Cerebrospinal Fluid Fractions Research Articles

Central nervous system (CNS) involvement in acute leukemia and lymphoma: indications for CNS assessment and advice for diagnostic strategy1)

AbstractInvolvement of the central nervous system (CNS) must be considered in leukemia and lymphoma to develop a promising treatment strategy. In particular, a relapse within the CNS compartment is associated with a dismal prognosis. Therefore, CNS involvement should be detected as early as possible during the disease course. The risk for CNS disease varies considerably between the diseases. For diseases with a proven high-risk for CNS involvement, such as acute lymphoblastic leukemia (ALL) or Burkitt’s lymphoma, prophylactic strategies are used. Although not in every entity, evidence-based therapeutic standards do exist, the detection of tumor cells in the cerebrospinal fluid (CSF) will have an impact on the therapeutic concept. This holds true for the preemptive situation with low tumor burden as well as for the therapeutic situation with a symptomatic patient. Whereas parenchymatous lesions are detected by magnetic resonance imaging, CSF analysis is the method of choice for the diagnosis of neoplastic meningitis. Cytomorphology of CSF cells, if used by an experienced cytopathologist, is of sufficient specificity but limited sensitivity for the diagnosis of leukemia. In lymphoma, cytomorphology has only modest specificity, because rather seldom conclusions can be drawn about the lineage commitment of the lymphoma cells and there is no possibility to prove clonal disease. Furthermore, it is often very difficult to discriminate between neoplastic meningitis and reactive pleocytosis. By combining cytomorphology with flow cytometry, sensitivity as well as specificity can be improved. Careful acquisition of the CSF sample by lumbar puncture is important. Only the last of several CSF fractions should be assessed by cytomorphology and flow cytometry to avoid contamination with peripheral blood. It is further mandatory that essential clinical information is provided by the clinician to the laboratory, so that a well-suited antibody panel can be chosen. This will save material and laboratory time and will further accelerate analysis. Owing to the limited number of cells within the CSF, analysis should always start with a basic antibody panel, which can then be extended on demand.

Microparticles and microscopic structures in three fractions of fresh cerebrospinal fluid in schizophrenia: Case report of twins

Schizophrenia is a diagnosis with a set of symptoms of aberrant psychological phenomena. Here, we discuss that there may be an imbalance of proteostasis of neurons in the brain leading to increase in membrane shedding and buildup of microparticles (MPs) appearing in the cerebrospinal fluid. The number of MPs can be determined and their phenotypes verified by size and membrane expression with flow cytometry. This is the first report of specified MPs in cerebrospinal fluid (CSF) in schizophrenia. Two 56-year-old Swedish-born female monozygotic twins of Caucasian ethnicity with onset of schizophrenia more than 30years ago were studied. Three fractions of fresh CSF were examined for microparticles by flow cytometry analysis, which measure the specific binding of antibodies to CD42a (platelet-MP; 33 GPIX), CD144 (endothelial-MP; Ve-cadherin), CD45 (leukocyte-MP; pan-leukocyte antigen) and of phosphatidylserine to lactadherin. The patients with schizophrenia displayed more phosphatidylserine-positive MPs in CSF compared with healthy control subjects. The scanning electron microscopic (SEM) structures in CSF studied over a 3-year period in twins with schizophrenia were of similar appearance at both time points. The increased number of MPs in fresh CSF may be a sign of enhanced membrane shedding in the central nervous system. Such MPs can be investigated for both human and non-human DNA, RNA and microRNA that may activate different immune signaling systems in patients with schizophrenia.

Microscopic Particles in Two Fractions of Fresh Cerebrospinal Fluid in Twins with Schizophrenia or Bipolar Disorder and in Healthy Controls

BackgroundUsing scanning electron microscopy, microscopic structures have been identified in fresh cerebrospinal fluid (CSF) in patients with schizophrenia and bipolar disorder, but only rarely in control subjects. However, it has not been determined whether these microscopic particles represent state or trait markers, i.e. if their presence is related to clinical manifestations of the disease or if they also can be found in as yet asymptomatic individuals with a genetic liability. This question can be addressed by studying twins discordant or concordant for schizophrenia or bipolar disorder.Methodology/Principal FindingsWe investigated microscopic structures in CSF in 102 individuals: 21 monozygotic and 16 dizygotic twins affected or not affected with schizophrenia, schizoaffective disorder or bipolar disorder and in 65 healthy singleton controls. A first and a second fraction of CSF was freshly applied on filters and examined by scanning electron microscopy technique. Spherical particles with lipid appearance averaging between 0.1 to 8.0 µm in diameter were detected in the center of the filter as well as located in the margins of larger aggregates binding in a viscous state. Structures were found in 12 of 17 probands, 5 of 12 healthy co-twins and 3 of 73 healthy controls. Thus, a positive microscopic finding significantly increased the likelihood of belonging to the proband group (OR = 48, 95% CL: 8.2–550, p<0.0001) and the co-twin-group (OR = 16, 95% CL: 2.0–218, p = 0.006). Age, sex, history of alcohol abuse or anxiety syndrome, somatic disorder and markers of acute inflammatory activity did not account for group differences; nor did exposure to psychotropic medication.ConclusionPresence of microscopic particles in CSF may possibly reflect trait dependent genetic or environmental vulnerability in patients with schizophrenia, schizoaffective disorder or bipolar disorder.

Leukocyte-derived microparticles and scanning electron microscopic structures in two fractions of fresh cerebrospinal fluid in amyotrophic lateral sclerosis: a case report

IntroductionAmyotrophic lateral sclerosis is a progressive neurodegenerative disorder characterized by degeneration of motoneuron cells in anterior spinal horns. There is a need for early and accurate diagnosis with this condition. In this case report we used two complementary methods: scanning electron microscopy and fluorescence-activated cell sorting. This is the first report to our knowledge of microparticles in the cerebrospinal fluid of a patient with amyotrophic lateral sclerosis.Case presentationAn 80-year-old Swedish man of Caucasian ethnicity presented to our facility with symptoms of amyotrophic lateral sclerosis starting a year before his first hospital examination, such as muscle weakness and twitching in his right hand progressing to arms, body and leg muscles. Electromyography showed classical neurophysiological findings of amyotrophic lateral sclerosis. Routine blood sample results were normal. A lumbar puncture was performed as a routine investigation and his cerebrospinal fluid was normal with regard to cell count and protein levels, and there were no signs of inflammation. However, scanning electron microscopy and fluorescence-activated cell sorting showed pronounced abnormalities compared to healthy controls. Flow cytometry analysis of two fractions of cerebrospinal fluid from our patient with amyotrophic lateral sclerosis was used to measure the specific binding of antibodies to CD42a, CD144 and CD45, and of phosphatidylserine to lactadherin. Our patient displayed over 100 times more phosphatidylserine-positive microparticles and over 400 times more cell-derived microparticles of leukocyte origin in his cerebrospinal fluid compared to healthy control subjects. The first cerebrospinal fluid fraction contained about 50% more microparticles than the second fraction. The scanning electron microscopy filters used with cerebrospinal fluid from our patient were filled with compact aggregates of spherical particles of lipid appearance, sticking together in a viscous batter. The quantitative increase in scanning electron microscopy findings corresponded to the flow cytometry result of an increase in leukocyte-derived microparticles.ConclusionsMicroparticles represent subcellular arrangements that can influence the pathogenesis of amyotrophic lateral sclerosis and may serve as biomarkers for underlying cellular disturbances. The increased number of leukocyte-derived microparticles with normal cell counts in cerebrospinal fluid may contribute to the amyotrophic lateral sclerosis inflammatory process by formation of immune complexes of prion-like propagation, possibly due to misfolded proteins. The two complementary methods used in this report may be additional tools for revealing the etiology of amyotrophic lateral sclerosis, for early diagnostic purposes and for evaluation of clinical trials, long-term follow-up studies and elucidating the pathophysiology in amyotrophic lateral sclerosis.

Diffuse axonal injury in mild traumatic brain injury: a 3D multivoxel proton MR spectroscopy study.

Since mild traumatic brain injury (mTBI) often leads to neurological symptoms even without clinical MRI findings, our goal was to test whether diffuse axonal injury is quantifiable with multivoxel proton MR spectroscopic imaging ((1)H-MRSI). T1- and T2-weighted MRI images and three-dimensional (1)H-MRSI (480 voxels over 360 cm(3), about 30 % of the brain) were acquired at 3 T from 26 mTBI patients (mean Glasgow Coma Scale score 14.7, 18-56 years old, 3-55 days after injury) and 13 healthy matched contemporaries as controls. The N-acetylaspartate (NAA), choline (Cho), creatine (Cr) and myo-inositol (mI) concentrations and gray-matter/white-matter (GM/WM) and cerebrospinal fluid fractions were obtained in each voxel. Global GM and WM absolute metabolic concentrations were estimated using linear regression, and patients were compared with controls using two-way analysis of variance. In patients, mean NAA, Cr, Cho and mI concentrations in GM (8.4 ± 0.7, 6.9 ± 0.6, 1.3 ± 0.2, 5.5 ± 0.6 mM) and Cr, Cho and mI in WM (4.8 ± 0.5, 1.4 ± 0.2, 4.6 ± 0.7 mM) were not different from the values in controls. The NAA concentrations in WM, however, were significantly lower in patients than in controls (7.2 ± 0.8 vs. 7.7 ± 0.6 mM, p = 0.0125). The Cho and Cr levels in WM of patients were positively correlated with time since mTBI. This (1)H-MRSI approach allowed us to ascertain that early mTBI sequelae are (1) diffuse (not merely local), (2) neuronal (not glial), and (3) in the global WM (not GM). These findings support the hypothesis that, similar to more severe head trauma, mTBI also results in diffuse axonal injury, but that dysfunction rather than cell death dominates shortly after injury.

α-Synuclein in human cerebrospinal fluid is principally derived from neurons of the central nervous system

The source of Parkinson disease-linked α-synuclein (aSyn) in human cerebrospinal fluid (CSF) remains unknown. We decided to measure the concentration of aSyn and its gradient in human CSF specimens and compared it with serum to explore its origin. We correlated aSyn concentrations in CSF versus serum (QaSyn) to the albumin quotient (Qalbumin) to evaluate its relation to blood–CSF barrier function. We also compared aSyn with several other CSF constituents of either central or peripheral sources (or both) including albumin, neuron-specific enolase, β-trace protein and total protein content. Finally, we examined whether aSyn is present within the structures of the choroid plexus (CP). We observed that QaSyn did not rise or fall with Qalbumin values, a relative measure of blood–CSF barrier integrity. In our CSF gradient analyses, aSyn levels decreased slightly from rostral to caudal fractions, in parallel to the recorded changes for neuron-specific enolase; the opposite trend was recorded for total protein, albumin and β-trace protein. The latter showed higher concentrations in caudal CSF fractions due to the diffusion-mediated transfer of proteins from blood and leptomeninges into CSF in the lower regions of the spine. In postmortem sections of human brain, we detected highly variable aSyn reactivity within the epithelial cell layer of CP in patients diagnosed with a range of neurological diseases; however, in sections of mice that express only human SNCA alleles (and in those without any Snca gene expression), we detected no aSyn signal in the epithelial cells of the CP. We conclude from these complementary results that despite its higher levels in peripheral blood products, neurons of the brain and spinal cord represent the principal source of aSyn in human CSF.

P.1.005 Season of sampling and season of birth determine serotonin metabolite levels in human cerebrospinal fluid

The rate of production of cerebrospinal fluid (CSF) and the caudorostral gradients of total CSF protein were measured in seven subjects with Down syndrome (DS) and compared to age-matched healthy normal volunteers. The CSF production rate in DS subjects (0.35 ± 0.02 mL/min.) did not differ significantly from normal subjects (0.37 ± 0.09 mL/min.). In addition, the caudorostral gradient of total protein was similar in DS and normal subjects, with more caudal fractions of lumbar CSF having higher total protein levels than more rostral fractions. These data suggest that there is no gross distrubance in CSF dynamics in DS.

Glutathione in the blood and cerebrospinal fluid: A study in healthy male volunteers

Glutathione (GSH) is an important regulator of intracellular redox homeostasis. In the brain, glutathione is considered a major antioxidant, which is also found at high concentrations in the extracellular environment. Altered GSH balance in plasma, blood and cerebrospinal fluid (CSF) has been observed in several disorders suggesting that an impaired antioxidant function is part of the pathophysiology. The aim of the present study was to investigate a possible relationship between glutathione in plasma and CSF. Blood samples were collected from 26 healthy male volunteers at 8a.m., noon, 4p.m. and 8 p.m. At 8a.m. the following morning, blood was drawn and three 6-ml fractions of CSF were collected by lumbar puncture. In CSF, a disrupted gradient was found showing the highest glutathione concentration in the second compared to the first and third fraction (P<0.002). Moreover, correlation and regression analyses between glutathione in plasma and CSF revealed an association between the third fraction CSF and plasma glutathione 8 p.m. the day before lumbar puncture. Thus, if carefully standardised due to the disrupted gradient in CSF, it might be possible to estimate glutathione levels in CSF by analysing plasma in healthy males.

A longitudinal study of alterations in cognitive function and brain metabolites among women receiving chemotherapy for primary breast cancer.

TPS240 Background: Cancer patients treated with chemotherapy frequently complain of cognitive deficits during treatment and often for some years thereafter. Many cytotoxic drugs commonly used in chemotherapy result in cognitive deficits via direct neurotoxicity to the cerebral parenchyma, neuronal axons, microglia, and/or oligodendrocytes. This chemotherapy-induced neurotoxicity may lead to changes in the levels of various brain metabolites. Previous studies using Magnetic Resonance Spectroscopy (MRS) suggest that cognitive impairment is accompanied by changes in specific brain metabolites such as N-acetyl aspartate (NAA), creatine, choline, and myo-inositol (Jung RE et al; 1999, Ross AJ et al; 2004). Because metabolic changes are likely to precede clinically detectable cognitive dysfunction, MRS may allow early identification of patients who will develop cognitive dysfunction associated with cancer treatments. Methods: The aims of this study are 1. to evaluate cognitive function using neuropsychological testing, 2. to collect MRS data from left thalamus, hippocampus, frontal and parietal lobes, 3. to characterize the temporal relationship between brain metabolite concentrations and cognitive functioning. Fifty newly diagnosed breast cancer patients, who are to receive adjuvant chemotherapy are evaluated with a battery of neuropsychological tests and MRS at three time points: before receiving chemotherapy (baseline), 4-8 weeks after completing their final treatment, and one year later. Patients also complete self-report questionnaires at all three time points. Exclusion criteria include CNS metastasis and depression. Neuropsychological tests are scored by standard procedures with adjustments for age, gender and education, and converted to Z-scores. Concentrations of NAA, creatine, choline, and myo-inositol are calculated from the MRS data for all time points by standard methods, with correction for the cerebrospinal fluid fraction in the voxel. Results of this study could lead to greater understanding of chemotherapy-induced cognitive impairment and improved prevention/treatment strategies in this population of cancer survivors.

Grey matter deficit in long‐term recovered anorexia nervosa patients

Cerebral grey matter (GM) reduction has repeatedly been shown in anorexia nervosa (AN). Evidence concerning completeness of GM restitution in recovered patients is contradictory. Five long-term recovered patients with AN were compared to symptomatic subjects and healthy controls using voxel-based morphometry. Whole brain GM, white matter and cerebrospinal fluid fractions were studied. Additionally, voxels that had shown significant GM reductions in symptomatic patients were investigated. Recovered subjects had been severely affected when symptomatic (mean body mass index: 12.1 kg/m²) and were in remission for a very long time period (>5 years). Whole brain tissue fractions did not differ from controls. Regional analysis showed persistent GM volume reduction, in particular of the precuneus. This study further supports the assumption that GM volume restitution is incomplete in subjects, who had previously been severely affected by anorexia nervosa. The meaning of GM reduction in long-term recovered AN patients, that is, its pathophysiological relevance, however, remains unclear. Furthermore, the precise aetiology of GM reduction remains an open question.

Individual Differences in Subconscious Motor Control Predicted by GABA Concentration in SMA

Subliminal visual stimuli affect motor planning, but the size of such effects differs greatly between individuals. Here, we investigated whether such variation may be related to neurochemical differences between people. Cortical responsiveness is expected to be lower under the influence of more of the main inhibitory neurotransmitter, GABA. Thus, we hypothesized that, if an individual has more GABA in the supplementary motor area (SMA)--a region previously associated with automatic motor control--this would result in smaller subliminal effects. We measured the reversed masked prime--or negative compatibility--effect, and found that it correlated strongly with GABA concentration, measured with magnetic resonance spectroscopy. This occurred specifically in the SMA region, and not in other regions from which spectroscopy measurements were taken. We replicated these results in an independent cohort: more GABA in the SMA region is reliably associated with smaller effect size. These findings suggest that, across individuals, the responsiveness of subconscious motor mechanisms is related to GABA concentration in the SMA.

Comparison of the performance of two affinity depletion spin filters for quantitative proteomics of CSF: Evaluation of sensitivity and reproducibility of CSF analysis using GeLC‐MS/MS and spectral counting

For biomarker discovery in cerebrospinal fluid (CSF), removal of major serum proteins is advantageous as more CSF proteins including brain-derived proteins can be identified. Our goal was to create a reproducible discovery workflow with acceptable throughput that can identify 500-1000 CSF proteins in small volumes of CSF. In this study, we compared the performance of two multi-affinity depletion methods in spin filter format: MARS Human 14 and Seppro-IgY-14. To this end, we analyzed depleted and bound CSF fractions isolated from 0.5 mL aliquots of the same CSF sample (n=3 per depletion method) by label-free GeLC-MS/MS-based proteomics and normalized spectral counting. The whole CSF dataset contained 884 proteins identified at high confidence. Depletion spin filter performance was assessed in terms of sensitivity and reproducibility of the CSF analysis. MARS and IgY-14 spin filters yielded comparable reproducibility of protein identification (71-74%) and quantification (CV 17-18%) but a significant difference in the total number of identified CSF proteins (767 and 703 proteins, respectively). The MARS filter compared to IgY-14 filter provides a CSF analysis with enhanced proteome coverage. We anticipate that this enhanced sensitivity will facilitate biomarker discovery in early stages of cancer or neurological disease.

Micrometer‐sized thread‐like and/or spherical particles in the first fraction of cerebrospinal fluid in patients with bipolar disorder

Scanning electron microscopy (SEM) is a powerful tool to identify pathogenic factors for which sensitive tests are lacking. The technique has been used to recognize structures in the cerebrospinal fluid (CSF) of patients with schizophrenia. The aim of this study was to use SEM to screen for potential particles in CSF in bipolar disorder. Fresh CSF samples from 56 euthymic bipolar patients, 31 bipolar I disorder and 25 bipolar II disorder, were compared to CSF samples from 20 controls. SEM of two portions of 200 microL filtered CSF was performed; the first 0.6 mL of CSF and the following 12 mL. The microscopic structures were identified and the quantity and patterns were rated by two independent researchers. Quantitative SEM examinations showed that of the 56 patients, 11 were free of any SEM structures in CSF, while 45 patients displayed morphological structures in the first 0.6 mL of CSF. By contrast, only 2 patients showed structures in the second CSF fraction drawn from the following mixed 12 mL of CSF. In total, 45 of the 56 patients had either thread-like, spherical, or both structures in the CSF, compared to none of the 20 controls. The identified particles in the first fraction of CSF have previously not been described in patients with bipolar disorder. Hypothetically, the amount of SEM structures in CSF, from none to many, might correlate to the degree of the alleged underlying disease processes in the central nervous system in patients with bipolar disease.

A positive correlation between fractional white matter volume and the response of Parkinson disease patients to subthalamic stimulation

Since optimal patient selection is essential for the success of subthalamic nucleus (STN) stimulation, the identification of reliable outcome predictors is important. The purpose of this study was to identify new imaging characteristics sufficiently reliable to predict treatment results. Using preoperative magnetic resonance imaging studies of 21 Parkinson disease (PD) patients treated by STN stimulation, we performed whole brain-based analysis of voxel-based morphometry (VBM) data. Intracranial structures segmented into the gray matter fraction (GMF), white matter fraction (WMF), and cerebrospinal fluid fraction (CSFF) were subjected to univariate and multivariate analysis of the correlation between fractional volumes and postoperative improvement rates using the Unified PD Rating Scale (UPDRS). At 3 months after surgery, the WMF was significantly correlated with improvement rated on the total UPDRS (p = 0.006), UPDRS part II (activities of daily living; p = 0.008), UPDRS part III (motor; p = 0.005). In contrast, there was no significant correlation between the effect of STN stimulation and GMF or the effect of stimulation and CSFF. The WMF also showed a significant correlation with postoperative scores in the "on" drug and "on" stimulation state (total UPDRS, p = 0.027; UPDRS part II, p = 0.019; UPDRS part III, p = 0.034). Our data indicate that patients with a larger white matter volume benefited from STN stimulation whereas the volume of other brain structures was not correlated with its effect. We posit that preserved connectivity between components of the basal ganglia-thalamocortical circuit may be required for the effectiveness of electrical stimulation. VBM may represent a powerful tool to predict the response of patients with advanced PD to STN stimulation.

Quantification of ethanol methyl 1H magnetic resonance signal intensity following intravenous ethanol administration in primate brain

In vivo 1H magnetic resonance spectroscopy (MRS) can be used to directly monitor brain ethanol. Previously, studies of human subjects have lead to the suggestion that the ethanol methyl 1H MRS signal intensity relates to tolerance to ethanol’s intoxicating effects. More recently, the ethanol 1H MRS signal intensity has been recognized to vary between brain gray matter (GM), white matter (WM), and cerebrospinal fluid (CSF) due to differences in T 2 within these environments. The methods presented here extend ethanol MRS techniques to non-human primate subjects. Twelve monkeys were administered ethanol while sedated and positioned within a 3T MRI system. Chemical shift imaging (CSI) measurements were performed following intravenous infusion of 1 g/kg ethanol. Magnetic resonance imaging (MRI) data were also recorded for each monkey to provide volume fractions of GM, WM, and CSF for each CSI spectrum. To estimate co-variance of ethanol MRS intensity with GM, WM, and CSF volume fractions, the relative contribution of each tissue subtype was determined following corrections for radiofrequency pulse profile non-uniformity, chemical shift artifacts, and differences between the point spread function in the CSI data and the imaging data. The ethanol MRS intensity per unit blood ethanol concentration was found to differ between GM, WM, and CSF. Individual differences in MRS intensity were larger in GM than WM. This methodology demonstrates the feasibility of ethanol MRS experiments and analysis in non-human primate subjects, and suggests GM may be a site of significant variation in ethanol MRS intensity between individuals.

Cerebrospinal fluid of brain trauma patients inhibitsin vitroneuronal network function via NMDA receptors

Neurological diseases frequently induce pathological changes of cerebrospinal fluid (CSF) that might secondarily influence brain activity, as the CSF-brain barrier is partially permeable. However, functional effects of CSF on neuronal network activity have not been specified to date. Here, we report that CSF specimens from patients with reduced Glasgow Coma Scale values caused by severe traumatic brain injury suppress synchronous activity of in vitro-generated neuronal networks in comparison with controls. We present evidence that underlying mechanisms include increased N-methyl-D-aspartate receptor activity mediated by a CSF fraction containing elevated amino acid concentrations. These proof-of-principle data suggest that determining effects of CSF specimens on neuronal network activity might be of diagnostic value.

Taurine in plasma and CSF: a study in healthy male volunteers

In order to explore the interrelationship between plasma and cerebrospinal fluid taurine concentrations, three consecutive 6-ml fractions of cerebrospinal fluid were drawn from 30 healthy male volunteers in the early morning after 8 h in the fasting condition. Repeated plasma samples were drawn over 24 h the day before lumbar puncture. Taurine in plasma and cerebrospinal fluid was determined by high performance liquid chromatography. The subjects were categorized as extensive or poor metabolizers with respect to the cytochrome P450 2D6 genotype. The taurine cerebrospinal fluid/plasma ratio at 8 a.m. was negatively influenced by the plasma taurine concentration at 4 p.m. the previous day. It was also negatively influenced by body mass index and positively by the intraspinal pressure. Three poor metabolizers of cytochrome P450 2D6 had higher plasma taurine areas under the curve than 27 extensive metabolizers. Hypothetically, cytochrome P450 2D6 influences the transport of taurine across the blood-brain barrier.

Cerebrospinal Fluid Amino Acids in Pathological Gamblers and Healthy Controls

Amino acids, such as valine, isoleucine and leucine compete with tyrosine and tryptophan for transport into the brain and might thus affect the central serotonin and catecholamine patterns. Furthermore, the excitatory amino acids glutamic acid, aspartic acid and glycine are known to act on the N-methyl-D-aspartate receptor, which is part of the reward system. Based on these facts, we have explored the role of cerebrospinal fluid (CSF) amino acids in pathological gambling. Concentrations of amino acids were determined in CSF obtained from one female and 11 pathological male gamblers and 11 healthy male controls. In an ANCOVA with best subset regression, pathological male gamblers had higher CSF levels of the excitatory glutamic and aspartic acids, as well as of phenylalanine, isoleucine, citrulline and glycine. A negative contribution of glycine in interaction with the neuraxis distance might mirror a reduced spinal supply or an altered elimination of glycine in pathological gamblers. A decreasing CSF gradient from the first (0–6 ml) to the third (13–18 ml) CSF fraction was found for glutamic acid, glycine, leucine, isoleucine, lysine, ornithine and glutamine in both pathological gamblers and healthy controls. A decreasing gradient was found, however, for aspartic acid and phenylalanine in pathological male gamblers. The altered pattern of CSF amino acids in pathological gamblers might exert an influence on central monoamines as well as on N-methyl-D-aspartate receptor function.

Separation of proteins including metallothionein in cerebrospinal fluid by size exclusion HPLC and determination of trace elements by HR-ICP-MS

A method to study the protein binding patterns of trace elements in human cerebrospinal fluid (CSF) is described. Proteins in CSF samples were separated by size exclusion chromatography combined with high performance liquid chromatography (SEC-HPLC). The column was calibrated to separate proteins in the molecular weight range 6–70 kDa. Fractions were then analyzed off-line for trace elements using high resolution inductively coupled plasma mass spectrometry (HR-ICP-MS). We were able to accurately determine more than 10 elements of clinical interest in the CSF fractions. Results are presented for Cd, Mn, Fe, Pb, Cu and Zn. The total concentrations of 16 trace elements in human plasma and CSF are also presented. The method was able to differentiate the relative contribution of metallothionein and other proteins towards metal binding in human CSF.

CSF cholecystokinin, γ-aminobutyric acid and neuropeptide Y in pathological gamblers and healthy controls

The sulphated cholecystokinin (CCK) octapeptide (CCK-8S), the CCK tetrapeptide (CCK-4), neuropeptide Y (NPY) and gamma-aminobutyric acid (GABA) were determined in cerebrospinal fluid (CSF) obtained from 11 pathological male gamblers and 11 healthy male controls. Compared with healthy controls, pathological male gamblers displayed higher concentrations of CCK-8S, CCK-4 and GABA (but not NPY). A gradient with decreasing concentrations from the first to the third 6-ml CSF fraction was found for CCK-8S, CCK-4 and NPY, but only in pathological gamblers. Disrupted gradients were found for GABA and for NPY in healthy controls. Given that CCK is a modulator of dopamine in the reward process, the increase in CCK-8S and CCK-4 is not unexpected. The high level of GABA in pathological gamblers is in conformity with a compensatory inhibitory action on noradrenergic neurons. The CSF gradient of CCK-8S and CCK-4 in pathological male gamblers (but not healthy controls) might indicate a difference in diurnal variation. The results obtained are in line with an altered CCK and GABA function in pathological gambling.