Radiolabeling of protected tryptophan with [18F]fluoromethyl tosylate: Formation of [18F]fluoromethyl ester of tryptophan instead of 1-N-[18F]fluoromethyl tryptophan methylester.

The reaction of [18F]fluoromethyl tosylate with methyl(tert-butoxycarbonyl)-l-tryptophanate results in formation the O-alkylated ester of the tryptophan instead of alkylation of the indole nitrogen of tryptophan as initially anticipated. Treatment of protected tryptophan with NaH in dimethyl formamide (DMF) along with [18F]fluoromethyl tosylate at 130°C results in the formation of [18F]fluoromethyl(tert-butoxycarbonyl)-l-tryptophanate. Preferential formation of the O-alkylated product is postulated to be due to the hydrolysis of the ester. Confirmation of the O-alkylation was obtained by synthesizing the [19F]fluoromethyl(tert-butoxycarbonyl)-l-tryptophanate insitu and examining its NMR characteristics using multiple NMR techniques. Similar results were also obtained when reacting Boc-tryptophan-N-carboxyanhydride precursor with fluoromethyl tosylate.

Challenges in the automated synthesis of [18F]-1-fluoroethyl tryptophan: Formation of both O- and N-alkylated products.

[18F]Fluoroethyl tosylate was synthesized using an automated "Synthra" module using ethylene di-tosylate and [18F]fluoride/K222/K2CO3 in acetonitrile. [18F]Fluoroethyl tosylate was purified by semi-preparative HPLC followed by reformulation using a C18 Sep-Pak cartridge and eluted with DMF. Using this [18F]fluoroethyl tosylate, we attempted to alkylate protected tryptophan aiming to obtain the N-[18F]fluoroethyl-t-Boc-tryptophan methyl ester. Initial attempts resulted in the formation of the O-alkylated, rather than N-alkylated product. Manual removal of the cartridge from the automated module, followed by an extended drying of the cartridge under high flow nitrogen, was required to form the desired N-alkylated product. This demonstrates that the drying process in automated modules requires modification for sensitive N-alkylation of compounds and may be essential for compounds like tryptophan methyl ester that have multiple potential sites of alkylation in their chemical structure.

Management of radioactive waste gases from PET radiopharmaceutical synthesis using cost effective capture systems integrated with a cyclotron safety system.

The emphasis on the reduction of gaseous radioactive effluent associated with PET radiochemistry laboratories has increased. Various radioactive gas capture strategies have been employed historically including expensive automated compression systems. We have implemented a new cost-effective strategy employing gas capture bags with electronic feedback that are integrated with the cyclotron safety system. Our strategy is suitable for multiple automated 18F radiosynthesis modules and individual automated 11C radiosynthesis modules. We describe novel gas capture systems that minimize the risk of human error and are routinely used in our facility.

Heteronuclear NMR spectroscopic investigations of hydrogen bonding in 2-(benzo[d]thiazole-2'-yl)-N-alkylanilines.

The 2-(benzo[d]thiazole-2'-yl)-N-alkylanilines have previously revealed the presence of a strong intramolecular hydrogen bond. This in turn gives rise to a more complicated multiplet for the protons attached to the carbon adjacent to the amino group. This intramolecular hydrogen bond was investigated by a deuterium exchange experiment using heteronuclear NMR spectroscopy (1H, 13C, 15N and 2H). We observed changes in the multiplet structure and chemical shifts providing further evidence that the deuterium replaces the hydrogen in the intramolecular hydrogen bond. A time course study of the D2O exchange confirmed the presence of a strong hydrogen bond. The comparison of the structures obtained by X-ray crystallography showed a very small difference in planarity between the two-substituted and four-substituted amino compounds. In both the cases, the phenyl ring is not absolutely coplanar with the thiazole unit. The existence of this intramolecular hydrogen bond in 2-(benzo[d]thiazole-2'-yl)-N-alkylanilines was further confirmed by single crystal X-ray crystallography.

Synthesis, characterization and (11) C-radiolabeling of aminophenyl benzothiazoles: structural effects on the alkylation of amino group.

Several aminophenyl benzothiazoles were prepared with a view to using them as amyloid binding agents for imaging ß-amyloid in Alzheimer's disease. These precursors were radiolabeled with (11) C-positron-emitting radioisotope using an automated synthesizer and selected radiolabeled compounds were further purified by HPLC. Our results demonstrate that changes in structure have a major influence on the radioactive yield and the ease with which the radiolabel can be introduced. Aminophenyl benzothiazoles with an attached isopropyl group resisted dialkylation perhaps due to steric hindrance caused by this group. Straight chain attachment of methyl, ethyl, butyl, and crotyl groups in the structure decreased the radiochemical yield. Notably, the o-aminophenyl benzothiazole derivatives were difficult to alkylate despite stringent experimental conditions. This reactivity difference is attributed to the hydrogen bonding characteristics of the o-amino group with the nitrogen atom of the thiazole ring.

Language skills of school-aged children prenatally exposed to antiepileptic drugs.

OBJECTIVES: Fetal exposure to some antiepileptic drugs (AEDs) carries increased risk of major birth defects, and may be associated with reduced intellectual abilities. The impact on language remains unclear. This study aimed to investigate the impact of fetal AED exposure on language skills. METHODS: Women with epilepsy and their children were recruited to this observational study through the Australian Pregnancy Register for Women with Epilepsy and Allied Disorders. Language skills of 102 AED-exposed children were assessed using the Clinical Evaluation of Language Fundamentals, fourth edition (CELF-4). Assessments were conducted blind to drug. Maternal epilepsy, pregnancy, and medical histories were obtained from prospectively collected records. RESULTS: Mean CELF-4 Core Language scores of children exposed to sodium valproate in monotherapy (mean 91.5, SD 17.5) or polytherapy (mean 73.4, SD = 22.3) were significantly below the standardized test mean of 100 (p < 0.05). Mean language scores of children exposed to carbamazepine or lamotrigine monotherapy, or polytherapy without sodium valproate, were not significantly different from normal. First-trimester sodium valproate dose was negatively correlated with language scores, and significantly predicted language scores after controlling for other group differences. CONCLUSIONS: Fetal exposure to sodium valproate increases the risk of language impairment. This should be taken into account when making treatment decisions for women with epilepsy of childbearing age.

Synthesis and characterization of 1- and 2-cinnamoyloxyacetonaphthones.

The synthesis of 1- and 2-cinnamoyloxyacetonaphthones was achieved in one step using hydroxyl acetonaphthones and substituted cinnamic acids in the presence of a catalytic amount of phosphoroxychloride. Structural characterization was accomplished using high-resolution nuclear magnetic resonance (NMR) spectroscopy. Chemical shifts of the compounds were compared and the change in the chemical shifts relative to electron-donating and -withdrawing groups is presented. Introduction of a thiophene ring instead of phenyl-substituted analogs caused shielding of the olefinic proton.

Accuracy of clinical signs, SEP, and EEG in predicting outcome of hypoxic coma: a meta-analysis.

OBJECTIVE: Accurate prediction of neurologic outcome after hypoxic coma is important. Previous systematic reviews have not used summary statistics to summarize and formally compare the accuracy of different prognostic tests. We therefore used summary receiver operating characteristic curve (SROC) and cluster regression methods to compare motor and pupillary responses with sensory evoked potential (SEP) and EEG in predicting outcome after hypoxic coma. METHODS: We searched PubMed, MEDLINE, and Embase (1966-2007) for reports in English, German, and French and identified 25 suitable studies. An SROC was constructed for each marker (SEP, EEG, M1 and M < or = 3), and the area under the curve (AUC), a measure of diagnostic accuracy, was determined. For comparison, we calculated the differences between the AUC for each test and M1 reference standard. RESULTS: The AUC for absent SEP was larger than those for M1, M < or = 3, absent pupillary response, and EEG when the examinations were performed within the first 24 hours. The difference between the AUC for SEP (AUC 0.891) and that for M1 (AUC 0.786) was small (0.105, 95% confidence interval 0.023-0.187), only reaching significance on day 1 after coma onset. The use of M < or = 3 improved the diagnostic accuracy of motor signs. CONCLUSIONS: This study demonstrated that sensory evoked potential (SEP) is marginally better than M1 at predicting outcome after hypoxic coma. However, the superiority of SEP diminishes after day 1 and when M < or = 3 is used. The findings therefore caution against the tendency to generalize that SEP is a better marker than clinical signs.

Hippocampal volume and depression: insights from epilepsy surgery.

BACKGROUND: Major depression is common after epilepsy surgery. It has previously been suggested that surgical removal of limbic system structures such as the hippocampus may contribute to this comorbidity. Recent magnetic resonance imaging studies have found smaller hippocampal volumes in depressed patients in comparison with controls. AIMS: The current study examined whether preoperative hippocampal volumes were associated with depression experienced after epilepsy surgery. Patients undergoing mesial (n = 26) and non-mesial (n = 16) temporal lobe resections were assessed preoperatively, and for 1 year postoperatively. Assessment included a clinical interview and the Beck Depression Inventory. Hippocampal volumes were measured on the preoperative T1-weighted magnetic resonance imaging scans of the patients and 41 neurologically normal controls. RESULTS: A similar proportion of mesial and non-mesial temporal patients had a preoperative history of major depression. Postoperatively, 42% of mesial and 19% of non-mesial temporal patients were depressed. There was no relationship between hippocampal volume and preoperative depression in either group. Depression after surgery was associated with significantly smaller hippocampal volumes contralateral to the resection in the mesial temporal group (p = 0.005). This effect was seen in mesial temporal patients who developed de novo depression (p = 0.006). Hippocampal volume was unrelated to postoperative depression in the non-mesial group. CONCLUSION: This study highlights the role of neurobiological factors in the development of postoperative depression. These initial findings have implications for understanding depression following epilepsy surgery as well as the pathogenesis of depression more generally.

Reduced striatal D1 receptor binding in autosomal dominant nocturnal frontal lobe epilepsy.

BACKGROUND: Mutations of the neuronal nicotinic acetylcholine (nACh) receptor identified in patients with autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) lead to increased sensitivity to ACh. As activation of presynaptic nicotinic receptors augments the release of dopamine in the striatum and the prefrontal regions, we tested the hypothesis that that the alpha4-Ser248Phe mutation affects dopaminergic transmission. METHODS: We measured D(1) receptor binding using [(11)C]-SCH23390 and PET in 12 subjects with the alpha4-Ser248Phe mutation (3 men, mean age 41 +/- 16 years) and 19 controls (8 men, mean age 36 +/- 13 years) matched for gender, smoking status, and age. Parametric images were produced using the simplified reference region method. Both MRI-based regions of interest and voxel based analyses were used. RESULTS: Reduced striatal [(11)C]-SCH23390 binding occurred with the mutation (controls 1.1 +/- 0.1; ADNFLE 0.97 +/- 0.2; p < 0.01). Statistical parametric mapping confirmed a region of reduced [(11)C]-SCH23390 binding in the right putamen in alpha4-Ser248Phe subjects compared to controls (309 voxels, local maxima 20 16 -2 mm; Z(score) 3.57, p < 0.05). CONCLUSIONS: Reduced D(1) receptor binding may represent increased extracellular dopamine levels or, more likely, receptor downregulation. Alterations in mesostriatal dopaminergic circuits may contribute to nocturnal paroxysmal motor activity in autosomal dominant nocturnal frontal lobe epilepsy.

Effect of levetiracetam on cortical excitability: a transcranial magnetic stimulation study.

BACKGROUND AND PURPOSE: We studied the effect of levetiracetam (LEV), an anticonvulsant with a novel mechanism of action, on cortical excitability, measured using transcranial magnetic stimulation (TMS). For this purpose, 38 healthy volunteers were assessed in two TMS sessions, before and after an oral dose of 3000 mg LEV. METHODS: Resting motor threshold (RMT), intracortical facilitation (ICF) and intracortical inhibition (ICI), cortical silent period (CSP) threshold and duration and motor-evoked potential (MEP) amplitude were calculated. RESULTS: After treatment with LEV, RMT was increased (mean +/- SD: 63 +/- 14% of the maximum stimulator output) compared with baseline (58 +/- 11%). CSP threshold was decreased after LEV (54 +/- 10%; baseline, 57 +/- 11%). CSP duration was increased after LEV (116 +/- 37 ms; baseline: 102 +/- 33 ms). LEV did not affect ICF or ICI or mean MEP amplitude significantly. CONCLUSIONS: Our results indicate that LEV modulates some aspects of cortical excitability. Whereas the increase in the RMT most probably reflects the effect of LEV on ion channel activity, effects on the CSP might represent a modulation of GABA receptors at cortical and spinal level.

Asymmetry of language activation relates to regional callosal morphology following early cerebral injury.

The anatomical factors underlying reorganization of language representation are yet to be elucidated, although correlations between asymmetric structures and language lateralization have been identified. Previous research has implicated the corpus callosum in the development of language lateralization. This study examined the relationship between callosal morphology and language asymmetry, using letter fluency functional magnetic resonance imaging, in 13 patients with focal epilepsy and 8 healthy controls. Regional callosal thickness was determined without relying on a priori delineation of callosal segments. We predicted that language asymmetry measured by fMRI activation laterality scores would be correlated with regional callosal thickness in both groups. However, only the degree of language activation asymmetry was significantly correlated with callosal thickness in the isthmus and the midbody of patients, and there was a significant interaction between the groups with respect to callosal thickness and language activation asymmetry. These data suggest that callosal pathways may be important for language reorganization in the context of early cerebral injury.

Hypoxic tissue in ischaemic stroke: persistence and clinical consequences of spontaneous survival.

In ischaemic stroke, expansion of the infarct core occurs at the expense of surrounding hypoxic, metabolically compromised tissue over a period of 24 h or more in a considerable proportion of patients. It is uncertain whether hypoxic tissue observed at later times after stroke onset retains the potential for survival or whether such survival has an impact on functional outcome. These factors may determine the effectiveness of therapeutic strategies aimed at salvaging this tissue. We tested the hypotheses that metabolically compromised hypoxic tissue observed within 48 h after onset of ischaemic stroke retains the potential for spontaneous survival and that the impact of such survival on functional outcome is time dependent. Consecutive patients presenting within 48 h of ischaemic stroke were studied with [(18)F]fluoromisonidazole, a ligand binding to hypoxic but viable tissue, and PET. Subjects were grouped into two time epochs, 12 h, based on the interval from stroke onset to the time of tracer injection, and had infarct volumes measured on CT/MRI at 7 days (n = 60). The total ischaemic volume (TIV) and the proportion of the TIV that spontaneously survived (surviving hypoxic volume ratio, SHVR) were defined from the co-registered CT/MRI images. These volumetric measures were correlated with neurological outcome assessed at day 7-10 by percentage change in the National Institutes of Health Stroke Scale (DeltaNIHSS), and at 3 months by Barthel Index (BI) and modified Rankin Score (mRS). Of 66 patients investigated, hypoxic tissue occurred in 33 and outcome data was available in 27. Hypoxic tissue constituted >20% of the TIV in 60% of studies 12 h. The spontaneously surviving proportion of the TIV (median 6.9%) or hypoxic tissue (median 45.9%) was not significantly different in patient subgroups studied 12 h after stroke onset. Spontaneous survival of hypoxic tissue (surviving hypoxic volume ratio) was associated with improved neurological outcome in both time epochs: 12 h, DeltaNIHSS (r = 0.59, P < 0.01) and day 90 mRS (r = -0.46, P < 0.05). The finding that similar proportions of hypoxic tissue survived spontaneously within each time epoch suggests that its fate is not predetermined. The favourable neurological outcome associated with spontaneous survival of hypoxic tissue, even 12-48 h after stroke onset, suggests that the volume of hypoxic tissue that progressed to infarction may represent a valuable target for therapeutic intervention.

Topography and temporal evolution of hypoxic viable tissue identified by 18F-fluoromisonidazole positron emission tomography in humans after ischemic stroke.

BACKGROUND AND PURPOSE: We sought to characterize the spatial and temporal evolution of human cerebral infarction. Using a novel method of quantitatively mapping the distribution of hypoxic viable tissue identified by 18F-fluoromisonidazole (18F-FMISO) PET relative to the final infarct, we determined its evolution and spatial topography in human stroke. METHODS: Patients with acute middle cerebral artery territory stroke were imaged with 18F-FMISO PET (n=19; <6 hours, 4; 6 to 16 hours, 4; 16 to 24 hours, 5; 24 to 48 hours, 6). The hypoxic volume (HV) comprised voxels with significant (P<0.05; >1 mL) uptake on statistical parametric mapping compared with 15 age-matched controls. Central, peripheral, and external zones of the corresponding infarct on the anatomically coregistered delayed CT were defined according to voxel distance from the infarct center and subdivided into 24 regions by coronal, sagittal, and axial planes. Maps ("penumbragrams") displaying the percentage of HV in each region were generated for each time epoch. RESULTS: Higher HV was observed in the central region of the infarct in patients studied within 6 hours of onset (analysis of covariance [ANCOVA]; P<0.05) compared with those studied later, in whom the HV was mainly in the periphery or external to the infarct. HV was maximal in the superior, mesial, and posterior regions of the infarct (ANCOVA; P<0.05). CONCLUSIONS: These observations suggest that infarct expansion occurs at the expense of hypoxic tissue from the center to the periphery of the ischemic region in humans, similar to that seen in experimental animal models. These findings have important pathophysiological and therapeutic implications.

Statistical parametric mapping of hypoxic tissue identified by [(18)F]fluoromisonidazole and positron emission tomography following acute ischemic stroke.

Positron emission tomography (PET) and the ligand [(18)F]fluoromisonidazole ((18)F-FMISO) have been used to image hypoxic tissue in the brain following acute stroke. Existing region of interest (ROI)-based methods of analysis are time consuming and operator-dependent. We describe and validate a method of statistical parametric mapping to identify regions of increased (18)F-FMISO uptake. The (18)F-FMISO PET images were transformed into a standardized coordinate space and intensity normalized. Then t statistic maps were created using a pooled estimate of variance. Statistical inference was based on the theory of Gaussian Random Fields. We examined the homogeneity of variance in normal subjects and the influence of normalization by mean whole brain activity versus mean activity in the contralateral hemisphere. Validity of the distributional assumptions inherent in parametric analysis was tested by comparison with a non-parametric method. The results of parametric analysis were also compared with those obtained with the existing ROI-based method. Variance in uptake at each voxel in normal subjects was homogeneous and not affected by mean voxel activity or distance from the centre of the image. The method of normalization influenced results significantly. Normalization by whole brain mean activity resulted in a smaller volume of tissue being classified as hypoxic compared to normalisation by mean activity in the contralateral hemisphere. The ROI-based method was subject to interobserver variability with a coefficient of variability of 16%. The volumes of hypoxic tissue identified by parametric and nonparametric methods were highly correlated (r = 0.99). These findings suggest that using a pooled variance and contralateral hemisphere normalisation, statistical parametric mapping can be used to objectively identify regions of increased (18)F-FMISO uptake following acute stroke in individual subjects.

Prolonged cortical silent period after transcranial magnetic stimulation in generalized epilepsy.

Transcranial magnetic stimulation (TMS) produces a cortical silent period (CSP) during a voluntary contraction. The duration of the CSP was used to assess the level of intracortical inhibition in patients with untreated idiopathic generalized epilepsy (IGE). Mean CSP duration was assessed at three TMS stimuli in 21 patients with IGE compared with 19 normal control subjects. Mean CSP duration was increased at all stimulus intensities, indicating that intracortical inhibition is increased in patients with IGE.

Lateralizing value of peri-ictal headache: A study of 100 patients with partial epilepsy.

To determine the lateralizing value of peri-ictal headache, the authors conducted a standardized interview of 100 patients with partial epilepsy, 60 with temporal lobe epilepsy (TLE) and 40 with extratemporal epilepsy (ETE). Peri-ictal headache occurred in 47 of 100 (47%) patients. Peri-ictal headache was more likely to be ipsilateral to the seizure onset in TLE (27 of 30 = 90%) than in ETE (two of 17 = 12%; p< 0.001). For both groups, peri-ictal headache usually conformed to the diagnostic criteria for common migraine (18 of 30 = 60% in TLE; 7 of 17 = 41% in ETE).

T2 relaxometry can lateralize mesial temporal lobe epilepsy in patients with normal MRI.

In unselected patients with intractable temporal lobe epilepsy (TLE), approximately 15% do not have detectable hippocampal atrophy on MRI. The purpose of this study was to evaluate whether T2 relaxometry can identify hippocampal pathology and lateralize the epileptic focus in patients with intractable TLE, who do not demonstrate hippocampal atrophy on volumetric MRI (MRIV). We selected 14 patients with unilateral TLE who had unilateral atrophy and 11 patients with unilateral TLE who had no evidence of atrophy on MRIV. Images were acquired on a 1.5 T MR scan using a dual echo sequence with 23 contiguous oblique coronal slices in all patients and in 14 healthy subjects. Fitting a single exponential decay equation to the imaging data generated T2 maps. Averages of six slices containing the head, body, and tail of the hippocampus were used to calculate hippocampal T2 relaxation times (HT2). The epileptic focus was defined by history, video-EEG, and surgical response. All TLE patients with hippocampal atrophy and 9/11 (82%) patients with normal MRI had abnormally high HT2 ipsilateral to the epileptic focus. Bilateral abnormal HT2 were found in 6/14 (43%) of patients with unilateral hippocampal atrophy and 2/11 (18%) of patients with normal MRI. However, this increase was always greater ipsilateral to the epileptic focus. Qualitative hippocampal pathology showed gliosis and neuronal loss in 10/14 operated patients with hippocampal atrophy on MRIV and in 5/7 operated patients with normal MRI. In conclusion, hippocampal T2 mapping provides evidence of hippocampal damage in the majority of patients with intractable TLE who have no evidence of atrophy on MRI and can correctly lateralize the epileptic focus in most patients.

Sensorimotor organization in patients who have undergone hemispherectomy: a study with (15)O-water PET and somatosensory evoked potentials.

To identify cortical structures that subserve residual motor and sensory function in patients with congenital hemiparesis due to a porencephalic cyst, we examined, using [(15)O]H2O, PET and somatosensory evoked potentials (SEPs) in three patients with left-sided hemiparesis who had undergone hemispherectomy. Motor stimulation of the affected hand produced ipsilateral activation in the premotor area in all patients, the SMA in two patients, and SII in two patients. Vibrotactile stimulation resulted in activation of the ipsilateral SII in all subjects. Median nerve stimulation of the affected hand produced ipsilateral long-latency SEPs in fronto-centro-parietal areas, whereas stimulation of the non-affected hand produced normal early cortical potentials in the contralateral hemisphere. Our results suggest that residual function in the paretic hand is warranted through non-primary motor and sensory areas, and higher order associative areas in the intact hemisphere.