[The role of citicoline in glaucoma]. / Bedeutung von Citicolin bei der Glaukomerkrankung.

BACKGROUND: In recent years, many experimental and clinical studies have shown that in glaucoma, neuronal degeneration occurs not only at the level of the retina and optic nerve, but also along the entire visual pathway and the brain. OBJECTIVE: This article presents the neuroprotective effects of citicoline and their mechanisms in glaucoma disease. MATERIALS AND METHODS: The relevance of citicoline is explained against the background of neuroanatomy, neuroimaging, and the pathogenesis of glaucoma. Data from experimental and clinical studies are presented and a conclusion is drawn for clinical application. RESULTS: Citicoline has a neuroprotective effect via mechanisms relevant to glaucoma. The neuroprotective effect of citicoline in open-angle glaucoma can be demonstrated functionally and morphologically. It is independent of the glaucoma damage and intraocular pressure, and usually occurs only after 1 year. The effects of oral citicoline occur at a daily dose of 500-1000 mg. Citicoline can be taken permanently or in cycles. No side effects occurred in the studies when taking citicoline. Citicoline can improve cognitive performance and thus treatment adherence as well as quality of life in glaucoma patients. CONCLUSION: This relatively old nootropic drug, which is now marketed as a food supplement, seems to be a valuable addition to conventional treatment and also a rational option for prophylaxis of open-angle glaucoma.

5-Aminolevulinic acid-mediated sonosensitization of rat RG2 glioma cells in vitro.

Sonodynamic therapy (SDT) is a promising technique based on the ability of certain substances, called sonosensitizers, to sensitize cancer cells to non-thermal effects of low-energy ultrasound waves, allowing their destruction. Sonosensitization is thought to induce cell death by direct physical effects such as cavitation and acoustical streaming as well as by complementary chemical reactions generating oxygen free radicals. One of the promising sonosensitizers is 5-aminolevulinic acid (ALA) which upon selective uptake by cancer cells is metabolized and accumulated as protoporphyrin IX. The objective of the study was to describe ALA-mediated sonodynamic effects in vitro on a rat RG2 glioma cell line. Glioma cells, seeded at the bottom of 96-well plates and incubated with ALA (10 µg/ml) for 6 h, were exposed to the sinusoidal US pulses with a resonance frequency of 1 MHz, 1000 µs duration, 0.4 duty-cycle, and average acoustic power varying from 2 W to 6 W. Ultrasound waves were generated by a flat circular piezoelectric transducer with a diameter of 25 mm. Cell viability was determined by MTT assay. Structural cellular changes were visualized with a fluorescence microscope. Signs of cytotoxicity such as a decrease in cell viability, chromatin condensation and apoptosis were found. ALA-mediated SDT evokes cytotoxic effects of low intensity US on rat RG2 glioma cells in vitro. This cell line is indicated for further preclinical assessment of SDT in in vivo conditions.

Impact of thermal effects induced by ultrasound on viability of rat C6 glioma cells.

In order to have consistent and repeatable effects of sonodynamic therapy (SDT) on various cancer cells or tissue lesions we should be able to control a delivered ultrasound energy and thermal effects induced. The objective of this study was to investigate viability of rat C6 glioma cells in vitro depending on the intensity of ultrasound in the region of cells and to determine the exposure time inducing temperature rise above 43 °C, which is known to be toxic for cells. For measurements a planar piezoelectric transducer with a diameter of 20 mm and a resonance frequency of 1.06 MHz was used. The transducer generated tone bursts with 94 µs duration, 0.4 duty-cycle and initial intensity ISATA (spatial averaged, temporal averaged) varied from 0.33 W/cm(2) to 8 W/cm(2) (average acoustic power varied from 1 W to 24 W). The rat C6 glioma cells were cultured on a bottom of wells in 12-well plates, incubated for 24h and then exposed to ultrasound with measured acoustic properties, inducing or causing no thermal effects leading to cell death. Cell viability rate was determined by MTT assay (a standard colorimetric assay for assessing cell viability) as the ratio of the optical densities of the group treated by ultrasound to the control group. Structural cellular changes and apoptosis estimation were observed under a microscope. Quantitative analysis of the obtained results allowed to determine the maximal exposure time that does not lead to the thermal effects above 43 °C in the region of cells for each initial intensity of the tone bursts used as well as the threshold intensity causing cell death after 3 min exposure to ultrasound due to thermal effects. The averaged threshold intensity was found to be about 5.7 W/cm(2).

Diverging oxidative damage and heat shock protein 72 responses to endurance training and chronic testosterone propionate treatment in three striated muscle types of adolescent male rats.

The aim of the study was to assess the effects of a combination of anabolic-androgenic steroid abuse and endurance training during adolescence on selected aspects of oxidative stress and antioxidant defenses in various striated muscle types. The effects were studied of testosterone propionate (TP) treatment (8 and 80 mg/kg/week, for 6 weeks), given alone or in combination with moderate-intensity endurance training, starting at adolescence, on thiobarbituric acid-reactive substances and heat shock protein 72 (Hsp72) contents, and androgen receptorm(AR) mRNA level in the heart left ventricle, soleus and extensor digitorum longus of male Wistar rats. TP treatment alone markedly elevated thiobarbituric acid-reactive substances only in the left ventricle and soleus; this effect was but marginally enhanced by endurance training. The training alone markedly elevated Hsp72 content in all muscles studied. TP treatment alone dose-dependently upregulated Hsp72, while the lower TP dose slightly curtailed the effect of the training. Low-dose TP treatment alone elevated, whereas high-dose TP treatment alone lowered androgen receptor mRNA level in the soleus and extensor digitorum longus. Endurance training alone elevated AR mRNA in all muscles studied, whereas TP treatment dose-dependently counteracted this effect. Exercise-associated rise in body temperature was significantly less in the TP-treated rats. We came to the conclusion that chronic suprapharmacological TP treatment might exert a protective effect on muscle cell proteins in adolescent sedentary rats, but it markedly enhanced lipid peroxidation. These effects were unlikely to result from an androgen receptor-mediated genomic action of testosterone. Exercise-related heat stress, and not oxidative stress, was mainly responsible for Hsp72 upregulation in striated muscles of chronic TP-treated endurance-trained adolescent male rats.

Metabolic changes in rat brain following intracerebroventricular injections of streptozotocin: a model of sporadic Alzheimer's disease.

A decrease in cerebral glucose metabolic uptake is an early and characteristic sign of Alzheimer's disease (AD). Streptozotocin (STZ) is a bacterial toxin which damages insulin-producing cells and insulin receptors. Intracerebroventricular (icv) application of STZ in rats has been found to chronically decrease cerebral glucose uptake and produce other effects that bear a resemblance to several other molecular and pathological features of AD. In the present experiments in vivo (1)H MR Spectroscopy with short echo time (3 ms) was used to non-invasively obtain a neurochemical profile of rat brains, 3 weeks and 2 months after double icv injections of STZ or vehicle. Seventeen metabolites were quantified from 27 microL tissue volume which included hippocampus and a part of cerebral cortex, using the LCModel and unsuppressed water signal as an internal reference. Three weeks after icv STZ several metabolites were significantly decreased, the most prominent changes noted in glycerophosphocholine and phosphocholine (-38 +/- 5%), glutathione (-37 +/- 4%), taurine (-30 +/- 19%), glutamate (-26 +/- 14%), phosphocreatine (-23 +/- 15%) and N-acetylaspartate (-16 +/- 6%). On the contrary, the concentration of N-acetylaspartylglutamate was found significantly increased (+38 +/- 18%). After 2 months some of these changes were even more pronounced. We conclude that in vivo (1)H MRS of rat brain following icv STZ injections provides a new input into a better understanding of the critical dependency of neural function and structure on brain glucose consumption, and may be of relevance in further studies of AD pathomechanism.

Effect of parenteral cladribine on relapse rates in patients with relapsing forms of multiple sclerosis: results of a 2-year, double-blind, placebo-controlled, crossover study.

OBJECTIVE: This randomized, 2-year, double-blind, placebo-controlled, crossover study evaluated cladribine for relapsing forms of multiple sclerosis. PATIENTS: (n = 84) received seven 5-day courses of subcutaneous cladribine at 5 mg/day (group A) or placebo (group B) in year 1; treatment was reversed in year 2. RESULTS: Cladribine was well tolerated and associated with a favorable safety profile. Mean Expanded Disability Status Scale scores remained stable. In group A, mean relapse rates were 0.15 in year 1 (cladribine) and 0.42 in year 2. In group B, relapse rates were 0.61 in year 1 and 0.50 in year 2 (cladribine). PATIENTS required fewer steroid courses during cladribine periods. The therapeutic efficacy of cladribine was associated with a sustained reduction in lymphocyte count.

Astrocytic activation in relation to inflammatory markers during clinical exacerbation of relapsing-remitting multiple sclerosis.

The study aimed to assay the cerebrospinal fluid (CSF) levels of protein S100B, a biomarker of astrocyte activation in relation to kynurenic acid (KYNA) and nitric oxide (NO) metabolites, nitrate/nitrite (NOx) concentrations in acute relapse multiple sclerosis (MS) patients. Twenty relapsing-remitting MS (RR-MS) patients and 10 controls were enrolled. RR-MS patients were assessed on the expanded disability status scale (EDSS) and underwent lumbar puncture. The CSF KYNA, NOx and S100B levels were significantly higher in RR-MS group compared to controls (p = 0.01, 0.001, 0.04, respectively). There was a significant correlation between CSF S100B and KYNA (p = 0.01) but not NOx (p > 0.05) in RR-MS. CSF KYNA, NOx or S100B concentrations did not correlate with disease characteristics of MS patients. Our study suggests the activation of the kynurenine pathway leading to the increase of neuroprotective KYNA in the CSF of MS patients during acute relapse what contrasts with chronic phases of the disease.

ESR and monolayer study of the localization of coenzyme Q10 in artificial membranes.

The data obtained from the ESR experiments show a complex, depth dependent effect of CoQ10 on the lipid molecules mobility in the bilayer. These effects depend both on its concentration and the temperature. CoQ10 disturbs not only the hydrophobic core of the membrane but also the region close to the hydrophilic headgroups of phospholipids. Both these effects could be explained by the fact that the high hydrophobicity of CoQ10 causes the molecules to position itself in the interior of the bilayer, but at the same time its water seeking headgroup is located close to the region of the polar headgrops of membrane lipids. The presence of CoQ10 in the hydrophobic core has further implications on the properties of membrane intrinsic domain. Results of monolayer experiments indicate that CoQ10 may form aggregates when mixed with PC molecules in the lipid hydrocarbon chain-length dependent manner. CoQ10 is not fully miscible with DMPC or DPPC but it is well miscible with the long-chain DSPC molecules. Our suggestion is that CoQ10 when present in long-chain phospholipid bilayer, interacts with saturated fatty acyl-chains and adapt the structure which allows such interactions: either parallel to the saturated acyl chains or "pseudo-ring" conformation resembling sterol structure.

Metabolite ratios measured with a short echo time 1H-MRS in the frontal lobe of young healthy subjects: an approach to generate reference 1H-MRS data.

Proton magnetic resonance spectroscopy (1H-MRS) performed with a semi-automated Elscint Prestige 2 Tesla tomograph/spectroscope was used to determine the ratios of the most prominent resonances recorded from the 2 x 2 x 2 cm voxels located in the frontal lobe of healthy young males and females. The values of major metabolite ratios (NAA/Cr, Cho/Cr and ml/Cr) were normally distributed and comparable to results reported by other groups using advanced automated equipment. The coefficients of variation were smaller when the composed metabolite ratios (i.e. the ratios of a given resonance signal to the sum of all signals considered) were calculated. The composed metabolite ratios approach may be more sensitive for discrimination between normal and pathologically changed brain.

The effects of citicoline and/or MK-801 on survival, neurological and behavioral outcome of mice exposed to transient hyperglycemia and oligemic hypoxia.

UNLABELLED: The effects of citicoline and/or low dose of MK-801 (sufficient to prevent the development of seizures) on survival, neurological and behavioral recovery following transient hyperglycemic-oligemic-hypoxic insult have been evaluated in mice. Neurological recovery was assessed semi-quantitatively on the third and the 10th day after the insult, and behavioral tests evaluating spontaneous locomotor activity, motor coordination and spontaneous alternation performance were performed on day 10. Neither drug given alone did influence survival rate, but the combination of MK-801 and higher citicoline dose decreased mortality on day 10. Behavioral performance was markedly compromised by the insult. Citicoline, but not MK-801, slightly but significantly improved behavioral outcome in all three tests. CONCLUSION: when brain ischemic insult is complicated with acute hyperglycemia, post-treatment with citicoline combined with MK-801 in low anti-convulsive dose improves survival and neurological recovery, and citicoline but not MK-801 enhances behavioral recovery.

2-deoxyglucose enhances epileptic tolerance evoked by transient incomplete brain ischemia in mice.

The aim of the study was to assess the influence of chronic treatment with a non-metabolisable glucose analogue, 2-deoxyglucose (2-DG) at a 150 mg/kg dose on long-term epileptic tolerance (ET) evoked by 30 min bilateral carotid artery clamping (BCCA) in mice. The effects of protein synthesis inhibition with cycloheximide (CHX), given in three daily doses of 2.5 mg/kg starting either 1 day before (peri-insult regimen) or 1 day after the priming insult (post-insult regimen), on ET development was also studied. Seizures were induced 14 days after BCCA with 3.5 mg/kg of bicuculline; this dose (CD97) evokes convulsions in 97% of normal untreated mice. BCCA resulted in decreased mortality, prolonged latency to the onset of generalised convulsions and decreased overall seizure score. CHX given in the post-insult regimen did not influence, while the peri-insult regimen abolished, all signs of BCCA-evoked ET. 2-DG treatment of sham-operated animals resulted in a moderate but significant decrease in mortality rate and a tendency toward a lower seizure score. BCCA combined with 2-DG treatment resulted in a marked decrease in mortality rate, as well as reduction in all indicators of seizure susceptibility. CHX abolished the antiepileptic effects of BCCA alone, as well as BCCA combined with 2-DG, while it did not influence the 2-DG-related decrease in mortality. We conclude that the development of BCCA-induced epileptic tolerance, as well as unmasking antiepileptic effects of 2-DG by BCCA, is dependent on protein synthesis.

CDP-choline, but not cytidine, protects hippocampal CA1 neurones in the gerbil following transient forebrain ischaemia.

The effects of CDP-choline (citicoline), cytidine monophosphate or cytidine on the number of CA1 hippocampal neurones surviving five-minute forebrain ischaemia have been evaluated in gerbils. The substances tested were given in daily doses equivalent on a molar basis to 500 mg/kg CDP-choline, starting immediately after ischaemia. On day five the brains were perfused, postfixed, cut into 10 microm slices and stained with cresyl violet, and the number of neurones in the CA1 sectors was counted manually under a light microscope at magnification x 400. The results indicate a significant degree of protection provided by citicoline, but no protection by cytidine monophosphate or cytidine. The choline moiety of CDP-choline appears to be essential for the neuroprotective properties of the drug.

Brain tolerance and preconditioning.

In this review article the authors describe a phenomenon of "brain tolerance" which represents transient resistance of brain tissue to a lethal insult established by preconditioning with a mild insult of short duration. Tolerance evoked by brief ischemia resembles transient ischemic attack(s) (TIA) often preceding full-blown ischemic stroke in a clinical setting. A series of recent studies have described another relevant phenomenon termed "chemical preconditioning". Several substances interfering with cellular energy metabolism applied in subtoxic doses may provide protection against lethal insults of a different type. For example, 3-nitropropionic acid (3-NP), antibiotics erythromycin and kanamycin, acetylsalicylic acid, and 2-deoxyglucose have been shown to evoke tolerance. Recently, we have reported that NMDA receptor antagonists and 2-deoxyglucose used at relatively low doses were potent agents to potentiate the protective anticonvulsant effect induced by transient brain mild ischemia. Further studies are expected to prove similar action of these drugs in other experimental models. Based on the accumulated experimental and clinical data the brain tolerance subsequently reinforced by pharmacological intervention might become a successful prophylactic strategy against serious brain insults in patients.

Hematological effects of intermittent 2-hour infusions of cladribine in multiple sclerosis patients: a comparison of 2 dosage patterns.

Cladribine is a lymphocytotoxic purine nucleoside with potential for treatment of autoimmune diseases. However, optimal administration regimens remain to be established. Twenty multiple sclerosis patients enrolled into this study were given 30 intermittent 2-hour cladribine infusions (0.07 mg/kg per infusion) each. Ten patients received cycles of 5 consecutive daily infusions at 5-week intervals (clustered dosage) on an inpatient basis; the other 10 patients received 1 infusion weekly (nonclustered dosage) on an outpatient basis. Red blood cell (RBC), platelet, and total white blood cell (WBC) counts were assessed at 5-week intervals during the treatment and at 13-week intervals during a 26-week follow-up period. Major WBC and lymphocyte subsets were assessed cytometrically at 15-week intervals during the treatment and at 13-week intervals thereafter. The clustered dosage produced a lasting decline in granulocyte count, a delayed decrease in monocyte count, and a transient decrease in RBC count. The nonclustered dosage caused a larger and persistent decline in RBC count, a smaller (P = .051. compared over the study period) decrease in monocyte count, and no change in granulocyte count. Both regimens transiently reduced natural killer and B-cell subsets (by 40%-60% and >80%, respectively) and caused lasting declines in CD4+ T-cell subsets (by >50%). No significant change was found in CD8+ T-cell subsets. These results show similar potency of these regimens with respect to major lymphocyte subsets, while suggesting that the nonclustered dosage is less toxic to myeloid precursors and more toxic to erythroid lineage precursors.

Cladribine with prednisone versus chlorambucil with prednisone as first-line therapy in chronic lymphocytic leukemia: report of a prospective, randomized, multicenter trial.

The efficacy and toxicity of cladribine (2-CdA) + prednisone (P) versus chlorambucil (Chl) + P were compared in previously untreated patients with progressive or symptomatic chronic lymphocytic leukemia (CLL) in a randomized, multicenter prospective trial. Eligible patients were assigned to either 2-CdA 0.12 mg/kg per day in 2-hour infusions and P 30 mg/m(2) per day for 5 consecutive days or Chl 12 mg/m(2) per day and P 30 mg/m(2) per day for 7 consecutive days. Three courses were administered at 28-day intervals or longer if myelosuppression developed. The therapy was finished if complete response (CR) was achieved. Of 229 available patients 126 received 2-CdA+P and 103 received Chl+P as a first-line treatment. CR and overall response rates were significantly higher in the patients treated with 2-CdA+P (47% and 87%, respectively) than in the patients treated with Chl+P (12% and 57%, respectively) (P = .001). Progression-free survival was significantly longer in the 2-CdA-treated group (P = .01), but event-free survival was not statistically different. Thirteen percent of patients were refractory to 2-CdA+P and 43% to Chl+P (P = .001). Drug-induced neutropenia was more frequently observed during 2-CdA+P (23%) than Chl+P therapy (11%) (P = .02), but thrombocytopenia occurred with similar frequency in both groups (36% and 27%, respectively). Infections were seen more frequently in the 2-CdA+P-treated group (56%) than in the Chl+P-treated group (40%; P = .02). Death rates have so far been similar in patients treated with 2-CdA (20%) and with Chl (17%). The probability of overall survival calculated from Kaplan-Meier curves at 24 months was also similar for both groups (78% and 82%, respectively). (Blood. 2000;96:2723-2729)

Kinetics of a nucleoside release from lactide-caprolactone and lactide-glycolide polymers in vitro.

We assessed the rate of release of a model nucleoside (adenosine, 5%, w/w) from nine different lactide-glycolide or lactide-caprolactone polymers. The polymer discs were eluted every second day with an artificial cerebrospinal fluid at the elution rate roughly approximating the brain extracellular fluid formation rate. Adenosine in eluate samples was assayed by HPLC. Three polymers exhibited a relatively constant release of adenosine for over four weeks, resulting in micromolar concentrations of nucleoside in the eluate. This points to the necessity of further development of polymers of this types as intracerebral nucleoside delivery systems for local treatment of brain tumors.

Immune abnormalities in aneurysmal subarachnoid haemorrhage patients: relation to delayed cerebral vasospasm.

Peripheral blood CD3+, CD19+, CD4+, CD8+ and CD45RO+ mononuclear cell subsets, T-cell proliferative responses to combinations of coimmobilized OKT3 antibody and an ECM protein (collagen I, collagen IV, fibronectin or elastin), and T-cell adhesion to collagen IV, fibronectin and elastin were studied in patients with aneurysmal subarachnoid haemorrhage. No significant difference was found in the major lymphocyte subsets between subarachnoid haemorrhage patients receiving no dexamethasone for brain oedema treatment and healthy blood donors. Compared with the latter, both the dexamethasone-untreated and -treated subarachnoid haemorrhage patients showed decreased relative proliferative responses of circulating T cells to OKT3 combinations with collagen IV and fibronectin, and an increased PHA-activated T-cell adhesion to elastin. CD45RO+, CD4+ and CD19+ peripheral blood cell subsets, CD4+/CD8+ cell ratio, PHA-activated T-cell adhesion to fibronectin and collagen IV, and OKT3-triggered T-cell costimulatory responses to elastin, collagen IV and fibronectin were significantly higher in subarachnoid haemorrhage patients presenting with delayed cerebral vasospasm (DCV) than in their DCV-free counterparts. The DCV-related differences in circulating lymphocyte subsets showed no apparent relationship to the glucocorticoid treatment, whereas the differences in the other indices were confined to the dexamethasone-untreated subarachnoid haemorrhage patients. The above results suggest that the CD4+/CD8+ ratio and T cell-ECM interactions play a role in the emergence of subarachnoid haemorrhage/DCV and may represent potential targets for subarachnoid haemorrhage therapy.

Cladribine with or without prednisone in the treatment of previously treated and untreated B-cell chronic lymphocytic leukaemia - updated results of the multicentre study of 378 patients.

Between January 1992 and January 1999, we treated 378 B-chronic lymphocytic leukaemia (CLL) patients with cladribine (2-CdA), and 255 of the patients were also treated with prednisone. A total of 194 patients were previously untreated, and 184 had relapsed or refractory disease after previous other therapy. Complete response (CR) was obtained in 111 (29.4%) and partial response (PR) in 138 (36.5%) patients, giving an overall response (OR) rate of 65.9%. CR and OR were achieved more frequently in patients in whom 2-CdA was a first-line treatment (45.4% and 82.5% respectively) than in the pretreated group (12.5% and 48.4% respectively) (P < 0.0001). The median duration of OR for previously untreated patients was 14.7 months and for pretreated patients 13.5 months (P = 0.09). The median survival evaluated from the beginning of 2-CdA treatment was shorter in the pretreated group (16.3 months) than in the untreated group (19.4 months) (P < 0.0001). A total of 117 (63.9%) patients died in the pretreated group and 63 (32.6%) in the untreated group. In pretreated patients, 2-CdA + prednisone (P) and 2-CdA alone resulted in similar OR (51.0% and 45.0% respectively; P = 0.4). In contrast, in untreated patients, 2-CdA + P produced a higher OR (85.4%) than 2-CdA alone (72.1%) (P = 0.04). Infections and fever of unknown origin, observed in 91 (49.4%) pretreated and 74 (38.1%) untreated patients (P = 0.03), were the most frequent toxic effects. Our results indicate that 2-CdA is an effective, relatively well-tolerated drug, especially in previously untreated CLL.

[The mechanisms of neuronal death and advances in the neural protection in epilepsy]. / Mechanizmy smierci neuronów i postepy neuroprotekcji w padaczce.

Epilepsias are chronic central nervous system diseases which manifest by recurrent seizures. They may be classified into channelopathies resulting from mutations of ionic channels or receptors for neurotransmitters, and epilepsias which are secondary to cortical damage and result from the reconstruction of cortical neural networks. Chronic or generalized seizures initiate processes called the "excititoxic cascade" the effector side of which, quite similar to that of the "ischemic cascade", may lead to neuronal death through either apoptosis or necrosis. Therefore, complementing pharmacological treatment of epilepsia with neuroprotective substances such as free radical scavenger or CDP-choline which blocks the activation of phospholipase A2 may be indicated.

Malignancy of brain tumors evaluated by proton magnetic resonance spectroscopy (1H-MRS) in vitro.

Biopsies of 6 malignant gliomas (grade 3 or 4) and 11 low-grade meningiomas were extracted with perchloric acid or methanol/water, and the fully-relaxed 1H-MRS spectra of the extracts containing water-soluble metabolites and a concentration and chemical shift standard were recorded at 11.4 T. The resonance signals assigned to inositol (Ino), glycerophospho- and phosphocholine (GPC + PC), choline (Cho), creatine and phosphocreatine (Cr + PCr), glutamate (Glu), acetate (Ac), alanine (Ala) and lactate (Lac) were integrated, and analyzed by two methods. First, the concentrations of the aforementioned substances in the bioptates were estimated from their resonance signals in the extracts. Second, these signals were normalized to the Cr + PCr resonance signal. The Mann-Whitney U-test was used to verify statistical significance between the data sets obtained for gliomas and meningiomas. When the first method of analysis was used, the only difference was in the Ala concentration, which in meningiomas was on average 4 times higher than in gliomas (P < 0.01). However, when the second method of analysis was applied, gliomas expressed lower normalized resonance signals of Ala and Glu (P < 0.001, ranges not overlapping), Lac (P < 0.005), as well as Ino and GPC + PC (P < 0.05). In proton MR spectra of brain tumor tissue extracts containing water soluble metabolites, the resonance signals normalized to that of total creatine may provide a very good discrimination between malignant gliomas and low-grade meningiomas.