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19th IEEE-NPSS Real Time Conference (RT) -- MAY 26-30, 2014 -- Nara, JAPAN, WOS: 000380588000133, This paper presents the design of the NEDA frontend electronics, a first attempt to involve the use of digital electronics in large neutron detector arrays. Among the electronic modules taking part, we emphasize on the front-end analog processing, the digitalization, digital pre-processing, communications firmware, as well as the integration of the Global Trigger and Synchronization system. The NEDA array will be available for measurements in 2015., Osaka Univ, Res Nucl Phys, IEEE Nucl & Plasma Sci Soc

The existence of Sertoli cell factors which modulate the rat Leydig cell function prompted us to study the biological activity of selected proteins called CMB proteins and produced by immature rat Sertoli cells. Percoll purified Leydig cells (10(5)) from 20 days-old rats have been incubated 5 h at 32 degrees C in 1 ml Ham F12/DME medium with increasing concentrations of partially purified CMB proteins (0-1,000 ng/ml) either in presence or absence of oLH (25 ng/ml). Among the CMB proteins tested, only CMB-21 produces a dose related increase of testosterone production: from 2 to 500 pg/ml of CMB-21, testosterone output is unchanged (51 pg/10(5) cells) but 1 to 1,000 ng/ml of this protein produces a linear increase of testosterone productions (86 to 870 pg). In the presence of oLH which induces a 10-fold increase of testosterone production (499 pg), increasing doses of CMB-21 further stimulate testosterone output (775 to 2.272 pg/10(5) cells). Whatever the concentration of oLH used (0 to 50 ng/ml), CMB-21 (500 ng/ml) leads to a further 2 fold augmentation of testosterone synthesis; similarly, in the presence of dbcAMP (1 mM), CMB 21 increases the testosterone production but no effect is observed when Leydig cells are incubated in the presence of 22R-hydroxycholesterol (30 microM). The cAMP levels which are increased more than 4 fold by oLH, remain unchanged in the presence of CMB-21 either alone or with oLH, as observed when Sertoli cell culture medium is used.(ABSTRACT TRUNCATED AT 250 WORDS)

The polypeptide diazepam binding inhibitor (DBI) and drug ligands for the mitochondrial peripheral-type benzodiazepine receptor (PBR) have been shown to regulate cholesterol transport, the rate-determining step in steroidogenesis, in hormone-responsive steroidogenic cells including the MA-10 Leydig tumor cells. The present study was designed to characterize the role of DBI and PBR in the R2C rat Leydig tumor constitutive steroid-producing cell model. Both DBI and PBR were present in R2C cells. R2C cell treatment with a cholesterol-linked phosphorothioate oligodeoxynucleotide antisense to DBI, but not sense, resulted in the reduction of DBI levels and a concomitant dramatic decrease of the amount of progesterone produced. These observations strongly suggested that DBI was important in maintaining constitutive steroidogenesis in R2C cells. Radioligand binding assays revealed the presence of a single class of PBR binding sites with an affinity 10 times higher (Kd approximately 0.5 nM) than that displayed by the MA-10 PBR (Kd approximately 5 nM). Photolabeling of R2C and MA-10 cell mitochondria with the photoactivatable PBR ligand [3H]1-(2-fluoro-5-nitrophenyl)-N-methyl-N-(1-methyl-propyl)-3- isoquinolinecarboxamide showed that the M(r) 18,000 PBR protein was specifically labeled. This indicates that the R2C cells express a PBR protein which has properties similar to the MA-10 PBR. Chemical crosslinking studies of purified metabolically radiolabeled DBI to mitochondria provided direct evidence that DBI specifically binds to the M(r) 18,000 PBR protein. Moreover, DBI and a PBR synthetic ligand were able to increase steroid production in isolated R2C cell mitochondria which express the 5 nM affinity receptor. However, mitochondrial PBR binding was increased by 6-fold upon addition of the post-mitochondrial fraction, suggesting that a cytosolic factor modulates the binding properties of PBR in R2C cells and is responsible for the 0.5 nM affinity receptor seen in intact cells. In conclusion, these data demonstrate that DBI plays a key role in maintaining R2C constitutive steroidogenesis by binding to the mitochondrial higher affinity PBR which promotes a continuous supply of cholesterol to the inner mitochondrial side chain cleavage cytochrome P450.

Previous studies demonstrated that the polypeptide diazepam binding inhibitor (DBI) and its receptor, the peripheral-type benzodiazepine receptor (PBR), are involved in the regulation of steroid biosynthesis and that one site of PBR action resides in mitochondria. In the present investigation, evidence is presented that a functional form of PBR is also present at the cell surface. First, PBR was immunolocalized in the rat testis using biotin-streptavidin peroxidase immunocytochemistry, and results revealed that PBR was present exclusively in the interstitial Leydig cells. Next, the distribution of PBR in MA-10 Leydig cells was further examined using confocal microscopy. MA-10 cells were either fixed and immunostained or fixed/permeabilized and immunostained for PBR, followed by generation of confocal microscope optical sections, three-dimensional reconstructions of these sections, and then generation of vertical confocal sections of the three-dimensional reconstruction. In the fixed/unpermeabilized cells, PBR immunostaining at the cell surface was clearly evident, whereas in the fixed/permeabilized cells, intracellular PBR distribution was more robust. These results suggest that the plasma membrane fraction of the receptor could mediate the action of extracellular PBR ligands on Leydig cell function. Next, we examined whether DBI, the naturally occurring PBR ligand, is secreted by testicular cells and whether it could activate the cell surface PBR. Immunolocalization of DBI demonstrated that it was present in both Leydig and Sertoli cells. Further, using an immunoblot assay, we demonstrated that DBI is present in rat testicular interstitial fluid. Metabolic labeling of cultured immature rat Sertoli cells and MA-10 mouse tumor Leydig cells, followed by immunoprecipitation of the secreted proteins with an anti-DBI antiserum, demonstrated that both Leydig and Sertoli cells secrete DBI and could serve as a cell source for the interstitial fluid DBI. Then, we partially purified the DBI present in conditioned medium and interstitial fluid by reverse phase chromatography and demonstrated it to be bioactive, based on displacement of a radiolabeled benzodiazepine (Ro5-4864)-specific ligand for PBR; pronase treatment of different preparations eliminated all bioactivity. We then examined the effects of DBI on Leydig cell function. DBI added to MA-10 cells affected DNA synthesis and cell growth in a biphasic manner; at low concentrations (1 nM), DBI was mitogenic, increasing [3H]thymidine incorporation and cell numbers by 30-40%, while at high concentrations (1 microM), DBI inhibited cell growth (30-40%). Similar effects on cell growth were obtained using the benzodiazepine Ro5-4864.