Serotonin Synthesis In Rat Brain Research Articles

Looking at the bigger PICture: understanding and counteracting the decline of persistent inward currents in older adults.

Looking at the bigger PICture: understanding and counteracting the decline of persistent inward currents in older adults.

Effect of chronic protein ingestion on tyrosine and tryptophan levels and catecholamine and serotonin synthesis in rat brain

ObjectivesPrevious studies have shown that brain tyrosine (TYR) levels and catecholamine synthesis rate increase in rats as chronic dietary protein content increases from 2 to 10% (% weight). A single protein, casein, was examined. The present study explores how TYR levels and catecholamine synthesis (and tryptophan (TRP) levels and serotonin synthesis) change when different proteins are ingested chronically over the same range of dietary protein contents.MethodsMale rats ingested for 8 days diets contain 2 or 10% protein (zein, gluten, casein, soy protein, or alpha-lactalbumin). On the last day, they were killed 2.5 hours into the dark period, 30 minutes after receiving an injection of m-hydroxybenzylhydrazine, an inhibitor of aromatic l-amino acid decarboxylase. Brain samples were analyzed for amino acids, including 5-hydroxytryptophan (index of serotonin synthesis rate) and dihydroxyphenylalanine (index of catecholamine synthesis rate), by HPLC-electrochemical detection.ResultsTYR levels and catecholamine synthesis rate in brain were unaffected by the particular protein ingested. However, TRP levels and serotonin synthesis rate varied markedly, depending on the protein ingested, with effects being most prominent in the 10% protein groups. The effect of dietary protein on brain TRP correlated very highly with its effect on serotonin synthesis.DiscussionThe results indicate that the protein ingested can chronically modify TRP levels and serotonin synthesis in brain, but not TYR levels or catecholamine synthesis, with effects most distinct at an adequate level of protein intake (10%).

Distribution of the C1473G polymorphism in tryptophan hydroxylase 2 gene in laboratory and wild mice

The neurotransmitter serotonin is implicated in the regulation of various forms of behavior, including aggression, sexual behavior and stress response. The rate of brain serotonin synthesis is determined by the activity of neuronal-specific enzyme tryptophan hydroxylase 2. The missense C1473G substitution in mouse tryptophan hydroxylase 2 gene has been shown to lower the enzyme activity and brain serotonin level. Here, the C1473G polymorphism was investigated in 84 common laboratory inbred strains, 39 inbred and semi-inbred strains derived from wild ancestors (mostly from Eurasia) and in 75 wild mice trapped in different locations in Russia and Armenia. Among all the classical inbred strains studied, only substrains of BALB/c, A and DBA, as well as the IITES/Nga and NZW/NSlc strains were homozygous for the 1473G allele. In contrast to laboratory strains, the 1473G allele was not present in any of the samples from wild and wild-derived mice, although the wild mice varied substantially in the C1477T neutral substitution closely linked to the C1473G polymorphism. According to these results, the frequency of the 1473G allele in natural populations does not exceed 0.5%, and the C1473G polymorphism is in fact a rare mutation that is possibly eliminated by the forces of natural selection.

Meal ingestion, amino acids and brain neurotransmitters: Effects of dietary protein source on serotonin and catecholamine synthesis rates

Carbohydrate ingestion raises tryptophan uptake and serotonin synthesis in rat brain. The addition of protein is generally believed only to block such increases. However, some recent evidence suggests dietary protein may not be limited to this action. In the present studies, we fed rats single meals containing one of 5 proteins (zein, wheat gluten, soy protein isolate, casein, lactalbumin, 17% by weight) or no protein, and killed them 2.5 h later, 30 min after the injection of m-hydroxybenzylhydrazine, to allow serotonin and catecholamine synthesis rates to be measured in brain. Blood and cerebral cortex samples were analyzed for tryptophan and other large, neutral amino acids; 5-hydroxytryptophan and dihydroxyphenylalanine were measured in hypothalamus, hippocampus and cerebral cortex as indices of serotonin and catecholamine synthesis, respectively. An 8-fold variation occurred in cortex tryptophan: a marked decline followed zein ingestion, and modest reductions after casein or gluten. A large rise in cortex tryptophan occurred after lactalbumin consumption, and smaller increases after soy protein or carbohydrate (no protein). In the brain regions examined, a 4–8-fold range in serotonin synthesis occurred which closely followed the tryptophan alterations. No effects were observed in regional catecholamine synthesis rates. Cortical concentrations of leucine showed small changes; leucine has been linked to mTOR (mammalian target of rapamycin) signaling in brain circuits regulating food intake. The data suggest that tryptophan concentrations and serotonin synthesis in brain neurons are remarkably sensitive to which protein is present in a meal. Conceivably, this relationship might inform the brain about the nutritional quality of the protein ingested.

Regional brain serotonin synthesis rates in male and female irritable bowel syndrome (IBS) patients

Regional brain serotonin synthesis rates in male and female irritable bowel syndrome (IBS) patients

The suitability of [11C]-alpha-methyl-L-tryptophan as a tracer for serotonin synthesis: studies with dual administration of [11C] and [14C] labeled tracer.

The tracer [11C]-alpha-methyl-L-tryptophan (alphaMTP) has been used to measure brain serotonin synthesis rates with positron emission tomography (PET). To address questions about the accuracy of the kinetic model, [14C]alphaMTP was used to directly measure conversion to [14C]-alpha-methyl-serotonin (alphaM5HT) in monkeys that had been previously studied with PET and [11C]alphaMTP. Four male, fasted, isoflurane-anesthetized rhesus monkeys were studied with [11C]alphaMTP and PET. Immediately after the initial 3-hour scan, a second dose of [11C]alphaMTP was coinjected with 1 mCi of [14C]alphaMTP, and additional PET data were collected. Approximately 90 minutes after the second alphaMTP administration, the animals were killed with an overdose of phenobarbital, and brain samples from 21 regions were taken and analyzed by HPLC. Minimal conversion of alphaMTP to alphaM5HT occurred; HPLC analysis of 14C radioactivity showed that greater than 96% of the total counts were in fractions corresponding to the alphaMTP peak. Brain concentrations of serotonin, tryptophan, 5-hydroxyindole-3-acetic acid, and alphaMTP also were determined fluorometrically using external quantification. Patlak plots generated from PET images acquired over 3 hours showed no time period of linear increase, and final slopes were not significantly different from zero, consistent with the finding of minimal conversion to [14C]alphaM5HT. These data indicate that in the 3-hour period after injection, [11C]alphaMTP is acting predominantly as a tracer of tryptophan uptake, not serotonin synthesis.

Biochemical and autoradiographic measurements of brain serotonin synthesis rate in the freely moving rat: a reexamination of the alpha-methyl-L-tryptophan method.

Biochemical approaches were used in freely moving rats to determine, under steady-state conditions, the brain/arterial plasma partition coefficients of L-tryptophan and alpha-[3H]methyl-L-tryptophan, from which the lumped constant for the alpha-methyl-L-tryptophan method of estimating the rate of brain serotonin synthesis is calculated. The lumped constants were significantly different in the various structures examined: 0.149 +/- 0.003 in the raphe dorsalis, 0.103 +/- 0.002 in the raphe centralis, 0.087 +/- 0.003 in the reticular formation, and 0.62 +/- 0.08 in the pineal gland. From these data we proposed a two-compartment model to calculate the rate of serotonin synthesis by quantitative autoradiography using a three-time point experiment. Rates of synthesis for the raphe dorsalis and the reticular formation (620 +/- 57 and 80 +/- 35 pmol/g of tissue/min, respectively) were similar to those measured simultaneously by biochemical means, but rates were 50% higher for the raphe centralis (568 +/- 90 vs. 381 +/- 31 pmol/g of tissue/min). The lack of dynamic equilibrium of the tracer between plasma and tissue pools may explain the discrepancy between the two methods. Our findings did not confirm previous data, indicating that the application of the autoradiographic method to measure the rate of brain serotonin synthesis using alpha-methyl-L-tryptophan as tracer has limitations.

Pet studies of serotonin synthesis in the human brain.

The method for measuring serotonin synthesis in human brain uses [11C]alpha-methyl-L-tryptophan as a tracer and positron emission tomography. The alpha-methyl-L-tryptophan is converted to alpha-methylserotonin, which is not a substrate for monoamine oxidase and therefore accumulates in the brain. In a pilot study published recently, rates of serotonin synthesis were found to be higher in men than in women. This was due to the lower plasma free tryptophan in the women under the experimental conditions used, and does not necessarily reflect the situation in all circumstances. Acute tryptophan depletion lowered brain serotonin synthesis by 90% or more. Patients with borderline personality disorder, who exhibit emotional lability and impulsivity, may have lower brain serotonin synthesis rates than healthy controls.

Brain Serotonin Synthesis Rates in Rhesus Monkeys Determined by [11C]α-Methyl-l-Tryptophan and Positron Emission Tomography Compared to CSF 5-Hydroxyindole-3-Acetic Acid Concentrations

Twelve male, fasted, anesthetized rhesus monkeys were studied with positron emission tomography (PET) and [11C]α-methyl-l-tryptophan (αMTP) to determine serotonin synthesis rates as described by Diksic et al. (1991). It was expected that the serotonin synthesis rates determined for the whole brain would be correlated with CSF 5-hydroxyindole-3-acetic acid concentrations, a measure of central serotonin turnover, because both measures were obtained at steady state. However, no significant correlation was found. During data analysis, it was noticed that the calculated serotonin synthesis rates were significantly correlated to free plasma tryptophan (TP) concentrations (r = 0.88, p < .001). From repeat scans conducted in six monkeys, it was determined that day-to-day variability in free plasma TP and the percentage of protein binding (average percent difference was 48 and 37%, respectively) produced most of the variability in the calculated serotonin synthesis rates (50%); repeat K images, obtained from the PET data alone, differed by only 11%. Calculated serotonin synthesis rates reported for [11C]αMTP PET studies of humans (Nishizawa et al. 1997) and dogs (Diksic et al. 1991) were also highly correlated to reported differences in plasma free TP concentrations. It seems that the [11C]αMTP model for the computation of serotonin synthesis rates is very dependent on plasma free TP concentration and that it may not accurately determine actual serotonin synthesis rates.

Validation of a less-invasive method for measurement of serotonin synthesis rate with alpha-[11C]methyl-tryptophan.

We tested in normal human subjects a less invasive method to obtain plasma input function required in the calculation of the brain serotonin synthesis rate measured with positron emission tomography (PET) and alpha-[11C]methyl-tryptophan (alpha-MTrp). The synthesis rates derived with the arterial input function were compared to those derived from venous plasma and venous sinus time-radioactivity curves obtained from dynamic PET images. Dynamic PET images were obtained for the lengths up to 90 minutes after an injection of alpha-MTrp (400 to 800 MBq). Input functions were generated from both artery and vein in three subjects, and from artery only in two subjects. Net unidirectional uptake constants of alpha-MTrp (K*; mL/g/min) were calculated in several brain regions graphically using data between 20 and 60 minutes after injection with different input functions. In the five subjects with arterial sampling, we tested two methods for correcting the input functions from the venous samples: (1) normalization to the mean exposure time at 20 minutes from arterial curve; and (2) the use of the venous sinus curve for the first 20 minutes. Venous curves coincided with the arterial ones after about 20 minutes. When the venous curves were used, there was an underestimation of the area under the curves up to 20 minutes, resulting in a 5% to 30% overestimation of K* values. Combined use of the sinus curve up to 20 minutes and venous curve from 20 to 60 minutes as an input function resulted in the K* (mL/g/min) values larger by 7.1 +/- 3.8% than the K* values estimated with the arterial input function. Normalization of the venous curve to the exposure time at 20 minutes obtained from the arterial plasma curve resulted in a bias in the K* of about -0.34 +/- 3.32%. The bias from the K* values was propagated to the serotonin synthesis rates. The use of a combination of the venous blood samples and venous sinus as the input function resulted in an acceptable bias in the serotonin synthesis rates from the tissue time-radioactivity curves generated by PET.

115. - Lumped constant for pharmacokinetic estimation of brain serotonin synthesis rates

115. - Lumped constant for pharmacokinetic estimation of brain serotonin synthesis rates

The Acute Effects of Reserpine and NSD-1015 on the Brain Serotonin Synthesis Rate Measured by an Autoradiographic Method

The rate of serotonin (5-HT) synthesis was measured in the discrete regions of the rat brain utilizing an autoradiographic method and α[14C] methyl-L-tryptophan as a tracer after an acute treatment with reserpine (10 mg/kg IP) or NSD-1015 (m-hydroxybenzylhydrazine) (100 mg/kg IP). Controls were injected with the same volume of solvent in place of reserpine or NSD-1015. Our results showed that reserpine induced a statistically significant (except for medial geniculate body) decrease in the rate of 5-HT synthesis in a large number of discrete brain structures. Reserpine had no influence on the plasma concentration of amino acids sharing the same carrier with tryptophan nor on the fraction of plasma-free tryptophan. NSD-1015 induced a statistically significant increase (p <. 05) in the rate of 5-HT synthesis in 20 out of 28 brain regions but produced a pronounced decrease in the rate of 5-HT synthesis in the pineal body. This decrease in the pineal body serotonin synthesis rate is most likely the result of the loss of the label in the form of 5-hydroxy-α[14C]methyl-L-tryptophan [5-OHMTrp] that is not metabolized further because aromatic amino acid decarboxylase was inhibited. The data showing that there was no loss of the 5-OHMTrp from other brain structures as result of reserpine are also given. NSD-1015 treatment also induced a time-dependent increase in the plasma concentration of free tryptophan that becomes significant 30 minutes after NSD-1015 injection. Our results suggest that reserpine induces a decrease in 5-HT synthesis probably via direct or indirect inhibition of tryptophan hydroxylase activity. Since NSD-1015 alone increased the rate of 5-HT synthesis, the measurement of 5-HT synthesis in previous experiments using NSD-1015 and measuring the rate of 5-hydroxytryptophan accumulation after NSD-1015 induced inhibition of decarboxylase activity should be interpreted with reservation.

The acute effects of reserpine and NSD-1015 on the brain serotonin synthesis rate measured by an autoradiographic method

The rate of serotonin (5-HT) synthesis was measured in the discrete regions of the rat brain utilizing an autoradiographic method and alpha[14C]methyl-L-tryptophan as a tracer after an acute treatment with reserpine (10 mg/kg IP) or NSD-1015 (m-hydroxybenzylhydrazine) (100 mg/kg IP). Controls were injected with the same volume of solvent in place of reserpine or NSD-1015. Our results showed that reserpine induced a statistically significant (except for medial geniculate body) decrease in the rate of 5-HT synthesis in a large number of discrete brain structures. Reserpine had no influence on the plasma concentration of amino acids sharing the same carrier with tryptophan nor on the fraction of plasma-free tryptophan. NSD-1015 induced a statistically significant increase (p < .05) in the rate of 5-HT synthesis in 20 out of 28 brain regions but produced a pronounced decrease in the rate of 5-HT synthesis in the pineal body. This decrease in the pineal body serotonin synthesis rate is most likely the result of the loss of the label in the form of 5-hydroxy-alpha[14C]methyl-L-tryptophan [5-OHMTrp] that is not metabolized further because aromatic amino acid decarboxylase was inhibited. The data showing that there was no loss of the 5-OHMTrp from other brain structures as result of reserpine are also given. NSD-1015 treatment also induced a time-dependent increase in the plasma concentration of free tryptophan that becomes significant 30 minutes after NSD-1015 injection. Our results suggest that reserpine induces a decrease in 5-HT synthesis probably via direct or indirect inhibition of tryptophan hydroxylase activity. Since NSD-1015 alone increased the rate of 5-HT synthesis, the measurement of 5-HT synthesis in previous experiments using NSD-1015 and measuring the rate of 5-hydroxytryptophan accumulation after NSD-1015 induced inhibition of decarboxylase activity should be interpreted with reservation.

Alpha-methyl tryptophan as a tracer for in vivo studies of brain serotonin system, from autoradiography to positron emission tomography

The procedure for measuring the brain serotonin synthesis rate in living brain using labelled α-methyl-L-tryptophan (α-MTrp), a tryptophan analog, is described. This tracer could also be used for the anterograde axonal transport measurement of serotonin. One advantage of this method is that α-MTrp is converted in situ into a serotonin tracer, α-methyl serotonin, and should be distributed into the same compartments as serotonin.

Serotonin Synthesis Rate Measured in Living Dog Brain by Positron Emission Tomography

In vivo measurements by positron emission tomography of the brain serotonin synthesis rates in the normal dog, in the dog with increased plasma tryptophan concentration, and in the dog under different arterial oxygen tensions are described. The method described here permits repeated measurements in the same brain for the first time. An increase in the plasma tryptophan concentration from 16.6 to 191.5 and then to 381 microM resulted in close to a linear increase in the brain serotonin synthesis rate. When PaO2 was raised from 76 +/- 2 to 106 +/- 1 mm Hg, the rate of serotonin synthesis in the dog brain increased from 39 +/- 8 to 54 +/- 10 pmol g-1 min-1. The estimates of the Michaelis-Menten constants, Kappm and Vmax, for the transport of tryptophan through the blood-brain barrier are 303 +/- 54 microM and 63 +/- 10 nmol g-1 min-1, respectively.

A New Method to Measure Brain Serotonin Synthesis in vivo. I. Theory and Basic Data for a Biological Model

We describe here an autoradiographic method to measure the in vivo rate of serotonin synthesis in rat brain. The method is based on the use of the L-tryptophan analogue alpha-methyl-L-tryptophan (alpha-MTrp), which is converted in vivo into alpha-methylserotonin (alpha-M5HT). Since alpha-M5HT is not a substrate for monoamine oxidase, it is accumulated in the brain tissue. Data are presented to confirm time-dependent conversion of alpha-MTrp into alpha-M5HT in the dorsal raphe nucleus and also in the pineal body, an organ outside the blood-brain barrier. It has also been shown that washing brain slices in 10% trichloroacetic acid results in less than 3% incorporation of alpha-MTrp into brain proteins. The rates of synthesis are calculated in several grossly dissected brain structures by using tracer kinetics and a three-compartment biological model. The half-life of the precursor pool is estimated to be approximately 20 min. The rate of serotonin synthesis is highest in the pineal body.

Brain 5-HT1 and 5-HT2 binding sites following portacaval shunt in the rat.

Brain serotonin 5-HT1 and 5-HT2 binding properties were investigated in experimental chronic portal-systemic encephalopathy (PSE). End-to-side portacaval shunted (PCS) rats were subjected to open field behavioral testing (spontaneous activity and exploration) 3 weeks after the shunt procedure. Each individual animal was then assayed for 5-HT1 and 5-HT2 binding properties (Bmax and KD) in the cortex + hippocampus by the use of radioligand binding and rapid filtration technique. (3H)serotonin was used to label 5-HT1 binding sites and (3H)ketanserin to label 5-HT2 binding sites. Results revealed that the PCS rats exhibited significant behavioral changes with decreased spontaneous activity and exploratory behavior as compared with sham-operated controls (sham). The affinity for, and the number of, 5-HT1 and 5-HT2 binding sites, respectively, were not different between PCS and sham rats. The brain 5-HT1 and 5-HT2 binding properties were within the range of Bmax and KD previously reported for normal rats when similar techniques are used. This first report in PCS rats on the subject of brain 5-HT1 and 5-HT2 binding properties demonstrates that no major alterations are likely to occur. This contrasts the knowledge of a markedly increased brain serotonin synthesis rate in the PCS rat, suggesting minor functional relevance of the perturbed brain serotonin metabolism associated with chronic PSE.

Plasma amino acids and insulin levels in obesity: Response to carbohydrate intake and tryptophan supplements

We assessed the plasma amino acids, glucose, and insulin responses of obese and lean control subjects to midafternoon carbohydrate snacks. After a standard 400 kcal lunch, eight lean and nine obese subjects received, at 2 pm, a 30 g sucrose snack; blood samples were obtained at hourly intervals until 6 pm. Each subject participated in four similar studies in which the carbohydrate snack was consumed alone or with 250, 500, or 1,000 mg of l-tryptophan (Trp), offered as a capsule. The obese group exhibited elevated plasma levels of the branched-chain amino acids, phenylalanine and tyrosine, and the levels of these amino acids declined much less in response to carbohydrate intake than in lean controls. As a consequence, the plasma ratio of Trp:large neutral amino acids (Trp/LNAA ratio), which normally rises after carbohydrate consumption, showed virtually no change in the obese group. The plasma Trp/LNAA response of this group did not reach control values even when carbohydrate intake was increased to 50 or 75 g. Peak plasma Trp concentrations and Trp/LNAA ratios after 250, 500, and 1,000 mg Trp doses were also significantly lower in the obese. Since brain Trp uptake is strongly correlated with the plasma Trp/LNAA ratio, which in turn determines the rate of brain serotonin synthesis, the blunted Trp/LNAA response to carbohydrate intake in the obese could contribute to alterations in the serotonin-mediated regulation of food intake.

Brain serotonin metabolism and behavior in rats with carbon tetrachloride-induced liver cirrhosis.

Increased brain serotonin metabolism has been suggested as an etiologic factor in the development of portal-systemic encephalopathy (PSE) in connection with liver disease. We therefore investigated brain serotonin metabolism and open-field behavior (spontaneous activity and exploration) in rats with carbon tetrachloride (CCl4)-induced liver cirrhosis. Brain serotonin metabolism was evaluated in rats pretreated with an amino acid decarboxylase inhibitor. The 5-hydroxyindoles were analyzed by high-performance liquid chromatography (HPLC) with electrochemical detection. The results revealed an increased serotonin synthesis rate in all investigated brain regions in rats with histologically verified diffuse micronodular cirrhosis of the liver. Slightly impaired open-field behavior (i.e., decreased spontaneous activity) in the cirrhotic rats could not be excluded. However, the elevated brain serotonin synthesis rate could not be correlated to any abnormalities in open-field behavior.

Stimulation of serotonin synthesis in rat brain after antiepilepsirine, an antiepileptic piperine derivative

Piperine and two of its derivatives, antiepilepsirine (AE or 3,4,methylendioxycynna-moylpiperine) and compound 7448 ( N-isopropyl 3 (4 chloro-phenyl) propenoylamide) are very effective in stimulating serotonin (5HT) synthesis. AE raises the ratio of free-to-bound tryptophan (TP) in plasma and induces a long-lasting increase of this aminoacid in brain. At the same time in striatum and limbic area it causes a lasting increase in 5 hydroxyindolacetic acid (5HIAA) a 5HT metabolite and to a lesser extent, an increase in the levels of the monoamine itself. Together with this action on 5HT metabolism we found that AE caused release of 3H-5HT from an in vitro synaptosomal preparation. It thus appears that piperine and its derivatives AE and compound 7148 affect the central serotonergic system.