A cavity ringdown spectrometer for methane isotope analysis using a 1.65 µm distributed feedback diode laser with fiber optical feedback loop.

The development of a 1.65 µm cavity ringdown methane spectrometer for methane isotope analysis is reported. In order to reduce the laser linewidth, simple optical feedback with an 11 m external fiber cavity using a retroreflector was implemented and it improved the sensitivity. The detection limit at the ppt level for both 12CH4 and 13CH4 concentrations at 100 Torr gas pressure was evaluated from the Allan-Werle plot calculated from the dataset obtained at the fixed laser frequency. In contrast, the detection limit estimated from the baseline noise on the absorption spectrum was a few ppb for both methane isotopologues due to the periodic background oscillations that remained even after baseline correction. The system demonstrated the direct measurement of ambient methane in atmospheric room air, and the estimated 13CH4 ratio as well as the methane concentration were in good agreement with the reference values of ambient air.

All-optical presynaptic plasticity induction by photoactivated adenylyl cyclase targeted to axon terminals.

Intracellular signaling plays essential roles in various cell types. In the central nervous system, signaling cascades are strictly regulated in a spatiotemporally specific manner to govern brain function; for example, presynaptic cyclic adenosine monophosphate (cAMP) can enhance the probability of neurotransmitter release. In the last decade, channelrhodopsin-2 has been engineered for subcellular targeting using localization tags, but optogenetic tools for intracellular signaling are not well developed. Therefore, we engineered a selective presynaptic fusion tag for photoactivated adenylyl cyclase (bPAC-Syn1a) and found its high localization at presynaptic terminals. Furthermore, an all-optical electrophysiological method revealed rapid and robust short-term potentiation by bPAC-Syn1a at brain stem-amygdala synapses in acute brain slices. Additionally, bPAC-Syn1a modulated mouse immobility behavior. These results indicate that bPAC-Syn1a can manipulate presynaptic cAMP signaling in vitro and in vivo. The all-optical manipulation technique developed in this study can help further elucidate the dynamic regulation of various cellular functions.

SIPA1L1/SPAR1 Interacts with the Neurabin Family of Proteins and is Involved in GPCR Signaling.

Signal-induced proliferation-associated 1 (SIPA1)-like 1 (SIPA1L1; also known as SPAR1) has been proposed to regulate synaptic functions that are important in maintaining normal neuronal activities, such as regulating spine growth and synaptic scaling, as a component of the PSD-95/NMDA-R-complex. However, its physiological role remains poorly understood. Here, we performed expression analyses using super-resolution microscopy (SRM) in mouse brain and demonstrated that SIPA1L1 is mainly localized to general submembranous regions in neurons, but surprisingly, not to PSD. Our screening for physiological interactors of SIPA1L1 in mouse brain identified spinophilin and neurabin-1, regulators of G-protein-coupled receptor (GPCR) signaling, but rejected PSD-95/NMDA-R-complex components. Furthermore, Sipa1l1-/- mice showed normal spine size distribution and NMDA-R-dependent synaptic plasticity. Nevertheless, Sipa1l1-/- mice showed aberrant responses to α2-adrenergic receptor (a spinophilin target) or adenosine A1 receptor (a neurabin-1 target) agonist stimulation, and striking behavioral anomalies, such as hyperactivity, enhanced anxiety, learning impairments, social interaction deficits, and enhanced epileptic seizure susceptibility. Male mice were used for all experiments. Our findings revealed unexpected properties of SIPA1L1, suggesting a possible association of SIPA1L1 deficiency with neuropsychiatric disorders related to dysregulated GPCR signaling, such as epilepsy, attention deficit hyperactivity disorder (ADHD), autism, or fragile X syndrome (FXS).SIGNIFICANCE STATEMENT Signal-induced proliferation-associated 1 (SIPA1)-like 1 (SIPA1L1) is thought to regulate essential synaptic functions as a component of the PSD-95/NMDA-R-complex. In our screening for physiological SIPA1L1-interactors, we identified G-protein-coupled receptor (GPCR)-signaling regulators. Moreover, SIPA1L1 knock-out (KO) mice showed striking behavioral anomalies, which may be relevant to GPCR signaling. Our findings revealed an unexpected role of SIPA1L1, which may open new avenues for research on neuropsychiatric disorders that involve dysregulated GPCR signaling. Another important aspect of this paper is that we showed effective methods for checking PSD association and identifying native protein interactors that are difficult to solubilize. These results may serve as a caution for future claims about interacting proteins and PSD proteins, which could eventually save time and resources for researchers and avoid confusion in the field.

Cooperation of LIM domain-binding 2 (LDB2) with EGR in the pathogenesis of schizophrenia.

Genomic defects with large effect size can help elucidate unknown pathologic architecture of mental disorders. We previously reported on a patient with schizophrenia and a balanced translocation between chromosomes 4 and 13 and found that the breakpoint within chromosome 4 is located near the LDB2 gene. We show here that Ldb2 knockout (KO) mice displayed multiple deficits relevant to mental disorders. In particular, Ldb2 KO mice exhibited deficits in the fear-conditioning paradigm. Analysis of the amygdala suggested that dysregulation of synaptic activities controlled by the immediate early gene Arc is involved in the phenotypes. We show that LDB2 forms protein complexes with known transcription factors. Consistently, ChIP-seq analyses indicated that LDB2 binds to > 10,000 genomic sites in human neurospheres. We found that many of those sites, including the promoter region of ARC, are occupied by EGR transcription factors. Our previous study showed an association of the EGR family genes with schizophrenia. Collectively, the findings suggest that dysregulation in the gene expression controlled by the LDB2-EGR axis underlies a pathogenesis of subset of mental disorders.

Impairment of spatial memory accuracy improved by Cbr1 copy number resumption and GABAB receptor-dependent enhancement of synaptic inhibition in Down syndrome model mice.

Down syndrome is a complex genetic disorder caused by the presence of three copies of the chromosome 21 in humans. The most common models, carrying extra-copies of overlapping fragments of mouse chromosome 16 that is syntenic to human chromosome 21, are Ts2Cje, Ts1Cje and Ts1Rhr mice. In electrophysiological analyses using hippocampal slices, we found that the later phase of the depolarization during tetanic stimulation, which was regulated by GABAB receptors, was significantly smaller in Ts1Cje and Ts2Cje mice than that in WT controls but not in Ts1Rhr mice. Furthermore, isolated GABAB receptor-mediated inhibitory synaptic responses were larger in Ts1Cje mice. To our knowledge, this is the first report that directly shows the enhancement of GABAB receptor-mediated synaptic currents in Ts1Cje mice. These results suggest that GABAB receptor-mediated synaptic inhibition was enhanced in Ts1Cje and Ts2Cje mice but not in Ts1Rhr mice. The Cbr1 gene, which is present in three copies in Ts1Cje and Ts2Cje but not in Ts1Rhr, encodes carbonyl reductase that may facilitate GABAB-receptor activity through a reduction of prostaglandin E2 (PGE2). Interestingly, we found that a reduction of PGE2 and an memory impairment in Ts1Cje mice were alleviated when only Cbr1 was set back to two copies (Ts1Cje;Cbr1+/+/-). However, the GABAB receptor-dependent enhancement of synaptic inhibition in Ts1Cje was unaltered in Ts1Cje;Cbr1+/+/- mice. These results indicate that Cbr1 is one of the genes responsible for DS cognitive impairments and the gene(s) other than Cbr1, which is included in Ts1Cje but not in Ts1Rhr, is responsible for the GABAB receptor-dependent over-inhibition.

Influence of High-Dose Folic Acid on Methotrexate Efficacies and Safety in Japanese Rheumatoid Arthritis Patients.

Backgrounds Folic acid dose at ≦5 mg/week has been recommended for rheumatoid arthritis (RA) patients to decrease risk of methotrexate adverse effects. However, higher doses of folic acid is used in some cases. We examined the influence of high-dose folic acid on methotrexate efficacies and safety in Japanese RA patients. Methods 502 RA patients of four hospitals prescribed methotrexate and folic acid were included. These patients were divided into two subgroups according to the threshold of folic acid dose by 5 mg/week. Basic patient characteristics, methotrexate doses, and the efficacies or adverse effects of methotrexate were retrospectively compared between the two patient subgroups. Results The frequency of folic acid use at doses higher than 5 mg/week was significantly different between the four hospitals (P<0.001). The prevalence of methotrexate adverse effects was not significantly different between the patients taking folic acid less and more than 5 mg/week. However, in the lower dose methotrexate subgroup (≦8 mg/week), the prevalence of patients exhibiting abnormal serum ALT concentrations in the patients using higher (>5 mg/week) dose of folic acid was significantly higher than that in the lower (≦5 mg/week) folic acid-treated subgroup (P=0.029). Folic acid dose between patients taking methotrexate less and more than 8 mg/week was not significantly different. Major conclusion Folic acid dose was dependent on the hospitals, while efficacies and hepatotoxicity of methotrexate was not basically different between patients taking less and more than 5 mg/week of folic acid.

Morphology-based investigation of metachronous inguinal hernia after negative laparoscopic evaluation - is it acquired indirect inguinal hernia?

PURPOSE: Patients who developed apparent metachronous contralateral inguinal hernia (MCIH) after negative laparoscopic diagnosis have been reported. We performed this study to investigate the morphological characteristics and etiology of this phenomenon. PATIENTS AND METHODS: A consecutive series of 1,747 patients (858 boys and 889 girls) with symptomatic unilateral inguinal hernia were studied. During laparoscopic percutaneous completely extraperitoneal closure, morphological appearances at the asymptomatic groin were inspected for contralateral patent processus vaginalis (CPPV) with definitive criteria. If positive CPPV was identified, it was closed by the same technique used for the affected side. The patients were reviewed for occurrence of metachronous contralateral hernia. RESULTS: A total of 755 patients (43.2%) had positive CPPV. Of the 992 patients whose CPPVs were evaluated as true negative, eight (seven boys, one girl) developed MCIH (time taken, three months to five years six months). During second-look operations, morphological appearances of the CPPV that was originally considered as true negative CPPV was found to be wide open. CONCLUSIONS: Despite a true negative evaluation by laparoscopy, there was a 0.8% chance of developing a MCIH. This phenomenon was male-oriented and may be acquired indirect inguinal hernia without preceding of CPPV.

PX-RICS-deficient mice mimic autism spectrum disorder in Jacobsen syndrome through impaired GABAA receptor trafficking.

Jacobsen syndrome (JBS) is a rare congenital disorder caused by a terminal deletion of the long arm of chromosome 11. A subset of patients exhibit social behavioural problems that meet the diagnostic criteria for autism spectrum disorder (ASD); however, the underlying molecular pathogenesis remains poorly understood. PX-RICS is located in the chromosomal region commonly deleted in JBS patients with autistic-like behaviour. Here we report that PX-RICS-deficient mice exhibit ASD-like social behaviours and ASD-related comorbidities. PX-RICS-deficient neurons show reduced surface γ-aminobutyric acid type A receptor (GABAAR) levels and impaired GABAAR-mediated synaptic transmission. PX-RICS, GABARAP and 14-3-3ζ/θ form an adaptor complex that interconnects GABAAR and dynein/dynactin, thereby facilitating GABAAR surface expression. ASD-like behavioural abnormalities in PX-RICS-deficient mice are ameliorated by enhancing inhibitory synaptic transmission with a GABAAR agonist. Our findings demonstrate a critical role of PX-RICS in cognition and suggest a causal link between PX-RICS deletion and ASD-like behaviour in JBS patients.

Development and Validation of a Novel Fibrosis Marker in Biliary Atresia during Infancy.

OBJECTIVES: Most biliary atresia (BA) patients suffer from liver fibrosis and often require liver transplantation. The aim of this study was to develop and validate a novel fibrosis marker for BA patients aged <1 year-the infant BA liver fibrosis (iBALF) score-subsequent to the previously reported fibrosis marker for BA patients aged ≥1 year. METHODS: From three institutions for pediatric surgery, BA patients and their native liver histology examinations performed at the age of <1 year were retrospectively identified and assigned to a development cohort (58 patients and 73 examinations) or validation cohort (92 patients and 117 examinations) according to their institutions. Histological fibrosis stages (F0-F4), blood test results, and clinical information at the time of liver histology examination were reviewed. The iBALF score was determined using multivariate ordered logistic regression analysis and was assessed for its associations with histological fibrosis stages. RESULTS: The iBALF score equation was composed of natural logarithms, including serum total bilirubin level, blood platelet counts, and days of age. The score revealed a strong correlation with fibrosis stage (r=0.80 and 0.73 in the development and validation cohorts, respectively; P<0.001). The areas under the receiver-operating characteristic curves for diagnosing each fibrosis stage were 0.86-0.94 in the development cohort and 0.86-0.90 in the validation cohort (P<0.001), indicating good diagnostic power. In addition, no patient with an iBALF score >6 (equivalent to F4) at the initial surgery survived with their native liver at 1 year of age (n=9). CONCLUSIONS: The iBALF score that was developed was a good noninvasive marker of native liver fibrosis for BA patients aged <1 year.

Yb:YAG thin-disk chirped pulse amplification laser system for intense terahertz pulse generation.

We have developed a 1 kHz repetition picosecond laser system dedicated for intense terahertz (THz) pulse generation. The system comprises a chirped pulse amplification laser equipped with a Yb:YAG thin-disk amplifier. At room temperature, the Yb:YAG thin-disk regenerative amplifier provides pulses having energy of over 10 mJ and spectral bandwidth of 1.2 nm. The pulse duration achieved after passage through a diffraction grating pair compressor was 1.3 ps. By employing this picosecond laser as a pump source, THz pulses having a peak frequency of 0.3 THz and 4 µJ of energy were generated by means of optical rectification in an Mg-doped LiNbO3 crystal.

Contact grating device with Fabry-Perot resonator for effective terahertz light generation.

A novel design for a contact grating device with an incorporated Fabry-Perot resonator is proposed for high-power terahertz (THz) light generation. We deposited a multilayer consisting of Ta(2)O(5) and Al(2)O(3) on a magnesium-doped stoichiometric LiNbO(3) substrate and fabricated grating grooves on the outermost layer. The multilayer was designed such that conditions for a Fabry-Perot resonator were satisfied for light diffracted by the grating. Consequently, the fraction of light transmitted into the LiNbO(3) substrate, i.e., the diffraction efficiency, was enhanced by the resonator. The diffraction efficiency of the fabricated device was 71%, which is close to the calculated value of 78% from the optimized design. THz light generation was also demonstrated with the contact grating device. The THz output of 0.41 µJ was obtained using near-infrared pump light of 2.7 mJ.

The mechanisms of the strong inhibitory modulation of long-term potentiation in the rat dentate gyrus.

The hippocampus is essential for the formation of certain types of memory, and synaptic plasticity such as long-term potentiation (LTP) is widely accepted as a cellular basis of hippocampus-dependent memory. Although LTP in both perforant path-dentate gyrus (DG) granule cell and CA3-CA1 pyramidal cell synapses is similarly dependent on activation of postsynaptic N-methyl-D-aspartate receptors, several reports suggest that modulation of LTP by γ-aminobutyric acid (GABA) receptor-mediated inhibitory inputs is stronger in perforant path-DG granule cell synapses. However, little is known about how different the mechanism and physiological relevance of the GABAergic modulation of LTP induction are among different brain regions. We confirmed that the action of GABA(A) receptor antagonists on LTP was more prominent in the DG, and explored the mechanism introducing such difference by examining two types of GABA(A) receptor-mediated inhibition, i.e. synaptic and tonic inhibition. As synaptic inhibition, we compared inhibitory vs. excitatory monosynaptic responses and their summation during an LTP-inducing stimulus, and found that the balance of the summated postsynaptic currents was biased toward inhibition in the DG. As tonic inhibition, or sustained activation of extrasynaptic GABA(A) receptors by ambient GABA, we measured the change in holding currents of the postsynaptic cells induced by GABA(A) receptor antagonists, and found that the tonic inhibition was significantly stronger in the DG. Furthermore, we found that tonic inhibition was associated with LTP modulation. Our results suggest that both the larger tonic inhibition and the larger inhibitory/excitatory summation balance during conditioning are involved in the stronger inhibitory modulation of LTP in the DG.

[Cancer therapy analysis from the viewpoint of DPC data].

DPC (Diagnosis Procedure Combination), Japanese specific prospective payment system, is gradually taking root in Japan. Hospitals under this system are required to submit nationally standardized electronic data to get reimbursed. DPC data include not only information for reimbursement but also details of medical records of each patient. We have tried to investigate actual situation of cancer treatment in Japan by analyzing DPC data. In this study, we have revealed actual situation of non-small cell lung cancer treatment in both Osaka prefecture and nationwide using girasol , the DPC-analysis-system, by Medical Architects Inc. Moreover, we report the significant variations in non-small cell lung cancer chemotherapy regimens and therapy protocols between the two hospitals in Osaka prefecture.

Laparoscopic completely extraperitoneal repair of inguinal hernia in children: a single-institute experience with 1,257 repairs compared with cut-down herniorrhaphy.

BACKGROUND: Conventional open herniorrhaphy in children has been reported to have 0.3-3.8% recurrence and 5.6-30% postoperative contralateral hernia rates. We developed a unique technique to achieve completely extraperitoneal ligation of PPV without any skip areas under laparoscopic control. This report introduces our technique and results compared with the cut-down herniorrhaphy. METHODS: A consecutive series of 1,585 children with inguinal hernia/hydrocele (1996-2006) was analyzed. In laparoscopic patent processus vaginalis (PPV) closure (LPC), an orifice of PPV was encircled with a 2-0 suture extraperitoneally by a specially devised Endoneedle and tied up from outside of the body achieving completely extraperitoneal ligation of the ring. The round ligament was included in the ligation, whereas the spermatic cord and testicular vessels were excluded by advancing the needle across them behind the peritoneum. Cut-down herniorrhaphy (CD), with or without diagnostic laparoscopy, or LPC was selected according to parental preference under informed consent. RESULTS: Parents gave more preference to LPC (LPC in 1,257 children, CD in 308, and miscellaneous in 20). Age ranges were equal for both groups. Sex distribution showed female preponderance in the LPC group (44.8% vs. 26.6%, p < 0.001) and umbilical hernia/cysts were predominantly included in the LPC group (11.9% vs. 2.9%, p < 0.001). Mean operation times were equal for both groups for unilateral repair (28.2 +/- 9.2 for LPC vs. 27.8 +/- 13.5 for CD) and were shorter for bilateral repair in the LPC group (35.8 +/- 11.6 vs. 46.7 +/- 17.7). The incidence of postoperative hernia recurrence and contralateral hernia in the LPC group was 0.2% and 0.8%. Two children in the CD group had injuries to their reproductive system during the operation (0.6%). CONCLUSIONS: The advantages of our technique include following: technically simple, short operation time, inspection of bilateral IIRs with simultaneous closure of cPPV, reproductive systems remain intact, routine addition of umbilicoplasty if desired, and essentially indiscernible wounds.

Hepatoduodenal ligament teratoma with hepatic artery running inside.

Teratoma in infants and children is not particularly rare. However, the teratoma in the hepatoduodenal ligament is extremely rare and the surgical treatment is challenging because of its anatomical complexity. We could find only six cases in the literature. In this report, we present the seventh case with special reference to the operative technique to save the hepatic artery for resection of the tumor using intraoperative ultrasonography.

Light guided procedure for congenital pyriform sinus fistula; new and simple procedure for impalpable fistula.

Pyriform sinus fistula causes acute suppurative thyroiditis, and there is a risk of recurrence if the sinus tract is not excised completely. The tract should be dissected as high as possible toward the pyriform fossa. We report our devised technique "the light guided procedure" for the impalpable fistula. A bronchoscope was inserted into the pyriform sinus fistula with the help of a gastrofiberscope positioned at the larynx. The tract of the fistula was identified with the help of the light from the bronchoscope. We were able to reach the proximal end of the fistula, directly. This procedure is easy, safe, and minimally invasive.

Bcr-Abl signaling through the PI-3/S6 kinase pathway inhibits nuclear translocation of the transcription factor Bach2, which represses the antiapoptotic factor heme oxygenase-1.

The malignant phenotype of chronic myeloid leukemia (CML) is due to the abnormal tyrosine kinase activity of the Bcr-Abl oncoprotein. We have previously reported that expression of the Bach2 transcription factor, which induces apoptosis in response to oxidative stress, is greatly reduced in CML cells. Because these cells are resistant to apoptosis, we tested whether Bach2 could also be regulated through posttranslational mechanisms that promote inhibition of the apoptotic response to mutagenic stimuli in CML. We found that Bach2 is phosphorylated on S521 via the phosphatidylinositol-3/S6 kinase pathway, and substitution of this site to alanine leads to nuclear accumulation of the protein, indicating that this phosphorylation is important for its subcellular localization. Ectopic expression of the S521 mutant imparts greater impairment to CML cell growth than the wild-type factor. Furthermore, we showed that Bach2 transcriptionally represses heme oxygenase-1, an antiapoptotic factor up-regulated in CML. Because CML cells are known to produce high levels of intracellular reactive oxygen species, overexpression of heme oxygenase-1 resulting from inhibition of Bach2 activity may contribute to their genomic instability and leukemic phenotype.

Photoexcitation of K-shell and L-shell hollow beryllium.

We have observed K-shell and L-shell hollow beryllium atoms (2s(2)2p3s and 1s3s(2)3p) created by photoexcitation using synchrotron radiation. Resonance shapes were fitted to the Fano profile and the parameters were deduced. A Dirac-Fock calculation was performed to identify the configuration of the peaks and to predict other hollow atomic peaks. The results of the calculation were in good agreement with the experimental data. The comparison of the transition strength has revealed that the three-electron photoexcitation to the 1s3s(2)3p configuration is stronger than the two-electron photoexcitation to the 2s(2)2p3s configuration. This is attributed to the large overlap between the 2s orbital of the ground state (1s(2)2s(2)) with the orbital of the L-shell hollow state (1s3s(2)3p).