CERAD-neuropsychological battery in screening mild Alzheimer's disease.

OBJECTIVES: The Consortium to Establish a Registry for Alzheimer's Disease (CERAD) neuropsychological battery (nb) is used as an evaluation tool for dementia. In Finland, CERAD-nb was introduced in 1999 and has been proposed to be used in primary health care. However, some of its parts need reassessment and focusing. The goal of this study was to examine the sensitivity and specificity of the subtests and their cut-off points most appropriate for identifying mild Alzheimer's disease (AD). MATERIALS AND METHODS: The study population consisted of 171 patients with mild AD and 315 cognitively normal elderly. Both groups underwent CERAD-nb investigation as a part of a wider examination procedure. RESULTS: The most efficient subtests to discriminate patients with mild AD from the normal elderly were Wordlist delayed recall and savings, Wordlist learning and Wordlist recognition and a new variable of Total recall. Optimal cut-off points for each subtest are suggested. The sensitivities of the verbal memory subtests varied between 0.75 and 0.94, the specificities between 0.80 and 0.93 and the areas under the receiver operating characteristics curve between 0.89 and 0.96. CONCLUSIONS: The CERAD-nb is capable of differentiating cases with mild AD from normal elderly individuals particularly with its verbal memory subtests. New cut-off scores for CERAD's subtests validated in the study further enhance the differentiating power, and with these clarifications, CERAD-nb is considered appropriate to be used as a screening tool for AD even in primary health care.

DARS2 mutations in mitochondrial leucoencephalopathy and multiple sclerosis.

BACKGROUND: Leucoencephalopathy with brain stem and spinal cord involvement and high brain lactate (LBSL) was first defined by characteristic magnetic resonance imaging and spectroscopic findings. The clinical features include childhood or juvenile onset slowly progressive ataxia, spasticity, and dorsal column dysfunction, occasionally accompanied by learning difficulties. Mutations in DARS2, encoding mitochondrial aspartyl-tRNA synthetase, were recently shown to cause LBSL. The signs and symptoms show some overlap with the most common leucoencephalopathy of young adults, multiple sclerosis (MS). OBJECTIVE: To clarify the molecular background of LBSL patients in Finland, and to look for DARS2 mutations in a group of MS patients. METHODS: Clinical evaluation of LBSL patients, DARS2 sequencing and haplotype analysis, and carrier frequency determination in Finland. RESULTS: All eight LBSL patients were compound heterozygotes for DARS2 mutations: all carried R76SfsX5 change, seven had M134_K165del, and one had C152F change. Axonal neuropathy was found in five of the eight patients. The carrier frequencies of the R76SfsX5 and M134_K165del mutations were 1:95 and 1:380, respectively. All patients shared common European haplotypes, suggestive of common European LBSL ancestors. No enrichment of the two common DARS2 mutations was found in 321 MS patients. CONCLUSION: All LBSL patients were compound heterozygotes, which suggests that DARS2 mutation homozygosity may be lethal or manifest as a different phenotype. The authors show here that despite identical mutations the clinical picture was quite variable in the patients. Axonal neuropathy was an important feature of LBSL. DARS2 mutations cause childhood-to-adolescence onset leucoencephalopathy, but they do not seem to be associated with MS.

Plasma Abeta42 and Abeta40 as markers of cognitive change in follow-up: a prospective, longitudinal, population-based cohort study.

BACKGROUND: Single measurements of plasma Aß are not useful in the diagnostics of Alzheimer's disease (AD). However, changes in plasma Aß levels during repeated testing may be helpful in the prediction and evaluation of progression of the incipient AD or mild cognitive impairment. OBJECTIVE: To examine the relation of baseline and serial plasma Aß levels to cognitive change in follow-up. METHODS: 269 subjects (52 cognitively impaired and 217 controls) from a population-based cohort were clinically followed up from 3 to 6 years. Serial plasma samples were available from 70 subjects who were followed up for 3 years and 43 subjects followed for 6 years. The plasma Aß levels were measured using ELISA. RESULTS: Subjects who declined cognitively during the follow-up had lower levels of plasma Aß42 at the baseline. Plasma Aß42 and the Aß42/Aß40 ratio decreased (-2.4 pg/ml for Aß42 in 6 years) in those who declined in follow-up, whereas Aß42 and the Aß42/Aß40 ratio increased in the subjects who remained cognitively stable or improved in follow-up. Subjects using acetylsalicylic acid, dipyridamole, antidiabetic or anticoagulant drugs as well as subjects with coronary heart disease had higher levels of Aß40. CONCLUSIONS: Low or decreasing plasma Aß42 during the follow-up is associated with cognitive decline. Serial measurement of plasma Aß42 may be useful in the detection of the subjects who are at risk for cognitive decline.

Use of cerebrospinal fluid biomarkers in diagnosis of dementia across Europe.

BACKGROUND AND PURPOSE: Cerebrospinal fluid (CSF) biomarkers have been reported to be useful in dementia diagnosis. Not much is known about their use in clinical practice in Europe. METHODS: We analyzed data from a survey on the use of CSF biomarkers in the diagnosis of dementia across Europe using a questionnaire which was filled out by representatives of the 25 member countries of the European Federation of Neurological Societies (EFNS). RESULTS: Cerebrospinal fluid beta-amyloid, total tau, and phosphorylated tau proteins are frequently evaluated in the majority of the countries (in 18 out of 23 countries). No major technical or ethical issues were found that would hamper the procedure's ability to become routine in early and differential diagnostics of Alzheimer's disease. Cut-off values for beta-amyloid (median 500, range 300-849 pg/ml), total tau (367; 195-450 pg/ml) and phosphorylated tau (60; 40-85 pg/ml) varied considerably amongst countries and even within every country. CONCLUSIONS: Cerebrospinal fluid analysis of beta-amyloid, tau, and phosphorylated tau is frequently used in Europe. However, the use of various cut off values seriously hampers comparability and yields a potential threat to an interpretation and balanced use in clinical practice. We recommend that each laboratory establishes normative data and that multi-centered studies should be organized to explore the reasons for any differences.

EFNS guidelines for the diagnosis and management of Alzheimer's disease.

BACKGROUND AND OBJECTIVES: In 2008 a task force was set up to develop a revision of the European Federation of the Neurological Societies (EFNS) guideline for the diagnosis and management of Alzheimer's disease (AD) and other disorders associated with dementia, published in early 2007. The aim of this revised international guideline was to present a peer-reviewed evidence-based statement for the guidance of practice for clinical neurologists, geriatricians, psychiatrists, and other specialist physicians responsible for the care of patients with AD. Mild cognitive impairment and non-Alzheimer dementias are not included in this guideline. METHODS: The task force working group reviewed evidence from original research articles, meta-analysis, and systematic reviews, published before May 2009. The evidence was classified and consensus recommendations graded (A, B, or C) according to the EFNS guidance. Where there was a lack of evidence, but clear consensus, good practice points were provided. RESULTS: The recommendations for clinical diagnosis, blood tests, neuropsychology, neuroimaging, electroencephalography, cerebrospinal fluid (CSF) analysis, genetic testing, disclosure of diagnosis, treatment of AD, behavioural and psychological symptoms in dementia, legal issues, counselling and support for caregivers were all revised as compared with the previous EFNS guideline. CONCLUSION: A number of new recommendations and good practice points are made, namely in CSF, neuropsychology, neuroimaging and reviewing non-evidence based therapies. The assessment, interpretation, and treatment of symptoms, disability, needs, and caregiver stress during the course of AD require the contribution of many different professionals. These professionals should adhere to these guideline to improve the diagnosis and management of AD.

Combined risk effects of IDE and NEP gene variants on Alzheimer disease.

Polymorphisms in genes encoding amyloid beta-peptide (A beta)-degrading enzymes neprilysin (NEP) and insulin-degrading enzyme (IDE) individually affect the susceptibility to Alzheimer disease (AD) among the Finnish population. Here we show that a combination of risk genotypes for NEP and IDE genes leads to a higher susceptibility to AD. Individuals with the combination of risk genotypes for NEP and IDE conferred a threefold higher susceptibility to AD when compared with individuals not carrying these genotypes. Although no significant interaction was observed between NEP and IDE genes, these data suggest that NEP and IDE exhibit an additive risk effect in AD.

Cerebrospinal fluid, serum and plasma protein oxidation in Alzheimer's disease.

OBJECTIVES: Many studies have shown differences in carbonylation and nitration of individual proteins in brain and body fluids of Alzheimer's disease (AD) patients. Therefore, we wanted to examine whether total levels of these oxidative stress markers of proteins were altered in AD. PATIENTS AND METHODS: Total levels of carbonyls and nitrotyrosine in cerebrospinal fluid, serum and plasma were measured in 22 AD patients and 18 age-matched controls using commercially available enzyme immunoassay kits. RESULTS: Protein carbonylation in cerebrospinal fluid did not differ between AD patients and controls but was decreased in APOE epsilon4 carriers as compared with non-carriers. Serum but not plasma levels of carbonyls tended to be decreased in AD patients as compared with aged controls. Nitrotyrosine concentrations did not differ between the groups. Surrogate cerebrospinal fluid markers for AD, beta-amyloid (1-42) and tau, correlated with blood carbonyl and nitrotyrosine levels. CONCLUSIONS: According to these preliminary data, changes in oxidative metabolism related to the pathogenesis of AD cannot be detected as increased cerebrospinal fluid, serum or plasma protein carbonylation or nitration.

Analyses of mortality risk in patients with dementia treated with galantamine.

OBJECTIVE: To analyze mortality data from patients with Alzheimer's disease (AD), Alzheimer's plus cerebrovascular disease (AD + CVD) or vascular dementia (VaD). METHODS: (1) Meta-analysis of mortality data from double-blind, placebo-controlled, randomized trials; and (2) recontact study to collect additional longer term mortality data from previous galantamine trial participants. RESULTS (META-ANALYSIS): Across 12 trials (< or =6 months duration), there was no increased risk of mortality associated with the use of galantamine (n = 4116) compared with that of placebo (n = 2386) (OR galantamine/placebo: 0.67, 95% CI 0.41-1.10). RESULTS (RECONTACT STUDY): Median survival was 79 months for patients with AD (n = 478) and 59 months for patients with AD + CVD (n = 180) or VaD (n = 145). Prolonged galantamine treatment (> vs < or =6 months) was not associated with decreased survival time (75 vs 61 months respectively; P = 0.02). Cox regression analyses were consistent with the Kaplan-Meier analyses. CONCLUSIONS: We found no short-term or longer term evidence of increased risk of mortality associated with the use of galantamine in patients with AD, AD + CVD or VaD.

A worldwide multicentre comparison of assays for cerebrospinal fluid biomarkers in Alzheimer's disease.

BACKGROUND: Different cerebrospinal fluid (CSF) amyloid-beta 1-42 (Abeta(1-42)), total Tau (Tau) and Tau phosphorylated at threonine 181 (P-Tau) levels are reported, but currently there is a lack of quality control programmes. The aim of this study was to compare the measurements of these CSF biomarkers, between and within centres. METHODS: Three CSF-pool samples were distributed to 13 laboratories in 2004 and the same samples were again distributed to 18 laboratories in 2008. In 2004 six laboratories measured Abeta(1-42), Tau and P-Tau and seven laboratories measured one or two of these marker(s) by enzyme-linked immunosorbent assays (ELISAs). In 2008, 12 laboratories measured all three markers, three laboratories measured one or two marker(s) by ELISAs and three laboratories measured the markers by Luminex. RESULTS: In 2004, the ELISA intercentre coefficients of variance (interCV) were 31%, 21% and 13% for Abeta(1-42), Tau and P-Tau, respectively. These were 37%, 16% and 15%, respectively, in 2008. When we restricted the analysis to the Innotest (N = 13) for Abeta(1-42), lower interCV were calculated (22%). The centres that participated in both years (N = 9) showed interCVs of 21%, 15% and 9% and intra-centre coefficients (intraCV) of variance of 25%,18% and 7% in 2008. CONCLUSIONS: The highest variability was found for Abeta(1-42). The variabilities for Tau and P-Tau were lower in both years. The centres that participated in both years showed a high intraCV comparable to their interCV, indicating that there is not only a high variation between but also within centres. Besides a uniform standardization of (pre)analytical procedures, the same assay should be used to decrease the inter/intracentre variation.

Interferon regulatory factor 5 (IRF5) gene variants are associated with multiple sclerosis in three distinct populations.

BACKGROUND: IRF5 is a transcription factor involved both in the type I interferon and the toll-like receptor signalling pathways. Previously, IRF5 has been found to be associated with systemic lupus erythematosus, rheumatoid arthritis and inflammatory bowel diseases. Here we investigated whether polymorphisms in the IRF5 gene would be associated with yet another disease with features of autoimmunity, multiple sclerosis (MS). METHODS: We genotyped nine single nucleotide polymorphisms and one insertion-deletion polymorphism in the IRF5 gene in a collection of 2337 patients with MS and 2813 controls from three populations: two case-control cohorts from Spain and Sweden, and a set of MS trio families from Finland. RESULTS: Two single nucleotide polymorphism (SNPs) (rs4728142, rs3807306), and a 5 bp insertion-deletion polymorphism located in the promoter and first intron of the IRF5 gene, showed association signals with values of p<0.001 when the data from all cohorts were combined. The predisposing alleles were present on the same common haplotype in all populations. Using electrophoretic mobility shift assays we observed allele specific differences in protein binding for the SNP rs4728142 and the 5 bp indel, and by a proximity ligation assay we demonstrated increased binding of the transcription factor SP1 to the risk allele of the 5 bp indel. CONCLUSION: These findings add IRF5 to the short list of genes shown to be associated with MS in more than one population. Our study adds to the evidence that there might be genes or pathways that are common in multiple autoimmune diseases, and that the type I interferon system is likely to be involved in the development of these diseases.

PET amyloid ligand [11C]PIB uptake and cerebrospinal fluid beta-amyloid in mild cognitive impairment.

BACKGROUND: In mild cognitive impairment (MCI), Alzheimer's disease (AD)-type cerebrospinal fluid (CSF) biomarker profiles predict rapid progression and conversion to AD. An increased brain amyloid burden in AD and MCI has been demonstrated with PET using [(11)C]PIB (Pittsburgh compound B). Little is known about the relationship between these biomarkers in MCI. METHODS: We studied 15 patients with amnestic MCI and 22 controls with PET using [(11)C]PIB. In MCI patients, CSF levels of Abeta42, pTAU, totalTAU and the Abeta42/pTAU ratio were measured. RESULTS: In MCI patients, CSF Abeta42 was abnormal in 53% of patients, totalTAU in 67%, pTAU in 64% and the Abeta42/pTAU ratio in 64%. A composite neocortical [(11)C]PIB uptake score was increased in 87% of the MCI patients. Only 54% of [(11)C]PIB-positive subjects showed AD-type Abeta42 values. During a 2-year follow-up, 6 MCI patients converted to AD, all of them had increased neocortical PIB scores at the MCI stage. Abnormal CSF Abeta42 was found in 3 patients, pTAU in 3 patients and Abeta42/pTAU ratio in 4 patients. CONCLUSION: Follow-up studies are needed to confirm whether [(11)C]PIB uptake might be more sensitive than CSF Abeta42 concentration in detecting increased amyloid burden in MCI, as suggested by the results of this study.

Reduction in water and metabolite apparent diffusion coefficients during energy failure involves cation-dependent mechanisms. A proton nuclear magnetic resonance study of rat cortical brain slices.

Proton (1H) nuclear magnetic resonance (NMR) diffusion spectroscopy was used to assess apparent diffusion coefficients (ADCs) in rat brain slices. Aglycemic hypoxia caused reductions in the ADC of N-acetylaspartate (NAA) (0.15 to 0.09 x 10(-3) mm2/s) and "slow" diffusion coefficient (D2) of tissue water (0.51 to 0.37 x 10(-3) mm2/s), together with a 32+/-11% increase in tissue water volume, attributable to tissue swelling. The ADC and D2 reductions were diminished, however, by removing external Ca2+, and under 10 mmol/L Mg2+, normoxic diffusion coefficients persisted until 40 minutes of hypoxia. The data suggest that the shift of water into the intracellular space alone cannot satisfactorily explain the reduced cerebral diffusion upon energy failure and that external Mg2+ and Ca2+ play crucial modulatory roles.

Alpha 1-antichymotrypsin and IL-1 beta are not increased in CSF or serum in Alzheimer's disease.

Chronic inflammation associated with the amyloid plaques may represent an acute phase response in the brain. We quantitated the levels of two inflammatory markers; alpha 1-antichymotrypsin (alpha 1-ACT) and interleukin 1 beta (IL-1 beta) in paired serum and cerebrospinal fluid (CSF) samples from 40 patients with Alzheimer's disease (AD), 20 patients with Parkinson's disease (PD), and 42 age-matched controls. No differences in serum or CSF levels of either alpha 1-ACT or IL-1 beta were found between the groups. However, some AD patients had increased alpha 1-ACT index, suggesting an intrathecal production of alpha 1-ACT. Although alpha 1-ACT or IL-1 beta might be involved in the pathogenesis of AD, our results show that their measurement in serum or CSF is not valuable to support the clinical diagnosis of AD.

Relationship between apoE genotype and CSF beta-amyloid (1-42) and tau in patients with probable and definite Alzheimer's disease.

We investigated the usefulness of cerebrospinal fluid (CSF) beta-amyloid42 (Abeta42), beta-amyloid40 (Abeta40) and tau analyses in the diagnosis of Alzheimer's disease (AD). The study included 41 definite AD cases, 80 patients with probable AD. 27 with other dementias and 39 neurological controls. Abeta42, Abeta340 and tau protein concentrations in CSF were measured of using ELISA assays. Abeta42 levels were decreased and tau increased in AD. Combination of Abeta42 and tau resulted a sensitivity of 50.4% for AD and specificities of 94.8% for controls and 85.2% for other dementias. Ninety-one percent of the patients with Abeta42 below the cutoff value (340 pg/ml) and tau above the cutoff value (380 pg/ml) had AD. AD patients carrying apoE epsilon4 allele had lower Abeta42 (P < 0.005) and higher tau (P < 0.05) levels than those without an E4 allele, and 18 (81.8%) of the 22 AD patients who had normal Abeta42 and tau levels were apoE e4 allele non-carriers. Low Abeta42 and high tau concentration in CSF strongly support the diagnosis of AD. Measurement of Abeta42 may help the early diagnosis of cases at risk for AD such as apoE E4 allele carriers.

Neuronal loss and beta-amyloid removal in the amygdala of people with Down syndrome.

The decrease in the number of neurons free of neurofibrillary changes, neurons with neurofibrillary degeneration, and the total volume of beta-amyloid (A beta) deposits in the amygdala of people with Down syndrome and in late stages of Alzheimer disease were estimated by using morphometry and regression analysis. This model predicts that the duration of neurofibrillary changes from the pretangle stage to ghost tangles is approximately 4.7 years. The correlation between the decrease in the number of neurons and the decrease in the amount of A beta indicates that amyloid deposition is associated with neurons and that loss of neurons causes decrease in A beta deposition. The presence of neurons only with neurofibrillary tangles, and the absence of the amyloid deposits predicted by regression analysis suggest that neurons with tangles are not engaged in amyloid deposition. The disappearance of amyloid by approximately 2.2 years after loss of neurons free of neurofibrillary changes indicates that A beta deposits are degradable and removable and that even in severely atrophic amygdala, there are mechanisms of amyloid resolution. This study shows that in normal aging in the amygdala, extracellular A beta appears later than neurofibrillary changes.

Apolipoprotein E genotype and amyloid load in Alzheimer disease and control brains.

We investigated the effect of apolipoprotein E (apoE) genotype on amyloid load in the frontal and cerebellar cortices of 24 patients with definite Alzheimer disease (AD) and 19 controls. Amyloid load was examined by using two methods: 1) acid-extractable amyloid beta-protein (A beta) and insoluble A beta levels of frontal and cerebellar cortices were measured by using enzyme-linked immunosorbent assay, and 2) all types of amyloid plaques and neurofibrillary tangles (NFT) in the frontal cortices were counted after silver staining. Acid-extractable A beta and insoluble A beta levels were higher in AD brains than controls, although there was an overlap between the groups. Acid-extractable A beta and insoluble A beta levels were higher from AD and controls with the apoE epsilon 4 alleles than those without such alleles. However, the differences did not reach statistical significance in AD group. There was no correlation between acid-extractable A beta or insoluble A3 levels and the number of amyloid plaques in AD and control brains. However, insoluble A beta levels correlated positively with the number of NFT in AD brains. Our results show that although apoE epsilon 4 influences the accumulation of A beta, multiple processes may be involved in deposition of A beta in the brain.

Plasma and cerebrospinal fluid levels of amyloid beta proteins 1-40 and 1-42 in Alzheimer disease.

BACKGROUND: In brains with AD, Abeta is a major component of diffuse plaques. Previous reports showed that CSF Abeta42 levels were lower in patients with AD than in controls. Although studies showed higher plasma Abeta42 levels in familial AD, a recent report has indicated that plasma Abeta42 levels were similar in a sporadic AD group and controls. However, no information is published on plasma Abeta40 and Abeta42 levels in relation to Apo E genotype or severity of dementia in sporadic AD. OBJECTIVE: To examine plasma and cerebrospinal fluid (CSF) levels of amyloid beta protein 1-40 (Abeta40) and 1-42 (Abeta42) levels in patients with probable Alzheimer disease (AD) and elderly nondemented control subjects in relation to the apolipoprotein E (Apo E) genotype and dementia severity. SETTING: Two university medical centers. PATIENTS AND METHODS: Levels of Abeta40 and Abeta42 were measured in plasma from 78 patients with AD and 61 controls and in CSF from 36 patients with AD and 29 controls by means of a sandwich enzyme-linked immunosorbent assay. RESULTS: Mean plasma Abeta40 levels were higher in the AD group than in controls (P = .005), but there was substantial overlap; Abeta42 levels were similar between the groups. Levels of Abeta40 and Abeta42 showed no association with sex or Mini-Mental State Examination scores. There was a significant relationship between age and Abeta40 level in controls but not in the AD group. Levels of Abeta40 were higher in patients with AD with the Apo E epsilon4 allele than in controls (P<.01). Cerebrospinal fluid Abeta40 levels were similar in the AD group and controls. However, Abeta42 levels were lower in the AD group than in controls (P<.001). The levels showed no association with severity of dementia. CONCLUSIONS: Although mean plasma Abeta40 levels are elevated in sporadic AD and influenced by Apo E genotype, measurement of plasma Abeta40 levels is not useful to support the clinical diagnosis of AD. Lower levels of CSF Abeta42 in the AD group are consistent with previous studies.

Cerebrospinal fluid concentrations of soluble amyloid beta-protein and apolipoprotein E in patients with Alzheimer's disease: correlations with amyloid load in the brain.

OBJECTIVE: To compare soluble amyloid beta-protein and apolipoprotein E levels in cerebrospinal fluid (CSF) and brain extracts from patients with definite Alzheimer's disease. SETTING: University medical center. PATIENTS: Nineteen patients with definite Alzheimer's disease. MAIN OUTCOME MEASURES: Soluble amyloid beta-protein and apolipoprotein E levels in CSF, in neutral and low-pH brain extracts, and in formic acid-treated sections of the frontal, temporal, and cerebellar cortices, measured using enzyme-linked immunosorbent assay. RESULTS: Soluble amyloid beta-protein and apolipoprotein E levels in CSF were significantly lower in patients with congophilic angiopathy than in those without angiopathy. The levels did not correlate with the number of amyloid plaques in the neocortex. There was, however, a tendency toward an inverse correlation between the amount of amyloid beta-protein in the frontal cortex extracts and the soluble amyloid beta-protein level in CSF. CONCLUSION: Soluble amyloid beta-protein levels in CSF may reflect amyloid accumulation in brain blood vessels.

Celiac disease, brain atrophy, and dementia.

We report 5 patients who developed dementia before age 60 and were subsequently found to have celiac disease (CD). Intellectual deterioration ranged from moderate to severe, and diffuse cerebral or cerebellar atrophy was found on brain CT. Diagnosis of CD was confirmed by findings of subtotal villous atrophy in jejunal biopsy specimens and positive serum reticulin and gliadin antibodies. Conspicuously, gastrointestinal symptoms were mild. The gluten-free diet failed to improve the neurologic disability except in 1 patient. CD is a multisystem disorder and may play a role in some cases of presenile dementia. Although the pathogenetic mechanisms are obscure, immunologic mechanisms are implicated.

Recovery of intracellular pH in cortical brain slices following anoxia studied by nuclear magnetic resonance spectroscopy: role of lactate removal, extracellular sodium and sodium/hydrogen exchange.

[31P]- and [1H]nuclear magnetic resonances recorded in an interleaved fashion were used in order to quantify high-energy phosphates, intracellular pH and lactate in cortical brain slices of the guinea-pig superfused in a CO2/HCO3(-)-buffered medium during and after anoxic insults. The volume-averaged intracellular pH and energy status of the preparation following anoxia were determined. In the presence of external Na+, intracellular pH normalized in 3 min and was significantly more alkaline from 10 to 12 min of recovery, but lactate remained elevated for 12 min of reoxygenation following anoxia. The amount of lactate removed was only 40% of the quantity of acid extruded showing operation of H+ neutralizing transmembrane mechanisms other than transport of lactic acid. Amiloride (1 or 2 mM) did not prevent the recovery of intracellular pH, but it blocked the "overshoot" of the alkalinization at 10-12 min of recovery. In a medium containing 70 mM K+, 60 mM Na+ and 0.1 mM Ca2+, the recovery of pH, but not lactate washout, was significantly delayed. Removal of external Na+ caused severe energetic failure, decreases both in oxygen uptake and in N-acetyl aspartate concentration, indicating loss of viable tissue. In Na(+)-free superfusion, lactic acidosis caused a more severe drop in intracellular pH than in the presence of Na+. Complexing of extracellular Ca2+ in the Na(+)-free medium inhibited the acidification by 0.38 pH units during anoxia which is as much as the acidification caused by lactate accumulation in the absence of Na+. In Na(+)-free medium intracellular pH recovered, however, from an anoxic level to a normoxic value in 6 min. Metabolic damage of the slice preparation induced by anoxia in the absence of Na+ was as profound in the presence as in the absence of Ca2+ showing that accumulation of Ca2+ is not the only reason for the damage. It is concluded that recovery of intracellular pH from lactic-acidosis can occur independently of energetic recovery and involves acid extrusion mechanism(s) that is(are) dependent on external Na+ and sensitive to high K+.