Probenecid-induced accumulation of 5-hydroxyindoleacetic acid and homovanillic acid in rat brain

The accumulation of 5-hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA) in rat brain has been examined after probenecid infusion over 8 h. At plasma probenecid concentrations of 200-400 micrograms mL-1 a steady state level in the accumulation of 5-HIAA and HVA was achieved, the increase above the endogenous levels being 135% and 65%, respectively. When the plasma concentration of probenecid rose above 400 micrograms mL-1 there was further accumulation of both 5-HIAA and HVA probably induced by increased neuronal activity or toxicity due to probenecid. The explanation for the plateau of 5-HIAA and HVA obtained over the plasma probenecid concentration interval of 200-400 micrograms mL-1 could be that the levels were reached when there was complete inhibition of active transport, and when the rate of formation of the metabolites equalled the rate of elimination by alternative routes i.e. bulk flow and diffusion. Therefore when probenecid is used to inhibit the active transport of acid monoamine metabolites across the blood-brain barrier, its plasma concentration should be within the range of 200-400 micrograms mL-1.     

Simultaneous determination of femtomole quantities of 5-hydroxytryptophan, serotonin and 5-hydroxyindoleacetic acid in brain using HPLC with electrochemical detection

A simple rapid method for the determination of 5-hydroxytryptophan, serotonin (5-hydroxytryptamine), and 5-hydroxyindoleacetic acid in brain is presented. Brain proteins are precipitated with Zn(OH)2. The indoles in the supernatant are separated by HPLC in less than than 8 min on a reverse phase column and detected electrochemically. As little as 38 fmol of hydroxyindole compound can be detected and quantitated. Because the method is rapid and uncomplicated many samples can be processed in a day.     

Regional rat brain levels of 3,4-dihydroxyphenylacetic acid and homovanillic acid: concurrent fluorometric measurement and influence of drugs

A concurrent semi-automatic fluorometric assay technique for 3, 4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), is described. The method is based on a rapid manually performed isolation of DOPAC and HVA on small columns of Sephadex G-10 followed by an automated fluorometric detection with a continuous flow system. DOPAC and HVA were measured in the corpus striatum, nucleus accumbens and olfactory tubercle of the rat, under normal conditions and after treatment with amphetamine, apomorphine, clozapine, haloperidol, morphine, oxotremorine, pargyline, probenecid, sulpiride and thioridazine. Clozapine, morphine, sulpiride and oxotremorine induced the most pronounced rise of DA metabolites in the nucleus accumbens. Probenecid produced a DOPAC accumulation in the nucleus accumbens. Striking differences were observed between the DOPAC/HVA ratios in the different structures of control animals. The concurrent assay enables a rapid screening of the action of drugs in regional DA metabolism.     

The estimation of 3,4-dihydroxyphenylacetic acid, homovanillic acid and homo-isovanillic acid in nervous tissue by gas-liquid chromatography and electron capture detection.

1 A gas chromatographic method using electron capture detection is described for the estimation of three acidic metabolites of dopamine, 4-hydroxy-3-methoxyphenylacetic acid (homovanillic acid, HVA), 3,4-dihydroxyphenylacetic acid (DOPAC) and 3-hydroxy-4-methoxyphenylacetic acid (homo-isovanillic acid, iso-HVA). The method is based on the formation of the trifluoroacetyl-hexafluoroisopropyl derivatives of the three acids. 2 The method has been applied to the estimation of DOPAC, HVA and iso-HVA in tissues from the central and peripheral nervous systems.     

Simultaneous determination of dopamine and 3,4-dihydroxyphenylacetic acid in mouse striatum using mixed-mode reversed-phase and cation-exchange high-performance liquid chromatography

The measurement of dopamine and 3,4-dihydroxyphenylacetic acid (DOPAC) is useful for an index of dopamine turnover. We developed a simultaneous determination method for dopamine and DOPAC with high-performance liquid chromatography-fluorescence detection. Mixed-mode reversed-phase and cation-exchange column (CAPCELL PAK CR column) which contained C18 silica particles and sulfonic acid cation-exchange particles was used for the separation of 3,4-dihydroxy-l-phenylalanine, catecholamines (norepinephrine, epinephrine, and dopamine) and DOPAC. The mobile phase was optimized for factors such as pH and counter ion concentration. With a mobile phase of 15 mM sodium acetate buffer (pH 4.5), separation was achieved within 22 min. The developed method was applicable to the determination of dopamine and DOPAC in mouse striatum. The concentrations of dopamine and DOPAC in mouse striatum were 4.98 ± 0.66 and 1.00 ± 0.11 μM, respectively (n = 10).     

Simultaneous assay for L-tryptophan, serotonin, 5-hydroxyindoleacetic acid, norepinephrine and dopamine in brain

A routine simultaneous assay for the title compounds is described, which uses a cation-exchange resin for separations, and standard fluorometric methods for analyses. Practicability of the ion-exchange chromatography is enhanced by means of a novel apparatus, and the procedure has the flexibility to permit extension to other endogenous compounds using published techniques.     

Enhancement of haloperidol-induced increase in rat striatal or mesolimbic 3,4-dihydroxyphenylacetic acid and homovanillic acid by pretreatment with chronic methamphetamine

After a drug-free period of 1 week following 2 weeks of haloperidol treatment, the increased response of striatal 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) to a challenge dose of haloperidol was significantly reduced. Tolerance to this effect was not, however, seen in the mesolimbic system. Pretreatment of the rats with methamphetamine (MAP) for 8 days prior to chronic haloperidol significantly enhanced the DOPAC and HVA increase produced by the challenge with haloperidol in both brain areas. The reduced response of striatal DOPAC or HVA after chronic haloperidol was prevented by pretreatment with MAP. The data suggest that the long-term dopamine receptor stimulation induced by MAP may antagonize the tolerance produced by chronic haloperidol treatment.     

Influence of convulsive therapy on 5-hydroxyindoleacetic acid and homovanillic acid in cerebrospinal fluid in endogenous depression

The cerebrospinal fluid (CSF) levels of 5-hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA) were determined in 20 patients with endogenous depression before and 2–6 days after a full course of convulsive therapy, the depressive symptoms being simultaneously rated.

1. The level of 5-HIAA was similar to that in previous series of healthy controls while the level of HVA appeared somewhat low.
2. The levels did not change after therapy in spite of considerable clinical improvement.
3. There was no relation between the levels and the severity of the depressive state, nor between changes of the levels and degree of clinical improvement.
4. There was no relation between the levels and the severity of the depressive state, nor between changes of the levels and degree of clinical improvement.

The validity of determination of acid monoamine metabolites in CSF relative to the cerebral turn-over of the amines may be increased, if the elimination of metabolites from CSF is blocked with probenecid.     

Effects of altered brain 5-hydroxytryptaminergic activity on brain tryptophan, 5-hydroxytryptamine and 5-hydroxyindoleacetic acid

The effects of procedures known to alter 5-hydroxytryptaminergic activity (raphe stimulation, raphe lesions and exposure to elevated temperature) on brain tryptophan, 5-hydroxytryptamine and 5-hydroxyindoleacetic acid concentrations were investigated. Electrical stimulation of the dorsal raphe nucleus of unanaesthetised unrestrained rats for 1 hr, using implanted electrodes, significantly increased 5-hydroxyindoleacetic acid concentrations in the hypothalamus, striatum, midbrain and cortex but not in the hippocampus. The level of 5-hydroxytryptamine fell proportionately with the rise of 5-hydroxyindoleacetic acid in the midbrain but did not alter significantly in any other region. Tryptophan concentration was not significantly altered except in the hypothalamus where it increased moderately. Rats kept at 40°C for 60 min had significantly increased plasma and brain tryptophan and brain 5-hydroxyindoleacetic acid. Lysergic acid diethylamide prevented only the increase in 5-hydroxyindoleacetic acid. Electrolytic lesions in the dorsal and median raphe nuclei markedly lowered brain 5-hydroxytryptamine and 5-hydroxyindoleacetic acid but had no effect on brain or plasma tryptophan. Therefore, evidence is against a major role for brain tryptophan changes in the regulation of 5-hydroxytryptamine synthesis following alteration of 5-hydroxytryptaminergic activity. Other possible factors involved in the relationship of 5-hydroxytryptamine metabolism to neuronal activity are discussed.     

Rapid method for the determination of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid in small regions of rat brain

A rapid and sensitive method for measuring 5-hydroxytryptamine and 5-hydroxyindoleacetic acid, using o-phthalaldehyde and L-cysteine, is presented, enabling both compounds to be measured in small areas of rat brain.     

Determinations of 5-hydroxyindoleacetic acid and homovanillic acid in human CSF with monitoring of probenecid levels in CSF and plasma

The accumulation of 5-HIAA and HVA in cerebrospinal fluid (CSF) was studied in eight healthy volunteers after oral administration of probenecid. Simulation indicated that a dose of 4.5 g probenecid should be used to achieve probenecid plasma concentrations between 200 and 400 g/ml. Almost complete inhibition of the active transport of the acidic metabolites was assumed to be obtained at these concentrations. Probenecid 4.5 g was administered in two doses (2.5 g and 2 g), separated by 4 h. Plasma samples were drawn at varying intervals over a period of 46 h and lumbar puncture (LP) was performed at either 14 h or 20 h after the first administration of probenecid. The concentration of probenecid, 5-HIAA and HVA in CSF was estimated and the probenecid-induced accumulation of 5-HIAA and HVA was compared with their baseline values. There were no statistically significant differences (P>0.05) in the accumulation of the monoamine metabolites between the two LP (14 h and 20 h), neither were there any differences in CSF concentrations of probenecid at the time of LP. There were only small differences in probenecid plasma concentrations, although statistically significant. Due to maximum blockade of the active transport system no correlation was observed between the CSF concentration of probenecid and the induced accumulation of 5-HIAA and HVA, respectively. The range of probenecid-induced accumulation for 5-HIAA and HVA in these volunteers was 156–429% and 183–600%, respectively. The suggested monitoring of probenecid plasma levels is proposed as a suitable model to investigate central neuronal activity of dopamine and serotonin in the central nervous system.     

Effects of prolintane on 3,4-dihydroxyphenylacetic acid concentration in rat brain after spiperone treatment

Prolintane (1-[alpha-propylphenethyl]-pyrrolidine) but not l-(alpha-methylphenethyl)-pyrrolidine markedly enhanced the increase in 3,4-dihydroxyphenylacetic acid concentration in the brains of rats treated with spiperone, a neuroleptic drug. This action and other properties of prolintane described in the literature place it in a group of stimulant drugs that includes methylphenidate, cocaine and amfonelic acid with properties that differ from those of amphetamine.     

Effect of short-term swimming stress and diazepam on 3,4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindoleacetic acid (5-HIAA) levels in the caudate nucleus: an in vivo voltammetric study

Summary The effects of swimming stress on dopaminergic and serotonergic neurons were studied by an in vivo voltammetry technique. 3,4-Dihydroxyphenylacetic acid (DOPAC) and 5-hydroxyindoleacetic acid (5-HIAA) levels in rat striatum were measured by differential pulse voltammetry with an electrochemically treated carbon fiber electrode. Exposure to swimming stress for 1 to 10 min to the animal increased the DOPAC and 5-HIAA peaks, which depended on the length of stress. Pretreatment of the rats with diazepam (10 mg/kg, i.p.) prevented completely the stress-induced increase in DOPAC levels but only partially reduced the increase in 5-HIAA levels.