Precision determination of absolute neutron flux.

A technique for establishing the total neutron rate of a highly-collimated monochromatic cold neutron beam was demonstrated using an alpha-gamma counter. The method involves only the counting of measured rates and is independent of neutron cross sections, decay chain branching ratios, and neutron beam energy. For the measurement, a target of 10B-enriched boron carbide totally absorbed the neutrons in a monochromatic beam, and the rate of absorbed neutrons was determined by counting 478 keV gamma rays from neutron capture on 10B with calibrated high-purity germanium detectors. A second measurement based on Bragg diffraction from a perfect silicon crystal was performed to determine the mean de Broglie wavelength of the beam to a precision of 0.024%. With these measurements, the detection efficiency of a neutron monitor based on neutron absorption on 6Li was determined to an overall uncertainty of 0.058%. We discuss the principle of the alpha-gamma method and present details of how the measurement was performed including the systematic effects. We also describe how this method may be used for applications in neutron dosimetry and metrology, fundamental neutron physics, and neutron cross section measurements.

Golden Retriever dogs with neuronal ceroid lipofuscinosis have a two-base-pair deletion and frameshift in CLN5.

We studied a recessive, progressive neurodegenerative disease occurring in Golden Retriever siblings with an onset of signs at 15 months of age. As the disease progressed these signs included ataxia, anxiety, pacing and circling, tremors, aggression, visual impairment and localized and generalized seizures. A whole genome sequence, generated with DNA from one affected dog, contained a plausibly causal homozygous mutation: CLN5:c.934_935delAG. This mutation was predicted to produce a frameshift and premature termination codon and encode a protein variant, CLN5:p.E312Vfs*6, which would lack 39 C-terminal amino acids. Eighteen DNA samples from the Golden Retriever family members were genotyped at CLN5:c.934_935delAG. Three clinically affected dogs were homozygous for the deletion allele; whereas, the clinically normal family members were either heterozygotes (n = 11) or homozygous for the reference allele (n = 4). Among archived Golden Retrievers DNA samples with incomplete clinical records that were also genotyped at the CLN5:c.934_935delAG variant, 1053 of 1062 were homozygous for the reference allele, 8 were heterozygotes and one was a deletion-allele homozygote. When contacted, the owner of this homozygote indicated that their dog had been euthanized because of a neurologic disease that progressed similarly to that of the affected Golden Retriever siblings. We have collected and stored semen from a heterozygous Golden Retriever, thereby preserving an opportunity for us or others to establish a colony of CLN5-deficient dogs.

Characterization of neuronal ceroid-lipofuscinosis in 3 cats.

Three young domestic shorthair cats were presented for necropsy with similar histories of slowly progressive visual dysfunction and neurologic deficits. Macroscopic examination of each cat revealed cerebral and cerebellar atrophy, dilated lateral ventricles, and slight brown discoloration of the gray matter. Histologically, there was bilateral loss of neurons within the limbic, motor, somatosensory, visual, and, to a lesser extent, vestibular systems with extensive astrogliosis in the affected regions of all 3 cases. Many remaining neurons and glial cells throughout the entire central nervous system were distended by pale yellow to eosinophilic, autofluorescent cytoplasmic inclusions with ultrastructural appearances typical of neuronal ceroid-lipofuscinoses (NCLs). Differences in clinical presentation and neurological lesions suggest that the 3 cats may have had different variants of NCL. Molecular genetic characterization in the 1 cat from which DNA was available did not reveal any plausible disease-causing mutations of the CLN1 (PPT1), CLN3, CLN5, CLN8, and CLN10 (CTSD) genes. Further investigations will be required to identify the mutations responsible for NCLs in cats.

Improved determination of the neutron lifetime.

The most precise determination of the neutron lifetime using the beam method was completed in 2005 and reported a result of τ(n)=(886.3±1.2[stat]±3.2[syst]) s. The dominant uncertainties were attributed to the absolute determination of the fluence of the neutron beam (2.7 s). The fluence was measured with a neutron monitor that counted the neutron-induced charged particles from absorption in a thin, well-characterized 6Li deposit. The detection efficiency of the monitor was calculated from the areal density of the deposit, the detector solid angle, and the evaluated nuclear data file, ENDF/B-VI 6Li(n,t)4He thermal neutron cross section. In the current work, we measure the detection efficiency of the same monitor used in the neutron lifetime measurement with a second, totally absorbing neutron detector. This direct approach does not rely on the 6Li(n,t)4He cross section or any other nuclear data. The detection efficiency is consistent with the value used in 2005 but is measured with a precision of 0.057%, which represents a fivefold improvement in the uncertainty. We verify the temporal stability of the neutron monitor through ancillary measurements, allowing us to apply the measured neutron monitor efficiency to the lifetime result from the 2005 experiment. The updated lifetime is τ(n)=(887.7±1.2[stat]±1.9[syst]) s.

Nonpneumonic, short-incubation-period Legionellosis (Pontiac fever) in men who cleaned a steam turbine condenser.

Pontiac fever affected ten men who had cleaned a steam turbine condenser with compressed air. Previous epidemics of Pontiac fever and Legionnaires' disease--both caused by Legionella Pneumophila (proposed sp. nov.)--involved "airborne spread" from air-conditioning cooling towers or evaporative condensers. Aerosols of contaminated water in heat-rejection systems appear to be important sources of epidemic legionellosis.

Australian Cattle Dogs with Neuronal Ceroid Lipofuscinosis are Homozygous for a CLN5 Nonsense Mutation Previously Identified in Border Collies.

BACKGROUND: Neuronal ceroid lipofuscinosis (NCL), a fatal neurodegenerative disease, has been diagnosed in young adult Australian Cattle Dogs. OBJECTIVE: Characterize the Australian Cattle Dog form of NCL and determine its molecular genetic cause. ANIMALS: Tissues from 4 Australian Cattle Dogs with NCL-like signs and buccal swabs from both parents of a fifth affected breed member. Archived DNA samples from 712 individual dogs were genotyped. METHODS: Tissues were examined by fluorescence, electron, and immunohistochemical microscopy. A whole-genome sequence was generated for 1 affected dog. A TaqMan allelic discrimination assay was used for genotyping. RESULTS: The accumulation of autofluorescent cytoplasmic storage material with characteristic ultrastructure in tissues from the 4 affected dogs supported a diagnosis of NCL. The whole-genome sequence contained a homozygous nonsense mutation: CLN5:c.619C>T. All 4 DNA samples from clinically affected dogs tested homozygous for the variant allele. Both parents of the fifth affected dog were heterozygotes. Archived DNA samples from 346 Australian Cattle Dogs, 188 Border Collies, and 177 dogs of other breeds were homozygous for the reference allele. One archived Australian Cattle Dog sample was from a heterozygote. CONCLUSIONS AND CLINICAL IMPORTANCE: The homozygous CLN5 nonsense is almost certainly causal because the same mutation previously had been reported to cause a similar form of NCL in Border Collies. Identification of the molecular genetic cause of Australian Cattle Dog NCL will allow the use of DNA tests to confirm the diagnosis of NCL in this breed.

The Fundamental Neutron Physics Facilities at NIST.

The program in fundamental neutron physics at the National Institute of Standards and Technology (NIST) began nearly two decades ago. The Neutron Interactions and Dosimetry Group currently maintains four neutron beam lines dedicated to studies of fundamental neutron interactions. The neutrons are provided by the NIST Center for Neutron Research, a national user facility for studies that include condensed matter physics, materials science, nuclear chemistry, and biological science. The beam lines for fundamental physics experiments include a high-intensity polychromatic beam, a 0.496 nm monochromatic beam, a 0.89 nm monochromatic beam, and a neutron interferometer and optics facility. This paper discusses some of the parameters of the beam lines along with brief presentations of some of the experiments performed at the facilities.

Measurement of the Neutron Lifetime by Counting Trapped Protons.

We measured the neutron decay lifetime by counting in-beam neutron decay recoil protons trapped in a quasi-Penning trap. The absolute neutron beam fluence was measured by capture in a thin (6)LiF foil detector with known efficiency. The combination of these measurements gives the neutron lifetime: τ n = (886.8 ± 1.2 ± 3.2) s, where the first (second) uncertainty is statistical (systematic) in nature. This is the most precise neutron lifetime determination to date using an in-beam method.

Hypercapnic acidosis and dimethyl amiloride reduce reperfusion induced cell death in ischaemic ventricular myocardium.

OBJECTIVE: The aim was to investigate the effects of slowing the recovery of ischaemia induced intracellular acidosis with hypercapnic acidosis or dimethyl amiloride (DMA) on the extent of reperfusion induced cell death. METHODS: Isolated arterially perfused rabbit papillary muscles and septa were suspended in a controlled atmosphere and perfused with a modified Tyrode solution containing erythrocytes and trypan blue (500 microM). Ischaemia was produced by arrest of perfusion and withdrawal of atmospheric O2. Extracellular pH of the muscle during reperfusion was controlled by adjusting the pH of the perfusate (pH 6.6 or pH 7.6 with and without DMA 20 microM) and changing the PCO2 of the chamber atmosphere. After 30 min of reperfusion following 30 min (group A) or 60 min (group B) of ischaemia, papillary muscles were fixed with paraformaldehyde. Cell death was assessed by trypan blue staining of nuclei in histological sections of the papillary muscles. RESULTS: The magnitude of cell death was greatest after reperfusion with pH 7.6 as measured by the percentage of nuclei staining with trypan blue (15.1% in group A; 41.8% in group B). By contrast, reperfusion at pH 6.6 reduced cell killing (group A, 3.6%; group B, 7.2%). Reperfusion at pH 7.6 with DMA (20 microM) also reduced trypan blue uptake (group A, 2.8%; group B, 3.8%). Despite the attenuation of cell death afforded by acidosis or Na+/H+ exchange inhibition, significant swelling of the extracellular space and microvascular injury was noted. CONCLUSIONS: Hypercapnic acidosis and Na+/H+ exchange inhibition during reperfusion attenuate lethal reperfusion injury to ventricular myocardium and extend to the intact myocardium the concept of the "pH paradox" in which recovery of intracellular pH after reperfusion is a precipitating factor in lethal cell injury.

Regulation of triple-negative breast cancer cell metastasis by the tumor-suppressor liver kinase B1.

Liver kinase B1 (LKB1), also known as serine/threonine kinase 11 (STK11), has been identified as a tumor suppressor in many cancers including breast. Low LKB1 expression has been associated with poor prognosis of breast cancer patients, and we report here a significant association between loss of LKB1 expression and reduced patient survival specifically in the basal subtype of breast cancer. Owing to the aggressive nature of the basal subtype as evidenced by high incidences of metastasis, the purpose of this study was to determine if LKB1 expression could regulate the invasive and metastatic properties of this specific breast cancer subtype. Induction of LKB1 expression in basal-like breast cancer (BLBC)/triple-negative breast cancer cell lines, MDA-MB-231 and BT-549, inhibited invasiveness in vitro and lung metastatic burden in an orthotopic xenograft model. Further analysis of BLBC cells overexpressing LKB1 by unbiased whole transcriptomics (RNA-sequencing) revealed striking regulation of metastasis-associated pathways, including cell adhesion, extracellular matrix remodeling, and epithelial-to-mesenchymal transition (EMT). In addition, LKB1 overexpression inhibited EMT-associated genes (CDH2, Vimentin, Twist) and induced the epithelial cell marker CDH1, indicating reversal of the EMT phenotype in the MDA-MB-231 cells. We further demonstrated marked inhibition of matrix metalloproteinase 1 expression and activity via regulation of c-Jun through inhibition of p38 signaling in LKB1-expressing cells. Taken together, these data support future development of LKB1 inducing therapeutics for the suppression of invasion and metastasis of BLBC.

Multiple enhancer elements mediate induction of c-fos in vascular smooth muscle cells.

Previous work from this and other laboratories has demonstrated that the vasoconstrictor peptide angiotensin II results in hypertrophy of rat aortic smooth muscle cells that is associated with an increase in transcription of the early growth response gene c-fos. To explore the molecular mechanism responsible for c-fos induction in rat aortic smooth muscle cells, we used a series of reporter constructs linked to the chloramphenicol acetyl transferase gene in transient transfection experiments in rat aortic smooth muscle cells. Constructs containing both the serum response element and cAMP response element exhibited a 20-fold increase in chloramphenicol acetyl transferase activity in response to either serum or angiotensin II, whereas no increase was seen in vehicle-treated cells. Mutations in either the serum response element or cAMP response element alone, which have been demonstrated to inactivate these elements in other cell types, had no effect on chloramphenicol acetyl transferase inducibility. In contrast, if both elements were mutated, inducibility was almost abolished. Electrophoretic mobility shift assays with oligonucleotides corresponding to either serum response element or cAMP response element demonstrated that these oligonucleotides are capable of forming specific complexes with proteins from rat aortic smooth muscle cell nuclear extracts. One of the proteins binding to the serum response element is the previously described serum response factor, since it was supershifted by a monospecific antibody. These studies demonstrate that c-fox induction in smooth muscle occurs by a dual mechanism that can activate transcription via the serum response element or cAMP response element. These elements appear to act equally and independently, involving a distinct set of transacting factors.

Nicotine-produced relearning deficit in C57BL/6J and DBA/2J mice.

Acute nicotine administration has been shown to influence the acquisition and retention of learning tasks. In order to investigate the many possible behavioral and pharmacological effects of nicotine, a modified 2 X 2 state-dependent learning design was used to assess nicotine's effects on active avoidance learning. Male and female mice of the C57BL/6J (C57) and DBA/2J (DBA) inbred strains were injected with a control solution or with 0.5, 1.0, or 2.0 mg/kg nicotine 5 min before the start of training and, following a 24-h period, 5 min before retraining. Nicotine had no effect on the acquisition of the learning task but, depending on strain and sex, did have an effect on relearning. Relearning in the C57 males was unaffected by nicotine injection, whereas the most prominent effect of nicotine in the C57 females and the DBA males and females was a retrieval deficit. The prevalence of a nicotine-induced retrieval deficit in the present experiment suggests that those mechanisms underlying the retrieval of previously learned information are, in part, mediated or modulated by perturbations within nicotine-sensitive areas of the central nervous system.

Long-term consequences of prenatal cocaine exposure on biogenic amines in the brains of mice: the role of sex.

Prenatal cocaine exposure leads to multiple abnormalities in the mature offspring. We explored the effects of gestational exposure to cocaine on neurotransmitter systems of adult mice. The subjects were the mature offspring of mice (a) prenatally fed cocaine between gestational day (G) 8 and G19, (b) pair-fed chow and water, or fed chow and water ad libitum. The forebrains of the mature offspring were assayed for monoamines and amino acids. Cocaine exposure particularly affected the dopaminergic system and in a sex-specific manner. In males dopamine concentrations were decreased and dopamine turnover was increased, whereas in females dopamine concentrations were increased and turnover was decreased. Neither norepinephrine, the serotonergic system, nor neuroactive amino acids (or their precursors) were affected by cocaine. Thus, in utero exposure to cocaine produces long-lasting, specific defects in the dopaminergic system.

Differential effects of MK-801 on cerebrocortical neuronal injury in C57BL/6J, NSA, and ICR mice.

1. Antagonists of the N-methyl-D-aspartate (NMDA) glutamate (Glu) receptor, including [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate], dizocilpine maleate (MK-801), injure pyramidal neurons in the posterior cingulate/retrosplenial (PC/RS) cortex when administered systemically to adult rats and mice. 2. These results have, to our knowledge, only been reported previously in Harlan Sprague Dawley albino rats and International Cancer Research (ICR) mice, an outbred albino strain. 3. Male Non-Swiss Albino (NSA) mice, an albino outbred strain, and male C57BL/6J (B6) mice, a pigmented inbred strain, were injected systemically with 1 mg/kg of MK-801 in the first experiment. This dose of MK-801 reliably produces cytoplasmic vacuoles in neurons in layers III and IV of the PC/RS cortex in 100% of ICR mice treated 4. There was a significant difference in the number of vacuolated neurons in B6 and NSA mice, as assessed by ANOVA. The NSA were not significantly different than previously examined ICR mice, but the B6 had fewer vacuolated neurons than either of the two outbred strains. 5. In the second experiment, male NSA, ICR, and B6 mice were injected systemically with a high dose, 10 mg/kg, of MK-801. This dose has been demonstrated to result in necrosis in the same population of neurons injured by lower doses of MK-801. 6. An ANOVA indicated that there was a significant difference among the three strains of mice, and a Fisher's protected t revealed that the B6 mice were significantly different from both the NSA and ICR, but that, with our test, those two strains were indistinguishable. 7. Male ICR, NSA, and B6 mice were tested in the holeboard food search task 5 hours after 1 mg/kg of MK-801. There were significant differences between the strains in performance both pre and posttreatment. The effect of the drug was not statistically significant. 8. These results suggest that there may be a genetically mediated difference in the reaction to NMDA receptor antagonists, a finding which may be important given the NMDA receptor hypofunction hypothesis for the etiology of schizophrenic symptoms.

Choristomas of the seventh and eighth cranial nerves.

Choristomas, masses of normal tissues in aberrant locations, contain smooth muscle fibers and fibrous tissues. We describe the MR imaging features of two choristomas located in the internal auditory canals and arising from the facial and vestibulocochlear nerves. Both lesions enhanced with contrast material. In one case, enhancement was seen in the geniculate ganglion and greater superficial petrosal nerve. In the other, a medial component enhanced less than the lateral component did.

Genetic influences on brain growth restriction induced by development exposure to alcohol.

Genetic factors have been implicated as contributing to the considerable variation in the severity of alcohol-related birth defects in offspring of women who drink heavily during pregnancy. Two animal models of alcohol-related developmental effects incorporated different behavior genetic approaches to examine genetic influences on brain and body growth following alcohol exposure during development. The first, extensively developed in Sprague-Dawley rats, examined the effects of three doses of alcohol administered to two inbred rat strains (MR and M520) via artificial-rearing procedures during the early postnatal brain growth spurt. In both strains, alcohol produced a dose-dependent restriction of brain weight (but not body weight) on postnatal day 10, compared to artificially reared controls. The MR strain was more susceptible to cerebellar growth restriction than the M520 strain, an effect not attributable to strain differences in blood alcohol concentrations. In the second model, pregnant female Long-sleep and Short-sleep mice, selectively bred for differences in initial sensitivity to the hypnotic effects of acute alcohol administration, were intubated with ethanol from gestational days 7-18. Controls included either sucrose or maltose/dextrin intubation controls and non-intubated controls. The LS offspring showed growth deficits and brain weight reductions in adulthood, while the SS offspring were resistant to these detrimental effects of the prenatal alcohol exposure. Thus, differences in either maternal or fetal genotype may contribute to individual differences in the severity of the effects of alcohol exposure during development.

Differential effects of ethanol concentration on blood pH, PCO2 and PO2 in LS and SS mice.

Concentration-dependent effects of ethanol upon behavior and upon physiological regulatory mechanisms have been suggested. In a previous study, we found that the concentration of an acute ethanol injection confounded dose-response relationships for measures of blood pH, PCO2, and PO2. Two lines of mice that differ in CNS sensitivity to the hypnotic effects of ethanol (long sleep, LS; short sleep, SS) have also been found to differ in sensitivity to the physiological depressant effects of this drug. Therefore, we designed the present study to examine how intraperitoneal (IP) injections of varying ethanol concentrations differentially affect blood parameters (pH, PCO2, and PO2) and respiration rate in the LS and SS mouse lines. Different groups of LS female mice were injected IP with 165.0, 198.8, 248.1, or 330.0 mg/ml ethanol at a constant dose of 3.3 g/kg. Groups of SS female mice received 205.0, 247.0, 308.3, or 410.0 mg/ml ethanol at a dose of 4.1 g/kg. Blood parameters and respiration rate were measured at 60 min post-injection. In both the LS and SS mice, increasing concentrations of ethanol caused a progressive decline in respiration rate and blood pH. Blood PCO2 values were greater than control only at the highest ethanol concentration. Concentration-dependent effects of ethanol on blood PO2 values were not found in either line. However, LS PO2 was significantly elevated from the control value at all ethanol concentrations. These results suggest that a dose-response relationship may be obtained by varying ethanol concentration for some physiological measures, but not for others. Thus, attention should be paid to differences in concentration as well as amount of ethanol when dose-response curves are to be constructed.

Acute ethanol effects on blood pH, PCO2, and PO2 in LS and SS mice.

The effects of ethanol on blood pH, PCO2, and PO2 were measured in LS and SS mice in an attempt to ascertain whether these lines of mice, which differ in CNS sensitivity to the behavioral effects of ethanol, also differ in sensitivity to physiological effects of this drug. Long-sleep (LS) female mice were injected intraperitoneally with 1.8, 2.5, 3.3, or 3.8 g/kg ethanol; short-sleep (SS) female mice were administered 2.5, 3.3, 4.1, or 4.7 g/kg. Blood pH, PCO2, and PO2 were assessed at 15, 30, 60, 120, or 180 min after injection of the 2.5 and 4.1 g/kg doses or at 60 min after injection of the 1.8, 2.5, 3.3, 3.8, 4.1, and 4.7 g/kg doses. Opposite effects on blood pH and PCO2 over time were obtained in LS and SS mice at the 2.5 g/kg dose. Acidosis characterized the LS line, whereas alkalosis characterized the SS. The results obtained with SS mice at 4.1 g/kg dose were similar to those obtained with LS mice at the 2.5 g/kg dose. The dose-response curve for the SS mice generated at 60 min post-injection lies to the right of that for the LS mice. The effects of high ethanol doses on SS mice resemble the effects of low doses on LS animals. Thus, the two lines of mice differ in response to the effects of ethanol on these parameters related to respiration. The difference in sensitivity to the respiratory depressant effects of ethanol may contribute to the differences in behavioral sensitivity between the two lines.

Circadian and genetic influences on tissue sensitivity and sleep time to ethanol in LS and SS mice.

Circadian variations in response to ethanol were studied in long-sleep (LS) and short-sleep (SS) mice. Each LS animal received a 2.5 g/kg intraperitoneal ethanol injection, while the SS animals were injected with 4.1 or 5.0 g/kg. Different groups of mice were assessed for sleep time, waking blood alcohol concentration (BAC), and waking brain ethanol concentration (BREC) at 03.00, 09.00, 15.00, or 21.00 hr. Sleep times, waking BACs, and waking BRECs showed circadian variations in the LS mice. SS animals given the 4.1 g/kg dose showed circadian variations for waking BAC and waking BREC, but not for sleep time. The observed variations in the physiological parameters for these animals may have been confounded by a short sleep time so that they reflected circadian variations in drug absorption and/or distribution rather than in CNS sensitivity. SS mice given the 5.0 g/kg dose slept longer than those given the 4.1 g/kg dose and did not show circadian variations for sleep time, waking BAC, or waking BREC. These results suggest both circadian and genetic influences on tissue sensitivity to ethanol.

Fetal alcohol effects in long- and short-sleep mice: activity, passive avoidance, and in utero ethanol levels.

Genetic differences in susceptibility to fetal alcohol effects (FAE) have been suggested by both human and animal studies. The Long-Sleep (LS) and Short-Sleep (SS) mouse lines, selectively bred for differences in ethanol-induced narcosis, provide a model for studying differential alcohol sensitivity in the etiology of FAE. LS and SS mice were intubated with either 2.9 g/kg (20% w/v) ethanol (E) or an isocaloric amount of sucrose (S) twice per day (6 hr apart) on Days 7 through 15 of pregnancy. An untreated control group (C) was maintained for each line. Offspring were fostered to lactating Rockland-Swiss mice at birth. LS offspring prenatally exposed to ethanol exhibited increased open-field activity relative to LS controls, but this effect was due to the overactivity of one litter. Activity for SS mice prenatally exposed to ethanol did not differ from control levels. Ethanol content in blood (280 mg/dl), amniotic fluid (258 mg/dl), and fetal tissue (230 mg/dl) did not differ in similarly treated LS and SS dams. In a second experiment, females were treated from Days 7 through 18 of gestation, and their offspring were tested for either open-field activity or passive avoidance learning. There were no group differences in open-field activity, but LS mice prenatally exposed to alcohol took more trials to reach a passive avoidance criterion than their controls, whereas similarly treated SS mice did not differ from controls. These results suggest that genetically-mediated sensitivity to ethanol influences susceptibility to FAE and that this may be task specific.