TRYPTOPHAN LOADING: CONSEQUENT EFFECTS ON THE SYNTHESIS OF KYNURENINE AND 5-HYDROXYINDOLES IN RAT BRAIN

Abstract— Tryptophan loading of rats resulted in a continuous non-linear uptake of l -tryptophan from plasma into the brain. The optimum tryptophan load for increasing cerebral 5-hydroxytryptamine (5-HT) level was 25 mg/kg. Above this, there was a gradual decrease both in the levels and synthesis of 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) as assessed from simultaneous intraperitoneal or intraventricular injections of l [¹⁴C]tryptophan. A 5–10 fold increase in cerebral tryptophan produced a limited stimulation of 5-HT synthesis. When the cerebral tryptophan level reached 1 ± 10 -4 , substrate inhibition in vivo of the tryptophan monooxygenase (tryptophan-5-hydroxylase) but not of the indoleamine-2,3-dioxygenase occurred. Cerebral synthesis of kynurenine increased linearly with increasing tryptophan load. At a plasma ratio of 50:1 tryptophan to kynurenine, tryptophan loading interfered with the entry of peripheral kynurenine. Tryptophan loading also increased the efflux of 5-hydroxyindoles from the brain. One hour after intraperitoneal injection of l -kynurenine sulfate (5 mg/kg) into rats, there was a shift in the plasma ratio of l -tryptophan to l -kynurenine to 4:1. In these rats, a 20% reduction of cerebral tryptophan was noted.     

Monitoring the Effect of a Tryptophan Load on Brain Indole Metabolism in Freely Moving Rats by Simultaneous Cerebrospinal Fluid Sampling and Brain Dialysis

Rats were given L-tryptophan, 50 mg/kg i.p., and its concentration in the CNS was monitored in individual freely moving animals using repeated sampling of cisternal CSF and concurrent striatal dialysis. The 5-hydroxytryptamine metabolite 5-hydroxyindoleacetic acid (5-HIAA) was also measured. Results were compared with changes of central tryptophan and 5-HIAA concentrations in brains of rats killed at various times after administration of L-tryptophan, 50 mg/kg i.p. Tryptophan changes in CSF were proportionate to those in whole brain and followed essentially identical time courses. Results for the striatal dialysate and whole striatum also paralleled each other. Similarly, results for 5-HIAA showed proportionality between CSF and brain and between dialysate and striatum. The data obtained were used to determine pharmacokinetic data for individual rats, i.e., areas under curves for both tryptophan and 5-HIAA and half-lives for the decline of tryptophan. Kinetic parameters varied considerably from rat to rat. However, mean half-lives for tryptophan in CSF, brain, dialysate, and striatum were all comparable. Results in general show the value of repeated CSF sampling and intracerebral dialysis for concurrent monitoring of changes of indole metabolism in the whole brain and a specific brain region, respectively. The methods should be suitable for the continuous monitoring of changes of central transmitter metabolism in parallel with observation of behavior following environmental or dietary changes or drug administration. They also should be of use in the investigation of drug kinetics in the CNS.     

Potential role for nonesterified fatty acids in beta-adrenoceptor-induced increases in brain tryptophan

We tested the hypothesis that beta2- and beta3-adrenergic receptor-mediated increases in brain tryptophan are due to the liberation of fatty acids, which in turn displace tryptophan from its albumin-binding site and thus facilitate its entry into the brain. Male CD-1 mice were injected with subtype-selective beta-adrenergic agonists 1h before brain samples were collected for analysis of tryptophan content by HPLC with electrochemical detection, and blood samples were collected for analysis of total and free tryptophan and nonesterified fatty acid (NEFA) concentrations. The beta2-selective agonist, clenbuterol (0.1 mg/kg), increased concentrations of tryptophan in all brain regions studied and decreased plasma total tryptophan, but had no effect on plasma free tryptophan or NEFAs. The beta3-selective agonists, BRL 37344 (0.2 mg/kg) or CL 316243 (0.01 mg/kg), increased brain tryptophan, plasma NEFAs and free tryptophan. Pretreatment with nicotinic acid (500 mg/kg), an inhibitor of lipolysis, almost completely prevented the increase in plasma free tryptophan and NEFAs, and attenuated the increase in brain tryptophan induced by CL 316243. These results suggest that beta2- and beta3-adrenergic agonists increase brain tryptophan by a mechanism other than the liberation of NEFAs. Nonetheless, beta3-adrenergic agonists appear to increase brain tryptophan by a mechanism that may depend partially on elevations of plasma NEFAs.     

Injected tryptophan increases brain but not plasma tryptophan levels more in ethanol treated rats

In previous studies, long term treatment with ethanol has been shown to enhance brain 5-hydroxytryptamine 5-(HT) metabolism by increasing the activity of the regulatory enzyme tryptophan hydroxylase and or availability of circulating tryptophan secondarily to an inhibition of hepatic tryptophan pyrrolase. In the present study ethanol treatment given for two weeks decreased hepatic apo-tryptophan pyrrolase but not total tryptophan pyrrolase activity in rats. Tryptophan levels in plasma and brain did not increase significantly. But there was a marked increase of 5-HT but not 5-hydroxyindoleacetic acid (5-HIAA) concentration in brain, suggesting a possible increase in the activity of tryptophan hydroxylase. The effect of a tryptophan load on brain 5-HT metabolism was therefore compared in controls and ethanol treated rats. One hour after tryptophan injection (50 mg/kg i.p.) plasma concentrations of total and free tryptophan were identical in controls and ethanol treated rats, but the increases of brain tryptophan 5-HT and 5-HIAA were considerably greater in the latter group. The results are consistent with long term ethanol treatment enhancing brain serotonin metabolism and show that brain uptake/utilization of exogenous tryptophan is increased in ethanol treated rats and may be useful to understand the role and possible mechanism of tryptophan/serotonin involvement in mood regulation.     

Injected tryptophan increases brain but not plasma tryptophan levels more in ethanol treated rats

In previous studies, long term treatment with ethanol has been shown to enhance brain 5-hydroxytryptamine 5-(HT) metabolism by increasing the activity of the regulatory enzyme tryptophan hydroxylase and or availability of circulating tryptophan secondarily to an inhibition of hepatic tryptophan pyrrolase. In the present study ethanol treatment given for two weeks decreased hepatic apo-tryptophan pyrrolase but not total tryptophan pyrrolase activity in rats. Tryptophan levels in plasma and brain did not increase significantly. But there was a marked increase of 5-HT but not 5-hydroxyindoleacetic acid (5-HIAA) concentration in brain, suggesting a possible increase in the activity of tryptophan hydroxylase. The effect of a tryptophan load on brain 5-HT metabolism was therefore compared in controls and ethanol treated rats. One hour after tryptophan injection (50 mg/kg i.p.) plasma concentrations of total and free tryptophan were identical in controls and ethanol treated rats, but the increases of brain tryptophan 5-HT and 5-HIAA were considerably greater in the latter group. The results are consistent with long term ethanol treatment enhancing brain serotonin metabolism and show that brain uptake/utilization of exogenous tryptophan is increased in ethanol treated rats and may be useful to understand the role and possible mechanism of tryptophan/serotonin involvement in mood regulation.     

Oxindole, a Sedative Tryptophan Metabolite, Accumulates in Blood and Brain of Rats with Acute Hepatic Failure

Abstract: Rats treated with oxindole (10–100 mg/kg i.p.), a putative tryptophan metabolite, showed decreased spontaneous locomotor activity, loss of the righting reflex, hypotension, and reversible coma. Brain oxindole levels were 0.05 ± 0.01 nmol/g in controls and increased to 8.1 ± 1.7 or 103 ± 15 nmol/g after its administration at doses of 10 or 100 mg/kg i.p., respectively. To study the role that oxindole plays in the neurological symptoms associated with acute liver failure, we measured the changes of its concentration in the brain after massive liver damage, and we investigated the possible metabolic pathways leading to its synthesis. Rats treated with either thioacetamide (0.2 and 0.4 g/kg i.p., twice) or galactosamine (1 and 2 g/kg i.p.) showed acute liver failure and a large increase in blood or brain oxindole concentrations (from 0.05 ± 0.01 nmol/g in brains of controls to 1.8 ± 0.3 nmol/g in brains of thioacetamide-treated animals). Administration of tryptophan (300–1,000 mg/kg p.o.) caused a twofold increase, whereas administration of indole (10–100 mg/kg p.o.) caused a 200-fold increase, of oxindole content in liver, blood, and brain, thus suggesting that indole formation from tryptophan is a limiting step in oxindole synthesis. Oral administration of neomycin, a broad-spectrum, locally acting antibiotic agent able to reduce intestinal flora, significantly decreased brain oxindole content. Taken together, our data show that oxindole is a neurodepressant tryptophan metabolite and suggest that it may play a significant role in the neurological symptoms associated with acute liver impairment.     

Acute effects of caffeine injection on neutral amino acids and brain monoamine levels in rats

The injection of caffeine (100 mg/kg, i.p.) into male rats acutely increased brain levels of trytophan, serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA). Blood levels of glucose, nonesterified fatty acids (NEFA) and insulin also increased, while those of the aromatic and branched-chain amino acids fell. Serum tryptophan levels either did not fall, or increased. Consequently, the serum ratio of trypthopahn to the sum of other large neutral amino acids (LNAA) increased. Less consistently noted were increases in serum free tryptophan levels. Brain tyrosine levels were not appreciably altered by caffeine, nor was the serum tyrosine ratio. In dose-response studies, 25 mg/kg of caffeine was the minimal effective dose needed to raise brain tryptophan, but only the 100 mg/kg dose elevated all three indoles in brain. In no experiments did caffeine, at any time or dose, alter brain levels of dopamine or norepinephrine. Caffeine thus probably raises brain tryptophan levels by causing insulin secretion, and thereby changing plasma amino acid levels to favor increased tryptophan uptake into brain. The rises in brain 5-HT and 5-HIAA may follow from the increase in brain tryptophan, although further data are required clearly to establish such a mechanism.     

Serotonergic activity of HP 184: Does spontaneous release have a role?

Examination of HP 184, [N-n-propyl)-N-(3-fluoro-4-pyridinyl)-1H-3-methylindodel-1-amine hydrochloride], in a variety of tests for serotonergic activity revealed some unique properties of this compound. We report here that 100 μM HP 184 enhanced spontaneous release of [³H]serotonin (5-HT) from rat hippocampal slices. This release was independent of the uptake carrier. In vivo assays confirmed that HP 184 (20 mg/kg, i.p.) lacked significant interactions at the norepinephrine (NE) or 5-HT uptake carrier itself. Notably, HP 184 (15 mg/kg, i.p.) reduced drinking behavior in schedule-induced polydipsic (SIP) rats. We previously reported that some selective 5-HT reuptake inhibitors decrease SIP 30–40% after a 14–21 day treatment. In the current study, HP 184 decreased SIP beginning with the first treatment, and this reduction (30%) was maintained for 28 days. We further investigated HP 184 and serotonin metabolite levels. One hour after i.p. administration of 30 mg/kg HP 184, the ratio of whole brain 5-hydroxyindolacetic acid (5-HIAA) to 5-HT was increased, suggesting serotonergic activation. Under these conditions, the brain: plasma ratio of HP 184 was approximately 2∶1, with brain concentrations of 1.6 μg/gram. We speculate that the spontaneous release effects of HP 184 may be responsible for the behavioral effects observed.     

β-Adrenergic Effects on Plasma and Brain Large Neutral Amino Acids Are Unaltered by Chronic Administration of Antidepressants

Effects of isoproterenol (3 mg kg-1, i.p. for 60 min) and salbutamol (3, 10 mg kg-1, i.p. for 60 min) on large neutral amino acid concentrations in rat plasma and brain were assessed. Phenylalanine, leucine, isoleucine, and valine were measured by gas chromatography with electron-capture detection; tyrosine and tryptophan were measured by HPLC with electrochemical detection. These drugs induced increases in brain tryptophan, tyrosine, phenylalanine, and valine and decreases in plasma tryptophan, tyrosine, leucine, isoleucine, and valine. Effects of salbutamol (3 mg kg-1, i.p. for 60 min) were assessed following chronic administration of phenelzine sulfate and desipramine.HCl (each drug 10 mg kg-1 per day, s.c. via Alzet 2ML4 osmotic minipumps for 28 days). There were no effects of these antidepressants on basal levels of large neutral amino acids in brain and plasma. In both brain and plasma, salbutamol-induced changes in large neutral amino acids were unaffected by these antidepressants. The results indicate that beta-adrenoceptor-regulated availability of plasma and brain large neutral amino acids is unaffected by chronic administration of tricyclic or monoamine oxidase inhibitor antidepressants.     

Effects of imipramine on tyrosine and tryptophan are mediated by beta-adrenoceptor stimulation

Imipramine (IMI; 20 mg/kg) in rats decreased the plasma tyrosine concentration by 21% (90 min), whereas norepinephrine (NE; 1.25 mg/kg) raised it by 72% (40 min). Since NE raised plasma tyrosine by stimulating alpha-adrenoceptors, as shown by phenoxybenzamine (PB) completely abolishing this increase, an experiment was done to find out whether IMI lowered plasma tyrosine by blocking alpha-adrenoceptors. In contrast to PB, IMI pretreatment failed to alter the NE-induced elevation in plasma tyrosine, suggesting that at this dose IMI is not an effective alpha-adrenergic antagonist in vivo. Thus, IMI would not appear to reduce plasma tyrosine by blocking alpha-adrenoceptors. In a separate experiment, propranolol blocked the ability of IMI to lower plasma tyrosine. Propranolol also prevented a 17% elevation in brain tryptophan levels induced by IMI but did not alter the 29% decrease in plasma tryptophan. PB by itself decreased plasma tyrosine, but this decrease was not greater by additionally treating with IMI. Salbutamol (10 mg/kg), a beta 2 agonist, lowered plasma tyrosine to 76% and raised brain tryptophan to 143% of control. These results suggest that IMI decreases tyrosine concentrations in plasma and raises tryptophan in brain by stimulating beta-adrenoceptors.     

Brain noradrenergic neuronal activity affects 3,4-dihydroxyphenylethyleneglycol (DHPG) levels

Brain regional DHPG levels were determined following pharmacological manipulations that are known to alter brain noradrenergic neuronal activity. In rats given the α-adrenergic antagonist yohimbine (1, 5 and 10 mg/kg, i.p.) 2 h prior to sacrifice, there was a dose-dependent increase in cortical, midbrain, pons + medulla, hypothalamic and spinal total DHPG and MHPG concentrations. In contrast, cortical and spinal total DHPG and MHPG concentrations were markedly decreased 2 h following the α-adrenergic agonist, clonidine (10 and 250 μg/kg, i.p.). These findings indicate that rat brain DHPG formation is also sensitive to changes in brain noradrenergic neuronal impulse flow.     

Rat Brain Norepinephrine Metabolism: Substantial Clearance Through 3,4-Dihydroxyphenylethyleneglycol Formation

To assess whether the metabolic clearance of rat brain norepinephrine (NE) through 3,4-dihydroxyphenylethyleneglycol (DHPG) formation is quantitatively comparable or greater than through 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG) production, we studied the accumulation rates of conjugated DHPG and MHPG following probenecid administration in whole brain as well as in several brain regions. Administration of increasing doses of probenecid (100-500 mg/kg, i.p.) 1.5 h before sacrifice produced a dose-dependent increase of conjugated DHPG and MHPG levels. The maximum increment of these conjugated metabolites occurred at a dose of 300 mg/kg or higher. During the first hour following probenecid administration (300 mg/kg, i.p.), rat brain conjugated DHPG and MHPG levels accumulated linearly at a rate of 646 and 319 pmol/g/h, respectively. With the probenecid technique, the estimated appearance rates of conjugated DHPG significantly exceeded those of conjugated MHPG in hypothalamus, midbrain, brainstem, hippocampus, and cerebral cortex. These results clearly indicate that under resting conditions, formation and efflux of conjugated DHPG is the major route of metabolic clearance of rat brain NE.     

Effect of Tryptophan on Extracellular Concentrations of Tryptophan and 5-Hydroxyindoleacetic Acid in the Striatum and Cerebellum

The effects of L-tryptophan (50 mg/kg i.p.) on extracellular concentrations of tryptophan and the 5-hydroxytryptamine (5-HT) metabolite 5-hydroxyindoleacetic acid (5-HIAA) were determined in the rat striatum and cerebellum, regions with rich and poor 5-HT innervation, respectively. Determinations were on perfusates from dialysis probes in the brains of conscious, freely moving rats. The pharmacokinetic profiles of dialysate tryptophan after tryptophan load (peak concentration, time to peak concentration, area under curve, and half-life) in the two regions did not differ significantly. The dialysate 5-HIAA concentration in the striatum rose two- to threefold after the administration of tryptophan. Therefore, as 5-HIAA was undetectable in the cerebellum either before or after the administration of tryptophan, the increase of 5-HIAA in the striatum is unlikely to depend appreciably on its production within the cerebral vasculature or outside the brain or on its entering the striatum through a blood-brain barrier damaged by placement of the dialysis probe. Overall, the findings strengthen previous evidence that extracellular 5-HIAA concentrations determined by cerebral dialysis are a valid measure of the metabolism of 5-HT of brain neuronal origin.     

Tryptophan and serotonin turnover rate in the brain of genetically hyperammonemic mice

An investigation was made into the effects of hyperammonemia on the metabolism of brain serotonin (5-HT). The animal model used was the sparse fur (spf) mouse, which possesses an inborn error of the urea cycle, i.e. an abnormal form of ornithine transcarbamylase. Several indoles were measured in brain and plasma using liquid chromatography with electrochemical detection coupled to an u.v. detection (LCEC-u.v.). In the mutant mice, plasma total tryptophan (TRP) was higher when compared with the controls, while plasma free-TRP portion was unchanged. In these animals, brain TRP was increased whilst the 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) levels were significantly higher in the hypothalamus and midbrain. Experiments with NSD-1015 (100 mg/kg i.p.) indicated that the 5-hydroxytryptophan (5-HTP) synthesis rate was increased in the hyperammonemic mice. Pargyline experiments (100 mg/kg i.p.) confirmed the enhanced brain 5-HT turnover rate in the spf mice. In addition, these experiments led to the conclusion that hyperammonemia does not affect the various rate constants. After administration of NSD-1015, TRP level slightly increased in the spf mouse brains, while it was stationary in those of the controls. This result could indicate an increased activity of hepatic TRP-pyrrolase in the hyperammonemic mice. Valine (VAL) administration (200 mg/kg i.p.) reduced brain TRP content in the two kinds of mice, but its effect was of shorter duration in the spf when compared with the control. Comparison of brain tryptamine level indicated a slight but not significant increase in the mutant mice. The data reported here indicate that hyperammonemia may affect peripheral TRP metabolism with consequences upon brain 5-HT synthesis, which could promote certain neurologic disorders.     

Effects of desipramine on regional serotonin synthesis in the rat brain: acute and chronic autoradiographic studies

Various studies have implicated the involvement of noradrenaline (NA) and/or serotonin (5-hydroxytryptamine (5-HT)) in the pathogenesis and treatment of depression. The aim of the present study was to investigate the effects of acute and 7 days of administration of desipramine, a NA re-uptake inhibitor, on the rate of 5-HT synthesis in the rat brain. The study was done by an autoradiographic method using alpha-[14C]-methyl-L-tryptophan as a tracer. The acute (10mg/kg, i.p., 2h before i.v. infusion of the tracer) or 7 days of desipramine (10mg/kg per day, i.p.) did not affect plasma tryptophan (Trp) concentrations, as compared to control (saline treated) rats. Acute treatment with desipramine decreased the rate of 5-HT synthesis in the brain regions that contain 5-HT cell bodies between 19 and 28%, and increased the rate of 5-HT synthesis in the majority of areas containing 5-HT terminals between 21 and 65%. In contrast to the acute treatment, a 7-day administration increased 5-HT synthesis rates in the dorsal raphe (24%), but decreased it in raphe magnus (35%), superior olive (45%), caudate (31%), superior (38%) and inferior (53%) colliculus, and in the auditory cortex (35%). This suggests that the effect of desipramine on 5-HT synthesis rate is time-dependent and differs in the cell bodies and structures containing 5-HT nerve terminals.     

Determination of acteoside in Cistanche deserticola and Boschniakia rossica and its pharmacokinetics in freely-moving rats using LC-MS/MS

A sensitive LC-MS/MS method with a simple solid-phase extraction for the determination of acteoside in rat plasma and tissue homogenates was established for the investigation of bioavailability and brain distribution in freely-moving rats. Acteoside in Cistanche deserticola and Boschniakia rossica was also determined. Acteoside and internal standard were separated on a RP-select B column (125mmx4.6mm i.d., particle size 5microm). The mobile phase consisted of 35% methanol and 65% acetic acid-water (1:100, v/v) at a flow-rate of 1mL/min. Acteoside and the internal standard were monitored using the multiple-reaction monitoring (MRM) mode at m/z transitions of 623-->161 and 609-->301, respectively. The acteoside content was 38.4+/-2.4mg/kg (n=3) for B. rossica, which is obviously lower than 21134.2+/-805.5mg/kg (n=3) of C. deserticola. The protein binding in rat plasma was 75.5+/-1.8%. The brain distribution result indicated that acteoside was evenly distributed in brain tissues (brain stem, cerebellum, the rest of the brain, cortex, hippocampus and striatum) which was about 0.45-0.68% of that in plasma (4.5+/-0.5microg/mL) after 15min of acteoside administration (10mg/kg, i.v.). After acteoside was given (3mg/kg, i.v.; 100mg/kg, p.o.), the oral bioavailability (AUC(p.o.)/dose(p.o.))/(AUC(i.v.)/dose(i.v.)) was only 0.12%.     

Indolic Substances in Plasma, Cerebrospinal Fluid, and Frontal Cortex of Human Subjects Infused with Saline or Tryptophan

Psychiatric patients undergoing the psychosurgical operation of stereotactic subcaudate tractotomy were infused intravenously with either saline or L-tryptophan (15 mg/kg/h). Plasma, lumbar cerebrospinal fluid (CSF), ventricular CSF and a specimen of frontal cortex were collected. The relationships of plasma concentrations of substances claimed to influence brain tryptophan concentration (total tryptophan, free tryptophan, large neutral amino acids) with the concentration of tryptophan in the cortex and CSF were investigated. Tryptophan infusion resulted in plasma tryptophan values comparable to those found after oral doses used in treating depression or insomnia, and about sixfold increases of tryptophan in the cerebral cortex. Increased brain 5-hydroxytryptamine synthesis was indicated by significant rises of CSF 5-hydroxyindoleacetic acid. The concentration of plasma free tryptophan was a better predictor than plasma total tryptophan of cortex tryptophan concentration. As all correlation coefficients of plasma versus brain or plasma versus ventricular CSF tryptophan concentrations were decreased when allowance was made for differences of concentration of large neutral amino acids, the results suggest that the role of these substances within their physiological range as inhibitors of tryptophan transport to the brain may previously have been overemphasised.     

Transport of alovudine (3'-fluorothymidine) into the brain and the cerebrospinal fluid of the rat, studied by microdialysis

Extracellular unbound concentrations of alovudine were sampled by microdialysis in order to study the transport of alovudine between the blood and the brain and the cerebrospinal fluid (CSF) in the rat. The AUC (area under the curve) ratio CSF/blood was higher than the brain/blood ratio after i.v. infusion of alovudine 25mg/kg/hr after a loading dose of 25 mg/kg in 5 minutes (n=4). Neither i.v. infusion of thymidine (25 mg/kg/hr, n=5; 100 mg/kg/hr, n=2) nor acetazolamide (50 mg/kg i.p. bolus followed by 25 mg/kg i.p. every second hour, n=3) influenced the brain/blood AUC ratio after alovudine 25 mg/kg s.c. injection compared to controls (n=5). Finally, perfusion through the microdialysis probe with thymidine (1000 microM, n=3) had also no effect on the brain/blood AUC ratio after alovudine 25 mg/kg s.c. Because neither thymidine nor acetazolamide has significant influence on the ability of alovudine to penetrate the blood-brain barrier in the rat, neither thymidine transport nor carboanhydrase dependent CSF production appear to be major determinants of the blood-brain concentration gradient. Thus, it is concluded that alovudine reaches the extracellular fluid of the brain not by cerebrospinal fluid, but via the cerebral capillaries and that the existence of a concentration gradient over both blood-brain and CSF-brain barrier can probably be explained by the presence of an active process pumping alovudine out from the brain.     

Depletion of epinephrine in rat hypothalamus by a dopamine agonist,pergolide

The i.p. injection of pergolide mesylate, a dopamine agonist, at doses of 0.3–0.6 mg/kg led to a decrease in epinephrine concentration in rat hypothalamus. After a 0.6 mg/kg dose of pergolide mesylate, epinephrine concentration in hypothalamus decreased within 2 hr, reached a minimum concentration at about 8 hrs, and then returned toward control values. Norepinephrine N-methyltransferase activity was not decreased after pergolide injection in vivo nor was it inhibited by pergolide added in vitro at concentrations as high as 10^–3 M. Higher i.p. doses of less potent dopamine agonists, apomorphine (10 mg/kg) and lergotrile (3 mg/kg), also decreased epinephrine concentration in hypothalamus. The pergolideinduced decrease in hypothalamic epinephrine concentration was prevented by pretreatment with haloperidol or spiperone., antagonists of dopamine receptors. Activation of dopamine receptors appears to result in a decrease in epinephrine concentration in rat brain, possibly due to, enhanced release of epinephrine.     

THE EFFECT OF A LOW PROTEIN DIET AND EXOGENOUS INSULIN ON BRAIN TRYPTOPHAN AND ITS METABOLITES IN THE WEANLING RAT

—Male Wistar rats aged 24 days were divided into three groups. Two groups were given a high protein (250 g/kg casein) and a low protein (30 g/kg casein) diet respectively. The third group was given an amount of the high protein diet containing the same amount of energy as that consumed by the low protein diet rats. The plasma of the animals on low protein contained 20% of the concentration of tryptophan of animals on the other two diets. In these animals the concentration of tryptophan was reduced in the forebrain, cerebellum and brain stem, and the concentrations of 5-HT and 5-hydroxyindoleacetic acid were reduced in the forebrain and brain stem. The low protein diet decreased the total uptake of l -[G-³H]tryptophan into the brain and its incorporation into brain protein. Plasma insulin concentrations were reduced in the low protein and ‘restricted high protein’ animals and the plasma corticosterone concentration was raised in the low protein animals. Exogenous insulin did not raise the plasma tryptophan concentration in the low protein animals but it increased the uptake of l -[G-³H]tryptophan into the brain and its incorporation into protein. Rehabilitation for 7 days restored the plasma and brain tryptophan concentrations and those of brain 5-HT and 5-hydroxyindoleacetic acid to control values.