A prospective diagnostic accuracy study evaluating rotational thromboelastometry and thromboelastography in 100 patients with von Willebrand disease.

INTRODUCTION: Rotational thromboelastometry (ROTEM® ) and thromboelastography (TEG® ) are increasingly used in the perioperative and emergency assessment of bleeding tendencies. The diagnostic value of ROTEM and TEG for von Willebrand disease (VWD) remains to be established. AIM: To investigate whether ROTEM and TEG can discriminate patients with VWD from healthy controls. METHODS: Rotational thromboelastometry and TEG whole blood coagulation profiles were compared between VWD patients (n = 100) and healthy controls (n = 89). Measures of diagnostic accuracy were calculated, including sensitivity, specificity and receiver operating characteristic (ROC) curve. RESULTS: Prolonged TEG R-time had a positive and negative predictive value (PPV, NPV) of 0.84 and 0.68 respectively. TEG clotting index (CI) had a PPV of 1.00 and an NPV of 0.60. Both R-time and CI had a high specificity and accurately discriminated VWD patients from healthy controls, with an ROC area under the curve of 0.85 and 0.99 respectively. In multivariate analysis, low FVIII levels, but not von Willebrand factor (VWF) antigen or activity, determined hypocoagulable TEG R (R2 = 0.35) and CI levels (R2 = 0.51). The ROTEM coagulation profiles of VWD patients did not differ from healthy controls. CONCLUSIONS: Thromboelastography R and CI accurately discriminated VWD patients from healthy controls, partly through the detection of low FVIII levels. The test's performance may be improved through adjustment of the test thresholds to a local reference population. Both intrinsic pathway-activated (INTEM) and tissue factor pathway-activated (EXTEM) ROTEM were of limited diagnostic value in VWD.

CC chemokine ligands in patients presenting with stable chest pain: association with atherosclerosis and future cardiovascular events.

BACKGROUND: CC chemokine ligands (CCLs) are elevated during acute coronary syndrome (ACS) and correlate with secondary events. Their involvement in plaque inflammation led us to investigate whether CCL3-5-18 are linked to the extent of coronary artery disease (CAD) and prognostic for primary events during follow-up. METHODS: We measured CCL3-5-18 serum concentrations in 712 patients with chest discomfort referred for cardiac CT angiography. Obstructive CAD was defined as ≥50 % stenosis. The extent of CAD was measured by calcium score and segment involvement score (number of coronary segments with any CAD, range 0-16). Patients were followed up for all-cause mortality, ACS and revascularisation, for a mean 26 ± 7 months. RESULTS: Patients with obstructive CAD had significantly higher CCL5 (p = 0.02), and borderline significantly elevated CCL18 plasma levels as compared with patients with <50 % stenosis (p = 0.06). CCL18 levels were associated with coronary calcification (p = 0.002) and segment involvement score (p = 0.007). Corrected for traditional risk factors, only CCL5 provided independent predictive value for obstructive CAD: odds ratio (OR) 1.27 (1.02-1.59), p = 0.04. CCL5 provided independent predictive value for primary events during follow-up: OR 1.62 (1.03-2.57), p = 0.04. CONCLUSIONS: While CCL18 serum levels correlated with extent of CAD, CCL5 demonstrated an independent association with the presence of obstructive CAD, and occurrence of primary cardiac events.

Cortical regulation of dopamine depletion-induced dendritic spine loss in striatal medium spiny neurons.

The proximate cause of Parkinson's disease is striatal dopamine depletion. Although no overt toxicity to striatal neurons has been reported in Parkinson's disease, one of the consequences of striatal dopamine loss is a decrease in the number of dendritic spines on striatal medium spiny neurons (MSNs). Dendrites of these neurons receive cortical glutamatergic inputs onto the dendritic spine head and dopaminergic inputs from the substantia nigra onto the spine neck. This synaptic arrangement suggests that dopamine gates corticostriatal glutamatergic drive onto spines. Using triple organotypic slice cultures composed of ventral mesencephalon, striatum, and cortex of the neonatal rat, we examined the role of the cortex in dopamine depletion-induced dendritic spine loss in MSNs. The striatal dopamine innervation was lesioned by treatment of the cultures with the dopaminergic neurotoxin 1-methyl-4-phenylpyridinium (MPP+) or by removing the mesencephalon. Both MPP+ and mesencephalic ablation decreased MSN dendritic spine density. Analysis of spine morphology revealed that thin spines were preferentially lost after dopamine depletion. Removal of the cortex completely prevented dopamine depletion-induced spine loss. These data indicate that the dendritic remodeling of MSNs seen in parkinsonism occurs secondary to increases in corticostriatal glutamatergic drive, and suggest that modulation of cortical activity may be a useful therapeutic strategy in Parkinson's disease.

Endogenous morphine and codeine. Possible role as endogenous anticonvulsants.

Exogenously administered morphine can have both convulsive or anticonvulsive effects, depending on the dose and species. The levels of the endogenous opiate alkaloids morphine and codeine were significantly elevated in specific rat brain regions by the convulsive drug, pentylenetetrazole, as well as by the anticonvulsant drugs, carbamazepine and phenytoin. Morphine and codeine levels in peripheral tissues (heart, lung, spleen and adrenal) were unaffected by these drugs. Maximal increases in morphine levels were seen in the hypothalamus and striatum (2-10-fold), while lesser increases occurred in the midbrain and brain stem (2-4-fold). Codeine levels were also markedly increased in hypothalamus (5-10 fold), In contrast to morphine, codeine levels were also increased in the hippocampus (2-10-fold), but were unchanged in the striatum. These studies suggest that the endogenous alkaloids morphine and codeine are involved in the modulation of convulsions and that morphine and/or codeine may act as an endogenous anticonvulsant.

Anxiolytic-like effects of DAIZAC, a selective high-affinity 5-HT(3) receptor antagonist, in the mouse elevated plus-maze.

Behavioral effects of desamino-3-iodozacopride (DAIZAC) [(S)-5-chloro-3-iodo-2-methoxy-N-(1-azabicyclo[2.2.2]oct-3-yl)benzamide], a selective high-affinity 5-HT(3) receptor antagonist (K(D) 0.14 nM), were evaluated in the mouse elevated plus-maze using the anxiolytic benzodiazepine, diazepam, as a positive control. DAIZAC treatment produced a significant dose-related increase in the time spent in the open arm. The increased total time in the open arm resulted from a significant dose-dependent increase in the number of entries into that arm. The minimum dose of DAIZAC associated with a statistically significant increase in entries and time spent in the open arm was 0.05 mg/kg ip, consistent with its high affinity for the 5-HT(3) receptor. DAIZAC did not affect the amount of time spent in the open arm after each entry. Thus, DAIZAC reduced apparent avoidance of the open arm when the animal was in the central compartment, without affecting active avoidance of that arm when the animal was in the exposed condition. The increase in the open-arm entries was accompanied by a corresponding reduction in the number of entries into the closed arm with a consequent reduction in the time spent in the closed arm. The time spent in the closed arm after each entry was not altered by DAIZAC administration. As such, the sole apparent effect of DAIZAC was to alter the choice of arm to enter when the animal was in the central compartment. Diazepam also significantly increased total time in the open arm; however, the increase was not attributable to a single behavioral factor. The anxiolytic-like effects of DAIZAC reached maximum by 20-30 min and returned to baseline levels by 90 min. Ex vivo binding studies found that levels of DAIZAC-like activity assayed in brains of mice 25 min after DAIZAC injection were significantly correlated with the behavioral parameters associated with anxiolysis. These results indicate that DAIZAC produces dose-dependent anxiolytic-like behavioral changes in the mouse elevated plus-maze that are correlated with brain DAIZAC-like activity.

Rapid bioanalysis of vancomycin in serum and urine by high-performance liquid chromatography tandem mass spectrometry using on-line sample extraction and parallel analytical columns.

A novel high-performance liquid chromatography tandem mass spectrometry (LC/MS/MS) method is described for the determination of vancomycin in serum and urine. After the addition of internal standard (teicoplanin), serum and urine samples were directly injected onto an HPLC system consisting of an extraction column and dual analytical columns. The columns are plumbed through two switching valves. A six-port valve directs extraction column effluent either to waste or to an analytical column. A ten-port valve simultaneously permits equilibration of one analytical column while the other is used for sample analysis. Thus, off-line analytical column equilibration time does not require mass spectrometer time, freeing the detector for increased sample throughput. The on-line sample extraction step takes 15 seconds followed by gradient chromatography taking another 90 seconds. Having minimal sample pretreatment the method is both simple and fast. This system has been used to successfully develop a validated positive-ion electrospray bioanalytical method for the quantitation of vancomycin. Detection of vancomycin was accurate and precise, with a limit of detection of 1 ng/mL in serum and urine. The calibration curves for vancomycin in rat, dog and primate were linear in a concentration range of 0.001-10 microg/mL for serum and urine. This method has been successfully applied to determine the concentration of vancomycin in rat, dog and primate serum and urine samples from pharmacokinetic and urinary excretion studies.

Plasma cholecystokinin levels in Prader-Willi syndrome and obese subjects.

The cardinal feature of individuals with Prader-Willi syndrome (PWS) is severe hyperphagia-mediated obesity resulting from a faulty satiety mechanism. PWS is the most common genetic cause of marked obesity. Cholecystokinin (CCK) is a 33-amino-acid peptide found in high levels in the gut and brain involved in mediating the satiety response to meals. Free fatty acids (FFA) are responsible for the stimulation of CCK release after a fatty meal, and CCK and plasma FFA levels rise in tandem in normal individuals. Fasting plasma CCK levels were measured by radio-immunoassay in 33 PWS subjects with a mean age of 22.2 years +/- 8.1 years and 24 obese control subjects without a known cause of their obesity with a mean age of 28.7 years +/- 12.9 years. Consistent with previous findings, neither fasting plasma FFA levels (617.5 versus 486.8 microm/mL) or CCK levels (21.0 versus 19.1 pg/mL) were significantly different in PWS or control subjects, respectively. However, there was a significant correlation between fasting plasma FFA and CCK levels in obese subjects (r = 0. 64, P < 0.01), this correlation was completely lacking in PWS subjects (r = -0.06, P = 0.79). This difference in correlation coefficients constitutes a large effect. There were no significant effects observed for genetic subtypes (15q11-q13 deletion or maternal disomy 15), body mass index, percentage of fat, plasma levels of insulin, C-peptide, glucagon or leptin, age, or gender on CCK levels in our PWS subjects. These results suggest that differences in the peripheral CCK response to FFA levels may be a factor contributing to the altered satiety response in PWS subjects.

Evaluation of plasma time-activity curves of technetium-99m-mebrofenin for measurement of hepatic function in dogs.

In this study, plasma time-activity curves of 99mTc-mebrofenin were used to quantify hepatic function in dogs before and after induction of hepatic damage using a hepatotoxic agent. Nine dogs were determined to be healthy on the basis of physical examination, laboratory data and hepatic imaging. Plasma samples were collected 1, 3, 5, 7, 9, 15, 20, 30, 40, 50, and 60 minutes following a peripheral venous injection of 111-222 MBq (3-6 mCi) of 99mTc-mebrofenin. The area under the plasma time-activity curve (AUC) was calculated using two different methods and compared to direct measurement of the hepatic extraction efficiency. First pass hepatic extraction efficiency of 99mTc-mebrofenin was calculated from differential equation analysis of a two-compartment model following mesenteric venous injection of the radiopharmaceutical. In 7 of the original 9 dogs and 2 additional healthy dogs, plasma clearance and hepatic extraction efficiency determination were repeated following induction of hepatic injury by thiacetarsamide (3 mg/kg IV twice daily for 1 day). In one additional dog, hepatic injury was induced using carbon tetrachloride (0.3 ml/kg IP). Plasma time-activity curves of 99mTc-mebrofenin had kinetics of a two compartment model. Area under the curve was highly correlated with hepatic extraction efficiency. The AUC integrated from 1-60 minutes (AUC60) had the best correlation with hepatic extraction efficiency (r2 = 0.978, p < 0.001). A formula for calculation of hepatic extraction efficiency was derived using linear regression analysis: hepatic extraction efficiency = 105.583 - 3.099 x 10(5) x AUC60. Plasma clearance of a peripheral venous injection of 99mTc-mebrofenin is a simple, non-invasive, convenient method to quantify hepatic function which can be performed without a gamma camera.

Fractional oxidation of chylomicron-derived oleate is greater than that of palmitate in healthy adults fed frequent small meals.

Differences in oxidation of individual dietary fatty acids could contribute to the effect of dietary fat composition on risk factors for non-insulin-dependent diabetes mellitus and cardiovascular disease. Using a novel stable isotope technique, we compared fractional oxidation of chylomicron-derived oleate and palmitate in 10 healthy adults in a crossover study. 1-(13)C-labeled oleate or palmitate was emulsified into a eucaloric formula diet administered each 20 min for 7 h to produce a plateau in excretion of (13)C label in breath CO(2). Unlabeled oleate and palmitate each provided 16% of dietary energy, and other fatty acids provided 8% of energy. Total dietary fat was 40% of energy, carbohydrate was 46%, and protein was 14%. Diet without tracer was fed for 2 h before beginning tracer administration to establish a baseline fed state. Relative oxidation of oleate versus palmitate was defined as fractional oxidation of oleate divided by fractional oxidation of palmitate. Relative oxidation averaged 1.21 (99.5% confidence interval = 1.03;-1.39), indicating that fractional oxidation of oleate was significantly greater than that of palmitate.

Salt-induced immobilization of affinity ligands onto epoxide-activated supports.

The immobilization of affinity ligands onto epoxy-activated stationary phases is enhanced at high concentrations of certain salts, such as ammonium sulfate and potassium phosphate. This enhancement is thought to occur because of a salt-induced hydrophobic interaction between the affinity ligand and the surface of the stationary phase. The increase in concentration of the affinity ligand near the reactive epoxy groups leads to an increase in the rate of reaction between the nucleophilic groups on the affinity ligand and the epoxide. The salt-induced enhancement is applicable to proteins and nucleotides at neutral pH and to small affinity ligands at elevated pH. In most cases, the hydrolysis of the epoxy groups does not limit the amount of affinity ligand immobilized. This review discusses the use of high salt concentrations to immobilize proteins, oligonucleotides and peptides to epoxy-activated silica and polymer supports. These modified supports can be used in affinity applications such as affinity chromatography or immunoassays.

Biogenic amine activity in response to fluoxetine and desipramine in differentially reared rhesus monkeys.

BACKGROUND: It has been hypothesized that adverse early experience may be a mechanism by which children become vulnerable to later psychopathology via alteration of neurochemical or hormonal systems associated with such disorders. Such effects may in turn affect later responses to pharmacologic agents that act on these systems. METHODS: In this study, 18 mother-reared (MR) and 18 peer-reared (PR) rhesus monkeys experienced six 1-week separations from cagemates interspersed with 1-week reunions, while housed in like-reared groups of 3. Within rearing groups, equal numbers of animals received either fluoxetine (2 mg/kg), desipramine (5 mg/kg) or placebo delivered daily beginning 4 weeks before the first separation. Levels of norepinephrine (NE), the NE metabolite MHPG, the dopamine metabolites DOPAC and HVA, and the serotonin metabolite 5HIAA were measured in CSF samples collected approximately every 2 to 3 weeks during these procedures. RESULTS: Following treatment, DMI increased NE and decreased MHPG in the DMI-treated groups, while 5HIAA was decreased in the fluoxetine-treated groups following treatment. The increase in NE was followed by a sharp decline over the course of treatment, which was accompanied by an increase in MHPG. The rearing groups did not show a differential response to the drug treatments, and the separation manipulation itself had few effects. The mother-reared group showed higher levels of NE and DOPAC over all samples and higher levels of HVA in most samples. CONCLUSIONS: These rearing effects on biogenic amine activity were observed even in the presence of pharmacologic treatments that effectively altered the activity of these systems, and are consistent with previous findings from the same subject. The higher NE values observed in mother-reared infants over separations and reunions may have been due to higher basal levels of NE than peer-reared monkeys or to greater responsiveness to the stress of repeated social disruption or both. These findings agree with other primate studies showing that rearing differences persist beyond the infancy period and add to growing evidence of the important influence of the early social environment on neurobiologic development in primates.

Characterization of (S)-des-4-amino-3-[125I]iodozacopride ([125I]DAIZAC), a selective high-affinity radioligand for 5-hydroxytryptamine3 receptors.

The 5-hydroxytryptamine(HT)3 receptor subtype is present in the central nervous system (CNS) in low abundance, and few selective radiolabeled antagonists with high specific activity are available to study these sites. DAIZAC [desamino-3-iodo-(S)-zacopride; (S)-5-chloro-3-iodo-2-methoxy-N-(1-azobicyclo-[2.2. 2]oct-3-yl)benzamide] is a compound with high affinity and selectivity for the 5-HT3 receptor. Scatchard analysis of specific binding to NCB-20 cell membranes gave a Bmax of 340 +/- 58 fmol/mg protein and a KD of 0.14 +/- 0.03 nM, which is in agreement with the value previously reported in rat brain (KD = 0.15 nM). Nonspecific binding of [125I]DAIZAC in NCB-20 cells was <1% of total binding at the KD for DAIZAC compared with 17% in the rat brain preparation. Unlabeled DAIZAC (10 microM) showed minimal ability to displace binding of radiolabeled ligands selected for their affinities for other CNS receptor and uptake carrier binding sites. The discrimination ratio of DAIZAC for the 5-HT3 receptor over the M1 muscarinic binding site, the non-5-HT3 site at which it was most potent, was >2800. Serotonergic antagonists at every other known CNS serotonergic binding sites (3-30 microM) were ineffective in displacing [125I]DAIZAC binding in rat brain membranes. Similarly, antagonists (3-30 microM) for other nonserotonergic receptors and uptake sites were ineffective in displacing [125I]DAIZAC binding. Autoradiographic studies showed highest specific binding in area postrema and nucleus solitarius, with intermediate levels of binding in entorhinal cortex and hippocampus. DAIZAC inhibited 5-HT3 receptor-mediated inward cation current in NCB-20 cells with an IC50 of 0.24 nM. [125I]DAIZAC is a potent and highly selective ligand for in vitro studies of the 5-HT3 receptor.

Effects of the immunostimulant, levamisole, on opiate withdrawal and levels of endogenous opiate alkaloids and monoamine neurotransmitters in rat brain.

This report present evidence that the immunostimulant drug levamisole, (-)-(S)-2,3,5,6-tetrahydro-6-phenylimidazo[2,1-b] thiazole monohydrochloride, produced a significant elevation of endogeneous morphine and codeine levels in brain regions and peripheral organs and attenuated the effects of naltrexone-induced withdrawal in morphine-addicted rats. Levamisole also significantly altered the metabolism of norepinephrine, dopamine, and serotonin in specific brain regions. These results suggest that levamisole's attenuation of opiate withdrawal may be related to its ability to increase endogeneous opiate alkaloid levels and/or to alter central monoaminergic function. Levamisole does not have significant affinity for opiate receptors. These results raise the intriguing possibility that agents such as levamisole, which elevate the levels of the endogenous opiate alkaloids, might be useful for treating narcotic withdrawal. The mechanism for the immunostimulatory properties of agents such as levamisole and muramyl dipeptide (MDP) have not been established. We suggest that the ability of MDP and levamisole to increase endogenous opiate alkaloids may be related to their immunostimulatory properties.

Protein affinity map of chemical space.

Affinity fingerprinting is a quantitative method for mapping chemical space based on binding preferences of compounds for a reference panel of proteins. An effective reference panel of <20 proteins can be empirically selected which shows differential interaction with nearly all compounds. By using this map to iteratively sample the chemical space, identification of active ligands from a library of 30,000 candidate compounds has been accomplished for a wide spectrum of specific protein targets. In each case, <200 compounds were directly assayed against the target. Further, analysis of the fingerprint database suggests a strategy for effective selection of affinity chromatography ligands and scaffolds for combinatorial chemistry. With such a system, the large numbers of potential therapeutic targets emerging from genome research can be categorized according to ligand binding properties, complementing sequence based classification.

Lack of beta adrenoceptor desensitization in brain following the dual noradrenaline and serotonin reuptake inhibitor venlafaxine.

Venlafaxine, a dual amine reuptake inhibitor, was utilized to delineate the role of the individual aminergic components of the 'serotonin/noradrenaline link' in modifying receptor-linked second messenger cascades. Venlafaxine (20 mg/kg i.p. bid for 10 days) failed to alter in normal animals either the density of beta adrenoceptors or the response of the beta adrenoceptor-coupled adenylate cyclase system to noradrenaline but significantly decreased the cyclic AMP response to noradrenaline in the brain of rats with selective depletion of brain serotonin by p-chlorophenylalanine. The studies provide evidence for a cross-talk between noradrenergic and serotonergic receptor cascades at the level of mechanisms involved in the desensitization of the beta adrenoceptor-coupled adenylate cyclase system.

Characterization of desamino-5-[125I]iodo-3-methoxy-zacopride ([125I]MIZAC) binding to 5-HT3 receptors in the rat brain.

1. Antagonists at 5-HT3 receptors have shown activity in animal models of mental illness, however, few radiolabeled 5-HT3 ligands are available for preclinical studies. MIZAC, an analogue of the selective 5-HT3 antagonist, zacopride, binds with high affinity (1.3-1.5 nM) to CNS 5-HT3 sites. The authors report here the selectivity of MIZAC for these sites in rat brain homogenates. 2. Ninety-seven percent of total specific binding of [125I]MIZAC (0.1 nM) of was displaced by bemesetron (3 microM), a selective 5-HT3 antagonist. Competition studies using ligands with known affinities for 5-HT3 sites give a high correlation with reported pKi values (r2 0.98). Bemesetron displaceable binding has a regional distribution consistent with that of the 5-HT3 receptor, i.e. highest in cortex and hippocampus, and lowest in striatum and cerebellum. 3. Potent antagonists present at concentrations sufficient to occupy 95% of other 5-HT receptor populations (1A, 1B, 1D, 2A, 2B, 2C, 5A, 5B, 6, and 7) showed minimal ability to displace [125I]MIZAC binding (3 nM). Specificity studies using radioligand binding assays selective for 5-HT4, 5-HT6, and 5-HT7 receptors, and for binding sites of other neurotransmitters indicate a high degree of selectivity of [125I]MIZAC for the 5-HT3 receptor. 4. [125I]MIZAC binds to an apparent low affinity (benzac) site having a unique pharmacology. Low affinity binding was displaceable by benztropine, but not by other muscarinic agents nor inhibitors of dopamine uptake. The regional distribution of the low affinity site differed markedly from that of the high affinity site. The apparent affinity of [125I]MIZAC for the benzac site is two orders of magnitude lower than for the 5-HT3 receptor. Given its high selectivity for 5-HT3 binding sites, [125I]MIZAC appears to be a promising ligand for labeling 5-HT3 receptors in vitro and in vivo.

Otolaryngological manifestations of Creutzfeldt-Jakob disease.

Creutzfeldt-Jakob disease (CJD) is one of the uniformly fatal spongiform encephalopathies that is characterized clinically by an unrelenting progression of myoclonus, dementia, and ataxia. Since many of these patients will develop cerebellar abnormalities, some may present to the otolaryngologist with dizziness. Hearing loss, however, to our knowledge, has not been reported. We describe a patient with CJD who presented with hearing loss and vague symptoms of imbalance, and whose disease progressed rapidly and fatally despite an extensive initial workup that was otherwise unrevealing. A review and discussion of the otolaryngological manifestations of CJD is presented. The otolaryngologist should be aware that CJD can present with otolaryngological manifestations, and with proper diagnosis extensive workups may be avoided.

Synthesis and 5-HT-3 receptor binding of 2- and 3-(halo)alkoxyl substituted (S)-N-(1-azabicyclo[2.2.2]oct-3-yl)-5-chlorobenzamides as potential radioligands.

In an effort to develop selective, high-affinity radioligands for the 5-HT-3 receptor, a series of homologues of 5-chloro-2,3-dimethoxy-N-(1-azabicyclo[2.2.2]oct-3-yl)benzamide (2b) was prepared in which individual methoxy groups were replaced by ethoxyl, (2-fluoroethoxyl), allyloxyl, propargyloxyl, or (3-iodoallyl)oxyl groups. Affinities for the 5-HT-3 receptor were determined by displacement of the binding of [125I]MIZAC (2a), a selective 5-HT-3 receptor antagonist radioligand, in rat brain homogenates. The 3-substituted homologues were more potent than the lead compound, 2b. The homologue having the largest 3-substituent, i.e., E-(S)-N-(1-azabicyclo[2.2.2]oct-3-yl)-5-chloro-3-(3-iodo-2-propenyl)oxy- 2-methoxybenzamide (3b, THIZAC), had one of the highest affinities, Ki 0.08 nM. The 2-substituted homologues were equipotent with 2b, having Ki 0.2-0.3 nM, regardless of the size of the substituent. The corresponding iodoallyl derivative, E-(S)-N-(1-azabicyclo[2.2.2]oct-3-yl)-5-chloro-2-(3-iodo-2-propenyl)oxy- 3-methoxybenzamide (4, LIZAC), displayed a Ki of 0.29 nM. Saturation binding of [125I]-4 gave a KD of 0.31 +/- 0.04 nM and a Bmax of 2.36 +/- 0.10 fmol/mg of entorhinal cortex. In vivo biodistribution of [125I]-4 in the rat brain showed increased accumulation in hippocampus relative to that in cerebellum. Both the high-affinity ligands [125I]-3b and [125I]-4 are potentially useful radioligands for studying the 5-HT-3 receptor.

Elevated plasma gamma-aminobutyric acid (GABA) levels in individuals with either Prader-Willi syndrome or Angelman syndrome.

Plasma gamma-aminobutyric acid (GABA) levels were measured in 14 subjects with Prader-Willi syndrome, 9 subjects with Angelman syndrome, and matched control subjects. Mean levels in both patient groups were 2 to 3 times higher than in nonretarded moderately obese or retarded nonobese control subjects. Levels in each patient group differed significantly from both control groups. Neither the two patient groups nor the two control groups differed. GABA levels seemed unrelated to genetic status (chromosome 15 deletion or disomy). These preliminary findings of elevated plasma GABA levels possibly represent a compensatory increase in presynaptic GABA release in response to hyposensitivity of a subset of GABA receptors and could produce increased postsynaptic activation of other normal GABA receptor subtypes, resulting in complex alterations of GABAergic function throughout the brain.

Attenuation of 1-methyl-4-phenylpyridinium (MPP+) neurotoxicity by deprenyl in organotypic canine substantia nigra cultures.

Systemic administration of MPTP to experimental animals induces neurodegeneration of dopaminergic neurons in the central nervous system. MPTP crosses the blood-brain barrier where it is taken up by astrocytes and converted to MPP+ by monamine oxidase-B (MAO-B). Subsequently, MPP+ is selectively taken up by dopaminergic neurons upon which it exerts intracellular neurotoxic effects. Systemic administration of the selective MAO-B inhibitor deprenyl prevents the conversion of MPTP to MPP+ and by this mechanism is able to protect against MPTP neurotoxicity. Deprenyl has also been reported to exert neuroprotective effects that are independent of its MAO-B inhibitory properties, but since MPP+ itself does not cross the blood-brain barrier it is difficult to directly study the MAO-B independent in vivo effects of MPP+ itself. One approach is to use organotypic tissue cultures of the canine substantia nigra (CSN) which permit administration of precise concentrations of pharmacological agents directly to mature, well-developed and metabolically active dopaminergic neurons. These neurons as well as other components of the cultures exhibit morphological and biochemical characteristics identical to their in vivo counterparts. This study was undertaken to evaluate the neuroprotective effects of deprenyl in MPP(+)-treated cultures by measuring changes in the levels of HVA as an indicator of dopamine release and metabolism by dopaminergic neurons and to correlate this indication of dopaminergic function with morphological evidence of survival or loss of dopaminergic neurons in mature CSN cultures. Mature CSN cultures, at 44 days in vitro (DIV), were exposed to either MPP+ alone, deprenyl alone or simultaneously to both deprenyl and MPP+ or to MPP+ following 4 day pretreatment with deprenyl. Exposure to MPP+ alone caused significant reduction in HVA levels, evidence of widespread injury and ultimate disappearance of large neurons in the cultures. These effects were attenuated by simultaneous exposure to MPP+ and deprenyl and the destructive effects of MPP+ appeared to be prevented by pretreatment with deprenyl. Thus the neuroprotective effects of deprenyl on MPP(+)-induced reduction of HVA levels in living cultures appears similar to the effects of deprenyl on dopamine levels and tyrosine hydroxylase activity reported by others in cultures previously exposed to deprenyl and MPP+. These studies also confirm that the neuroprotective effects of deprenyl against MPP+ in dopaminergic neurons are, at least in part, independent of deprenyl's inhibition of MAO-B.