Raltegravir Cerebrospinal Fluid Concentrations in HIV-1 Infection

Introduction

Raltegravir is an HIV-1 integrase inhibitor currently used in treatment-experienced HIV-1-infected patients resistant to other drug classes. In order to assess its central nervous system penetration, we measured raltegravir concentrations in cerebrospinal fluid (CSF) and plasma in subjects receiving antiretroviral treatment regimens containing this drug.

Methods

Raltegravir concentrations were determined by liquid chromatography tandem mass spectrometry in 25 paired CSF and plasma samples from 16 HIV-1-infected individuals. The lower limit of quantitation was 2.0 ng/ml for CSF and 10 ng/ml for plasma.

Results

Twenty-four of the 25 CSF samples had detectable raltegravir concentrations with a median raltegravir concentration of 18.4 ng/ml (range, <2.0–126.0). The median plasma raltegravir concentration was 448 ng/ml (range, 37–5180). CSF raltegravir concentrations correlated with CSF:plasma albumin ratios and CSF albumin concentrations.

Conclusions

Approximately 50% of the CSF specimens exceeded the IC₉₅ levels reported to inhibit HIV-1 strains without resistance to integrase inhibitors. In addition to contributing to control of systemic HIV-1 infection, raltegravir achieves local inhibitory concentrations in CSF in most, but not all, patients. Blood-brain and blood-CSF barriers likely restrict drug entry, while enhanced permeability of these barriers enhances drug entry.     

Relationship Between Serotoninergic Measures in Blood and Cerebrospinal Fluid Simultaneously Obtained in Humans

The relationships between the concentration of serotonin (5-HT) and related metabolites in human blood and CSF have been studied. Plasma tryptophan (TP), 5-HT, 5-hydroxyindoleacetic acid (5-HIAA), and indoleacetic acid (IAA), whole-blood 5-HT, and CSF TP, 5-HT, 5-HIAA, IAA, homovanillic acid, and 3-methoxy-4-hydroxyphenylethylene glycol were determined in 35 unmedicated outpatients who underwent minor surgical operations and had no history of psychiatric or neurological illnesses. Significant correlations were found between the serotoninergic parameters analyzed in blood and CSF. Plasma free 5-HT correlated significantly with CSF 5-HT (r = 0.411, p less than 0.02), and plasma 5-HIAA correlated with the CSF 5-HIAA/5-HT ratio (r = 0.508, p less than 0.004). The concentration of 5-HIAA in CSF correlated with the plasma 5-HIAA/5-HT ratio (r = 0.405, p less than 0.026) (which can be taken as an index of monoamine oxidase type A activity in peripheral tissues) and with the platelet 5-HT/plasma 5-HT ratio (r = 0.375, p less than 0.05). The concentrations of IAA in CSF and plasma were strongly correlated (r = 0.899, p less than 0.001). The significance of these results and their relationship to the use of "in vivo" measures of 5-HT and related metabolites in plasma and platelets as an index of serotoninergic function in affective disorders are discussed.     

Plasma and Cerebrospinal Fluid Amino Acids in Epileptic Patients

Altered plasma and cerebrospinal fluid amino acid levels may be associated with human epilepsy. We studied three groups of patients, those with a generalized epileptic syndrome, juvenile myoclonic epilepsy, patients with refractory localization-related epilepsies, and patients with acute seizures (within 24 h). Plasma levels of amino acids were studied in all patient groups, as were those in the cerebrospinal fluid (CSF) of patients with acute seizures. After acute seizures, the amino acid changes in the CSF were limited to a reduction in the level of taurine, whereas the levels of most amino acids in plasma were decreased. On the other hand, levels of the excitatory amino acids glutamate and aspartate were increased. The most notable finding in the juvenile myoclonic epilepsy patients was an increase in glutamate level in the plasma. Our study supports the conception of an altered metabolism of glutamate in generalized epilepsies.     

Relationship Between Plasma and Cerebrospinal Fluid Norepinephrine and Dopamine Metabolites in a Nonhuman Primate

Major and minor pathways of metabolism in the mammalian CNS result in the formation of 3-methoxy-4-hydroxyphenylethylene glycol (MHPG) and normetanephrine (NMN) from norepinephrine (NE), and homovanillic acid (HVA) and 3-methoxytyramine (3-MT) from dopamine (DA), respectively. The correlational relationships between HVA and 3-MT and between MHPG and NMN in primate CSF and plasma have not been described. These relationships may help to elucidate the usefulness of CSF and plasma metabolites as indices of CNS NE and DA activity. In addition, because NMN is unlikely to cross the blood-brain barrier. CSF NMN concentrations would not be confounded by contributions from plasma, which is a major issue with CSF MHPG. We have obtained repeated samples of plasma and CSF from drug-naive male squirrel monkeys and have measured the concentrations of MHPG, HVA, NMN, and 3-MT to define their correlational relationships. For the NE metabolites, significant correlations were obtained for CSF MHPG and NMN (r = 0.806, p less than 0.001), plasma MHPG and CSF NMN (r = 0.753, p less than 0.001), and plasma and CSF MHPG (r = 0.776, p less than 0.001). These results suggest that CSF and plasma MHPG and CSF NMN may reflect gross changes in whole brain steady-state noradrenergic metabolism. Only a single significant relationship was demonstrated for the DA metabolites, with CSF 3-MT correlating with plasma HVA (r = 0.301, p less than 0.025). The results for the DA metabolites probably reflect regional differences in steady-state brain dopaminergic metabolism.     

Dexamethasone,Cerebrospinal Fluid Matrix Metalloproteinase Concentrations and Clinical Outcomes in Tuberculous Meningitis

Background

Adjunctive dexamethasone reduces mortality from tuberculous meningitis, but how it produces this effect is not known. Matrix metalloproteinases (MMPs) are important in the immunopathology of many inflammatory CNS diseases thus we hypothesized that that their secretion is important in TBM and might be influenced by dexamethasone.

Methodology/Principal Findings

The kinetics of cerebrospinal fluid (CSF) MMP and tissue inhibitors of MMPs (TIMPs) concentrations were studied in a subset of HIV uninfected adults (n = 37) with TBM recruited to a randomized, placebo-controlled trial of adjuvant dexamethasone. Analysis followed a pre-defined plan. Dexamethasone significantly reduced CSF MMP-9 concentrations in early follow up samples (median 5 days (range 3–8) of treatment), but had no significant influence on other MMPs/TIMPs. Additionally CSF MMP-9 concentration was strongly correlated to concomitant CSF neutrophil count.

Conclusions/Significance

Dexamethasone decreased CSF MMP-9 concentrations early in treatment and this may represent one mechanism by which corticosteroids improve outcome in TBM. The strong correlation between CSF MMP-9 and neutrophil count suggests that polymorphonuclear leukocytes may play a central role in the early pathogenesis of TBM.     

Homeostasis of Brain and Cerebrospinal Fluid Calcium Concentrations During Chronic Hypo-and Hypercalcemia

Three-week-old rats were made hypocalcemic or hypercalcemic by being fed diets low or high in Ca. Both total and ionized [Ca]s in the plasma decreased about 40% and remained depressed for 4 weeks in rats fed a low-Ca diet. Plasma [Ca]s in rats fed a high-Ca diet increased by 30% and remained elevated for 7 weeks. After 8 weeks on the diets, cerebrospinal fluid (CSF) [Ca] changed by less than 30% whereas brain [Ca] changed by less than 20% of the chronic changes in plasma ionized [Ca]. Assuming a brain extracellular volume of 20% and noting that brain extracellular volume equilibrates freely with CSF, the findings demonstrate only small perturbations in the Ca content of the brain cellular compartment during sustained hypo or hypercalcemia. Partial regulation of CSF and brain extracellular Ca suggests a role for the blood-brain barrier in regulating CNS [Ca] during chronic changes in plasma [Ca].     

Evidence Disputing the Link Between Seizure Activity and High Extracellular Glutamate

Abstract: As seizures in experimental models can be induced by the activation and suppressed by the inhibition of glutamate receptors, it is often proposed that a high extracellular glutamate level subsequent to excessive presynaptic release and/or altered glutamate uptake is epileptogenic. The purpose of this study was to ascertain the link between seizure activity and high extracellular glutamate. To assist the detection of any putative rise in extracellular glutamate during seizures, microdialysis was coupled to enzyme-amperometric detection of glutamate, which provides maximal sensitivity and time resolution. Electrical activity and field potential were also recorded through the dialysis membrane to confirm that epileptic activity was present at the sampling site. No increase in dialysate glutamate content was detected during picrotoxin-induced seizures, even when the K⁺ concentration in the perfusion medium was raised to 50% above that measured previously during paroxysmal activity. In addition, sustained inhibition of glutamate uptake by l - trans -pyrrolidine-2,4-dicarboxylate increased the extracellular glutamate level >20-fold but did not produce electrophysiological changes indicative of excessive excitation. These findings indicate that seizures are not necessarily accompanied by an increased extracellular glutamate level and that increased glutamatergic excitation in epilepsy may result from other abnormalities such as increased density of glutamate receptors, enhanced activation subsequent to reduced modulation, or sprouting of glutamatergic synapses.     

Metal Concentrations in Cerebrospinal Fluid and Blood Plasma from Patients with Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is a progressive and fatal degenerative disorder of motor neurons. The cause of this degeneration is unknown, and different causal hypotheses include genetic, viral, traumatic and environmental mechanisms. In this study, we have analyzed metal concentrations in cerebrospinal fluid (CSF) and blood plasma in a well-defined cohort (n?=?17) of ALS patients diagnosed with quantitative electromyography. Metal analyses were performed with high-resolution inductively coupled plasma mass spectrometry. Statistically significant higher concentrations of manganese, aluminium, cadmium, cobalt, copper, zinc, lead, vanadium and uranium were found in ALS CSF compared to control CSF. We also report higher concentrations of these metals in ALS CSF than in ALS blood plasma, which indicate mechanisms of accumulation, e.g. inward directed transport. A pattern of multiple toxic metals is seen in ALS CSF. The results support the hypothesis that metals with neurotoxic effects are involved in the pathogenesis of ALS.     

The Significance of Homovanillic Acid and 3,4-Dihydroxyphenylacetic Acid Concentrations in Human Lumbar Cerebrospinal Fluid

The concentrations of the acidic dopamine (DA) catabolites homovanillic acid (HVA) and 3,4-dihydroxyphenylacetic acid (DOPAC) measured in human CSF are supposed to reflect the "turnover" of DA in the brain. The notion of "turnover" is, however, not synonymous with impulse nerve activity in the dopaminergic systems. Significant amounts of DOPAC and HVA could, indeed, be demonstrated in brain structures wherein dopaminergic innervation has not been documented. It must also be noted that DA is not only a neurotransmitter itself, but also a precursor of norepinephrine and epinephrine. Furthermore, in lumbar CSF, levels of biogenic amine catabolites partially reflect metabolism in the spinal cord and may have limited relevance to neurotransmission in the brain. To elucidate these points further, we determined the concentrations of DOPAC and HVA in 22 areas of six human brains and eight levels of six human spinal cords. The data were correlated with the concentration of DA. Quantitative determinations were done using HPLC with electrochemical detection, after solvent and ion-pair extraction. In this study, significant amounts of both DOPAC and HVA were demonstrated in brain structures not previously associated with dopaminergic innervation. The relatively lower DA concentration in these structures suggests that in these regions, the DOPAC and HVA concentrations are unrelated to dopaminergic neurotransmission. The possible role of capillary walls and glial cells in the catabolism of DA must be further evaluated. The demonstration of DOPAC and HVA in the spinal cord is another argument against the hypothesis that CSF levels of HVA and DOPAC reflect closely the activity of the dopaminergic systems in the brain.     

Ventricular Cerebrospinal Fluid Monoamine Transmitter and Metabolite Concentrations Reflect Human Brain Neurochemistry in Autopsy Cases

Concentrations of dopamine (DA), its metabolites 3-methoxytyramine and homovanillic acid (HVA), noradrenaline (NA), its metabolites normetanephrine (NM) and 3-methoxy-4-hydroxyphenylglycol (MHPG), 5-hydroxytryptamine (5-HT, serotonin), and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) were measured in 14 brain regions and in CSF from the third ventricle of 27 human autopsy cases. In addition, in six cases, lumbar CSF was obtained. Monoamine concentrations were determined by reversed-phase liquid chromatography with electrochemical detection. Ventricular/lumbar CSF ratios indicated persistence of rostrocaudal gradients for HVA and 5-HIAA post mortem. Ventricular CSF concentrations of DA and HVA correlated positively with striatal DA and HVA. CSF NA correlated positively with NA in hypothalamus, and CSF MHPG with levels of MHPG in hypothalamus, temporal cortex, and pons, whereas CSF NM concentration showed positive correlations with NM in striatum, pons, cingulate cortex, and olfactory tubercle. CSF 5-HT concentrations correlated positively with 5-HT in caudate nucleus, whereas the concentration of CSF 5-HIAA correlated to 5-HIAA levels in thalamus, hypothalamus, and the cortical areas. These data suggest a specific topographic origin for monoamine neurotransmitters and their metabolites in human ventricular CSF and support the contention that CSF measurements are useful indices of central monoaminergic activity in man.     

Relationship Between Leucocyte and Plasma Ascorbic Acid Concentrations

Leucocyte and plasma ascorbic acid values were measured in healthy students, adult factory employees, and old people not receiving supplementary vitamin C and in healthy old people receiving 500 mg of vitamin C daily. Significant positive correlations between leucocyte and plasma ascorbic acid were found in all the groups. The regression lines differed significantly between one another within the sexes, but the pooled lines for each sex did not differ significantly in the unsupplemented groups. The relationship between plasma and leucocyte ascorbic acid values in the supplemented group differed significantly from that in the pooled unsupplemented groups.There was a limited range of variation in leucocyte ascorbic acid values compared with the range in plasma values in the supplemented group, whereas there was a wider range of variation in the leucocyte values in the unsupplemented groups. Leucocytes can therefore achieve a saturation level of ascorbic acid. Measurement of leucocyte ascorbic acid concentrations alone does not provide a reliable guide for the estimation of tissue status of ascorbic acid in normal individuals. Leucocyte concentrations provide a measure of the availability of ascorbic acid for storage, and plasma levels give an indication of its metabolic turnover rate. When these values are related the regression lines provide information about the storage and metabolism of ascorbic acid in normal individuals.     

Low Levels of γ-Aminobutyric Acid in Cerebrospinal Fluid of Dogs with Epilepsy

gamma-Aminobutyric acid (GABA) levels were determined in cisternal cerebrospinal fluid (CSF) of 19 epileptic dogs with generalized tonic-clonic (grand mal) seizures using a radioreceptor assay. Thirty-four healthy age-matched dogs served as controls. The average CSF GABA level in epileptic dogs (40 pmol/ml) was significantly lower than that determined in controls (66 pmol/ml). Treatment with phenobarbital or primidone seemed not to affect CSF GABA levels.     

The Choroid Plexuses and the Barriers Between the Blood and the Cerebrospinal Fluid

1. The fluid homeostasis of the brain depends both on the endothelial blood–brain barrier and on the epithelial blood–cerebrospinal fluid (CSF) barrier located at the choroid plexuses and the outer arachnoid membrane.2. The brain has two fluid environments: the brain interstitial fluid, which surrounds the neurons and glia, and the CSF, which fills the ventricles and external surfaces of the central nervous system.3. CSF acts as a fluid cushion for the brain and as a drainage route for the waste products of cerebral metabolism.4. Recent findings suggest that CSF may also act as a third circulation conveying substances secreted into the CSF rapidly to many brain regions.     

Concentrations of Homovanillic Acid and 5-Hydroxyindoleacetic Acid in Cerebrospinal Fluid from Human Infants in the Perinatal Period

To assess maturation of central serotonin and catecholamine pathways at birth, we measured lumbar CSF homovanillic acid (HVA) and 5-hydroxyindoleacetic acid (5-HIAA), stable acid metabolites of dopamine and serotonin, using HPLC with electrochemical detection. CSFs from 57 neonates (38 premature and 19 at term) and 13 infants 1-6 months old were studied. HVA levels increased with maturity (p less than 0.05; ANOVA), whereas 5-HIAA levels were similar in all these subjects. HVA/5-HIAA ratios increased markedly from 1 +/- 0.12 in the most premature neonates to 1.98 +/- 0.17 in the older infants (p less than 0.01; t test). There were no sex differences for these values.     

Relationship between Cerebrospinal Fluid Biomarkers for Inflammation,Demyelination and Neurodegeneration in Acute Optic Neuritis

Background

Various inflammatory biomarkers show prognostic potential for multiple sclerosis (MS)-risk after clinically isolated syndromes. However, biomarkers are often examined singly and their interrelation and precise aspects of their associated pathological processes remain unclear. Clarification of these relationships could aid the appropriate implementation of prognostic biomarkers in clinical practice.

Objective

To investigate the interrelation between biomarkers of inflammation, demyelination and neurodegeneration in acute optic neuritis and to assess their association to measures of MS risk.

Material and Methods

A prospective study at a tertiary referral centre from June 2011 to December 2012 of 56 patients with optic neuritis as a first demyelinating symptom and 27 healthy volunteers. Lumbar puncture was performed within 28 (median 16) days of onset. CSF levels of CXCL13, matrix metalloproteinase (MMP)-9, CXCL10, CCL-2, osteopontin and chitinase-3-like-1, myelin basic protein (MBP) and neurofilament light-chain (NF-L) were determined. MS-risk outcome measures were dissemination in space (DIS) of white matter lesions on cerebral MRI, CSF oligoclonal bands and elevated IgG-index.

Results

In the interrelation analysis the biomarkers showed close correlations within two distinct groups: Biomarkers of leukocyte infiltration (CXCL13, MMP-9 and CXCL10) were strongly associated (p<0.0001 for all). Osteopontin and chitinase-3-like-1 were also tightly associated (p<0.0001) and correlated strongly to tissue damage markers (NF-L and MBP). The biomarkers of leukocyte infiltration all associated strongly with MS-risk parameters, whereas CHI3L1 and MBP correlated with MRI DIS, but not with CSF MS-risk parameters and osteopontin and NF-L did not correlate with any MS-risk parameters.

Conclusions

Our findings suggest two distinct inflammatory processes: one of leukocyte infiltration, represented by CXCL13, CXCL10 and MMP-9, strongly associated with and potentially predicting MS-risk; the other represented by osteopontin and CHI3L1, suggesting tissue damage-related inflammation, potentially predicting residual disabilities after attack and perhaps cumulative damage over time. These hypotheses should be further addressed in follow-up studies.     

γ-Glutamylglutamine and Taurine Concentrations Are Decreased in the Cerebrospinal Fluid of Drug-Naive Patients with Schizophrenic Disorders

Abstract: HPLC and gas chromatography-mass spectrometry analyses of 18 amino acids, N -acetylaspartate, N -acetylaspartyglutamate, and 5-hydroxyindoleacetic acid, derived from serotonin, and homovanillic acid, derived from dopamine, were performed in CSF collected from a group of patients with schizophrenia who either had been drug free for at least 1 year (n = 5) or were drug naive for psychotropic drugs (n = 21) and in 15 control subjects. Significant differences were found only for taurine (15% lower in the patients) and isoleucine (7% higher). A number of unidentified substances were detected, one of which proved to be markedly reduced (16%) among the schizophrenic patients. Liquid chromatography-mass spectrometry with continuous flow-fast atom bombardment interface allowed us to identify this substance as γ-glutamyglutamine. The decreased level of γ-glutamylglutamine may reflect a deficiency in the γ-glutamyltransferase system, a system probably involved in glutamate uptake, or a deficiency in glutamine, an important precursor of releasable glutamate. Although glutamate was nonsignificantly reduced in the patients, it was one of the five substances (including γ-glutamylglutamine) that were necessary for the best discrimination between the schizophrenic patients and the controls. These findings support the notion that the glutamatergic system is affected in schizophrenic disorders. In addition, they underscore the need to apply rigid bioanalytical techniques and use drug-naive patients to gain in-depth information on the pathophysiology of brain disorders such as schizophrenia.     

Cerebrospinal Fluid Glucose: Turnover and Metabolism

The turnover of cerebrospinal fluid (CSF) glucose was studied in cats during steady-state perfusion. In all experiments, the perfusion fluid contained either tracer [¹⁴C]glucose alone or tracer glucose along with 4.45 mM unlabeled glucose. In some studies, serum glucose was lowered with insulin. The concentration of glucose and [¹⁴C]glucose in the effluent fluid was measured, and the distribution of ¹⁴C between glucose and lactate was determined by chromatography. From these values, the extraction of glucose and the metabolism of glucose to lactate were calculated. From the decrease in the specific activity of glucose in the perfusion fluid, the influx of glucose from serum was also determined. During steadystate perfusion, 71% of the radioactivity was recovered in the effluent fluid: 50% in the form of glucose, 6% in the form of lactate, and 15% in forms that were not identified. Thus, 50% of the perfusion fluid glucose was cleared, of which 29% was extracted and 21% metabolized. The influx of glucose was proportional to the serum glucose when the latter was about 2.5 mM or 10.0 mM. During perfusion with tracer glucose only, the concentration of glucose in the effluent fluid was 25% that of serum. The transport of glucose from serum was independent of the glucose concentration gradient between serum and perfusion fluid. However, when perfusion fluid glucose concentration was greater than that of serum, transport was inhibited. These studies suggest that in maintaining CSF glucose at a lower concentration than serum glucose, with equal amounts of glucose entering and leaving the CSF, 50% of CSF glucose concentration cleared is replaced by 25% of serum glucose concentration.     

Artifactual Increases in the Concentration of Free GABA in Samples of Human Cerebrospinal Fluid Are Due to Degradation of Homocarnosine

Abstract: Samples of untreated human cerebrospinal fluid (CSF) were kept at room temperature (20±1°C) up to 72 h, and changes in γ-aminobutyric acid (GABA) and homocarnosine contents were measured. The concentration of free GABA increased with time, and concomitantly a similar decrease occurred in the concentration of homocarnosine. Total GABA after hydrolysis (present in human CSF at concentrations 40–100 times that of free GABA) did not change. After 2 h the increase in CSF GABA for seven subjects ranged from 42 to 244 pmol/ml. The rate of increase in CSF GABA was positively correlated with the initial homocarnosine concentration. Approximately 5% per h of the initial homocarnosine content was degraded during the first 7 h at room temperature; thereafter the rate gradually decreased. No free GABA was formed in CSF frozen at −70°C for 10 days. When this CSF was restored to room temperature, the formation of free GABA from homocarnosine occurred at essentially the same rate as that observed in fresh CSF. These results demonstrate that the well-known artifactual increase in GABA concentration of untreated human CSF depends on the concentration of homocarnosine. The rapidity of this increase (up to 2 pmollmlimin) could account for disparities among CSF free GABA concentrations previously reported from normal subjects. It is suggested that measurement of concentrations of total GABA in the CSF would provide a better index of human brain GABA concentration than determination of CSF free GABA.