A compact detector, sized particularly for imaging a mouse, is described. The active area of the detector is approximately 46 mm x 96 mm. Two flat-panel Hamamatsu H8500 position-sensitive photomultiplier tubes (PSPMTs) are coupled to a pixellated NaI(TI) scintillator which views the animal through a copper-beryllium (CuBe) parallel-hole collimator specially designed for I-125. Although the PSPMTs have insensitive areas at their edges and there is a physical gap, corrections for scintillation light collection at the junction between the two tubes results in a uniform response across the entire rectangular area of the detector. The system described has been developed to optimize both sensitivity and resolution for in-vivo imaging of small animals injected with iodinated compounds. We demonstrate an in-vivo application of this detector, particularly to SPECT, by imaging mice injected with approximately 10-15 mu Ci of I-125.

Proper functioning of the adult nervous system is critically dependent on neurons adopting the correct neurotransmitter phenotype during early development. Whereas the importance of cell‐cell communication in fate determination is well documented for a number of neurotransmitter phenotypes, the contributions made by early lineage to this process remain less clear. This is particularly true for γ‐aminobutyric acid (GABA)ergic and glutamatergic neurons, which are present as the most abundant inhibitory and excitatory neurons, respectively, in the central nervous system of all vertebrates. In the present study, we have investigated the role of early lineage in the determination of these two neurotransmitter phenotypes by constructing a fate map of GABAergic and glutamatergic neurons for the 32‐cell stage Xenopus embryo with the goal of determining whether early lineage influences the acquisition of these two neurotransmitter phenotypes. To examine these phenotypes, we have cloned xGAT‐1, a molecular marker for the GABAergic phenotype in Xenopus, and described its expression pattern over the course of development. Although we have identified isolated examples of a blastomere imparting a statistically significant bias, when taken together, our results suggest that blastomere lineage does not impart a widespread bias for subsequent GABAergic or glutamatergic fate determination. In addition, the fate map presented here suggests a general dorsal‐anterior to ventral‐posterior patterning progression of the nervous system for the 32‐cell stage Xenopus embryo. J. Comp. Neurol. 495:645–657, 2006. © 2006 Wiley‐Liss, Inc.

Microarray technology has become an important tool for studying large-scale gene expression for a diversity of biological applications. However, there are a number of experimental settings for which commercial arrays are either unsuitable or unavailable despite the existence of sequence information. With the increasing availability of custom array manufacturing services, it is now feasible to design high-density arrays for any organism having sequence data. However, there have been relatively few reports discussing gene selection, an important first step in array design. Here we propose an in silico strategy for custom microarray gene selection that is applicable to a wide range of organisms, based on utilizing public domain microarray information to interrogate existing sequence data and to identify a set of homologous genes in any organism of interest. We demonstrate the utility of this approach by applying it to the selection of candidate genes for a custom Xenopus laevis microarray. A significant finding of this study is that 3%-4% of Xenopus expressed sequence tags (ESTs) are in an orientation contrary to that indicated in the public database entry (http://mssaha.people.wm.edu/suppMSS.html).

A compact detector, sized particularly for imaging a mouse, is described. The active area of the detector is approximately 48 mmtimes96 mm. Two flat-panel Hamamatsu H8500 position sensitive photomultiplier tubes (PSPMT) are coupled to a pixellated NaI(Tl) scintillator which views the animal through a copper-beryllium (CuBe) parallel-hole collimator. Because the PSPMTs have insensitive areas at their edges, particular care has been given to bridging the gap where the two tubes join so as to maintain uniform response across the entire rectangular area of the detector. The system described has been developed to optimize both sensitivity and resolution for in vivo imaging of small animals injected with iodinated compounds. We demonstrate an in vivo application of this detector by imaging a mouse injected with 10 muCi of 125I

A rotating cylindrical gantry apparatus holding a pair of Hamamatsu R3292 position sensitive photomultiplier tubes (PSPMTs), each of which can include either a parallel-hole collimator or a pinhole collimator, has been used to gather planar images and single photon emission computer tomography (SPECT) data of an entire mouse. In addition, in a number of experiments a magnified view of a specific organ was obtained with pinhole collimation. The scintillators used were pixelated NaI(Tl) or CsI(Tl) with pixels 1/spl times/1/spl times/3 mm and were air coupled to the PSPMTs. The active area of each detector was a circle about 110 mm in diameter. The parallel hole detector had an intrinsic resolution in planar images of 2 mm and the pinhole collimator was used in a magnification which yielded 1 mm spatial resolution. A rigid, lightweight Lexan tube is used to contain a heated mouse bed formed in a semicylindrical shape. Warmed, humidified air as well as gas anesthetic can be introduced and exhausted via the tube. The arrangement facilitates accurate centering and positioning of the SPECT cameras to the animal. The simultaneous acquisition of whole body data and a magnified image of the gut and pancreas within the animal has been applied in a study of /sup 125/I tagged insulin in hyperglycemic and normal mice. In a study of the effectiveness of KI blocking on uptake of iodine in a mouse thyroid, images a phantom of the thyroid and submaxillary glands were of considerable interest.

The authors have developed a multimodality system for imaging the biodistribution of biologically interesting ligands tagged with /sup 125/I. By incorporating a small fluoroscope as an additional modality, they have enhanced their small animal nuclear imaging system to include both X-rays and images from two Hamamatsu R3292 5" diameter position sensitive photomultiplier tubes (PSPMT) viewing pixel...

As the major excitatory neurotransmitter in the vertebrate nervous system, glutamate not only plays an essential role in adult neural signaling, but has also been implicated as a trophic factor in neuronal cell maturation, differentiation, and survival. An essential component of the glutamatergic neurotransmission system is the family of glutamate transporters, a multigene family that codes for plasma membrane-bound as well as vesicle-bound proteins responsible for the removal of glutamate from the cleft and its re-uptake into the synaptic vesicle. Here we describe the spatial and temporal expression of the vesicular glutamate transporter (xVGlut1) during the early developmental stages of the amphibian Xenopus laevis. RNAse protection analysis and in situ hybridization reveal that xVGlut1 is first expressed at late neurula stages in the developing spinal cord and trigeminal nerve. By tailbud stages xVGlut1 transcripts are detected in several of the cranial nerves, the pineal gland, and medial forebrain. By hatching stages xVGlut1 expression reappears in localized tracts within the spinal cord. Expression levels increase throughout development into adulthood.

The detector group at the Thomas Jefferson National Accelerator Facility (Jefferson Lab) and the Biology, Physics and Applied Sciences Departments at the College of William and Mary are collaborating on the development of a miniature dual modality SPECT-CT system for mouse imaging. The detector heads of the SPECT subsystem are capable of imaging the gamma- and X-ray emissions (28-35 keV) of the radioactive isotope iodine-125 (Ie 125). Two different sets of I-125 imaging detectors are configured on a gantry that has an open-barrel type design. One set of detector heads is based on the 1 inch square Hamamatsu R5900-M64 position sensitive photomultiplier tube coupled to crystal scintillator arrays. The other detector heads configured on the gantry are two 5-inch diameter Hamamatsu R3292-based compact gamma cameras. The X-ray radiographic projections will be obtained using a LIXI Inc. model LF-85-503-OS X-ray imaging system that has an active area of 5.5 cm in diameter. The open-barrel shaped gantry facilitates the positioning of various mini gamma-ray imaging detectors and the X-ray system. The data acquisition and gantry control is interfaced through a Macintosh G3 workstation. SPECT reconstruction results using the R5900 based detector are presented

We have developed an economical dual-modality nuclear imaging system comprised of two Hamamatsu 125 mm diameter position sensitive photomultiplier tubes (PSPMT) viewing pixelated scintillators and a small fluoroscopic X-ray system (Lixi, Inc.). Collimators placed between the animal and the scintillators can be readily interchanged and include CuBe parallel-hole collimators with a range of resolution/sensitivity combinations as well as brass pinhole collimators with various pinhole diameters. The small X-ray fluoroscope provides 5 cm diameter images, several of which can readily be combined to provide structural information from the animal under study. The system has been used to follow the metabolism of compounds tagged with /sup 125/I. Biological information has been obtained on the uptake of tagged insulin and tumor necrosis factor-alpha (TNF/spl alpha/) thus demonstrating the applicability of this system for in vivo analysis of diseases such as diabetes.

Approximately 1% of the Xenopus laevis genome consists of highly repetitive DNA known alternatively as OAX (for Oocyte Activation in Xenopus), Satellite I, or Repetitive HindIII Monomer 2. Present as tandemly repeated units of approximately 750 base pairs, OAX encodes a family of small RNA species transcribed by RNA polymerase III. Although the subject of many of the classic studies on early embryonic gene regulation, reports on OAX expression remain contradictory and incomplete. Using whole-mount in situ hybridization and RNase protection assays, we have therefore examined in detail the expression pattern of OAX in Xenopus embryos of various stages. OAX is initially expressed during gastrula stages; by tailbud stages embryos display discrete zones of expression at the dorsal boundary of the cement gland, in the developing somites and differentiating skeletal muscle, as well as in the dorsal aspect of the neural tube. These data demonstrate that OAX is expressed in a dynamic pattern under tight spatial and temporal regulation.

Required to supply nutrients and oxygen to the growing embryo, the vascular system is the first functional organ system to develop during vertebrate embryogenesis. Although there has been substantial progress in identifying the genetic cascade regulating vascular development, the initial stages of vasculogenesis, namely, the origin of vascular endothelial cells within the early embryo, remain unclear. To address this issue we constructed a fate map for specific vascular structures, including the aortic arches, endocardium, dorsal aorta, cardinal veins, and lateral abdominal veins, as well as for the red blood cells at the 16-cell stage and the 32-cell stage ofXenopus laevis.Using genetic markers to identify these cell types, our results suggest that vascular endothelial cells can arise from virtually every blastomere of the 16-cell-stage and the 32-cell-stage embryo, with different blastomeres preferentially, though not exclusively, giving rise to specific vascular structures. Similarly, but more surprisingly, every blastomere in the 16-cell-stage embryo and all but those in the most animal tier of the 32-cell-stage embryo serve as progenitors for red blood cells. Taken together, our results suggest that during normal development, both dorsal and ventral blastomeres contribute significantly to the vascular endothelial and red blood cell lineages.