Development of wavelength calibration techniques for high-resolution x-ray imaging crystal spectrometers on the EAST tokamak.

Newly developed large-area pixelated two-dimensional detector and two-crystal assemblies were deployed for the first time on tokamaks to enable time-resolved Bragg-diffracted x-ray imaging with good framing rate and water-cooling capabilities for in-vacuum long-pulse operations. High-quality helium-like (He-like) and hydrogen-like (H-like) argon spectra have been observed simultaneously for the first time on a single detector for a wide range of plasma parameters to infer both ion temperature and rotation profiles and support studies on spontaneous rotation, impurity transport, and RF physics. Since tokamak plasmas rotate in both the poloidal (θ) and toroidal (ϕ) directions, a reliable wavelength calibration is needed to account for the correct Doppler shift as well as to compute the spectrometer's instrumental function. Lyα lines emitted from Cd x-ray tubes are proposed to be used as "markers" to provide an in situ calibration of the EAST's X-ray imaging crystal spectrometer systems measuring He- and H-like argon spectra. The first lab test indicated that the X-ray tube can excite strong Lyα lines at 15 kV voltage and 1 mA current when the crystal is shined for 10 min. Other indirect calibration methods using locked-mode discharge scenarios were also studied as complementary methods.

Upgrade of X-ray crystal spectrometer for high temperature measurement using neon-like xenon lines on EAST.

A two-crystal X-ray spectrometer system has been implemented in the EAST tokamak to simultaneously diagnose high- and low-temperature plasmas using He- and H-like argon spectra. But for future fusion devices like ITER and Chinese Fusion Engineering Test Reactor (CFETR), argon ions become fully stripped in the core and the intensity of the H-like lines will be significantly at high temperatures (Te > 5 keV). With increasing auxiliary heating power on EAST, the core plasma temperature could also reach 5 keV and higher. In such conditions, the use of a xenon puff becomes an appropriate choice for both ion-temperature and flow-velocity measurements. A new two-crystal system using a quartz 110 crystal (2d = 4.913 Å) to view He-like argon lines and a quartz 011 crystal (2d = 6.686 Å) to view Ne-like xenon spectra has been deployed on a poloidal X-ray crystal spectrometer. While the He-like argon spectra will be used to measure the plasma temperature in the edge plasma region, the Ne-like xenon spectra will be used for measurement in the hot core. The new crystal arrangement allows a wide temperature measurement ranging from 0.5 to 10 keV or even higher, being the first tests for burning plasmas like ITER and CFETR. The preliminary result of lab-tests, Ne-like xenon lines measurement will be presented.

Fast-ion Dα spectrum diagnostic in the EAST.

In toroidal magnetic fusion devices, fast-ion D-alpha diagnostic (FIDA) is a powerful method to study the fast-ion feature. The fast-ion characteristics can be inferred from the Doppler shifted spectrum of Dα light according to charge exchange recombination process between fast ions and probe beam. Since conceptual design presented in the last HTPD conference, significant progress has been made to apply FIDA systems on the Experimental Advanced Superconducting Tokamak (EAST). Both co-current and counter-current neutral beam injectors are available, and each can deliver 2-4 MW beam power with 50-80 keV beam energy. Presently, two sets of high throughput spectrometer systems have been installed on EAST, allowing to capture passing and trapped fast-ion characteristics simultaneously, using Kaiser HoloSpec transmission grating spectrometer and Bunkoukeiki FLP-200 volume phase holographic spectrometer coupled with Princeton Instruments ProEM 1024B eXcelon and Andor DU-888 iXon3 1024 CCD camera, respectively. This paper will present the details of the hardware descriptions and experimental spectrum.

Measurement of helium-like and hydrogen-like argon spectra using double-crystal X-ray spectrometers on EAST.

A two-crystal assembly was deployed on the tangential X-ray crystal spectrometer to measure both helium-like and hydrogen-like spectra on EAST. High-quality helium-like and hydrogen-like spectra were observed simultaneously for the first time on one detector for a wide range of plasma parameters. Profiles of line-integrated core ion temperatures inferred from two spectra were consistent. Since tungsten was adopted as the upper divertor material, one tungsten line (W XLIV at 4.017 Å) on the short-wavelength side of the Lyman-α line (Lα1) was identified for typical USN discharges, which was diffracted by a He-like crystal (2d = 4.913 Å). Another possible Fe XXV line (1.85 Å) was observed to be located on the long-wavelength side of resonance line (w), which was diffracted from a H-like crystal (2d = 4.5622 Å) on the second order. Be-like argon lines were also observable that fill the detector space between the He-like and H-like spectra.

The prediction of human oral absorption for diffusion rate-limited drugs based on heuristic method and support vector machine.

Support vector machine (SVM), as a novel machine learning technique, was used for the prediction of the human oral absorption for a large and diverse data set using the five descriptors calculated from the molecular structure alone. The molecular descriptors were selected by heuristic method (HM) implemented in CODESSA. At the same time, in order to show the influence of different molecular descriptors on absorption and to well understand the absorption mechanism, HM was used to build several multivariable linear models using different numbers of molecular descriptors. Both the linear and non-linear model can give satisfactory prediction results: the square of correlation coefficient R(2) was 0.78 and 0.86 for the training set, and 0.70 and 0.73 for the test set respectively. In addition, this paper provides a new and effective method for predicting the absorption of the drugs from their structures and gives some insight into structural features related to the absorption of the drugs.

Treatment with atypical antipsychotics: new indications and new populations.

Atypical antipsychotics have revolutionized the treatment of schizophrenia, becoming the treatment of choice for patients not only during their first episode, but also throughout their life course. Of note, as of 1999 more than 70% of prescriptions for these drugs are being prescribed for conditions other than schizophrenia, such as bipolar disorder and geriatric agitation. While there have been very few controlled trials that have established the efficacy of the atypical antipsychotics for these "off-label" uses, there have been a large number of open trials and case reports. The few controlled trials suggest that the atypical antipsychotics may be useful for affective disorders (both mania and depression), geriatric conditions such as senile dementia and aggression, as well as a variety of other disorders. Atypical agents may be particularly helpful for elderly, child, or adolescent patients who are especially susceptible to the side effects of medications and whose risk of tardive dyskinesia is high but further controlled studies are necessary.

Psychopharmacologic treatment strategies for depression, bipolar disorder, and schizophrenia.

Patients with serious psychiatric disorders are frequently treated by primary care physicians, who may have difficulty keeping up with recent advances in psychiatry. This paper presents an updated synopsis for three major psychiatric illnesses: major depression, bipolar disorder, and schizophrenia. Current definitions, updated diagnostic criteria, short- and long-term treatment strategies with algorithms, and special challenges for the clinician are discussed for each of these illnesses. On the basis of each illness's distinct characteristics, five treatment principles are emphasized: 1) Treatment strategies should be long-term and should emphasize adherence, 2) treatment choice should be empirical, 3) combinations of medications may be helpful, 4) a combination of psychosocial and pharmacologic treatments may be more useful than either alone, and 5) the family or "significant others" as well as a consumer organization need to be involved. Some of the new directions in dinical research to refine these strategies and meet these challenges are also described.

Targeted disruption of the Kvlqt1 gene causes deafness and gastric hyperplasia in mice.

The KvLQT1 gene encodes a voltage-gated potassium channel. Mutations in KvLQT1 underlie the dominantly transmitted Ward-Romano long QT syndrome, which causes cardiac arrhythmia, and the recessively transmitted Jervell and Lange-Nielsen syndrome, which causes both cardiac arrhythmia and congenital deafness. KvLQT1 is also disrupted by balanced germline chromosomal rearrangements in patients with Beckwith-Wiedemann syndrome (BWS), which causes prenatal overgrowth and cancer. Because of the diverse human disorders and organ systems affected by this gene, we developed an animal model by inactivating the murine Kvlqt1. No electrocardiographic abnormalities were observed. However, homozygous mice exhibited complete deafness, as well as circular movement and repetitive falling, suggesting imbalance. Histochemical study revealed severe anatomic disruption of the cochlear and vestibular end organs, suggesting that Kvlqt1 is essential for normal development of the inner ear. Surprisingly, homozygous mice also displayed threefold enlargement by weight of the stomach resulting from mucous neck cell hyperplasia. Finally, there were no features of BWS, suggesting that Kvlqt1 is not responsible for BWS.

Cbl-b regulates the CD28 dependence of T-cell activation.

Whereas co-stimulation of the T-cell antigen receptor (TCR) and CD28 triggers T-cell activation, stimulation of the TCR alone may result in an anergic state or T-cell deletion, both possible mechanisms of tolerance induction. Here we show that T cells that are deficient in the adaptor molecule Cbl-b (ref. 3) do not require CD28 engagement for interleukin-2 production, and that the Cbl-b-null mutation (Cbl-b(-/-)) fully restores T-cell-dependent antibody responses in CD28-/- mice. The main TCR signalling pathways, such as tyrosine kinases Zap-70 and Lck, Ras/mitogen-activated kinases, phospholipase Cgamma-1 and Ca2+ mobilization, were not affected in Cbl-b(-/-) T cells. In contrast, the activation of Vav, a guanine nucleotide exchange factor for Rac1/Rho/CDC42, was significantly enhanced. Our findings indicate that Cbl-b may influence the CD28 dependence of T-cell activation by selectively suppressing TCR-mediated Vav activation. Mice deficient in Cbl-b are highly susceptible to experimental autoimmune encephalomyelitis, suggesting that the dysregulation of signalling pathways modulated by Cbl-b may also contribute to human autoimmune diseases such as multiple sclerosis.

Altered thymic positive selection and intracellular signals in Cbl-deficient mice.

Cbl is the product of the protooncogene c-cbl and is involved in T cell antigen receptor (TCR)-mediated signaling. To understand the role of Cbl for immune system development and function, we generated a Cbl-deficient mouse strain. In Cbl-deficient mice, positive selection of the thymocytes expressing major histocompatibility complex class II-restricted transgenic TCR was significantly enhanced. Two factors may have contributed to the altered thymic selection. First, Cbl deficiency markedly up-regulated the activity of ZAP-70 and mitogen-activated protein kinases. The mitogen-activated protein kinase pathway was shown previously to be involved in thymic positive selection. Second, Cbl-deficient thymocytes expressed CD3 and CD4 molecules at higher levels, which consequently may increase the avidity of TCR/major histocompatibility complex/coreceptor interaction. Thus, Cbl plays a novel role in modulating TCR-mediated multiple signaling pathways and fine-tunes the signaling threshold for thymic selection.

Mice with a fluorescent marker for interleukin 2 gene activation.

Production of interleukin (IL)-2 by T lymphocytes is one of the earliest events during immune response. A mutant mouse strain was generated by replacing the IL-2 gene with a cDNA encoding green fluorescent protein (GFP). In this model, GFP fluorescence is readily detectable upon T cell activation and is mostly coexpressed with IL-2 at the single cell level. Thus, individual activated T cells can express the IL-2 gene biallelically. Upon stimulation through the T cell antigen receptor, CD4+ cells separate into distinct GFP+ and GFP- populations, both of which are capable of differentiating into either Th1 or Th2 effectors. These mice allow noninvasive detection of IL-2 production by single cells and analysis of the subsequent differentiative fate of these cells as an immune response develops.

Somatic mutation of TSSC5, a novel imprinted gene from human chromosome 11p15.5.

We previously reported the isolation of a 2.5 Mb tumor-suppressing subchromosomal transferable fragment (STF) from 11p15.5 and the identification of nine known genes and four novel genes within this STF. We now report the isolation of a fifth novel cDNA, tumor-suppressing STF cDNA 5, designated TSSC5, located within the STF. TSSC5 encodes a predicted protein of 424 amino acids. Sequence analysis suggests that TSSC5 is a membrane protein with 10 transmembrane segments, and it is located between two imprinted genes, p57KIP2 and TSSC3. Northern blot hybridization revealed a 1.6-kb transcript in multiple adult tissues and in fetal liver and kidney, consistent with a potential role in embryonal tumors. We also found that TSSC5 is imprinted with preferential expression from the maternal chromosome. Reverse transcription-PCR analysis of TSSC5 revealed frequent occurrence of aberrant RNA splicing, which deleted exons 4, 5, and 6 in Wilms' tumors. Mutational analysis of TSSC5 by direct DNA sequencing of exons revealed a base substitution of G1120A in a Wilms' tumor, matched normal kidney, and the patient's mother, changed Arg at codon 309 to Gln. The G1120A substitution thus represents either a rare polymorphism or a tumor-predisposing mutation, because the mutant allele was of maternal origin and preferentially expressed in the patient's tissue. A second base substitution, C892T, was found in a lung cancer, changing Ser at codon 233 to Phe. This substitution was absent from the matched normal tissue and thus represented a somatic mutation. We also found loss of heterozygosity in the lung cancer, suggesting that TSSC5 may be a conventional tumor suppressor gene in the adult human lung and an imprinted tumor suppressor gene in the fetal kidney.

Syntenic organization of the mouse distal chromosome 7 imprinting cluster and the Beckwith-Wiedemann syndrome region in chromosome 11p15.5.

In human and mouse, most imprinted genes are arranged in chromosomal clusters. Their linked organization suggests co-ordinated mechanisms controlling imprinting and gene expression. The identification of local and regional elements responsible for the epigenetic control of imprinted gene expression will be important in understanding the molecular basis of diseases associated with imprinting such as Beckwith-Wiedemann syndrome. We have established a complete contig of clones along the murine imprinting cluster on distal chromosome 7 syntenic with the human imprinting region at 11p15.5 associated with Beckwith-Wiedemann syndrome. The cluster comprises approximately 1 Mb of DNA, contains at least eight imprinted genes and is demarcated by the two maternally expressed genes Tssc3 (Ipl) and H19 which are directly flanked by the non-imprinted genes Nap1l4 (Nap2) and Rpl23l (L23mrp), respectively. We also localized Kcnq1 (Kvlqt1) and Cd81 (Tapa-1) between Cdkn1c (p57(Kip2)) and Mash2. The mouse Kcnq1 gene is maternally expressed in most fetal but biallelically transcribed in most neonatal tissues, suggesting relaxation of imprinting during development. Our findings indicate conserved control mechanisms between mouse and human, but also reveal some structural and functional differences. Our study opens the way for a systematic analysis of the cluster by genetic manipulation in the mouse which will lead to animal models of Beckwith-Wiedemann syndrome and childhood tumours.

A 2.5-Mb transcript map of a tumor-suppressing subchromosomal transferable fragment from 11p15.5, and isolation and sequence analysis of three novel genes.

11p15.5 is an important tumor-suppressor gene region, showing loss of heterozygosity in Wilms tumor, rhabdomyosarcoma, adrenocortical carcinoma, and lung, ovarian, and breast cancer. We previously mapped directly by genetic complementation a subtransferable fragment (STF) harboring an embryonal tumor-suppressor gene and spanning about 2.5 Mb. We have now mapped the centromeric end of this STF between D11S988 and D11S12 and its telomeric end between D11S1318 and TH. We have isolated a complete contig of PAC, P1, BAC, and cosmid genomic clones spanning the entire 2.5-Mb region defined by this STF, as well as more than 200 exons from these genomic clones using exon trapping. We have isolated genes in this region by directly screening DNA libraries as well as by database searching for ESTs. Nine of these genes have been reported previously by us and by others. However, the initial mapping of most of those genes was based on FISH or somatic cell hybrid analysis, and here we precisely define their physical location. These genes include RRM1, GOK (D11S4896E), Nup98, CARS, hNAP2 (NAP1L4), p57KIP2 (CDKN1C), KVLQT1 (KCNA9), TAPA-1, and ASCL2. In addition, we have identified several novel genes in this region, three of which, termed TSSC1, TSSC2, and TSSC3, are reported here. TSSC1 shows homology to Rb-associated protein p48 and chromatin assembly factor CAF1, and it is located between GOK and Nup98. TSSC2 is homologous to Caenorhabditis elegans beta-mannosyl transferase, and it lies between Nup98 and CARS. TSSC3 shows homology to mouse TDAG51, which is implicated in FasL-mediated apoptosis, and it is located between hNAP2 and p57KIP2. Thus, these genes may play a role in malignancies that involve this region.

[3H]RY 80: A high-affinity, selective ligand for gamma-aminobutyric acidA receptors containing alpha-5 subunits.

The radiochemical synthesis and pharmacological properties are described of [3H]RY 80 (ethyl-8-acetylene-5, 6-dihydro-5-methyl-6-oxo-4H-imidazo[1,5a][1, 4]benzodiazepine-3-carboxylate, [ethyl-3H]). This compound is one of a series of 8-substituted imidazobenzodiazepines that exhibits both high affinity and selectivity for gamma-aminobutyric acid (GABA)A receptors containing alpha-5 subunits. Saturable, high-affinity (Kd approximately 0.7 nM) binding of [3H]RY 80 was observed in hippocampal membranes. The maximum number (Bmax) of [3H]RY 80 binding sites was approximately 18% of that obtained with [3H]flunitrazepam, a radioligand that labels all "diazepam-sensitive" GABAA receptors. This value is consistent with previous estimates (10-20%) of the proportion of rat hippocampal GABAA receptors containing alpha-5 subunits determined by immunoprecipitation with selective antibodies and competition experiments using an alpha-5-selective ligand. In recombinant GABAA receptors composed of alpha-5 beta-3 gamma-2 subunits, the Kd of [3H]RY 80 (approximately 0.5 nM) was consistent with the value obtained in hippocampus, whereas the Bmax value was not significantly different from that obtained with [3H]flunitrazepam. The potencies of several benzodiazepine site ligands to inhibit [3H]RY 80 binding to hippocampal membranes were in agreement with the values obtained in recombinant (alpha-5 beta-3 gamma-2) GABAA receptors. [3H]RY 80 was used both in a "GABA shift" assay to correctly predict the in vivo actions of a novel, alpha-5-selective ligand and to characterize a population of GABAA receptors containing alpha-5 subunits in neonatal rat cortex. These findings demonstrate that [3H]RY 80 can be used as a radioligand to examine the properties of GABAA receptors containing alpha-5 subunits.

Human KVLQT1 gene shows tissue-specific imprinting and encompasses Beckwith-Wiedemann syndrome chromosomal rearrangements.

Genomic imprinting is an epigenetic chromosomal modification in the gamete or zygote causing preferential expression of a specific parental allele in somatic cells of the offspring. We and others have identified three imprinted human genes on 11p15.5, IGF2, H19, and p57KIP2, although the latter gene is separated by 700 kb from the other two, and it is unclear whether there are other imprinted genes within this large interval. We previously mapped an embryonal tumour suppressor gene to this region, as well as five balanced germline chromosomal rearrangement breakpoints from patients with Beckwith-Wiedemann syndrome (BWS), a condition characterized by prenatal overgrowth and cancer. We isolated the upstream exons of the previously identified gene KVLQT1, which causes the familial cardiac defect long-QT (LQT) syndrome. We found that KVLQT1 spans much of the interval between p57KIP2 and IGF2, and that it is also imprinted. We demonstrated that the gene is disrupted by chromosomal rearrangements in BWS patients, as well as by a balanced chromosomal translocation in an embryonal rhabdoid tumour. Furthermore, the lack of parent-of-origin effect in LQT syndrome appears to be due to relative lack of imprinting in the affected tissue, cardiac muscle, representing a novel mechanism for variable penetrance of a human disease gene.

A novel human homologue of yeast nucleosome assembly protein, 65 kb centromeric to the p57KIP2 gene, is biallelically expressed in fetal and adult tissues.

Three genes on 11p15.5 are known to undergo genomic imprinting. The gene for insulin-like growth factor II (IGF2) is normally expressed from the paternal allele, while H19 and p57KIP2, a cyclin-dependent kinase inhibitor, are expressed from the maternal allele. Five germline balanced chromosomal rearrangement breakpoints from patients with Beckwith-Wiedemann syndrome (BWS) have been mapped to 11p15.5 between p57KIP2 and IGF2, and all are derived from the maternal chromosome. By positional cloning from BWS breakpoints, we have isolated a gene 100 kb and 65 kb centromeric to the proximal end of this BWS breakpoint cluster and p57KIP2, respectively. This gene is homologous to yeast nucleosome assembly protein (NAP1) and to a human homologue of NAP1, and we designate it hNAP2 (human nucleosome assembly protein 2). hNAP2 diverges in its expression pattern from IGF2, H19, and p57KIP2, and it shows biallelic expression in all tissues tested. Thus, hNAP2 is functionally insulated from the imprinting domain of 11p15.

Synthesis and pharmacological properties of novel 8-substituted imidazobenzodiazepines: high-affinity, selective probes for alpha 5-containing GABAA receptors.

The synthesis and pharmacological properties of imidazobenzodiazepines with both high affinity and selectivity for alpha 5-containing GABAA receptors are described. Four of these compounds (5, 6, 8, and 9) inhibited [3H]flunitrazepam binding to recombinant alpha 5 beta 2 gamma 2 GABAA receptors with IC50 values between approximately 0.4 and 5 nM. These compounds were > or = 24-75-fold more selective for recombinant receptors containing alpha 5 subunits compared to other, "diazepam-sensitive" GABAA receptors containing either alpha 1, alpha 2, or alpha 3 subunits. Imidazobenzodiazepine 9 (used as the prototypical alpha 5 selective ligand) inhibited [3H]flunitrazepam binding to hippocampal membranes with high- and low-affinity components (IC50 0.6 +/- 0.2 and 85.6 +/- 13.1 nM, respectively), representing approximately 16% and approximately 84% of the receptor pool. Inhibition of [3H]flunitrazepam binding to cerebellar membranes with imidazobenzodiazepine 9 was best fitted to a single population of sites with an IC50 of 79.8 +/- 18.3 nM. These imidazobenzodiazepines behaved as GABA negative ligands in recombinant GABAA receptors expressed in Xenopus oocytes and were convulsant in mice after parenteral administration. The relative potencies of flumazenil and zolpidem in blocking convulsions induced by 9 and DMCM, respectively, indicated that occupation of alpha 5-containing GABAA receptors substantially contributed to the convulsant properties of acetylene analog 9. These 8-substituted imidazobenzodiazepines (5, 6, 8 and 9) should prove useful in examining the physiological roles of GABAA receptors bearing an alpha 5 subunit and may also lead to the development of novel, subtype selective therapeutic agents.