Choroid Plexus Calcification Correlates with Cortical Microglial Activation in Humans: A Multimodal PET, CT, MRI Study.

BACKGROUND AND PURPOSE: The choroid plexus (CP) within the brain ventricles is well-known to produce cerebrospinal fluid (CSF). Recently, the CP has been recognized as critical in modulating inflammation. MRI-measured CP enlargement has been reported in neuroinflammatory disorders like MS as well as with aging and neurodegeneration. The basis of MRI-measured CP enlargement is unknown. On the basis of tissue studies demonstrating CP calcification as a common pathology associated with aging and disease, we hypothesized that previously unmeasured CP calcification contributes to MRI-measured CP volume and may be more specifically associated with neuroinflammation. MATERIALS AND METHODS: We analyzed 60 subjects (43 healthy controls and 17 subjects with Parkinson's disease) who underwent PET/CT using 11C-PK11195, a radiotracer sensitive to the translocator protein expressed by activated microglia. Cortical inflammation was quantified as nondisplaceable binding potential. Choroid plexus calcium was measured via manual tracing on low-dose CT acquired with PET and automatically using a new CT/MRI method. Linear regression assessed the contribution of choroid plexus calcium, age, diagnosis, sex, overall volume of the choroid plexus, and ventricle volume to cortical inflammation. RESULTS: Fully automated choroid plexus calcium quantification was accurate (intraclass correlation coefficient with manual tracing = .98). Subject age and choroid plexus calcium were the only significant predictors of neuroinflammation. CONCLUSIONS: Choroid plexus calcification can be accurately and automatically quantified using low-dose CT and MRI. Choroid plexus calcification-but not choroid plexus volume-predicted cortical inflammation. Previously unmeasured choroid plexus calcium may explain recent reports of choroid plexus enlargement in human inflammatory and other diseases. Choroid plexus calcification may be a specific and relatively easily acquired biomarker for neuroinflammation and choroid plexus pathology in humans.

Anterior cingulate cortex γ-aminobutyric acid deficits in youth with depression.

Abnormally low γ-aminobutyric acid (GABA) levels have been consistently reported in adults with major depressive disorder (MDD). Our group extended this finding to adolescents, and documented that GABA deficits were associated with anhedonia. Here we aimed to confirm our prior finding of decreased brain GABA in youth with depression and explore its associations with clinical variables. Forty-four psychotropic medication-free youth with MDD and 36 healthy control (HC) participants (12-21 years) were studied. Participants represent a combined sample of 39 newly recruited youth (MDD=24) and 41 youth from our previously reported study (MDD=20). GABA levels and the combined resonances of glutamate and glutamine (Glx) were measured in vivo in the anterior cingulate cortex using proton magnetic resonance spectroscopy. Youth with depression exhibited significantly lower GABA levels than HC in both the newly reported (P=0.003) and the combined (P=0.003) samples. When depressed participants were classified based on the presence of anhedonia, only the anhedonic MDD subgroup showed reduced GABA levels compared to HC (P=0.002). While there were no associations between any clinical measures and GABA or Glx levels in the new sample, GABA was negatively correlated with only anhedonia severity in the combined MDD group. Furthermore, in the combined sample, hierarchical regression models showed that anhedonia, but not depression severity, anxiety or suicidality, contributed significant variance in GABA levels. This report solidifies the evidence for a GABA deficit early in the course of MDD, which correlates specifically with anhedonia in the disorder.

Relationships among Cortical Glutathione Levels, Brain Amyloidosis, and Memory in Healthy Older Adults Investigated In Vivo with 1H-MRS and Pittsburgh Compound-B PET.

BACKGROUND AND PURPOSE: Oxidative stress has been implicated as an important pathologic mechanism in the development of Alzheimer disease. The purpose of this study was to assess whether glutathione levels, detected noninvasively with proton MR spectroscopy, are associated with brain amyloidosis and memory in a community-dwelling cohort of healthy older adults. MATERIALS AND METHODS: Fifteen cognitively healthy subjects were prospectively enrolled in this study. All subjects underwent 1H-MR spectroscopy of glutathione, a positron-emission tomography scan with an amyloid tracer, and neuropsychological testing by using the Repeatable Battery for the Assessment of Neuropsychological Status. Associations among glutathione levels, brain amyloidosis, and memory were assessed by using multivariate regression models. RESULTS: Lower glutathione levels were associated with greater brain amyloidosis in the temporal (P = .03) and parietal (P = .05) regions, adjusted for apolipoprotein E ε4 carrier status. There were no significant associations between glutathione levels and cognitive scores. CONCLUSIONS: This study found an association between cortical glutathione levels and brain amyloidosis in healthy older adults, suggesting a potential role for 1H-MR spectroscopy measures of glutathione as a noninvasive biomarker of early Alzheimer disease pathogenesis.

Increased ventricular cerebrospinal fluid lactate in depressed adolescents.

BACKGROUND: Mitochondrial dysfunction has been increasingly examined as a potential pathogenic event in psychiatric disorders, although its role early in the course of major depressive disorder (MDD) is unclear. Therefore, the purpose of this study was to investigate mitochondrial dysfunction in medication-free adolescents with MDD through in vivo measurements of neurometabolites using high-spatial resolution multislice/multivoxel proton magnetic resonance spectroscopy. METHODS: Twenty-three adolescents with MDD and 29 healthy controls, ages 12-20, were scanned at 3T and concentrations of ventricular cerebrospinal fluid lactate, as well as N-acetyl-aspartate (NAA), total creatine (tCr), and total choline (tCho) in the bilateral caudate, putamen, and thalamus were reported. RESULTS: Adolescents with MDD exhibited increased ventricular lactate compared to healthy controls [F(1,41)=6.98, P=0.01]. However, there were no group differences in the other neurometabolites. Dimensional analyses in the depressed group showed no relation between any of the neurometabolites and symptomatology, including anhedonia and fatigue. CONCLUSIONS: Increased ventricular lactate in depressed adolescents suggests mitochondrial dysfunction may be present early in the course of MDD; however it is still not known whether the presence of mitochondrial dysfunction is a trait vulnerability of individuals predisposed to psychopathology or a state feature of the disorder. Therefore, there is a need for larger multimodal studies to clarify these chemical findings in the context of network function.

A pilot in vivo proton magnetic resonance spectroscopy study of amino acid neurotransmitter response to ketamine treatment of major depressive disorder.

The N-methyl-D-aspartate receptor antagonist ketamine can improve major depressive disorder (MDD) within hours. To evaluate the putative role of glutamatergic and GABAergic systems in ketamine's antidepressant action, medial prefrontal cortical (mPFC) levels of glutamate+glutamine (Glx) and γ-aminobutyric acid (GABA) were measured before, during, and after ketamine administration using proton magnetic resonance spectroscopy. Ketamine (0.5 mg kg(-1) intravenously) was administered to 11 depressed patients with MDD. Glx and GABA mPFC responses were measured as ratios relative to unsuppressed voxel tissue water (W) successfully in 8/11 patients. Ten of 11 patients remitted (50% reduction in 24-item Hamilton Depression Rating Scale and total score ⩽10) within 230 min of commencing ketamine. mPFC Glx/W and GABA/W peaked at 37.8%±7.5% and 38.0%±9.1% above baseline in ~26 min. Mean areas under the curve for Glx/W (P=0.025) and GABA/W (P=0.005) increased and correlated (r=0.796; P=0.018). Clinical improvement correlated with 90-min norketamine concentration (df=6, r=-0.78, P=0.023), but no other measures.

Motor cortex glutathione deficit in ALS measured in vivo with the J-editing technique.

This study compared in vivo levels of the antioxidant glutathione (GSH) in the motor cortex of 11 ALS patients with those in 11 age-matched healthy volunteers (HV). Using the standard J-edited spin-echo difference MRS technique, GSH spectra were recorded on a 3.0 T GE MR system from a single precentral gyrus voxel. GSH levels expressed as ratios to the unsuppressed voxel tissue water (W) were 31% lower in ALS patients than in HV (p=.005), and 36% lower in ALS than in HV (p=.02) when expressed as ratios to the total creatine peak (tCr), supporting a role for oxidative stress in ALS. Levels of the putative neuronal marker N-acetylaspartate (NAA) relative to W did not differ between ALS and HV (p=.26), but were lower by 9% in ALS than in HV (p=.013) when expressed as ratios relative to tCr. This discrepancy is attributed to small but opposite changes in NAA and tCr in ALS that, as a ratio, resulted in a statistically significant group difference, further suggesting caution in using tCr as an internal reference under pathological conditions.

Assessment of disease severity in late infantile neuronal ceroid lipofuscinosis using multiparametric MR imaging.

BACKGROUND AND PURPOSE: LINCL is a uniformly fatal lysosomal storage disease resulting from mutations in the CLN2 gene that encodes for tripeptidyl peptidase 1, a lysosomal enzyme necessary for the degradation of products of cellular metabolism. With the goal of developing quantitative noninvasive imaging biomarkers sensitive to disease progression, we evaluated a 5-component MR imaging metric and tested its correlation with a clinically derived disease-severity score. MATERIALS AND METHODS: MR imaging parameters were measured across the brain, including quantitative measures of the ADC, FA, nuclear spin-spin relaxation times (T2), volume percentage of CSF (%CSF), and NAA/Cr ratios. Thirty MR imaging datasets were prospectively acquired from 23 subjects with LINCL (2.5-8.4 years of age; 8 male/15 female). Whole-brain histograms were created, and the mode and mean values of the histograms were used to characterize disease severity. RESULTS: Correlation of single MR imaging parameters against the clinical disease-severity scale yielded linear regressions with R2 ranging from 0.25 to 0.70. Combinations of the 5 biomarkers were evaluated by using PCA. The best combination included ADC, %CSF, and NAA/Cr (R2=0.76, P<.001). CONCLUSIONS: The multiparametric disease-severity score obtained from the combination of ADC, %CSF, and NAA/Cr whole-brain MR imaging techniques provided a robust measure of disease severity, which may be useful in clinical therapeutic trials of LINCL in which an objective assessment of therapeutic response is desired.

Metabolic response of glioblastoma to superselective intra-arterial cerebral infusion of bevacizumab: a proton MR spectroscopic imaging study.

BACKGROUND AND PURPOSE: SIACI of bevacizumab has emerged as a promising novel therapy in the treatment of recurrent GB. This study assessed the potential of (1)H-MRS as an adjunctive technique in detecting metabolic changes reflective of antiproliferative effects of targeted infusion of bevacizumab in the treatment of GB. MATERIALS AND METHODS: Eighteen patients enrolled in a phase I/II study of SIACI of bevacizumab for treatment of recurrent GB were included. Concurrent MR imaging and (1)H-MRS scans were performed before and after treatment. Five distinct morphologic ROIs were evaluated for structural and metabolic changes on MR imaging and (1)H-MRS, which included enhancing, nonenhancing T2 hyperintense signal abnormality, and multiple control regions. Pre- and post-SIACI of bevacizumab peak areas for NAA, tCho, tCr, as well as tCho/tCr and tCho/NAA ratios, were derived for all 5 ROIs and compared using the Wilcoxon signed-rank test. RESULTS: A significant median decrease of 25.99% (range -55.76 to 123.94; P = .006) in tCho/NAA was found post-SIACI of bevacizumab relative to pretreatment values in regions of enhancing disease. A trend-level significant median decrease of 6.45% (range -23.71 to 37.67; P = .06) was noted in tCho/NAA posttreatment in regions of nonenhancing T2-hyperintense signal abnormality. CONCLUSIONS: The results of this (1)H-MRS analysis suggest that GB treatment with SIACI of bevacizumab may be associated with a direct antiproliferative effect, as demonstrated by significant reductions of tCho/NAA after the intervention.

Natural history of MELAS associated with mitochondrial DNA m.3243A>G genotype.

OBJECTIVE: To describe the natural history of clinical and laboratory features associated with the m.3243A>G mitochondrial DNA point mutation. Natural history data are needed to obtain prognostic information and for clinical trial planning. METHODS: We included 85 matrilineal relatives from 35 families with at least 2 visits in this prospective cohort study. Thirty-one were fully symptomatic with mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS), and 54 were carrier relatives. Evaluations included standardized questionnaires (medical history and daily living functioning), physical examination, neuropsychological testing, and a battery of imaging and laboratory tests. We evaluated changes in clinical and laboratory features over time and survival. Outcomes are reported over a follow-up period of up to 10.6 years (mean 3.8 ± 2.2 years for patients and 5.5 ± 3.0 for carrier relatives). RESULTS: Neurologic examination, neuropsychological testing, and daily living scores significantly declined in all patients with MELAS, whereas no significant deterioration occurred in carrier relatives. Cerebral MRI scores declined significantly in patients with MELAS. Magnetic resonance spectroscopy estimates of lactate in the lateral ventricles increased over time, and high lactate was associated with increased mortality. Symptom onset in childhood often was associated with worse outcome. Patients with MELAS had a greater death rate than carrier relatives. CONCLUSIONS: Patients with MELAS carrying the m.3243A>G mutation show a measurable decline in clinical and imaging outcomes. It is hoped that these data will be helpful in anticipating the disease course and in planning clinical trials for MELAS.

A pilot study of the effects of chronic paroxetine administration on hippocampal N-acetylaspartate in generalized anxiety disorder.

The neural basis of generalized anxiety disorder (GAD) is poorly characterized. The effect of chronic administration (12 weeks) of paroxetine, a selective serotonin reuptake inhibitor, on N-acetylaspartate (NAA), a marker of neuronal viability, was evaluated in adults with GAD using proton magnetic resonance spectroscopic imaging ((1)H MRSI) at 1.5 T. We hypothesized that, pretreatment abnormalities in hippocampal NAA/creatine (NAA/Cr) would normalize with symptomatic improvement. Nine GAD patients (mean age = 41.7 year; 4 females) received 12 weeks of open-label paroxetine treatment, flexibly dosed up to 60 mg/day. Clinical outcome was assessed with the Hamilton Anxiety Rating Scale (HAM-A). Multislice ( 1)H MRSI scans were performed at unmedicated baseline and following 6 and 12 weeks of treatment. Ten untreated healthy volunteers (HVs) (mean age = 37.1 year; 4 females) received scans at the same intervals. All patients achieved remission (HAM-A

Neurological deterioration in late infantile neuronal ceroid lipofuscinosis.

BACKGROUND: Late infantile neuronal ceroid lipofuscinosis (LINCL) is associated with progressive degeneration of the brain and retina starting in early childhood. METHODS: Thirty-two individual neurologic, ophthalmologic, and CNS imaging (MRI and MRS) assessments of 18 children with LINCL were analyzed. Disease severity was followed by two rating scales, one previously established but modified to solely assess the brain and exclude the retinal disease (modified Hamburg LINCL scale), and a newly developed scale, with expanded evaluation of the CNS impairment (Weill Cornell LINCL scale). RESULTS: For the 18 children, the Weill Cornell scale yielded a closer correlation with both age and time since initial clinical manifestation of the disease than did the modified Hamburg scale. There were no significant differences as a function of age or time since initial manifestation of the disease in the rating scales among the most frequent CLN2 mutations (G3556C, 56% of all alleles or C3670T, 22% of all alleles). Measurements of cortical MRS N-acetyl-aspartate content, MRI ventricular, gray matter and white matter volume, and cortical apparent diffusion coefficient correlated to a variable degree with the age of the children and the time since initial clinical manifestation of the disease. All imaging measurements correlated better with the Weill Cornell CNS scale compared to the modified Hamburg LINCL scale. CONCLUSION: The data suggest that the Weill Cornell late infantile neuronal ceroid lipofuscinosis (LINCL) scale, together with several of the MRI measurements, may be useful in the assessment of severity and progression of LINCL and for the evaluation of novel therapeutic strategies.

Assessing disease severity in late infantile neuronal ceroid lipofuscinosis using quantitative MR diffusion-weighted imaging.

BACKGROUND AND PURPOSE: Late infantile neuronal ceroid lipofuscinosis (LINCL), a form of Batten disease, is a fatal neurodegenerative genetic disorder, diagnosed via DNA testing, that affects approximately 200 children in the United States at any one time. This study was conducted to evaluate whether quantitative data derived by diffusion-weighted MR imaging (DWI) techniques can supplement clinical disability scale information to provide a quantitative estimate of neurodegeneration, as well as disease progression and severity. MATERIALS AND METHODS: This study prospectively analyzed 32 DWI examinations from 18 patients having confirmed LINCL at various stages of disease. A whole-brain apparent diffusion coefficient (ADC) histogram was fitted with a dual Gaussian function combined with a function designed to model voxels containing a partial volume fraction of brain parenchyma versus CSF. Previously published whole-brain ADC values of age-matched control subjects were compared with those of the LINCL patients. Correlations were tested between the peak ADC of the fitted histogram and patient age, disease severity, and a CNS disability scale adapted for LINCL. RESULTS: ADC values assigned to brain parenchyma were higher than published ADC values for age-matched control subjects. ADC values between patients and control subjects began to differ at 5 years of age based on 95% confidence intervals. ADC values had a nearly equal correlation with patient age (R2=0.71) and disease duration (R2=0.68), whereas the correlation with the central nervous system disability scale (R2=0.27) was much weaker. CONCLUSION: This study indicates that brain ADC values acquired using DWI may be used as an independent measure of disease severity and duration in LINCL.

Quantitative objective markers for upper and lower motor neuron dysfunction in ALS.

OBJECTIVE: To investigate the value of objective biomarkers for upper (UMN) and lower (LMN) motor neuron involvement in ALS. METHODS: We prospectively studied 64 patients with ALS and its subsets using clinical measures, proton MR spectroscopic imaging ((1)H MRSI), diffusion tensor imaging, transcranial magnetic stimulation, and the motor unit number estimation (MUNE) at baseline and every 3 months for 15 months and compared them with control subjects. RESULTS: (1)H MRSI measures of the primary motor cortex N-acetyl-aspartate (NAA) concentration were markedly reduced in ALS (p = 0.009) and all UMN syndromes combined (ALS, familial ALS [fALS], and primary lateral sclerosis; p = 0.03) vs control values. Central motor conduction time to the tibialis anterior was prolonged in ALS (p < 0.0005) and combined UMN syndromes (p = 0.001). MUNE was lower in ALS (p < 0.0005) and all LMN syndromes combined (ALS, fALS, and progressive muscular atrophy; p = 0.001) vs controls. All objective markers correlated well with the ALS Functional Rating Scale-Revised, finger and foot tapping, and strength testing, suggesting these markers related to disease activity. Regarding changes over time, MUNE changed rapidly, whereas neuroimaging markers changed more slowly and did not significantly differ from baseline. CONCLUSIONS: (1)H MR spectroscopic imaging measures of the primary motor cortex N-acetyl-aspartate (NAA) concentration and ratio of NAA to creatine, central motor conduction time to the tibialis anterior, and motor unit number estimation significantly differed between ALS, its subsets, and control subjects, suggesting they have potential to provide insight into the pathobiology of these disorders.

Dichloroacetate causes toxic neuropathy in MELAS: a randomized, controlled clinical trial.

OBJECTIVE: To evaluate the efficacy of dichloroacetate (DCA) in the treatment of mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). BACKGROUND: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, correlate with more severe neurologic impairment. The authors hypothesized that chronic cerebral lactic acidosis exacerbates neuronal injury in MELAS and therefore, investigated DCA, a potent lactate-lowering agent, as potential treatment for MELAS. METHODS: The authors conducted a double-blind, placebo-controlled, randomized, 3-year cross-over trial of DCA (25 mg/kg/day) in 30 patients (aged 10 to 60 years) with MELAS and the A3243G mutation. Primary outcome measure was a Global Assessment of Treatment Efficacy (GATE) score based on a health-related event inventory, and on neurologic, neuropsychological, and daily living functioning. Biologic outcome measures included venous, CSF, and 1H MRSI-estimated brain lactate. Blood tests and nerve conduction studies were performed to monitor safety. RESULTS: During the initial 24-month treatment period, 15 of 15 patients randomized to DCA were taken off study medication, compared to 4 of 15 patients randomized to placebo. Study medication was discontinued in 17 of 19 patients because of onset or worsening of peripheral neuropathy. The clinical trial was terminated early because of peripheral nerve toxicity. The mean GATE score was not significantly different between treatment arms. CONCLUSION: DCA at 25 mg/kg/day is associated with peripheral nerve toxicity resulting in a high rate of medication discontinuation and early study termination. Under these experimental conditions, the authors were unable to detect any beneficial effect. The findings show that DCA-associated neuropathy overshadows the assessment of any potential benefit in MELAS.

Objective tests for upper motor neuron involvement in amyotrophic lateral sclerosis (ALS).

OBJECTIVE: To develop objective markers for upper motor neuron (UMN) involvement in ALS, the value of single-voxel MR spectroscopy (MRS) and transcranial magnetic stimulation (TMS) was studied. METHODS: Test results of 164 ALS patients who had MRS only (n = 91), TMS only (n = 13), or both (n = 60) were analyzed; also, 11 autopsy examinations were evaluated. RESULTS: Abnormal test results consistent with UMN involvement were found in 134 patients with clinical UMN signs: 86% on MRS, 77% on TMS, and 70% on MRS and TMS together. Among 30 patients with solely LMN signs (progressive muscular atrophy), UMN results were found in 63% on MRS, 63% on TMS, and 46% on both tests together. There was a significant association of the degree of abnormal N-acetyl aspartate/creatine ratios with UMN signs (p = 0.01). The sensitivity to detect UMN involvement was 0.86 for MRS (specificity 0.37) and 0.77 for TMS (specificity 0.38). At autopsy, all 11 patients had pathologic UMN abnormalities, including 4 with normal MRS and 1 with normal TMS in life. CONCLUSIONS: MRS is highly sensitive, somewhat more than TMS, and shows good correlation with clinical UMN signs. Combining MRS and TMS results in the same patient with further refinement may help in the early diagnosis of ALS.

Cerebral lactic acidosis correlates with neurological impairment in MELAS.

OBJECTIVE: To evaluate the role of chronic cerebral lactic acidosis in mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). METHODS: The authors studied 91 individuals from 34 families with MELAS and the A3243G point mutation and 15 individuals from two families with myoclonus epilepsy and ragged red fibers (MERRF) and the A8344G mutation. Subjects were divided into four groups. Paternal relatives were studied as controls (Group 1). The maternally related subjects were divided clinically into three groups: asymptomatic (no clinical evidence of neurologic disease) (Group 2), oligosymptomatic (neurologic symptoms but without the full clinical picture of MELAS or MERRF) (Group 3), and symptomatic (fulfilling MELAS or MERRF criteria) (Group 4). The authors performed a standardized neurologic examination, neuropsychological testing, MRS, and leukocyte DNA analysis in all subjects. RESULTS: The symptomatic and oligosymptomatic MELAS subjects had significantly higher ventricular lactate than the other groups. There was a significant correlation between degree of neuropsychological and neurologic impairment and cerebral lactic acidosis as estimated by ventricular MRS lactate levels. CONCLUSIONS: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, are associated with more severe neurologic impairment.

Measurement of intramyocellular lipid levels with 2-D magnetic resonance spectroscopic imaging at 1.5 T.

Intramyocellular lipid (IMCL) plays an important role in the study of metabolism in vivo. Magnetic resonance spectroscopy (MRS) studies of IMCL are usually performed with clinical 1.5-T magnetic resonance imaging (MRI) systems and have employed the single-voxel MRS technique. The present study reports the results of our systematic evaluation of the ability of single- and multi-voxel MRS to yield high-quality, contamination-free IMCL levels from the tibialis anterior (TA) muscle. A clinical, 1.5-T, whole-body MRI scanner was used to measure IMCL with a standard knee coil, head coil, or a 3-cm receive-only surface coil with a body coil transmit. Excellent IMCL spectra were obtained in healthy males in only 8 min from multiple 0.25-cm(3) voxels using the surface coil receive/body coil transmit in conjunction with the standard PRESS spectroscopic imaging (SI) technique. This approach provided the spatial resolution and voxel placement flexibility permitting optimal separation of IMCL and extramyocellular lipid. Our findings demonstrate the potential of the SI approach.

In vitro activities of ertapenem (MK-0826) against recent clinical bacteria collected in Europe and Australia.

Ertapenem (MK-0826, L-749,345) is a 1-beta-methyl carbapenem with a long serum half-life. Its in vitro activity was determined by broth microdilution against 3,478 bacteria from 12 centers in Europe and Australia, with imipenem, cefepime, ceftriaxone, and piperacillin-tazobactam used as comparators. Ertapenem was the most active agent tested against members of the family Enterobacteriaceae, with MICs at which 90% of isolates are inhibited (MIC(90)s) of < or =1 microg/ml for all species. Ertapenem also was more active than imipenem against fastidious gram-negative bacteria and Moraxella spp.; on the other hand, ertapenem was slightly less active than imipenem against streptococci, methicillin-susceptible staphylococci, and anaerobes, but its MIC(90)s for these groups remained < or =0.5 microg/ml. Acinetobacter spp. and Pseudomonas aeruginosa were also much less susceptible to ertapenem than imipenem, and most Enterococcus faecalis strains were resistant. Ertapenem resistance, based on a provisional NCCLS MIC breakpoint of > or =16 microg/ml, was seen in only 3 of 1,611 strains of the family Enterobacteriaceae tested, all of them Enterobacter aerogenes. Resistance was also seen in 2 of 135 anaerobes, comprising 1 Bacteroides fragilis strain and 1 Clostridium difficile strain. Ertapenem breakpoints for streptococci have not been established, but an unofficial susceptibility breakpoint of < or =2 microg/ml was adopted for clinical trials to generate corresponding clinical response data for isolates for which MICs were as high as 2 microg/ml. Of 234 Streptococcus pneumoniae strains tested, 2 required ertapenem MICs of 2 microg/ml and one required an MIC of 4 microg/ml, among 67 non-Streptococcus pyogenes, non-Streptococcus pneumoniae streptococci, single isolates required ertapenem MICs of 2 and 16 microg/ml. These streptococci also had diminished susceptibilities to other beta-lactams, including imipenem as well as ertapenem. The Etest and disk diffusion gave susceptibility test results in good agreement with those of the broth microdilution method for ertapenem.

Metabolism of alternative substrates and the bioenergetic status of EMT6 tumor cell spheroids.

In order to evaluate the ability of EMT6/Ro multicellular spheroids to utilize various pathways of energy production, (13)C and (31)P MRS have been employed to monitor the metabolism of glucose, glutamine, acetate and propionate. EMT6/Ro spheroids perfused with culture medium containing 5.5 mM glucose maintain stable levels of nucleotide triphosphates (NTP) and phosphocreatine (PCr) for up to 48 h, even in the absence of glutamine. The metabolism of 1-(13)C-glucose was almost entirely to 3-(13)C-lactate (88 +/- 12%, n = 7), even though the perfusion medium was equilibrated with 95% O(2). Labeling was also observed in other glycolytic metabolites, primarily alanine and alpha-glycerolphosphate. A low level of (13)C labeling in glutamate, indicative of mitochondrial oxidative metabolism (TCA cycle), was consistently detected when spheroids were perfused with 1-(13)C-glucose, almost exclusively in the C4 position of glutamate. Labeling of glutamate C2 and C3 was always less than 20% of the labeling in C4 and was usually undetectable. No evidence of adjacent carbon labeling in individual glutamate molecules (indicative of multiple cycles of label incorporation) was found, even in high-resolution (13)C NMR spectra of extracts from cells or spheroids. Despite the predominantly glycolytic metabolism of glucose, the mitochondrial substrate glutamine (2 mM, in the presence of < or =0.5 mM glucose from fetal bovine serum), supported stable levels of NTP and PCr in the tumor cells for up to 12 h. In the presence of 2.5 mM acetate, the bioenergetic status of cells in EMT6 spheroids declined slowly but measurably, and no incorporation of label from 2-(13)C-acetate into other metabolites was detected either in intact perfused spheroids or in high-resolution spectra of extracts. In contrast, when the anaplerotic TCA cycle substrate 3-(13)C-propionate replaced acetate, the high-energy phosphate levels in EMT6/Ro spheroids were somewhat reduced, but stabilized at a new lower level. Incubation of spheroids with 3-(13)C-propionate (with natural abundance glucose and glutamine) resulted in label detectable in the C2 and C3 of glutamate, but the primary labeled compound was methylmalonate, an intermediate in propionate metabolism. Addition of vitamin B(12), a cofactor for methylmalonyl CoA reductase, to the growth medium 24 h prior to perfusion with propionate resulted in the elimination of the methylmalonate resonance. A variety of 2- and 3-labeled metabolites were detected, including succinate, malate and glutamate. Labeling of C2 and C3 of lactate implicated cytoplasmic malic enzyme activity.

Hippocampal pathology in schizophrenia: magnetic resonance imaging and spectroscopy studies.

The hippocampus is a site of previously reported structural and functional abnormalities in schizophrenia. We used magnetic resonance imaging (MRI) and proton magnetic resonance spectroscopy (MRS) to measure gray matter volumes, the neuronal marker N-acetylaspartate (NAA), and the combination of glutamate (Glu), glutamine (Gln), and gamma-aminobutyric acid (GABA), designated Glx. Measurements were obtained of the medial temporal lobe, centered on the hippocampus, in 10 male patients with schizophrenia (3 neuroleptic-medicated and 7 medication-free), and 10 matched normal volunteers. MRI volumetric measurements and MRS data obtained with short echo time (TE=20 ms) one-dimensional STEAM chemical shift imaging (CSI) on a GE 1.5 Tesla Signa system were analyzed. A laterality index ¿(L-R)/(L+R) was generated from the ratio of Glx to choline-containing compounds (Cho) to test asymmetry changes. Reliability of the MRS measures was assessed with five test-retest studies of healthy volunteers and showed coefficients of variation (CV) in the range of 36-44% for the MRS ratios and standard deviations (S.D.) of 0.15-0.17 for the laterality indices. The Glx/Cho laterality index showed a relative right-sided excess in this region in the patients (-0.23+/-0.20) compared to the controls (+0.06+/-0.20), which was not confounded by tissue composition or placement variability of the MRS voxels. Hippocampal volume deficit and asymmetry were not significant, and other MRS measures showed no differences between patients and controls. The preliminary finding of a lateralized abnormality in Glx is consistent with postmortem findings of asymmetric neurochemical temporal lobe abnormalities in schizophrenia.