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PubMed1996

Specific expression in brain and liver driven by the hemopexin promoter in transgenic mice.

Transgenic mice harboring the human hemopexin promoter sequences linked to the lacZ reporter gene were generated and analyzed for temporal and spatial distribution of beta-galactosidase. Upstream sequences spanning from -1800, -700 and -500 bp to the transcription start point direct regulated beta-galactosidase expression specifically to the liver and to the brain of transgenic mice. These results suggest that the 500 bp DNA fragment flanking the 5'end of the human hemopexin gene contains the cis-acting elements required for tissue and developmental stage-specific expression in vivo and provide evidence for a new extrahepatic site of expression of the hemopexin gene.

Acute-Phase Reaction↗

PubMed1995

Pollen fertility restoration by nuclear gene Fr in CMS bean: nuclear-directed alteration of a mitochondrial population.

Two nuclear genes, Fr and Fr2, have been identified that restore pollen fertility to cytoplasmic male sterile (CMS) common bean (Phaseolus vulgaris L.) by apparently distinct mechanisms. Whereas Fr2 appears to suppress the expression of a male sterility associated mitochondrial sequence (designated pvs), Fr restores pollen fertility by causing the elimination of this unusual mitochondrial DNA-segment. To further investigate the mechanism of Fr action, Fr and Fr2 were cointroduced into the nucleus of a bean line containing the sterility inducing cytoplasm. When the effect of pvs was suppressed by Fr2, the presence of Fr no longer directed the elimination of the mitochondrial pvs sequence. This result suggests that the Fr function is dependent on proper expression of the pvs sequence. To evaluate the temporal and spatial patterns of Fr action, we undertook a polymerase chain reaction-based approach to trace the fate of the pvs sequence in different tissues of F2 and F3 fertile-restored plants derived from a genetic cross between a cytoplasmic male sterile line of common bean, CMS-Sprite (frfr), and fertility restorer line R351 (FrFr). We demonstrate that the Fr-directed disappearance of pvs sequence occurs during flower development. Elimination of the pvs sequence from developing megaspores results in permanent fertility restoration in the following generations. Genetic analysis demonstrated that permanent fertility restoration, that is, the complete elimination of pvs from reproductive tissues requires two doses of the Fr allele or the absence of fr in F2 individuals.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗

PubMed2006

Phylogeography and spatial genetic structure of the Southern torrent salamander: implications for conservation and management.

The Southern torrent salamander (Rhyacotriton variegatus) was recently found not warranted for listing under the US Endangered Species Act due to lack of information regarding population fragmentation and gene flow. Found in small-order streams associated with late-successional coniferous forests of the US Pacific Northwest, threats to their persistence include disturbance related to timber harvest activities. We conducted a study of genetic diversity throughout this species' range to 1) identify major phylogenetic lineages and phylogeographic barriers and 2) elucidate regional patterns of population genetic and spatial phylogeographic structure. Cytochrome b sequence variation was examined for 189 individuals from 72 localities. We identified 3 major lineages corresponding to nonoverlapping geographic regions: a northern California clade, a central Oregon clade, and a northern Oregon clade. The Yaquina River may be a phylogeographic barrier between the northern Oregon and central Oregon clades, whereas the Smith River in northern California appears to correspond to the discontinuity between the central Oregon and northern California clades. Spatial analyses of genetic variation within regions encompassing major clades indicated that the extent of genetic structure is comparable among regions. We discuss our results in the context of conservation efforts for Southern torrent salamanders.

Animals↗

PubMed1994

Three-dimensional laser-scanning confocal microscopy of in situ hybridization in the skin.

In situ hybridization is an important tool in molecular and developmental biology to detect specific nucleic acid sequences (either mRNA or DNA) within cells. This technique is especially applicable to tissue sections since it provides information about the spatial distribution of DNA or mRNA sequences. However, previous studies utilizing in situ hybridization in the skin were hampered by a high degree of nonspecific background, which has made interpretation of the results difficult. In this paper, we demonstrate how refinements in in situ hybridization techniques, combined with laser-scanning confocal microscopy, significantly reduce nonspecific background and produce improved resolution of in situ hybridization in skin specimens. The sensitive detection method of laser-scanning confocal microscopy allows three-dimensional localization of S35 radioactive-labeled riboprobes within the emulsion of specimens, which is not possible with conventional bright or dark field light microscopy.

Dendritic Cells↗

PubMed1997

Impaired spatial information processing in aged monkeys with preserved recognition memory.

Spatial information processing was examined in a non-human primate model of cognitive aging, using procedures formally similar to tasks designed for rats. The test apparatus was a large open field containing eight reward locations. Monkeys rapidly learned to visit each location once per trial, and probe manipulations confirmed that young animals navigated according to the distribution of cues surrounding the maze. In contrast, aged monkeys solved the task using a response sequencing strategy, independent of extramaze spatial information. Object recognition memory was normal in the aged group. The results reveal substantial correspondence in the cognitive effects of aging across rat and primate models, and they establish appropriate procedures for testing the long-standing proposal that the role of the hippocampus in normal spatial learning is similarly conserved.

Aging↗

PubMed1993

A myelin proteolipid protein-LacZ fusion protein is developmentally regulated and targeted to the myelin membrane in transgenic mice.

Transgenic mice were generated with a fusion gene carrying a portion of the murine myelin proteolipid protein (PLP) gene, including the first intron, fused to the E. coli LacZ gene. Three transgenic lines were derived and all lines expressed the transgene in central nervous system white matter as measured by a histochemical assay for the detection of beta-galactosidase activity. PLP-LacZ transgene expression was regulated in both a spatial and temporal manner, consistent with endogenous PLP expression. Moreover, the transgene was expressed specifically in oligodendrocytes from primary mixed glial cultures prepared from transgenic mouse brains and appeared to be developmentally regulated in vitro as well. Transgene expression occurred in embryos, presumably in pre- or nonmyelinating cells, rather extensively throughout the peripheral nervous system and within very discrete regions of the central nervous system. Surprisingly, beta-galactosidase activity was localized predominantly in the myelin in these transgenic animals, suggesting that the NH2-terminal 13 amino acids of PLP, which were present in the PLP-LacZ gene product, were sufficient to target the protein to the myelin membrane. Thus, the first half of the PLP gene contains sequences sufficient to direct both spatial and temporal gene regulation and to encode amino acids important in targeting the protein to the myelin membrane.

Animals↗

PubMed1985

Electron microscopic heteroduplex mapping identifies regions of the engrailed locus that are conserved between Drosophila melanogaster and Drosophila virilis.

Physical localization of mutations in the engrailed (en) gene suggested that at least 70 kilobases (kb) of genomic sequences contribute to the normal function of this gene. Molecular characterization has suggested that en function is encoded in a small, 4.5-kb primary transcript. To identify functional regions within the 70 kb of the en locus of D. melanogaster, we identified sequences conserved in the D. virilis genome (estimated divergence time, 60 million years). Based on homology to D. melanogaster, we isolated en DNA from a D. virilis genomic library. Electron microscopic heteroduplex analysis indicated that in 70 kb there is 20 kb of conserved DNA in 33 different regions dispersed throughout the en locus, including two which encode parts of the major embryonic transcript. The conserved regions are in the same linear order and are spaced by similar lengths of nonconserved sequences in the D. virilis and D. melanogaster DNAs. What functional constraints have enforced conservation of sequences throughout the entire 70 kb and protected the region from divergence of size and arrangement? Our working hypothesis is that sequences necessary for the complex spatial and temporal pattern of en expression are dispersed throughout the 70-kb en locus and that selection for proper regulation restricts evolutionary divergence.

Animals↗

PubMed1995

A novel gene expressed during zebrafish gastrulation identified by differential RNA display.

Vertebrate gastrulation is a dynamic period of development characterized by extensive cell migrations. This stage of development is likely to require the expression of a new genetic repertoire to initiate and direct these dramatic changes. The differential RNA display has been used to identify genes specifically expressed during the gastrula stage of a model vertebrate, the zebrafish. One of the genes isolated by the differential display technique has been sequenced and characterized for its spatial and temporal expression. This gene, called G12, is expressed during a narrow window of time during gastrulation and is restricted to a single cell type. At this time of development the zebrafish embryo consists of three cell types: the yolk cell, EVL cells and deep cells. Interestingly, both EVL and deep cells derive early in development from common progenitor cells but G12 expression is restricted only to the EVL lineage. Comparison of the amino acid sequence from this gene with the Genbank database indicates similarity to two previously reported mammalian genes. The similarity between these three genes suggests that they may serve a common function. The G12 gene is the first example of restricted gene expression in EVL cells of the zebrafish. The G12 gene should prove to be a useful model for the study of regulated gene expression during gastrulation.

Amino Acid Sequence↗

PubMed2006

Three-dimensional spatial skill training in a simulated space station: random vs. blocked designs.

BACKGROUND: Astronauts floating inside a spacecraft must be able to recall the direction to surrounding visual landmarks, regardless of their viewing perspective. If 3D orientation skills are taught preflight, should perspective sequences be blocked or randomized? Can standard spatial skill tests predict performance? METHODS: Undergraduates (40 men and 40 women; ages 19-24) learned 3D spatial relationships among landmark pictures in a cubic chamber simulating a space station node. Subjects learned to predict picture directions when told one picture's direction (the one behind them) and the subject's simulated roll orientation, which was changed between trials by rotating pictures. The dependent variable was the proportion of correct predictions. A between group (n=40 per group) independent variable was training type (random vs. blocked sequencing of perspectives). Experiment phase (familiarization, training, transfer, and 2 retention phases) was a within group variable. Subjects also took three standard spatial skill tests: Card Rotation, Cube Comparison, and Group Imbedded Figures. RESULTS: As hypothesized, during training, performance for the random group (0.56) was worse than the blocked group (0.83); during transfer, the random group (0.75) was better than the blocked group (0.56); during retention-i, the random group (0.70) was better than the blocked group (0.55); and during retention-2, the random group (0.76) was better than the blocked group (0.65). Spatial skill tests correlated differently across the two groups, indicating that random sequencing elicits different skills. CONCLUSION: Random presentation enhances 3D spatial skill transfer and retention. Standard spatial tests can predict performance and have the potential to customize training.

Adult↗

PubMed1991

Positive and negative cis-acting DNA domains are required for spatial and temporal regulation of gene expression by a seed storage protein promoter.

Mutations affecting spatial and temporal regulation of a beta-phaseolin gene encoding the major storage protein of bean (Phaseolus vulgaris) were analyzed by stable and transient transformation approaches. The results substantiate the value of transient assays for rapid determination of the functionality of cis-acting sequences and the importance of stable transformation to identify tissue-specific determinants. Spatial information is specified primarily by two upstream activating sequences (UAS). UAS1 (-295 to -109) was sufficient for seed-specific expression from both homologous and heterologous (CaMV 35S) promoters. In situ localization of GUS expression in tobacco embryos demonstrated that UAS1 activity was restricted to the cotyledons and shoot meristem. A second positive domain, UAS2 (-468 to -391), extended gene activity to the hypocotyl. Temporal control of GUS expression was found to involve two negative regulatory sequences, NRS1 (-391 to -295) and NRS2 (-518 to -418), as well as the positive domain UAS1. The deletion of either negative element caused premature onset of GUS expression. These findings indicate combinatorial interactions between multiple sequence motifs specifying spatial information, and provide the first example of the involvement of negative elements in the temporal control of gene expression in higher plants.

Cloning, Molecular↗

PubMed1997

Spatial distribution of field potential profiles in the cat cerebellar cortex evoked by peripheral and central inputs.

The present study was designed to characterize the spread of excitation within the frontal plane of the cat cerebellar cortex following different types of stimuli. In particular, experiments were performed to determine whether the spread of excitation evoked by mossy fibre inputs proceeds primarily along the parallel fibres ("beam-like" spread) or whether these inputs activate non-propagated foci ("patches") in the cerebellar cortex. Field potentials were recorded within a frontal plane as a medial to lateral array at different depths in parallel tracks. The recordings were made following electrical stimulation of different forelimb nerves and functionally related areas of the sensorimotor cortex as well as during passive paw movements. The resulting spatial grid of responses provides discrete spatio-temporal information reflecting the activation of specific cerebellar afferents and the neuronal interactions they evoke. The method employed demonstrates the spatial distribution of the temporal sequence of excitability changes throughout all the cerebellar cortical layers. In general, the characteristics of the responses in the intermediate cerebellar cortex depended on the source of the signals. Activity patterns evoked by peripheral nerve stimulation showed more clustered foci compared with those following electrical stimulation of functionally related areas of the sensorimotor cortex. The centrally evoked profiles were generally more homogeneous. The largest number of foci were observed following passive movements around the wrist joint. The spread of excitation in the vertical direction was evaluated by the spatial shift of the line of reversal of the N3/P2-potential (zero-isopotential line). Lines of reversal for peripherally-evoked activity patterns were approximately 90 microns closer to the molecular layer than those evoked by central stimulation in animals in which recordings have been performed in lobule Vc. The opposite was found for recordings in lobule Vb, where potential reversals following peripheral stimulation were located 40 microns deeper than those evoked following central stimulation. Cortical inputs resulted in a more proximal activation of lobule Vc Purkinje cell dendrites than in lobule Vb. This type of input processing thus seems to be lobule dependent. A beam-like spread of excitation could not be demonstrated. For both climbing fibre and mossy fibre afferent systems multiple foci were found in the frontal plane. The foci due to mossy fibre activation arose from the granular layer and expanded vertically to the molecular layer. For the climbing fibre system the foci were restricted to the molecular layer, where they merged to form a superficial band of activation. Although the data presented in this paper favour a focal distribution of activity, they do not exclude beam-like propagation along the parallel fibres, because of the difficulty of detecting this pattern in response to the stimuli. The "beam"- and "patch"-like hypotheses need not be mutually exclusive. Each could contribute to a specific stage of the temporal-spatial processing in the cerebellar cortex in a functional and task-specific manner.

Animals↗

PubMed2005

Removal of epileptogenic sequences from video material: the role of color.

BACKGROUND: After Pokémon viewing triggered an epidemic of seizures in Japan, many efforts have been made to design safety guidelines and systems to protect subjects with photosensitivity. The authors developed a new method based upon nonlinear diffusion techniques capable of filtering the epileptogenic content of a video sequence related to color without altering its spatial and luminance content. METHODS: The authors showed to 25 photosensitive patients (18 women, mean age: 22 years) the original Pokémon sequence and a modified one in an ABBA protocol using two television (TV) sets (100 and 50 Hz). RESULTS: Twenty-three patients had a photoparoxysmal response (PPR) according to Waltz classification with at least one of the scenes. The modified sequence triggered fewer and less severe PPRs than the original version in both TVs (p < 0.001). Original sequences elicited generalized PPRs in 56.5% of the trials for the 50 Hz TV and in 41.3% for the 100 Hz TV, whereas modified sequences elicited these responses in only 8.7% (50 Hz) and 4.3% (100 Hz TV) of the trials (p < 0.001). Sensitivity to the modified version on the 50 Hz TV correlated with pattern sensitivity (p < 0.05). CONCLUSION: Specific manipulations of the color modulation-depth could be enough to decrease dramatically the risk of triggering seizures in susceptible subjects exposed to provocative visual scenes. This new method can be implemented in protective devices able to filter out the epileptogenic video sequences in which color plays a fundamental role while leaving intact the spatial content, frequency, and average luminance.

Adolescent↗

PubMed1987

A purine-pyrimidine motif verifying an identical presence in almost all gene taxonomic groups.

A statistical parameter identifies, with a high degree of significance, a motif which is present in protein-coding sequences of eukaryotes, prokaryotes, chloroplasts, mitochondria, viral introns, ribosomal RNA genes, and transfer RNA genes. The random probability of occurrence of such a situation is 10(-12). This motif has the following properties: (i) its significant presence in almost all present-day genes explains why it can be considered as primitive oligonucleotide, (ii) its nucleotide order is: YRY (N)6YRY, R being a purine base, Y a pyrimidine one and N any base, (iii) its length and its terminal trinucleotides YRY suggest a primordial function related to the spatial structure of the DNA sequences. This motif is found in some viral protein-coding genes, but not in eukaryotic introns.

Base Sequence↗

PubMed1992

Practical choices of fast spin echo pulse sequence parameters: clinically useful proton density and T2-weighted contrasts.

With the development of fast spin echo (FSE) MRI techniques, T2-weighted images of the brain may be obtained much more quickly than when using conventional spin echo techniques (CSE), because made the individual echoes on the FSE pulse sequence are phase encoded, allowing acquisition of the same spatial information as in CSE with less excitations. The pulse sequence parameters (echo train length, bandwidth, echo spacing) are discussed. Images were obtained on four volunteers using both CSE and FSE while varying repetition time, echo time and matrix. Comparison for signal intensity gray-white differentiation, fat and CSE signal, arifacts and vascular resolution showed that FSE images comparable in quality to those of CSE can be obtained in less than half the time. A practical choice of FSE parameters is recommended for clinical use. However, artifacts, possibly related to CSF and vascular pulsation, of which the radiologist should be aware, were identified on the FSE images.

Artifacts↗

PubMed1999

[A smooth pneumatic motion device for dynamic MRI imaging of the cervical spine].

PURPOSE: To develop and evaluate a new, pneumatically operated motion apparatus for the dynamic functional MR examination of the cervical spine. MATERIALS AND METHODS: A metal-free, pneumatically operated motion apparatus for functional dynamic MR imaging of the cervical spine was developed with respect to the geometry of a short-bore, superconducting high-field MR system. The examination protocol included a T2-weighted multi-slice turbospin echo sequence (T2-TSE, TR/TE = 2610/115, matrix 254/512) and a dynamic T2-weighted single-shot sequence (T2-TSE, TR/TE = 1110/110, matrix 255/256, acquisition time 1 s) in a sagittal plane. In order to evaluate the new motion apparatus and the examination protocol, 10 healthy subjects and 10 patients were examined. RESULTS: The new motion apparatus allowed us to perform a passive stepless inclination and reclination motion of the patient's cervical spine within the MR scanner without leaving the magnet bore. The maximum motion degrees of the cervical spine were within -35 degrees (reclination) and 30 degrees (inclination). Due to the T2-contrast and the high spatial resolution of the dynamic sequence, the myelon, the surrounding cerebrospinal fluid and the discoligamental structures were imaged with a good contrast and allowed a sufficient evaluation of the cervical spinal channel in all functional positions. CONCLUSIONS: This new motion apparatus allows a standardized and stepless dynamic functional MR examination in a short-bore high-field MR scanner. By the use of fast T2-weighted spin echo sequences it has been shown to be of high value for the evaluation of the cervical myelon, the anterior and posterior cerebrospinal fluid and the discoligamental structures.

Biomechanical Phenomena↗

PubMed1997

Spatial patterns of fasting and fed antropyloric pressure waves in humans.

1. Gastric mechanics were investigated by categorizing the temporal and spatial patterning of pressure waves associated with individual gastric contractions. 2. In twelve healthy volunteers, intraluminal pressures were monitored from nine side hole recording points spaced at 1.5 cm intervals along the antrum, pylorus and duodenum. 3. Pressure wave sequences that occurred during phase II fasting contractions (n = 221) and after food (n = 778) were evaluated. 4. The most common pattern of pressure wave onset along the antrum was a variable combination of antegrade, synchronous and retrograde propagation between side hole pairs. This variable pattern accounted for 42% of sequences after food, and 34% during fasting (P < 0.05). Other common pressure wave sequence patterns were: purely antegrade-29% after food and 42% during fasting (P < 0.05); purely synchronous-23% fed and 17% fasting; and purely retrograde-6% fed and 8% fasting. The length of sequences was shorter after food (P < 0.05). Some sequences 'skipped' individual recording points. 5. The spatial patterning of gastric pressure wave sequences is diverse, and may explain the differing mechanical outcomes among individual gastric contractions. 6. Better understanding of gastric mechanics may be gained from temporally precise correlations of luminal flows and pressures and gastric wall motion during individual gastric contraction sequences.

Adult↗

PubMed1996

High-density endocardial activation mapping of the right atrium in three dimensions by composition of multielectrode catheter recordings.

The use of radiofrequency ablation for treatment of complex arrhythmia substrates has prompted interest in transcatheter endocardial activation mapping. Technical constraints on catheter fabrication and the intention to use such maps to guide ablation both demand innovative approaches to mapping. A fluoroscopically based endocardial mapping technique is proposed to improve the ability of electrophysiologists to interpret large amounts of data acquired using multielectrode catheter arrays, improving their ability to visualize the data and act on its content. This technique addresses previous limitations imposed by the number of electrodes that can be deployed and by the difficulty in determining their relative spatial locations. It is based on the composition of multiple activation sequence mappings made in a single rhythm, with the spatial locations of recording electrode pairs determined in orthogonal fluoroscopic views referenced to stable intrathoracic markers. Rather than imposing a geometry determined primarily by the measurement apparatus, the spatial locations of only those electrodes in proximity to the endocardial surface, as determined by their ability to record bipolar electrograms, are measured. In this manner, the geometry of the endocardium may be approximated by measurements made of electrode position. Using this approach, the number of endocardial sites that can be sampled in a stable rhythm is theoretically unlimited, resulting in the realization of high-resolution activation maps. Spatiotemporal data may be used to create three-dimensional activation sequence maps, displayed as animated sequences. This technique was used in anatomically normal and diseased human right atria to create activation maps of sinus and paced rhythms, classic atrial flutter, and postoperative intraatrial reentrant tachycardia, using a median of 108 electrode positions (range, 27-197) in 25 maps. The activation sequences represented by these maps were diverse, but qualitatively concordant with known mechanisms of atrial activation. High-density catheter-based activation mapping of the right atrium is feasible and may improve understanding of complex arrhythmias and assist in the development of ablative techniques. Further research is needed on the spatial correlation between cardiac anatomy and fluorography, suppression of spatial artifact, and optimal mapping densities.

Body Surface Potential Mapping↗

PubMed1997

Magnetic resonance pulmonary angiography and direct imaging of embolus for the detection of pulmonary emboli.

RATIONALE AND OBJECTIVES: The authors developed a two-dimensional breathhold magnetic resonance (MR) technique for the direct imaging of pulmonary emboli. METHODS: In vitro MR imaging was performed to demonstrate the potential generation of clot-blood contrast by in vivo pulmonary embolism (PE). A two-dimensional magnetization prepared gradient-echo (turbo-FLASH) breathhold technique was designed to directly image intravascular emboli by the selective nulling of the blood signal. A turbo-FLASH pulmonary angiographic breathhold sequence was used to provide spatial localization of detected emboli. Thirteen patients with suspected PE were studied; 6 patients underwent conventional pulmonary angiography (CPA) and the remaining 7 had diagnoses based on findings from other studies. RESULTS: In vitro study of blood clot demonstrated an initial rise and then fall in T1 sufficient to generate clot-blood contrast after eight days of clot formation. All patients with CPA or alternative study evidence of PE were diagnosed as positive with direct embolus imaging MR. There were no false-positive diagnoses. Three additional emboli were detected using the MR technique compared with CPA. The MR pulmonary angiographic sequence provided a useful road map for localization of intravascular emboli but was less sensitive for PE detection than the embolus imaging technique. CONCLUSIONS: The direct imaging of PE is feasible using a simple two-dimensional breathhold technique.

Adult↗