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.

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.

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.

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.

A homozygous KCNJ10 mutation in Jack Russell Terriers and related breeds with spinocerebellar ataxia with myokymia, seizures, or both.

BACKGROUND: Juvenile-onset spinocerebellar ataxia has been recognized in Jack Russell Terriers and related Russell group terriers (RGTs) for over 40 years. Ataxia occurs with varying combinations of myokymia, seizures, and other signs of neurologic disease. More than 1 form of the disease has been suspected. HYPOTHESIS/OBJECTIVES: The objective was to identify the mutation causing the spinocerebellar ataxia associated with myokymia, seizures, or both and distinguish the phenotype from other ataxias in the RGTs. ANIMALS: DNA samples from 16 RGTs with spinocerebellar ataxia beginning from 2 to 12 months of age, 640 control RGTs, and 383 dogs from 144 other breeds along with the medical records of affected dogs were studied. METHODS: This case-control study compared the frequencies of a KCNJ10 allele in RGTs with spinocerebellar ataxia versus control RGTs. This allele was identified in a whole-genome sequence of a single RGT with spinocerebellar ataxia and myokymia by comparison to whole-genome sequences from 81 other canids that were normal or had other diseases. RESULTS: A missense mutation in the gene coding for the inwardly rectifying potassium channel Kir4.1 (KCNJ10:c.627C>G) was significantly (P < .001) associated with the disease. Dogs homozygous for the mutant allele all had spinocerebellar ataxia with varying combinations of myokymia and seizures. CONCLUSIONS AND CLINICAL IMPORTANCE: Identification of the KCNJ10 mutation in dogs with spinocerebellar ataxia with myokymia, seizures, or both clarifies the multiple forms of ataxia seen in these breeds and provides a DNA test to identify carriers.

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.

Elevated sedimentation on coral reefs adjacent to a beach nourishment project.

An increasingly common method to restore eroding beaches is nourishment, a process by which lost sand is replaced with terrestrial or offshore sediments to widen beaches. The southeastern Florida coastline contains shore-parallel coral reef communities adjacent to eroding beaches. Scleractinian corals and other reef-associated organisms are known to demonstrate sensitivity to elevated sedimentation levels. Sediment traps were used to examine spatio-temporal sedimentation patterns and assess the effects of nourishment (dredge and fill) activities. Several environmental variables correlated with among-site spatial variability of sediment parameters. Intra-annual variability correlated with wind velocity and direction. Nourishment activities showed localized effects, with sites in close proximity to dredging areas exhibiting significantly higher collection rates and lower percent fines than control sites. A regional increase in sedimentation occurred while nourishment activities were ongoing. Due to concurrent impacts of hurricanes, only one during-construction sampling interval revealed substantially higher collection rates relative to corresponding pre-construction sampling intervals.

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.

Measurement of the neutron lifetime using a proton trap.

We report a new measurement of the neutron decay lifetime by the absolute counting of in-beam neutrons and their decay protons. Protons were confined in a quasi-Penning trap and counted with a silicon detector. The neutron beam fluence was measured by capture in a thin 6LiF foil detector with known absolute efficiency. The combination of these simultaneous measurements gives the neutron lifetime: tau(n)=(886.8+/-1.2[stat]+/-3.2[syst]) s. The systematic uncertainty is dominated by uncertainties in the mass of the 6LiF deposit and the 6Li(n,t) cross section. This is the most precise measurement of the neutron lifetime to date using an in-beam method.

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.

Affective verbal memory in patients with temporal lobe epilepsy.

Eighteen epileptic patients with unilateral temporal lobe epilepsy (9 left, 9 right) were evaluated with a verbal memory task involving recall of 2 stories, 1 with affective content and 1 that was neutral. A trend for better performance by the group with intact left hemispheres was found for a quantitative score of number of story units recalled. For a qualitative score of number of symbolic distortions, a main effect of affective load was found, such that more distortions were made for the story with affective than neutral content. This effect remained significant when the left temporal lobe epilepsy patients were analyzed separately and was not found for the right temporal lobe epilepsy patients alone. Additional analyses for the subset of 5 patients with left and 6 patients with right temporal lobectomies involving removal of the hippocampus and amygdala were in the same direction as the analyses for all 18 participants. These findings are consistent with other reports of material-specific memory deficits, such that verbal memory deficits are associated with left temporal lobe epilepsy. The differences between performance on the affective and neutral stories for the left and right temporal lobe epilepsy patients are discussed and related to the role of the amygdala in affective processing.

Operant-self-administration of ethanol in mice prenatally exposed to cocaine.

Prenatal drug exposure may affect postnatal response to the reinforcing effects of a broad array of drugs. This study investigated the effects of prenatal cocaine exposure on operant self-administration of ethanol. Eighteen male, C57BL/6J mice were divided into three groups. The first had been prenatally exposed to 30 mg/kg of cocaine twice per day while the other groups were offspring of mothers which were either pair fed and saline intubated or untreated. Once adults, the mice were trained and subsequently tested for reinforcement from ethanol. The prenatal cocaine group responded less than the two control groups, with the largest decrease during a progressive ratio schedule of reinforcement. There were no differences in responding as a function of ethanol concentrations. These findings suggest that prenatal exposure to cocaine may not affect reinforcement per se, but may decrease motivation, drive state or propensity to work for ethanol.

Cytoplasmic factors do not contribute to a maternal effect on ethanol teratogenesis.

Both maternal and fetal genetic factors influence variations in response to prenatal ethanol exposure. To assess the effect of maternal genotype on the incidence of ethanol teratogenesis, a reciprocal cross study was conducted in an animal mode using the relatively susceptible C57BL/6J (B6) and the relatively resistant DBA/2J (D2) inbred mice. This mating pattern produced four embryonic genotypes: true-bred B6B6 and D2D2 litters and hybrid B6D2 and D2B6 litters. To examine the role of maternal egg cytoplasm as the source of variation that could account for a maternal effect, B6D2 and D2B6 F1 females were mated back to B6 males, which produced two additional embryonic genotypes: B6D2.B6 and D2B6.B6. Dams were intubated with either 5.8 g/kg of ethanol or an isocaloric amount of maltose-dextrin on day 9 of pregnancy. On day 18 of pregnancy, dams were sacrificed, fetuses were removed, weighed, sexed, and examined for gross morphological malformations. Every other fetus within a litter was prepared for either skeletal or soft tissue analysis. Results showed a higher rate of teratogenesis in the B6D2 group compared to the genetically similar D2B6 group, which indicates an influence of maternal genotype on susceptibility to ethanol teratogenesis. The percentage of affected male and female fetuses did not differ, which suggests that sex-linked factors are not responsible for the maternal effect. The backcross B6D2.B6 and D2B6.B6 litters did not differ significantly for any measure of teratogenesis, suggesting that differences in maternally transmitted cytoplasmic material are not the cause of the maternal effect. Factors that could account for the maternal effect are differences in the maternal uterine environment and genomic imprinting. Separating maternal from fetal-mediated mechanisms responsible for susceptibility to ethanol teratogenesis is needed for identifying mothers and infants at risk.

Differential biological activities of mammalian Id proteins in muscle cells.

Id proteins are helix-loop-helix (HLH) transcription factors that lack DNA-binding domains. These proteins form inactive heterodimers with basic HLH (bHLH) factors, inhibiting their DNA-binding and transcriptional activities. Consistent with a proposed role for Id proteins as inhibitors of terminal differentiation, Id1 and Id3 have been shown to negatively regulate myogenesis in cultured muscle cells. Here we have investigated the possibility that Id2 and/or Id4 can act in a similar manner. Surprisingly, while overexpression of Id2 resulted in inhibition of differentiation of Sol 8 myoblast cells, overexpression of Id4 did not. Sol 8 cells stably transfected with Id4 showed no apparent changes in expression of muscle-specific genes upon differentiation. DNA-binding activities present at the muscle creatine kinase (MCK) enhancer E-box and transcription of the MCK enhancer were not altered in Id4-overexpressing cells, compared with vector-transfected cells. Id2 is also a more potent inhibitor of protein/DNA complex formation at the MCK-R enhancer E-box than Identified in vitro. Therefore, our data support the notion that members of the Id family might be involved in the regulation of distinct developmental pathways.

Ethanol teratogenesis in the C57BL/6J, DBA/2J, and A/J inbred mouse strains.

Research has shown variations in susceptibility to alcohol-related birth defects in humans. Genetic differences are one reason for this variability. This study compared three inbred mouse strains to determine whether they differ in their susceptibilities to ethanol teratogenesis because previous studies have generated conflicting data. Pregnant C57BL/6J (B6), DBA/2J (D2), and A/J (A) dams were intubated intragastrically with either an acute dose of ethanol (5.8 g/kg) or an isocaloric amount of maltose-dextrine on day 9 of pregnancy. Litters were removed on day 18 of pregnancy and examined for gross, soft-tissue, and skeletal malformations. Results showed that ethanol-exposed B6 litters had a higher percentage of digit (19%), kidney (24%), and skeletal (32%, mostly vertebral) malformations than their maltose-exposed controls (7% or below). Prenatal exposure to ethanol increased skeletal (68%, both rib and vertebral) malformations for A litters when compared to their maltose-exposed controls (4%), but did not increase digit or kidney malformations. Ethanol-exposed D2 litters did not differ from maltose-exposed controls. Maternal blood ethanol levels did not differ among the B6, D2, and A strains. These results provide additional evidence suggesting a genetic component to ethanol teratogenesis.

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.

Effects of acute prenatal ethanol administration in a reciprocal cross of C57BL/6J and short-sleep mice: maternal effects and nonmaternal factors.

An animal model was used to see if maternal genetic factors contribute to ethanol-induced fetal malformations. Susceptible C57BL/6J (B6) and resistant Short-Sleep (SS) mice were used in a reciprocal cross-breeding design. This design produced four fetal genotypes: true-bred B6B6 and SSSS liters and hybrid B6SS and SSB6 litters. Dams were intubated with either 5.8 g/kg of ethanol or an isocaloric amount of maltose-dextrin on day 9 of pregnancy. Fetuses were removed on day 18 of pregnancy and assessed for soft tissue and gross malformations. Results show different prenatal treatment effects on malformations depending on the fetal genotype. Mean percentage of total malformations in ethanol-treated groups were 61% (B6B6), 28% (B6SS), 3% (SSB6), and 16% (SSSS), respectively. Litters exposed to maltose-dextrin showed low (< or = 4%) malformation rates regardless of fetal genotype. The difference in teratogenic response between genetically identical hybrid litters (B6SS and SSB6) suggests a maternal genetic contribution to susceptibility. Of the two major types of malformations, forelimb but not kidney malformations showed a pattern characteristic of a maternal genetic effect. It was concluded that maternal genetic effects can be distinguished from fetal genetic effects responsible for ethanol teratogenesis. Genetically based physiological responses to alcohol in the mother may confer varying degrees of risk for prenatal alcohol effects in the child.

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.