Cerebrospinal fluid beta-glucuronidase activity in rabbits with experimental allergic encephalomyelitis

Using a microassay, cerebrospinal fluid (CSF) β-glucuronidase activity increased precipitously in rabbits developing paralytic signs and characteristic lesions of experimental allergic encephalomyelitis (EAE). CSF protein concentration also increased in animals with EAE, but striking dissociation between protein and enzyme values in occasional rabbits suggests that each parameter represents an independent response to nervous tissue injury.     

Cerebrospinal fluid signs of neuronal damage after antiretroviral treatment interruption in HIV-1 infection

Background  

The neurofilament is a major structural component of myelinated axons. Increased cerebrospinal fluid (CSF) concentrations of the light chain of the neurofilament protein (NFL) can serve as a sensitive indicator of central nervous system (CNS) injury. To assess whether interrupting antiretroviral treatment of HIV infection might have a deleterious effect on the CNS, we measured NFL levels in HIV-infected subjects interrupting therapy.     

Interaction of acetylcholinesterase inhibitor donepezil with ionic channels of the neuronal membrane

The effects of donepezil on voltage-dependent Ca²⁺-and low-threshold K⁺-current were studied on isolated molluscan neurons using two-electrode voltage clamp technique. Donepezil reduced the amplitude of voltage-dependent Ca²⁺-current (IC₅₀=7.9 μM) and shifted the current-voltage relationships toward hyperpolarization. Donepezil in low concentration (5 μM) increased, while in higher concentrations (≥10 μM) decreased the low-threshold K⁺-current. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 142, No. 10, pp. 364–368, October, 2006     

Cerebrospinal fluid activity of tissue plasminogen activator in patients with neurological diseases.

AIM: To study cerebrospinal fluid (CSF) activity of tissue plasminogen activator (tPA) in patients with neurological diseases. METHODS: CSF tPA and urokinase (uPA) activities were studied using an immunocapture assay and zymography in 44 patients with neurological disease and 20 reference subjects. The patient group comprised three patients with meningitis, 21 with encephalitis, nine with acute lymphoblastic (n = 7) and myeloid (n = 2) leukaemia, seven with multiple sclerosis, three with facial paresis, and one with polyradiculitis. RESULTS: Raised tPA activities were observed in patients with multiple sclerosis, leukaemia and encephalitis. In contrast, there were no differences in the mean activities of tPA in patients with meningitis or other diseases compared with the reference subjects. The highest tPA activities were found in patients with multiple sclerosis. The mean activity in patients with leukaemia was higher than in those with meningitis and polyradiculitis, but not encephalitis and facial paresis. Although the CSF tPA activity correlated positively with age in reference subjects, no correlation was observed in patients. Samples were qualitatively screened for both tPA and uPA activity by zymography and positive samples were quantitated. Some of the samples had quantifiable levels of uPA activity: three of seven multiple sclerosis samples, 10 of 21 samples from patients with encephalitis and five of nine leukaemic samples. The highest activities were recorded in patients with leukaemia. uPA was not detected in the CSF of the patients with meningitis, facial paresis or polyradiculitis. CONCLUSIONS: Plasminogen activator activity can be measured reliably in CSF and the assessment of tPA activity may be useful for studying the pathogenesis of neurological diseases.     

Comparative effects of huperzine A, donepezil and rivastigmine on cortical acetylcholine level and acetylcholinesterase activity in rats

The cholinesterase inhibitors huperzine A, donepezil and rivastigmine were compared for their effects on extracellular acetylcholine concentration and acetylcholinesterase activity in the rat cortex. After i.p. injection, huperzine A (0.25-0.75 micromol/kg), donepezil (2-6 micromol/kg) and rivastigmine (0.75-1.5 micromol/kg) dose-dependently elevated the concentration of acetylcholine. The duration of huperzine A was longest. The time courses of cortical acetylcholinesterase inhibition with middle doses of these agents mirrored the increases of acetylcholine at the same doses. However, acetylcholinesterase inhibition was disproportionately greater after middle dose of rivastigmine than doses of huperzine A and donepezil that increased acetylcholine to a similar extent. Muscle fasciculation appeared only after donepezil with a dose-dependent incidence and intensity. In molar terms, huperzine A was 8- and 2-fold more potent than donepezil and rivastigmine, respectively, in increasing cortical acetylcholine levels, with a longer-lasting effect.     

Catalytic antibodies with acetylcholinesterase activity

We describe three catalytic cholinesterase-like catalytic antibodies (Ab1), as well as anti-idiotypic (Ab2) and idiotypic (Ab3) antibodies, to one of the Ab1s. The Ab1s were raised against the human erythrocyte acetylcholinesterase (AChE), and are unusual in that they both recognise and resemble acetylcholinesterase in their catalytic activity. No contamination of the antibody preparations with either acetylcholinesterase or butyrylcholinesterase (BChE) was found. None of the Ab2s showed catalytic activity, whereas four Ab3s did (an incidence of 1.26% of all Ab3s). Although there is considerable resemblance between Ab1s and Ab3s, there are significant differences between the two groups. All the antibodies were inhibited by phenylmethylsulphonyl fluoride (PMSF), indicating the presence of a serine residue in their active sites, and were inhibited by the cholinesterase active site inhibitors iso-OMPA and pyridostigmine, suggesting the similarity of the sites to those of cholinesterases. The Ab3s resemble the Ab1s in their ability to hydrolyse both acetyl and butyrylthiocholine (BTCh). However, the Ab3s appear to be better catalysts, having significantly reduced K(m) values (for acetyl, but not for butyrylthiocholine) and increased turnover numbers (K(cat)), rate enhancements (K(cat)/K(uncat)) and K(cat)/K(m) ratios, for both substrates, although these values by no means approach those of the natural enzymes. The Ab1s appear to have structures resembling the anionic sites of cholinesterases, as shown by their reaction with the anionic site inhibitors (edrophonium and tetramethylammonium). No such reactions were observed in the Ab3s. None of the antibodies show evidence of the sites resembling the peripheral anionic site (PAS) of acetylcholinesterase. All the antibodies recognise, to varying degrees, the peripheral anionic site of acetylcholinesterase. This was shown by their ability to inhibit acetylcholinesterase, to compete with peripheral site inhibitors, and to block acetylcholinesterase-mediated cell adhesion, a property of this site. The results indicate idiotypic mimicry of a catalytic antibody's active site, and suggest that the development of the catalytic activity in the anti-acetylcholinesterase antibodies may be related to the structural features of the peripheral anionic site of acetylcholinesterase.     

Cerebrospinal fluid and plasma apolipoproteins in patients with multiple sclerosis

Apolipoprotein (apo) E is synthesized by cells of the central and peripheral nervous systems, and its synthesis and secretion in the peripheral nervous systems, and its synthesis and secretion in the peripheral nervous system of animals are greatly stimulated following Wallerian degeneration. It has been suggested that apo E functions in the metabolism of myelin lipids, but its physiologic role in nervous tissue has not been elucidated. To determine if apo E might play a role in demyelinating neuropathy, the concentrations were examined of apos E and A-1 in the cerebrospinal fluid (CSF) and plasma of patients with multiple sclerosis during clinical remission, and in patients with no neurologic disease. Serum and CSF albumin concentrations were measured to account for the possible influences of serum apo E concentration and/or altered blood-brain barrier permeability on the CSF apo E pool. A CSF index for apo E and apo A-1 was calculated in the same manner presently used for calculation of immunoglobulin G (IgG) production in the nervous system of patients with multiple sclerosis (MS). The results showed that the concentrations of apo E, apo Al, and albumin in the CSF of the MS patients were not significantly altered. The concentration of apo E in the serum, however, was significantly (p less than 0.001) decreased by 44 percent in the MS patients. The difference was relatively specific for serum apo E because the serum apo Al and albumin concentrations were unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mecamylamine interactions with galantamine and donepezil: effects on learning,acetylcholinesterase, and nicotinic acetylcholine receptors

Patch-clamp recordings of single ion channel activity demonstrated that donepezil, but not galantamine, could be blocked by the nicotinic cholinergic antagonist mecamylamine, suggesting that galantamine acted at a separate (allosteric) site. The aim of this experiment was to demonstrate at a whole organism, behavioral level that galantamine, but not donepezil, could reverse mecamylamine-induced learning impairment. Forty-four young female rabbits received 15 sessions in the 750-ms delay eyeblink classical conditioning procedure, after one of five drug treatments: 0.5 mg/kg mecamylamine, 3.0 mg/kg donepezil, 0.5 mg/kg mecamylamine plus 3.0 mg/kg galantamine, 0.5 mg/kg mecamylamine plus 3.0 mg/kg donepezil, or sterile saline vehicle. An additional 24 young female rabbits were tested in the explicitly unpaired condition after treatment with the same mecamylamine plus galantamine or donepezil combinations or with vehicle. In a previous study we demonstrated that 3.0 mg/kg galantamine facilitated learning in young rabbits. Donepezil (3.0 mg/kg) did not facilitate learning in this experiment. However, both galantamine and donepezil reversed the deleterious effects of mecamylamine on learning. Significant differences in plasma and brain acetylcholinesterase levels were detected among the drug treatment groups. Fifteen daily injections did not produce statistically significant changes in nicotinic receptor binding in any of the five treatment groups. One possible interpretation of these results is that donepezil affected nicotinic acetylcholine receptors by raising the synaptic level of acetylcholine and hence, the probability of receptor activation, whereas galantamine bound to distinct allosteric sites not blocked by mecamylamine.     

Cerebrospinal fluid interleukin 1 like activity during chronic relapsing experimental allergic encephalomyelitis.

Cerebrospinal fluid (CSF) and plasma samples were taken from strain 13 guinea pigs in various stages of chronic relapsing experimental allergic encephalomyelitis (CREAE). The samples were assayed for interleukin 1 (IL-1) in the C3H/Hej mouse thymocyte assay. After removal of inhibitors, IL-1 was detectable in low amounts in plasma (5 U/ml) throughout the course of the disease but was raised in the acute phase (12 U/ml). CSF IL-1 was, however, only present in low amounts (6 U/ml) during the acute phase but was elevated (18 U/ml) during the chronic stages of CREAE. During the relapse phase levels of IL-1 correlated with the total leucocyte count in the CSF. On gel filtration of CSF, IL-1 activity eluted at approximately 15 kD and could not be attributed to leakage of plasma IL-1 during CSF puncture or IL-2 activity.     

Cerebrospinal fluid lysozyme in bacterial and viral meningitis

The concentration of lysozyme (LZM) in cerebrospinal fluid was determined in 25 patients with bacterial meningitis, in 18 patients with viral meningitis and in 25 control patients who had other fibrile illnesses. The concentration of LZM was less than 1.5 microgram/ml in all control patients, and slightly to markedly raised in 10 patients with viral meningitis and in 11 out of 13 patients with untreated bacterial meningitis. The concentration of LZM was significantly different in the viral and bacterial meningitis patients (p less than 0.001). Most raised concentrations of cerebrospinal fluid LZM persisted for at least one week after the start of antibiotic treatment. The concentrations of LZM correlated well with concentrations of lactic dehydrogenase. These results show that the determination of cerebrospinal fluid LZM is a useful tool in the differential diagnosis of meningitis, particularly when the prehospital treatment with antibiotics may be responsible for a diagnostically misleading negative bacterial culture of the cerebrospinal fluid and altered cerebrospinal fluid cytology.     

Cerebrospinal fluid cytoskeleton proteins in patients with subcortical white-matter dementia

The cerebrospinal fluid (CSF) levels of two cytoskeleton proteins, tau and the light subunit of neurofilament protein (NFL), both considered to reflect cerebral white-matter components, were investigated in a group of patients with a subtype of vascular dementia called 'subcortical white-matter dementia' (SWD). The group consisted of 25 demented patients with frontosubcortical brain syndromes, white-matter changes on computed tomography or magnetic resonance imaging and vascular disease or pronounced vascular risk factors. CSF-NFL was increased, whereas CSF-tau was normal, suggesting a differential involvement of the cytoskeleton in this patient group. The albumin ratio and the apolipoproteinE4 (ApoE4) allele status were also investigated. The albumin ratio was increased, indicating damage to the vessel walls with breakdown of the blood-brain barrier. No relationship was found between ApoE4 alleles and CSF levels of tau or NFL in this patient group. Besides presenting original data, the disease status of SWD is also discussed.     

Cerebrospinal fluid protein fractions in health and disease

Samples of lumbar cerebrospinal fluid from 20 healthy subjects and 20 patients with a variety of neurological disorders have been subjected to cellulose acetate electrophoresis and quantitative analyses carried out on the seven separated protein fractions. Normal percentage and absolute concentration limits for each fraction have been determined and compared with previous reports. It is concluded that absolute values are more valuable than percentage values in the discrimination of disease from health, but that the mechanisms governing protein abnormalities are difficult to ascertain from spinal fluid electrophoresis alone.     

Differential increase in cerebrospinal fluid-acetylcholinesterase after treatment with acetylcholinesterase inhibitors in patients with Alzheimer's disease

The clinical significance and the effects of pharmacological treatment of patients with Alzheimer's disease (AD) were evaluated by measurement of acetylcholinesterase (AChE) in the cerebrospinal fluid (CSF). CSF–AChE of AD patients was lower, not significantly, compared with controls. However, CSF–AChE was significantly increased after treatment of AD patients with AChE inhibitors (donepezil and galantamine). The increase was higher in patients treated with donezepil than in those treated with galantamine, which might be related to different mechanisms for the substances. The increase was also dose-dependent, and was especially marked in patients showing a clinical response. These data suggest that CSF biomarkers are capable not only of identifying a biochemical effect of drugs, but also of differentiating between different compounds in a dose-dependent manner.     

Determination of cholinesterase and acetylcholinesterase in amniotic fluid

The determination of acetylcholinesterase (AChE) has been shown to be as specific as alphafetoprotein (AFP) for the prenatal detection of open neural tube defects although AFP remains the method of choice. This paper describes a semi-automated technique for the analysis of acetylcholinesterase in amniotic fluid that: A) reduces the cost of the procedure; B) allows for a larger number of samples to be run at a time; and C) provides for more accurate and reproducible procedures and results. Six fetuses with neural tube defects (2 with gastroschisis and 3 where one twin was dead) were detected and found to have elevated AChE, TChE and 2 bands by electrophoresis. Quality control procedures using both pure enzyme and amniotic fluid with low and high levels of the enzyme are described. The analysis of 340 amniotic fluids of normal pregnancies indicates that the normal value for AChE is 5.17 +/- 2.63 mU/ml (97% confidence interval for the mean 4.84-5.49 mU/ml. A group of 27 abnormal pregnancies provides evidence that fetal vomiting and regurgitation, fetal demise, multiple cysts syndrome, idiopathic IUGR, arthrogryposis multiplex, hydrocephaly (stenosis of aqueductus), trisomy 21, trisomy 18, hydronephrosis, pyloric stenosis, heart malformation, ectopia cordis and multiple gestation produce elevated levels of pseudocholinesterase (PChE) in amniotic fluid. The use of pseudocholinesterase levels in amniotic fluid for prenatal diagnosis is proposed and discussed in view of its elevated levels in abnormal pregnancies where AChE is normal. The normal values for PChE are 23.86 mU/ml (mean) and 5.83 for standard deviation. Electrophoretic analysis was performed on all samples with values higher than one standard deviation above the mean.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dopaminergic supersensitivity after long-term bromopride treatment

The effects of bromopride administration on open-field and apomorphine-induced stereotyped behavior of rats were studied. Bromopride induced a displacement to the right of the control dose-response curve constructed for apomorphine stereotypes. Withdrawal from long-term bromopride treatment induced not only a significant increase in ambulation and rearing frequencies for male rats observed in the open-field but also an increased sensitivity of both male and female animals to apomorphine. The results are interpreted as a consequence of the development of supersensitivity of central dopaminergic receptors, probably of the mesostriatal pathways.     

Cerebrospinal fluid interleukin-6 activity in HIV infection and inflammatory and noninflammatory diseases of the nervous system

Interleukin-6 (IL-6) activity was measured in the cerebrospinal fluid (CSF) of patients at different stages of human immunodeficiency (HIV) virus infection and of patients with multiple sclerosis (MS) or other inflammatory (OID) and noninflammatory neurological diseases (OND). In the advanced stages of HIV infection and in OID, IL-6 was detected more frequently (80 and 75% of the cases) and at higher concentrations than in the early stages of HIV infection. MS and OND (44, 48, and 44% of cases). Analysis of CSF and paired sera indicated that IL-6 production can be compartmentalized to either of the fluids. Evidence that altered blood-brain barrier functions can, at least in part, influence the CSF IL-6 levels was found in OID patients. No association was evident between intrathecal immunoglobulin synthesis and CSF IL-6 levels. Interleukin-1 (IL-1) levels were detectable in a minority of the samples from neurological patients; one OID patient had high levels of both CSF IL-1 and IL-6.     

Cerebrospinal fluid biomarkers of brain injury,inflammation and synaptic autoimmunity predict long-term neurocognitive outcome in herpes simplex encephalitis

ObjectivesThe aim was to investigate the correlation between biomarkers of brain injury and long-term neurocognitive outcome, and the interplay with intrathecal inflammation and neuronal autoimmunity, in patients with herpes simplex encephalitis (HSE).MethodsA total of 53 adult/adolescent HSE patients were included from a prospective cohort in a randomized placebo-controlled trial investigating the effect of a 3-month follow-up treatment with valaciclovir. Study subjects underwent repeated serum/cerebrospinal fluid (CSF) sampling and brain magnetic resonance imaging in the first 3 months along with cognitive assessment using the Mattis Dementia Rating Scale (MDRS) at 24 months. CSF samples were analysed for biomarkers of brain injury, inflammation and synaptic autoimmunity. The predefined primary analysis was the correlation between peak CSF neurofilament protein (NFL), a biomarker of neuronal damage, and MDRS at 24 months.ResultsImpaired cognitive performance significantly correlated with NFL levels (rho = –0.36, p = 0.020). Development of IgG anti-N-methyl-D-aspartate receptor (NDMAR) antibodies was associated with a broad and prolonged proinflammatory CSF response. In a linear regression model, lower MDRS at 24 months was associated with previous development of IgG anti-N-methyl-D-aspartate receptor (NMDAR) (beta = –0.6249, p = 0.024) and age (z-score beta = –0.2784, p = 0.024), but not CSF NFL, which however significantly correlated with subsequent NMDAR autoimmunization (p = 0.006).DiscussionOur findings show that NFL levels are predictive of long-term neurocognitive outcome in HSE, and suggest a causative chain of events where brain tissue damage increases the risk of NMDAR autoimmunisation and subsequent prolongation of CSF inflammation. The data provides guidance for a future intervention study of immunosuppressive therapy administered in the recovery phase of HSE.     

Cerebrospinal fluid HIV RNA and drug levels with combination ritonavir and saquinavir.

Combination antiretroviral therapy with ritonavir and saquinavir has established potent and durable activity on plasma viremia. CNS HIV infection may be sequestered from drug therapy that does not penetrate the blood-brain barrier. Penetration of these protease inhibitors into the cerebrospinal fluid (CSF) and CSF HIV RNA levels on such therapy has not been well described. DESIGN/METHODS: In a cross-sectional study, 28 HIV1-infected study subjects were evaluated either before initiation of or before maximal response to ritonavir-saquinavir therapy, during maximal plasma virologic response, and after virologic failure. Simultaneous samples of plasma and cerebrospinal fluid were obtained from 24 study subjects to measure HIV RNA and protease inhibitor levels. RESULTS: Across the treatment groups, a strong correlation was found between plasma and CSF HIV RNA levels (r = 0.870; p < .001). In each study subject with plasma HIV RNA levels below assay limit (80 copies/ml), the CSF HIV RNA level was also below the limit of quantitation. Low levels of saquinavir (<2 ng/ml) and ritonavir (<25 ng/ml) in the CSF were observed, with a CSF:plasma drug concentration ratio of < or = 0.005 (0.5%) in all study subjects evaluated (n = 11). The plasma:CSF HIV RNA ratio was high before or early in treatment (median, 38; interquartile range [IQR], 13,97), but low (median, 0.29; IQR, 0.17, 7.5) in those failing therapy (group C, p < .001). CONCLUSIONS: CSF ritonavir and saquinavir levels are consistent with the estimated known fraction of unbound drug in plasma (<2%). Across these treatment response groups, suppression of plasma viremia can predict low CSF HIV RNA levels. This correlation may represent HIV RNA transport and equilibrium between CSF and plasma, or it may represent CNS anti-HIV activity of protease inhibitors. The low drug levels and inverted ratio of HIV RNA in the CSF compared with plasma early in plasma virologic breakthrough suggests CSF virologic failure may contribute to failure of plasma virologic response.