A pilot in vivo proton magnetic resonance spectroscopy study of amino acid neurotransmitter response to ketamine treatment of major depressive disorder.

The N-methyl-D-aspartate receptor antagonist ketamine can improve major depressive disorder (MDD) within hours. To evaluate the putative role of glutamatergic and GABAergic systems in ketamine's antidepressant action, medial prefrontal cortical (mPFC) levels of glutamate+glutamine (Glx) and γ-aminobutyric acid (GABA) were measured before, during, and after ketamine administration using proton magnetic resonance spectroscopy. Ketamine (0.5 mg kg(-1) intravenously) was administered to 11 depressed patients with MDD. Glx and GABA mPFC responses were measured as ratios relative to unsuppressed voxel tissue water (W) successfully in 8/11 patients. Ten of 11 patients remitted (50% reduction in 24-item Hamilton Depression Rating Scale and total score ⩽10) within 230 min of commencing ketamine. mPFC Glx/W and GABA/W peaked at 37.8%±7.5% and 38.0%±9.1% above baseline in ~26 min. Mean areas under the curve for Glx/W (P=0.025) and GABA/W (P=0.005) increased and correlated (r=0.796; P=0.018). Clinical improvement correlated with 90-min norketamine concentration (df=6, r=-0.78, P=0.023), but no other measures.

Relationship of resting brain hyperconnectivity and schizophrenia-like symptoms produced by the NMDA receptor antagonist ketamine in humans.

N-methyl-D-aspartate glutamate receptor (NMDA-R) antagonists produce schizophrenia-like positive and negative symptoms in healthy human subjects. Preclinical research suggests that NMDA-R antagonists interfere with the function of gamma-aminobutyric acid (GABA) neurons and alter the brain oscillations. These changes have been hypothesized to contribute to psychosis. In this investigation, we evaluated the hypothesis that the NMDA-R antagonist ketamine produces alterations in cortical functional connectivity during rest that are related to symptoms. We administered ketamine to a primary sample of 22 subjects and to an additional, partially overlapping, sample of 12 subjects. Symptoms before and after the experimental session were rated with the Positive and Negative Syndrome Scale (PANSS). In the primary sample, functional connectivity was measured via functional magnetic resonance imaging almost immediately after infusion began. In the additional sample, this assessment was repeated after 45 min of continuous ketamine infusion. Global, enhanced functional connectivity was observed at both timepoints, and this hyperconnectivity was related to symptoms in a region-specific manner. This study supports the hypothesis that pathological increases in resting brain functional connectivity contribute to the emergence of positive and negative symptoms associated with schizophrenia.

NMDA antagonist effects on striatal dopamine release: positron emission tomography studies in humans.

Previous brain imaging studies with [(11)C]raclopride have suggested that the psychotogenic effects of the noncompetitive N-methyl-D-aspartate antagonist ketamine in humans might be mediated by increased dopamine (DA) release and increased stimulation of DA D(2) receptors in the striatum. The goal of the present study was to assess the effect of ketamine on D(2) receptor availability in subregions of the striatum (dorsal caudate, DCA; dorsal putamen, DPU; ventral striatum, VST) in humans. Ten healthy subjects were studied twice. In a first group of five subjects, PET scanning was obtained twice for 90 min during bolus plus constant infusion of [(11)C]raclopride. No significant differences were observed in [(11)C]raclopride specific-to-nonspecific activity ratios (V(")(3)) measured during an early interval (30-50 min) and late interval (70-90 min), confirming that a state of sustained equilibrium had been established from 30-90 min (end of infusion). In a second group of five subjects, a similar experiment was performed twice, except that ketamine was administered beginning at 50 min (0.12 mg/kg i.v. bolus followed by 0.65 mg/kg/h i.v. infusion for 70 min). Raclopride V(")(3) measured before ketamine (30-50-min interval) was compared to [(11)C]raclopride V(")(3) measured during ketamine infusion (70-90-min interval). Ketamine induced a robust dissociative state. However, no significant differences were observed in D(2) receptor availability measured before and during the ketamine infusion (n = 10) in any of the regions examined (DCA, DPU, and VST). These data fail to demonstrate an effect of ketamine on [(11)C]raclopride BP and are consistent with microdialysis studies in rodents and nonhuman primates which reported only small effects of acute NMDA receptor blockade on extracellular striatal DA concentration.

Sex-related differences in nortriptyline-induced side-effects among depressed patients.

1. Men and women may differ in their pharmacokinetic responses to tricyclic antidepressants (TCAs), in a number of autonomic indices, and in various adrenergic receptor mediated responses. Emerging evidence also suggests that women may have a lower rate of serotonin synthesis in brain and a greater sensitivity to the depressant effects of tryptophan depletion, relative to men. However, sex-related differences in TCA-induced side-effects, including increases in heart rate (HR), dry mouth, constipation, and difficulty urinating, has not been systematically investigated. 2. The authors examined potential sex-related differences in the pattern of side-effects during treatment with nortriptyline (NT), a TCA that is still widely used. Seventy-eight healthy outpatients who met Research Diagnostic Criteria and DSM-III-R criteria for major depression participated in a double-blind, randomized parallel trial of NT versus placebo. 3. Each subject was acutely challenged with either placebo or 50 mg NT prior to and after a 6-week treatment with NT. NT doses were adjusted weekly to maintain therapeutic plasma levels. Patients were assessed at multiple time points to detect the presence of NT-induced side-effects. 4. The initial, single (50 mg) dose of NT significantly increased supine HR. Six-week treatment with NT was found to significantly increase supine and sitting HRs, irrespective of sex. In rechallenge with the single NT dose, there were no significant effects on HR. 5. When sex-related differences were examined, HR increases were greater in men than women during weeks 4 through 6 of the NT treatment, although no sex-related differences were present in plasma NT levels or metabolites. In addition, there was a significant NT to placebo difference in self-rated dry mouth for women during all 6-weeks of treatment, whereas men showed a significant NT-placebo difference during weeks 3 and 5. 6. The results suggest the presence of sex-related differences in elevated supine HR response during the course of 6-week NT treatment. Depressed men may be more susceptible to NT-induced increases in supine HR than women.

Separation methods for tricyclic antiviral drugs.

A review of the published analytical methodology for the tricyclic antiviral (TAV) drugs is presented. While amantadine and rimantadine are the only two approved drugs for the prophylaxis and treatment of the influenza A virus, amantadine has also been approved for the treatment of Parkinson's disease. In addition, a few structurally related compounds are finding important clinical applications in other central nervous system-related disorders. To effectively evaluate the pharmacokinetics, biotransformations, stability, and other critical parameters that are necessary for pre-clinical and clinical studies, analytical methodology that conforms to the rigors of regulatory requirements must be developed and made available. This review discusses the analytical methods used in the determination of amantadine, rimantadine, tromantadine and memantine and the pre-clinical and clinical application of these techniques.

Differential expression of carnosine, homocarnosine and N-acetyl-L-histidine hydrolytic activities in cultured rat macroglial cells.

N-acetyl-L-histidine (NAH) and N-acetyl-L-aspartate (NAA) are representatives of two series of substances that are synthesized by neurons and other cells in the vertebrate central nervous system (CNS). Histidine containing homologs of NAH are beta-alanyl-L-histidine or carnosine (Carn) and gamma-aminobutyrl-L-histidine or homocarnosine (Hcarn). A homolog of NAA is N-acetylaspartylglutamate (NAAG). These substances belong to a unique group of osmolytes in that they are synthesized in cells that may not to be able to hydrolyze them, and are released in a regulated fashion to a second compartment where they can be rapidly hydrolyzed. In this investigation, the catabolic activities for NAH, Carn, and Hcarn in cultured macroglial cells and neurons have been measured, and the second compartment for NAH and Hcarn has been identified only with astrocytes. In addition, oligodendrocytes can only hydrolyze Carn, although Carn can also be hydrolyzed by astrocytes. Thus, astrocytes express hydrolytic activity against all three substrates, but oligodendrocytes can only act on Carn. The cellular separation of these hydrolytic enzyme activities, and the possible nature of the enzymes involved are discussed.

Effects of ethanol and of alcohol dehydrogenase inhibitors on the reduction of N-acetylaspartate levels of brain in mice in vivo: a search for substances that may have therapeutic value in the treatment of Canavan disease.

N-Acetylaspartate (NAA) is an important osmolyte in the vertebrate brain that participates in an intercompartmental metabolic cycle. It is synthesized primarily in neurons from L-aspartate (Asp) and acetyl-CoA and, after its regulated release, it is hydrolysed by aspartoacylase in an oligodendrocyte compartment to produce Asp and acetate. NAA also gives a strong 1H magnetic resonance spectroscopic signal, which has led to its widespread use as a neuronal marker. Utilizing this noninvasive technique, the NAA concentrations in normal brain and in brains exhibiting a variety of CNS disease syndromes have been studied. In normal individuals, the concentration of NAA has been observed to be relatively stable over long periods. However, in many CNS disease processes there are long-term changes in the level of NAA that have been considered to signal changes in neuron density or function. We report that the concentration of NAA in brain is malleable and that, in addition to normal endogenous variation or changes due to disease processes, it can be modified by a variety of exogenous drugs and other substances. As a result of this investigation, we have also been able to identify a new class of NAA-active compounds--pyrazole and pyrazole derivatives--that have the ability to reduce brain NAA concentrations in normal mice. The importance of these findings in understanding the NAA intercompartmental cycle, and its role in Canavan disease, a genetic aspartoacylase deficiency disease, are discussed.

Disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments in rats.

BACKGROUND: This study was designed to determine the disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments, using multiple sampling time points in chronically prepared awake pregnant rats. METHODS: All animals received an intravenous infusion of bupivacaine at a rate of 0.33 mg. kg-1. min-1 over a period of 15 min. The fetuses were delivered either at the end of infusion or at 2 or 4 h after dosing. Maternal and fetal blood and tissue samples were obtained for the assays of bupivacaine and its metabolites using capillary gas chromatography-mass spectrometry. RESULTS: The elimination half-life of bupivacaine was 37.7 min. The major metabolite was 3'-hydroxybupivacaine. Bupivacaine and 3'-hydroxybupivacaine were present in all samples at the end of administration. The fetal to maternal concentration ratio of bupivacaine in plasma was 0.29, and in the placenta was 0.63. The amnion contained the highest bupivacaine concentration: threefold higher in the maternal and 11-fold higher than in the fetal plasma. At 4 h after dosing, bupivacaine was no longer detectable in any maternal and fetal samples, whereas 3'-hydroxybupivacaine was still present in all tissues except the fetal plasma and heart. CONCLUSIONS: These data indicate that a considerable amount of bupivacaine is taken up by both sides of the placenta, as well as the amnion and myometrium. 3'-Hydroxybupivacaine was present in all tissues except the fetal plasma and heart samples, even after the parent compound became no longer detectable. Whether this slow elimination of 3'-hydroxybupivacaine causes any adverse effects on the fetus-newborn needs to be explored.

Sensitive and selective liquid chromatographic assay of memantine in plasma with fluorescence detection after pre-column derivatization.

A procedure was developed for the determination of memantine in plasma using liquid chromatography with fluorescence detection. Following a liquid-liquid extraction from 1 ml of plasma containing the internal standard amantadine, the extract was derivatized at room temperature with dansyl chloride, and the highly fluorescent derivatives were chromatographed with a reversed-phase C18 column and a mobile phase of phosphate buffer and acetonitrile. Dansylated memantine and amantadine were eluted in less than 13 min with no interference from endogenous material. The calibration curve was linear over the concentration range of 3-400 ng/ml with inter- and intra-assay imprecision (C.V.) of less than 10%. The limit of quantitation was 3 ng/ml, and no major antidepressant, neuroleptic or their respective metabolites interfered with the quantitation of memantine. This method could also be applied to the quantitation of amantadine.

Phenylalanine kinetics are associated with tardive dyskinesia in men but not in women.

RATIONALE: An association between tardive dyskinesia (TD) and severely impaired metabolism of the large neutral amino acid (LNAA), phenylalanine (Phe) was defined in a group of mentally retarded patients. Subsequently, an altered kinetics of Phe was associated with TD in men with schizophrenia based on plasma analyses subsequent to the ingestion of a protein meal. METHODS: In the present study, a standardized oral challenge of pure Phe (100 mg/kg in 170 ml orange juice) was administered to psychiatric patients of both sexes (n = 312), with and without TD after an overnight fast. Plasma LNAA levels were assayed both fasting and 2 h subsequent to the ingestion of the challenge. The extent of the increase in plasma Phe levels 2 h following a standardized challenge is determined by the sum of the kinetic processes of plasma absorption, tissue distribution, metabolism and elimination. RESULTS: The study hypothesis, that TD would be associated with significantly higher post-challenge plasma Phe indices of an absolute plasma Phe level and plasma Phe/LNAA ratio (a brain availability measure), was verified for the study men (n = 209), but not for the study women (n = 103). CONCLUSIONS: The demonstrated altered kinetics of Phe in men with TD indicates a greater availability of Phe to the brain in these men. We suggest that the disorder may be related to the effects of this greater availability. Such effects could be the direct neurotoxic effects of Phe and its metabolites and/or the modulating effects of these compounds on the synthesis of the monoamine neurotransmitters. The fact that TD (Yes/No) group differences in post-challenge plasma Phe indices were not seen for the study women suggests the possibility of a sex difference in the biology of TD that we propose may be reflective of the young age of the study sample.

Branched chain amino acids decrease tardive dyskinesia symptoms.

RATIONALE: Prior studies had suggested (a) that a lessened ability to clear ingested forms of the large neutral amino acid (LNAA), phenylalanine (Phe), was associated with having tardive dyskinesia (TD), and (b) that greater availability of a group of LNAA, the branched chain amino acids (BCAA), concomitant with the lower availability of Phe to the brain are associated with a decrease in TD symptoms. The present study was then conducted to test whether increasing the daily intake of the BCAA would decrease the symptoms of TD. METHODS: A 2-week trial of a BCAA medical food administered three times a day was conducted in nine men with long neuroleptic treatment histories. Frequency counts of TD movements were collected by videotape throughout the trial and these tapes were analyzed in blind random sequence for both patient and time for TD symptom level changes subsequent to completion of the trial. Plasma levels of the LNAA were also collected throughout the trial. RESULTS: A statistically significant decrease in the level of TD symptoms was observed for the sample. The symptom changes were also clinically significant in that six of the nine subjects had symptom decreases of at least 58%, with all subjects having a decrease of at least 38%. BCAA administration increased plasma BCAA concentrations and BCAA/LNAA ratios and decreased plasma Phe concentrations and the Phe/LNAA ratio. Analyses indicated a strong significant correlation between the percent increase in the plasma BCAA values at the first administration and the percent improvement in TD over the trial in eight of the nine subjects. CONCLUSIONS: The BCAA show promise as a treatment for TD. The decrease in TD symptoms seen in the trial may have been modulated by the BCAA treatment-induced increased availability of the BCAA and decreased availability of Phe to the brain.

Expression of aspartoacylase activity in cultured rat macroglial cells is limited to oligodendrocytes.

N-acetyl-L-aspartate (NAA) is an important osmolyte in the vertebrate brain and eye, and its cyclical metabolism is accomplished in two separate compartments. In the brain, NAA is synthesized primarily in neurons, and after its regulated release, NAA is hydrolyzed by aspartoacylase, which is present in a glial-associated compartment. However, the precise nature of this hydrolytic compartment has remained obscure. It has been proposed that one role of aspartoacylase in the central nervous system (CNS) is as part of a molecular water pump (MWP) that uses the NAA intercompartmental cycle to remove nerve cell metabolic water against a water gradient and that oligodendrocytes comprise the second compartment in this metabolic sequence. The absence of aspartoacylase activity in Canavan disease (CD), a rare early onset genetic spongiform leukodystrophy, is associated with CNS edema, intramyelinic swelling and a progressive loss of oligdendrocytes. In order to evaluate the MWP hypothesis and its possible relationship to the etiology of CD further, both oligodendrocytes and astrocytes obtained from neonatal rat brain were grown in culture and tested for the presence of aspartoacylase activity. The results of this study show for the first time that aspartoacylase activity is expressed only in oligodendrocytes. The meaning of this observation in understanding the function of the NAA metabolic cycle is discussed.

Interactive effects of subanesthetic ketamine and subhypnotic lorazepam in humans.

Ketamine is an N-methyl-D-aspartate (NMDA) receptor antagonist with psychotogenic and dissociative effects in healthy humans. These cognitive and perceptual effects in humans are reportedly reduced by benzodiazepine premedication. This study assessed the interactive effects of a ketamine (i.v. bolus of 0.26 mg/kg followed by an infusion of 0.65 mg/kg per hour) and lorazepam 2 mg., PO, in humans. Twenty-three healthy subjects completed 4 test days involving the oral administration of lorazepam or matched placebo 2 h prior to the i.v. infusion of ketamine or placebo. Ketamine: 1) produced behaviors similar to the positive and negative symptoms of schizophrenia as assessed by the Brief Psychiatric Rating Scale (BPRS); 2) evoked perceptual alterations as measured by the Clinician-Administered Dissociative States Scale (CADSS); 3) impaired performance on the Wisconsin Card Sorting Test (WCST) and other tests sensitive to frontal cortical impairment; and 4) had amnestic effects. Lorazepam produced attention impairments, concrete proverb interpretations, and recall impairments. Lorazepam reduced ketamine-associated emotional distress and there was a non-significant trend for it to decrease perceptual alterations produced by ketamine. However, it failed to reduce many cognitive and behavioral effects of ketamine, including psychosis. Further, lorazepam exacerbated the sedative, attention-impairing, and amnestic effects of ketamine. There was no evidence of pharmacokinetic interaction between these medications. These data suggest that subhypnotic lorazepam and ketamine show a spectrum of interactive effects, ranging from antagonism to potentiation.

Enantiomeric determination of the phenylmorpholinol metabolite of bupropion in human plasma using coupled achiral-chiral liquid chromatography.

A coupled achiral-chiral stationary phase liquid chromatographic technique was developed to separate and quantitate the enantiomers of the phenylmorpholinol metabolite (2) of the antidepressant bupropion (1) in human plasma. At the retention time of 2, a switching valve loaded a portion of the eluting compound onto a protein-bonded chiral stationary phase which resolved 2 into the (+) and (-) stereoisomers using an aqueous mobile phase of potassium phosphate (pH = 6.25) and 5% 2-propanol. All eluting compounds were monitored using UV detection at 214 nm, and no plasma endogenous material or other commonly used psychotropic drugs were found to interfere. Within-day and between-day variation were less than 6% over the expected concentration range, and a limit of quantitation of about 125 ng/mL of 2 was observed. Steady-state plasma samples from 17 patients receiving 1 were found to contain the (-) enantiomer to the extent of about 96% of total 2. The potential clinical implications of these results are not known since all previous pharmacological studies were carried out with the racemic 2.

Effects of dopamine agonists on Cebus apella monkeys with previous long-term exposure to fluphenazine.

Sixty-one weeks after 48 weeks of treatment with fluphenazine decanoate or placebo, 37 socially living Cebus apella monkeys were evaluated for differences in dopaminergic sensitivity by exposure to 0.75 mg/kg, i.m. of amphetamine (AMPH) (indirect agonist) and apomorphine (APOM) (direct agonist). The fluphenazine-treated animals differed (p < or = 0.05) from control animals on some hourly measures of composite behavioral variables (CBVs). Animals exposed to fluphenazine showed a greater decrease in the aggressiveness CBV and a smaller decrease in self- and environment-directed behaviors than placebo animals. CBVs for normal locomotion and directs affiliation showed no significant differences. The fluphenazine-treated group showed greater agonist induction of stereotypic behavior (p < or = 0.01), and larger decreases in prolactin response to AMPH (p < or = 0.05). Our findings indicate that following extended treatment with an antipsychotic there is increased sensitivity to dopamine, as evidenced by stereotypies and possibly hypophyseal responsiveness.

2-Hydroxydesipramine and desipramine plasma levels: how are they related to antidepressant response?

Thirty-six outpatients aged 20 to 51 with RDC primary major depressive disorder (MDD) completed a 5-week trial of desipramine following a week of single-blind placebo. Five had a past history of hypomanic disorder. For all but one patient, daily dosage at bedtime was constant for the final 4 weeks, with a mean (S.D.) of 168.1 (46.5) mg. Plasma samples drawn at the three final weekly visits were assayed by high-performance liquid chromatography for 2-hydroxydesipramine (2-OH-DMI) and desipramine. Mean (S.D.) plasma levels were 59.8 (30.0) ng/ml for 2-OH-DMI and 142.9 (138.6) ng/ml for desipramine. Thirteen patients (36%) had a final 17-item Hamilton depression rating < and = 6 and were classified as responders. According to receiver operating characteristics analysis, patients with plasma 2-OH-DMI levels > and = 58 and < 92 ng/ml had a greater likelihood of responding than those with lower or higher levels (p = 0.005, Fisher's exact test), while patients with plasma desipramine levels > and = 64 ng/ml were more likely to respond than those with lower levels (p = 0.032, Fisher's exact test). Results using an alternate response criterion were similar. These findings suggest that in desipramine-treated outpatients with primary MDD the relationship between therapeutic response and plasma levels is curvilinear for 2-OH-DMI and linear for desipramine.

Determination of 3,4-dihydroxyphenyl glycol in plasma by gas chromatography-mass spectrometry and high-performance liquid chromatography methods.

Several modifications of GC-MS and HPLC methods for plasma level DHPG have been described. The effects of storage temperature and stabilizing agents on DHPG stability have been studied. The stabilizing agent has been found to play a more important role than low-temperature storage in preventing DHPG from decomposition during sample storage. A specific and sensitive GC-MS method (electron impact) has been established using stable isotope-labeled DHPG as an internal standard. HPLC has been improved by modifying the conditions, resulting in a good separation of DHPG and internal standard from solvent front other early eluting compounds. Comparison of the GC-MS and HPLC procedures demonstrates a strong correlation between these two methods.

Sensitive and selective liquid-chromatographic assay of fluoxetine and norfluoxetine in plasma with fluorescence detection after precolumn derivatization.

We determined fluoxetine (Prozac) and its major metabolite norfluoxetine in plasma by liquid chromatography with fluorescence detection. After liquid-liquid extraction from 1 mL of plasma, the extract was derivatized at room temperature with dansyl chloride, and the highly fluorescent derivatives were chromatographed with a reversed-phase C18 column and a mobile phase of phosphate buffer and acetonitrile. Dansylated fluoxetine, norfluoxetine, and the internal standard were eluted in less than 14 min with no interference from endogenous material. The calibration curve was linear over the concentration range 25-800 micrograms/L with inter- and intra-assay imprecision (CV) of less than 10%. Validity of the assay was checked by comparing results for 110 patients' samples with those by a liquid-chromatographic method with ultraviolet detection (r = 0.993 for fluoxetine, 0.957 for norfluoxetine). The identity of the dansylated derivatives was verified by positive chemical ionization mass spectroscopy. The lower limit of detection was approximately 3 micrograms/L. Because no major antidepressant, neuroleptic, or respective drug metabolites interfere with the quantification of fluoxetine and norfluoxetine, this is a useful procedure for pharmacokinetic studies and in clinical settings.