadditive with its activation induced by incubation in vitro of the enzyme under phosphorylating conditions and could be completely reversed with alkaline phosphatase. An administration of p-chlorophenylalanine, an irreversible inhibitor of TPH, decreased the duration of freezing in cataleptic rats. These findings indicate that hereditary predisposition to catalepsy is associated with increased TPH activity in the striatum due to local phosphorylation of the enzyme and suggest an essential role of the activation of striatal TPH in genetic predisposition to catalepsy.

Increase in cortical and midbrain tryptophan hydroxylase activity by intracerebroventricular administration of corticotropin releasing factor: block by adrenalectomy, by RU 38486 and by bilateral lesions to the central nucleus of the amygdala

Corticotropin releasing factor (CRF) infused bilaterally into the lateral ventricles of awake, chronically cannulated, male Sprague-Dawley rats produced a dose-dependent increase in the in vitro activity of cortical and midbrain tryptophan hydroxylase after 60 min. The maximal increase in enzyme activity of 60% over that of vehicle-treated controls was reached 45 min after an infusion of 3 μg CRF. The increase in enzyme activity after a single dose of CRF resembled that seen after exposure of rats to an acute sound stress: it was reversed by preincubation of the enzyme preparation with alkaline phosphatase and was nonadditive with the increase in activity obtained in the presence of phosphorylating conditions. The response to intracerebroventricularly administered CRF was abolished by bilateral adrenalectomy, but restored by repeated daily systemic administration of the synthetic glucocorticoid, dexamethasone (500 μg/day, i.p. for 3 days), to the adrenalectomized rats. Intracerebroventricular administration of the glucocorticoid antagonist, RU 38486 (200 μg/day for 4 days), also blocked the acute increase in tryptophan hydroxylase activity in response to CRF. Finally, bilateral lesions to the central nucleus of the amygdala, a region involved in mediating behavioral, endocrine and autonomic responses to stressful stimuli, abolished the increase in enzyme activity in response to intraventricular CRF. The glucocorticoid sensitivity of the response to CRF, as well as the involvement of the central nucleus of the amygdala support the view that CRF may have a role in mediating the enhancement of tryptophan hydroxylase activity by acute sound stress.

Circadian rhythm of tryptophan hydroxylase activity in chicken retina

1. Retinal tryptophan hydroxylase activity in chickens (1-4 weeks old and embryos) was estimated by determination of levels of 5-hydroxytryptophan (5HTP) in retinas at defined intervals after inhibition of aromatic L-amino acid decarboxylase with m-hydroxybenzylhydrazine (NSD1015). 2. The relationship of tryptophan hydroxylase activity to photoperiod was explored. In chickens maintained on a 12-hr light: 12-hr dark cycle, a diurnal cycle in tryptophan hydroxylase activity was observed. Activity during middark phase was 4.4 times that seen in midlight phase. Cyclic changes in tryptophan hydroxylase activity persisted in constant darkness with a period of approximately 1 day, indicating regulation of the enzyme by a circadian oscillator. The phase of the tryptophan hydroxylase rhythm was found to be determined by the phase of the light/dark cycle. The relationship of the tryptophan hydroxylase rhythm to the light/dark cycle mirrors previously described rhythms of melatonin synthesis and serotonin N-acetyltransferase (NAT) activity in the retina. 3. Light exposure for 1 hr during dark phase suppressed NAT activity by 82%, while tryptophan hydroxylase activity was suppressed by only 30%. 4. Based on the differential responses of retinal NAT activity and tryptophan hydroxylase activity to acute light exposure during dark phase, it was predicted that exposure to light during dark phase would divert serotonin in the retina from melatonin biosynthesis to oxidation by MAO. In support of this, levels of 5-hydroxyindole acetic acid (5HIAA) in retina were found to be elevated approximately two-fold in chickens exposed to 30 min of light during dark phase. In pargyline-treated chickens, 2 hr of light exposure during dark phase was found to increase retinal serotonin levels by 64% over pargyline-treated controls. 5. Cyclic changes in tryptophan hydroxylase activity and NAT activity persisted for 2-3 days in constant light. Tryptophan hydroxylase activity at mid-night gradually decreased on successive days in constant light; on the first day of constant light, tryptophan hydroxylase activity at mid-night was 70% of activity seen during middark phase of the normal light/dark cycle and decreased further on subsequent days. In contrast, on each of 3 days of constant light, NAT activity at mid-night was approximately 15% of normal middark phase activity. 6. Cycloheximide completely inhibited the nocturnal increase in tryptophan hydroxylase activity when given immediately before light offset. The nocturnal increase in NAT activity was inhibited in a similar fashion. 7. Like the development of the NAT rhythm, cyclic changes of tryptophan hydroxylase activity in the retinas of chickens began on or immediately before the day of hatching. hatching.(ABSTRACT TRUNCATED AT 400 WORDS)

Sound stress-induced Increase in tryptophan hydroxylase activity of cerebral cortex and midbrain is abolished by central administration of dehydroepiandrosterone

Sound stress-induced Increase in tryptophan hydroxylase activity of cerebral cortex and midbrain is abolished by central administration of dehydroepiandrosterone

Tryptophan hydroxylase activity in hypothalamus and brainstem of neonatal and adult rats treated with hydrocortisone or parachlorophenylalanine

The present study has been undertaken to determine whether glucocorticoid, and parachlorophenylalanine (PCPA, a tryptophan hydroxylase inhibitor) affects tryptophan hydroxylase (TPH) levels in brainstem and hypothalamus of neonatal and adult rats. Our results show that: (1) administration of hydrocortisone causes small but significant increases in TPH activity of neonatal brainstem: (2) treatment with PCPA plus glucocorticoid results in a marked decrease of TPH activity in brainstem and hypothalamus of both neonatal and adult rats.

Activation of cortical tryptophan hydroxylase by acute morphine treatment: blockade by 6-hydroxydopamine

Acute morphine produced a dose-dependent, naloxone-sensitive, reversible increase in tryptophan hydroxylase activity in low speed supernatants of midbrain, pons-medulla and cerebral cortex but not spinal cord. The increase in cortical enzyme activity was blocked by 6-hydroxydopamine pretreatment, could be reversed in vitro by incubation with alkaline phosphatase and was non-additive with the increase in enzyme activity induced in the presence of phosphorylating conditions. Morphine administration produced an increase in V max but no change in K m of cortical enzyme for substrate, tryptophan, or the artificial reduced pterin cofactor, 6-methyl-5,6,7,8-tetrahydropterin. The failure of morphine to increase spinal tryptophan hydroxylase activity despite enhancement of enzyme activity in medulla indicates regional differences in responsiveness of the enzyme to in vivo activation.

Agn 2979 [3-(3-methoxyphenyl)-3-(3-dimethylaminopropyl)-4,4-dimethylpiperidine-2, 6-dione]: An inhibitor of the activation of tryptophan hydroxylase

AGN 2979 [3-(3-methoxyphenyl)-3-(3-dimethylaminopropyl)-4,4-dimethylpiperidine-2,6-dione] blocked the increase in tryptophan hydroxylase activity that occurred when slices of brainstem were exposed to a depolarizing medium or to agents that mobilize intracellular pools of calcium, but it had no effect on the activity of enzyme prepared from slices of brainstem incubated in control medium. AGN 2979 also blocked the calcium-calmodulin-dependent activation of tryptophan hydroxylase that was seen when supernatant preparations of the enzyme were exposed to phosphorylating conditions but not the activation induced by calcium-dependent proteases that was triggered by millimolar calcium concentrations. An identical pattern of inhibition has been found with the antipsychotic drugs, haloperidol and fluphenazine [Boadle-Biber, Biochem. Pharmac. 31, 2495 (1982)]. The sensitivity to the same inhibitors of both the activation of tryptophan hydroxylase produced by pretreatment of brainstem slices and that induced by incubation of supernatant preparations of enzyme under phosphorylating conditions suggests involvement of a common mechanism of enzyme activation in response to these different treatments.

Increased tryptophan hydroxylase activity in serotonergic nerve terminals spared by 5,7-dihydroxytryptamine

Adult rats received intraventricular injections of 5,7-dihydroxytryptamine (5,7-DHT) to destroy serotonin (5-HT)-containing nerve terminals throughout the brain. When the animals were killed 3 or 21 days later, we observed a marked decrease in 5-HT content in septum and hippocampus and a parallel decline in in vitro high affinity 5-HT uptake. 5-Hydroxyindoleacetic acid (5-HIAA) concentrations also were reduced but by a much smaller extent, resulting in significant increases in the ratio of 5-HIAA to 5-HT. These changes were accompanied by similar increases in the ratio of tryptophan hydroxylase (TPH) activity to 5-HT content. The relative increases in TPH activity resulted from two temporally distinct processes, the first of which appeared to be an activation that could be mimicked in vitro by Ca 2+-dependent phosphorylation. We conclude that, after partial damage to 5-HT neurons, there is a compensatory increase in the synthesis and release of 5-HT from those terminals that remain.

Long-term local and distal increases in tryptophan hydroxylase activity following intracerebral kainic acid injections in the rat

The administration of kainic acid (1–2 μg) into the right striatum of adult rats resulted in a marked local increase in tryptophan hydroxylase activity (+ 54–106%). This change was significant as soon as on the second day after the treatment and persisted for at least 12 days. In addition, long-lasting elevations of tryptophan hydroxylase activity were also observed in the anterior raphe area, septum and ipsilateral hippocampus and cerebral cortex. In contrast, the intrahippocampal injection of kainic acid (1 μg) induced a long-term increase in tryptophan hydroxylase activity only in the injected structure. In all cases, the changes in tryptophan hydroxylase activity were associated with significant increases in the V max of the enzyme with no alteration of its apparent affinities for tryptophan and the pterin cofactor. Studies of the sensitivity of tryptophan hydroxylase from control and from kainic acid-treated rats to in vitro activating conditions (Ca +-dependent phosphorylation, partial trypsinization, exposure to sodium dodecyl sulfate) suggest that the intrastriatal injection of the neurotoxin induced a long-lasting activation of the enzyme. These findings indicate that intracerebral injections of kainic acid may be a valuable approach to explore further the mechanisms controlling tryptophan hydroxylase activity in vivo.

Blockade by haloperidol of the increase in tryptophan hydroxylase activity induced by incubation of slices of brain stem with dibutyryl cyclic AMP

Pretreatment of rat brain stem slices with dibutyryl cyclic AMP, caffeine, theophylline and 3-isobutyl-1-methylxanthine increased the activity of tryptophan hydroxylase in supernatant preparations of enzyme made from the slices. This effect does not appear to be mediated by a cyclic AMP sensitive mechanism since it was not reproduced by exposure of the slices to 8-bromo cyclic AMP, to papaverine, a nonxanthine phosphodiesterase inhibitor, or to other treatments known to raise tissue cyclic AMP levels. The ability of haloperidol to block this increase in enzyme activity is consistent with a role for calmodulin and calcium as mediators of the enzyme activation, particularly when this observation is considered in conjunction with the evidence that supernatant preparations of this enzyme was activated under phosphorylating conditions by a calcium-calmodulin dependent process [5]. Nevertheless, in view of the high concentrations of haloperidol employed in the present experiments, the possibility that this drug may produce its effects in the brain stem slices through some other action, unrelated to its ability to bind to calmodulin, should be kept in mind.

Increased serotonin turnover in corpus striatum following an injection of kainic acid: Evidence for neuronal feedback regulation of synthesis

Kainic acid, a cyclic analog of glutamate, has been reported by other investigators to reduce the number of serotonin (5HT) binding sites in rat striatum following an intrastriatal injection. We found that kainic acid injected into the striatum caused a dose-dependent increase in ipsilateral striatal 5-hydroxyindoleacetic acid (5HIAA) content, while not affecting the level of 5HT. There was no detectable change in 5HT metabolism in the contralateral striatum. The increase of 5HIAA was a reflection of increased 5HT turnover in the injected striatum as measured from the decline of 5HIAA after pargyline treatment or the rise of 5-hydroxytryptophan after benserazide treatment. There was also a concomitant increase of tryptophan hydroxylase activity. Kainic acid treatment resulted in an apparent decrease of the K mand increase of the V max for the pteridine cofactor and an increase of the V max for tryptophan by tryptophan hydroxylase. Kainic acid injection into the dorsal raphe nucleus caused a dose-dependent decrease in 5HT content of the dorsal raphe nucleus and in both striata, which are dorsal raphe nucleus projection areas. Our results suggest that 5HT formation in the striatum is normally modulated by an inhibitory neuronal feedback loop. Interruption of the loop by injecting kainic acid causes 5HT formation and tryptophan hydroxylase activity to increase in the ipsilateral but not contralateral striatum. Kainic acid apparently destroys 5HT neurons when injected close to the cell soma.

Acute effects of heroin and morphine on newly synthesized serotonin in rat brain

Heroin and morphine, in acute intraperitoneal doses of 2 and 10 mg/kg respectively, produced significant increments in the formation of newly formed brain serotonin from tritiated ( 3H)-L-tryptophan to 3H-serotonin. Opiate analgesia, Straub tail sign and catatonia, were observed during the increase in the synthesis of serotonin. The transport of radio-labelled tryptophan into the rat brain was not increased by the acute injection of the opiates, but brain levels of 3H-serotonin and of its main metabolite, 5-hydroxyindoleacetic acid, were significantly elevated. These opiates do not interfere with the accumulation of serotonin or with the transport of its metabolite in serotonergic neurons after inhibition of monoamine oxidases with Pargyline. An increase in the activity of tryptophan hydroxylases was more pronounced in the forebrain than in the brain stem. Stimulation of newly synthesized serotonin is probably mediated by an increase in tryptophan hydroxylase activity and not by an increase in the transport of tryptophan into the brain.