Overexpression of a Slit homologue impairs convergent extension of the mesoderm and causes cyclopia in embryonic zebrafish

Slit is expressed in the midline of the central nervous system both in vertebrates and invertebrates. In Drosophila, it is the midline repellent acting as a ligand for the Roundabout (Robo) protein, the repulsive receptor which is expressed on the growth cones of the commissural neurons. We have isolated cDNA fragments of the zebrafish slit2 and slit3 homologues and found that both genes start to be expressed by the midgastrula stage well before the axonogenesis begins in the nervous system, both in the axial mesoderm, and slit2 in the anterior margin of the neural plate and slit3 in the polster at the anterior end of the prechordal mesoderm. Later, expression of slit2 mRNA is detected mainly in midline structures such as the floor plate cells and the hypochord, and in the anterior margins of the neural plates in the zebrafish embryo, while slit3 expression is observed in the anterior margin of the prechordal plate, the floorplate cells in the hindbrain, and the motor neurons both in the hindbrain and the spinal cord. To study the role of Slit in early embryos, we overexpressed Slit2 in the whole embryos either by injection of its mRNA into one-cell stage embryos or by heat-shock treatment of the transgenic embryos which carries the slit2 gene under control of the heat-shock promoter. Overexpression of Slit2 in such ways impaired the convergent extension movement of the mesoderm and the rostral migration of the cells in the dorsal diencephalon and resulted in cyclopia. Our results shed light on a novel aspect of Slit function as a regulatory factor of mesodermal cell movement during gastrulation. (C) 2001 Academic Press.

The effects of interleukin-10 depletion in vivo on the immune response to Porphyromonas gingivalis in a murine model

Background: The immune response to Porphyromonas gingivalis in the mouse abscess model is known to be dependent upon CD4 T-cell activation and the regulatory role of cytokines. The role of interleukin-10 (IL-10) in this mouse model was examined in vivo. Methods: One-week-old, female BALB/c mice were divided into 4 groups. Groups 1 and 2 were given intraperitoneal (ip) injections of phosphate buffered saline (PBS) weekly for 5 weeks. Group 3 was given an ip injection of rat immunoglobulin. Group 4 was injected with rat anti-IL-10 antibodies. At week 6, group 1 was sham-immunized with PBS, and groups 2, 3, and 4 were injected with P gingivalis lipopolysaccharide (Pg-LPS) weekly for 2 weeks. One week after the final immunization, delayed-type hypersensitivity (DTH) was assessed by footpad swelling to Pg-LPS. The level of serum antibodies to Pg-LPS and IFN-gamma (IFN-gamma) was determined by enzyme-linked immunosorbent assay. Dorsal abscess formation induced by the injection of viable P gingivalis was examined daily for 30 days. Results: The footpad swelling of the anti-IL-10-treated group (group 4) was significantly higher than that of groups 1 to 3. Similarly, the serum IFN-gamma level in group 4 was much higher than that of the other experimental groups. There was no significant difference in serum IgG antibodies to Pg-LPS in any of the experimental groups. However, the level of IgM antibodies in group 4 mice was significantly lower than that in groups 2 and 3. In addition, serum IgG1 was suppressed in group 4 mice, while IgG2a antibodies were raised. However, there was no difference observed between the levels of IgG2b and IgG3 antibodies in any group of mice. The lesions in sham-immunized mice (group 1) persisted for 30 days, and those in group 2 and 3 were undetected by day 18 and 20, respectively. In sharp contrast, lesions in group 4 had healed completely by day 13. Conclusions: This study has shown that IL-10 depletion in vivo in P gingivalis LPS-induced immune response in mice led to an elevated DTH response, an increase in serum IFN-gamma levels, and raised levels of IgG and IgG2a antibodies. Treatment with anti-IL-10 antibodies resulted in suppressed IgG I and IgM responses and a more rapid healing of abscesses than in non-IL-10-depleted mice. These results suggest that IL-10 depletion in Pg-LPS-induced immune response in mice may lead to a Th1-like immune response and provide strong protection against a subsequent challenge with live P gingivalis in an abscess model.

Brain activity during the encoding, retention, and retrieval of stimulus representations

Studies of delayed nonmatching-to-sample (DNMS) performance following lesions of the monkey cortex have revealed a critical circuit of brain regions involved in forming memories and retaining and retrieving stimulus representations. Using event-related functional magnetic resonance imaging (fMRI), we measured brain activity in 10 healthy human participants during performance of a trial-unique visual DNMS task using novel barcode stimuli. The event-related design enabled the identification of activity during the different phases of the task (encoding, retention, and retrieval). Several brain regions identified by monkey studies as being important for successful DNMS performance showed selective activity during the different phases, including the mediodorsal thalamic nucleus (encoding), ventrolateral prefrontal cortex (retention), and perirhinal cortex (retrieval). Regions showing sustained activity within trials included the ventromedial and dorsal prefrontal cortices and occipital cortex. The present study shows the utility of investigating performance on tasks derived from animal models to assist in the identification of brain regions involved in human recognition memory.

The effect of mode of exposure to Beauveria bassiana on conidia acquisition and host mortality of Colorado potato beetle, Leptinotarsa decemlineata

The effects of the mode of exposure of second instar Colorado potato beetles to Beauveria bassiana on conidia acquisition and resulting mortality were investigated in laboratory studies. Larvae sprayed directly with a B, bassiana condial suspension, larvae exposed to B, bassiana-treated foliage, and larvae both sprayed and exposed to treated foliage experienced 76, 34, and 77% mortality, respectively. The total number of conidia and the proportion of germinating conidia were measured over time for four sections of the insect body: the ventral surface of the head (consisting mostly of ventral mouth parts), the ventral abdominal surface, the dorsal abdominal surface, and the legs. From observations at 24 and 36 h posttreatment, mean totals of 161.1 conidia per insect were found on sprayed larvae, 256.1 conidia on larvae exposed only to treated foliage, and 408.3 conidia on larvae both sprayed and exposed to treated foliage, On sprayed larvae, the majority of conidia were found on the dorsal abdominal surface, whereas conidia were predominantly found in the ventral abdominal surface and mouth parts on larvae exposed to treated foliage, Between 24 and 36 h postinoculation the percentage of conidia germinating on sprayed larvae increased slightly from 80 to 84%), On the treated foliage, the percentage of germinated conidia on larvae increased from 35% at 24 h to 50% at 36 h posttreatment, Conidia germination on sprayed larvae on treated foliage was 65% at 24 h and 75% at 36 h posttreatment, It is likely that the gradual acquisition of conidia derived from the continuous exposure to B. bassiana inoculum on the foliar surface was responsible for the increase in germination over time on larvae exposed to treated foliage, The density and germination of conidia were observed 0, 4, 8, 12, 16, 20, and 24 h after being sprayed with or dipped in conidia suspensions or exposing insects to contaminated foliage, Conidia germinated twice as fast on sprayed insects as with any other treatment within the first 12 h, This faster germination may be due to the pressure of the sprayer enhancing conidial lodging on cuticular surfaces. (C) 2001 Academic Press.

Transport of endosomal early antigen I in the rat sciatic nerve and location in cultured neurons

Early endosomal antigen I (EEAI) is known to be a marker of early endosomes and in cultured hippocampal neurons it preferentially localizes to the dendritic but not the axonal compartment. We show in cultured dorsal root ganglia and superior cervical ganglia neurons that EEAI localizes to the cell bodies and the neurites of both sensory and sympathetic neurons. We then show in vivo using a ligated rat sciatic nerve that EEAI significantly accumulates on the proximal side and not on the distal side of the ligation. This suggests that EEAI is transported in the anterograde direction in axons either as part of the homeostatic process or to the nerve ligation site in response to nerve injury. NeuroReport 12:281-284 (C) 2001 Lippincott Williams & Wilkins.

A comparison of the lateral and posterior retroperitoneoscopic approach for complete and partial nephroureterectomy in children

Objective To report the comparative results of a selective posterior or lateral retroperitoneoscopic approach (RPA) for nephroureterectomy in children. Patients and methods Following an established experience with RPA, 36 complete and 19 partial nephrouretectomies were prospectively randomized to a posterior and lateral retroperitoneoscopic approach. The patients were aged 4 months to 14 years, with a body weight at operation of 5.7-82 kg. For posterior RPA the child is positioned prone, with three access ports. The operating space was created with balloon dissection and maintained with CO2 insufflation. The child was then rotated 30 degrees with the kidney in the dependent position, and the operator and assistant standing on the affected side. In the lateral approach the child is in the lateral decubitus position with the operator and assistant facing the dorsal aspect of the patient. Results There was no significant difference in operative duration between the lateral and posterior approaches for nephrectomy (65 and 47 min) or partial nephrectomy (85 and 75 min). Two lateral nephrectomies required open conversion (one upper pole and one lower pole). Conclusion The posterior approach gives easy and quick access to the renal pedicle. It is preferable for complete nephrectomy alone and partial or polar excision. In children under 5 years old a near complete ureterectomy can be achieved. The lateral approach creates more inferomedial space, gives better access to ectopic kidneys and allows complete ureterectomy in all cases, Access to the pedicle in the normal position requires more frequent positioning of the kidney. Care must be taken as peritoneal tears are more common.

Monocular and binocular distance cues: insights from visual form agnosia I (of III)

The human nervous system constructs a Euclidean representation of near (personal) space by combining multiple sources of information (cues). We investigated the cues used for the representation of personal space in a patient with visual form agnosia (DF). Our results indicated that DF relies predominantly on binocular vergence information when determining the distance of a target despite the presence of other (retinal) cues. Notably, DF was able to construct an Euclidean representation of personal space from vergence alone. This finding supports previous assertions that vergence provides the nervous system with veridical information for the construction of personal space. The results from the current study, together with those of others, suggest that: (i) the ventral stream is responsible for extracting depth and distance information from monocular retinal cues (i.e. from shading, texture, perspective) and (ii) the dorsal stream has access to binocular information (from horizontal image disparities and vergence). These results also indicate that DF was not able to use size information to gauge target distance, suggesting that intact temporal cortex is necessary for learned size to influence distance processing. Our findings further suggest that in neurologically intact humans, object information extracted in the ventral pathway is combined with the products of dorsal stream processing for guiding prehension. Finally, we studied the size-distance paradox in visual form agnosia in order to explore the cognitive use of size information. The results of this experiment were consistent with a previous suggestion that the paradox is a cognitive phenomenon.

2,6,,9-trioxabicyclo[3.3.1]nona-3,7-dienes and 2,4,6,8-tetraoxaadamantanes: Novel chiral spacer units in macrocyclic polyethers

The unusual chiral heterocyclic systems, trioxabicyclo[3.3.1]nona-3,7-dienes (bridged bisdioxines), are incorporated as novel spacer molecules into macrocyclic polyether ring systems of various sizes (8, 9 as well as 11-15) by cyclocondensation reaction of the! bisacid chloride 4b or bisesters 6,7 and 10, with several ethylene glycols. The 2:2 macrocycles 12-14 are obtained in approximately 50:50 mixtures of diastereomers. These conclusions are mainly based on HPLC data presented in Table I as well as X-ray analyses of (1R,5R)-8c (space group Pbca, a = 10.163(3) Angstrom, b = 18.999(4) Angstrom, c = 36.187(10) Angstrom, V = 6987(3) Angstrom(3), Z = 8, d(calc) = 1.218 g cm(-3), 6974 reflections, R = 0.0553.), mesolrac-11 (space group P (1) over bar, a = 10.472(5) Angstrom, b = 16.390(5) Angstrom, c = 17.211(5) Angstrom, alpha = 98.69(2)degrees, beta = 93.04(2)degrees, gamma = 98.52(2)degrees, V = 2879.3(18) Angstrom(3), Z = 2, d(calc) = 1.173 g cm(-3), 11,162 reflections, R = 0.0945) and meso-12 (space group P2(1)/c, a = 9.927(2), b = 18.166(3), c = 17.820(3) Angstrom, beta = 96.590(10)degrees, V = 3192.3(10)Angstrom(3), Z = 4, D-c = 1.109 g cm(-3), 3490 reflections, R = 0.0646). The 1:1 macrocycles 8b,c are also formed by intramolecular transesterification of the open-chain bisesters 7b,c and their formation is favored by the use of metal ions as templates. The bridged bisdioxine moieties in 8b and 12 are converted into the corresponding chiral tetra-oxaadamantane spacers to afford macrocycles 16 and 17. Preliminary metal ion complexation studies with selected species (8c, 11-14) were also performed.

The ultrastructure of Macropodinium moiri and revised diagnosis of the Macropodiniidae (Litostomatea : Trichostomatia)

The ultrastructural features of Macropodinium moiri were investigated. The somatic cortex is composed of two lateral non-ciliated zones covered with trapezoidal plates and separated by a trough-like dorsoventral groove (DVG) which divides the cell into left and right halves. The somatic kineties occupy the margins of the DVG and are composed of monokinetids whose infraciliature shows a typical litostome pattern. The pellicular plates are lamellate, and separated by V-shaped grooves which are lined by thick-walled vacuoles. The DVG cortex is composed of electron-opaque U-shaped ribs which alternate with electron-lucent saccular structures. The DVG surface is composed of small regular pellicular sacs built up to form the ridges of the dorsal DVG. The vestibulum forms a laterally compressed cone with left/right differentiation. The basal section of its non-ciliated right side is internally lined (outer to innermost) by longitudinal fibres, nematodesmata and transverse microtubular ribbons. The left side bears the vestibular kineties and in its basal section is lined (outer to innermost) by small nematodesmata and transverse tubules. Cytoplasmic organelles include endoplasmic reticulum, starch granules and a single contactile vacuole surrounded by patches of nephridioplasm. Hydrogenosomes are absent and coccoid Gram-positive bacteria lie under the ciliated portions of the cell. This set of characteristics differs significantly from those of the all other trichostomes; Macropodiniidae is therefore designated Trichostomatia incertae sedis. A revised familial diagnosis of the Macropodiniidae is proposed.

N-terminal Slit2 promotes survival and neurite extension in cultured peripheral neurons

To investigate the effect of the N-terminal Slit2 protein on neuronal survival and development, recombinant human N-terminal Slit2 (N-Slit2) was assayed against isolated embryonic chick dorsal root ganglion sensory, ciliary ganglion and paravertebral sympathetic neurons. N-Slit2 promoted significant levels of neuronal survival and neurite extension in all of these populations. The protein was also assayed against postnatal mouse dorsal root ganglion neurons and found to promote neuronal survival in a similar manner. These findings suggest the Slit proteins may play an important role during development of the nervous system, mediating cellular survival in addition to the well documented role these proteins play in axonal and neuronal chemorepulsion.

Ultrastructure of the mature spermatozoon of the musky rat-kangaroo, Hypsiprymnodon moschatus (Potoroidae : Marsupialia)

It is currently accepted that Hypsiprymnodon moschatus is a basal macropod, retaining several primitive features from the ancestral phalangeroid that gave rise both to modern possums and macropods. Sperm ultrastructure is frequently found to provide informative characters for phylogenetic analysis as these features are not strongly selected for and are thus unlikely to be confounded by effects such as convergence. Caudal epididymal biopsies were taken from two male H. moschatus and prepared for transmission and scanning electron microscopy in order to study mature spermatozoan ultrastructure. Within the diprotodont group, several features were found to be unique to H. moschatus. These were an unusual acrosome covering nearly 100% of the dorsal nuclear surface, a midpiece fibre network which is loose, indistinct and extends to the anterior-most aspect of the midpiece, a nucleus that is very streamlined, while the principal piece is comparatively short, and a mitochondrial helix and annulus which are similar to those of dasyurids. Also reported is the presence of a fibrous network in die connecting piece, not previously reported for any marsupial.

Contribution of cholinergic systems to state-dependent modulation of respiratory control

Respiration is altered during different stages of the sleep-wake cycle. We review the contribution of cholinergic systems to this alteration, with particular reference to the role of muscarinic acetylcholine receptors (MAchRs) during rapid eye movement (REM) sleep. Available evidence demonstrates that MAchRs have potent excitatory effects on medullary respiratory neurones and respiratory motoneurones, and are likely to contribute to changes in central chemosensitive drive to the respiratory control system. These effects are likely to be most prominent during REM sleep, when cholinergic brainstem neurones show peak activity levels. It is possible that MAchR dysfunction is involved in sleep-disordered breathing, Such as obstructive sleep apnea. (C) 2002 Elsevier Science B.V. All rights reserved.

Spondylostrobus F. Mueller: operculate fruits of an extinct dicotyledon from the mid-Tertiary of Australia

Spondylostrobus F. Mueller, which accommodates operculate fruit-stones reported only from the mid-Tertiary of Australia, is redefined on the basis of type and other specimens of the type species, S. smythii F. Mueller, and of specimens included in S. rozefeldsii sp. nov. The globose to ellipsoidal fruits have 3-6 locules symmetrically disposed around a massive fibrous axis. Each locule has a single anatropous ovule, axile placentation, and a dorsal germination operculum that extends from near the base to the apex. In possessing these characters Spondylostrobus more closely resembles operculate fruits within the tribe Spondiadeae (Anacardiaceae) than operculate fruits of other dicotyledonous families. Spondylostrobus has widespread distribution in Oligocene-Miocene sediments of eastern Australia. At many localities it is associated with fruit-stones having affinities with extant taxa that now occur in rainforests, monsoonal forests, and fringing communities of northern Australia. (C) 2002 Elsevier Science B.V. All rights reserved.

Porous solids from layered clays by combined pillaring and templating approaches

The pore structure formation in bentonite, pillared with a mixed sol of silicon and titanium hydroxides and treated subsequently with quaternary ammonium surfactants, is investigated. The surfactant micelles act as a template, similar to their role in MCM41 synthesis. Because both the surfactant micelles and the sol particles are positively charged, it is greatly favorable for them to form meso-phase assembles in the galleries between the clay layers that bear negative charges. Besides, the sol particles do not bond the clay layers strongly as other kinds of pillar precursors do, so that the treatment with surfactants can result in radical structure changes in sol-pillared clays. This allows us to tailor the pore structure of these porous clays by choice of surfactant. The surfactant treatment also results in profound increases in porosity and improvement in thermal stability. Therefore, the product porous clays have great potential to be Used to deal with large molecules or at high operating temperatures. We also found that titanium in these samples is highly dispersed in the silica matrix rather than existing in the form of small particles of pure titania. Such highly dispersed Ti active centers may offer excellent activities for catalytic oxidation reactions such as alkanes into alcohols and ketones.

Exogenous Slit2 does not affect ureteric branching or nephron formation during kidney development

In an attempt to elucidate the role of Slit2 invertebrate kidney development, the effect of adding exogenous human Slit2 protein (hSlit2) to developing murine metanephric kidney explants was examined. To confirm the activity of the recombinant Slit2 protein, neurons from 8 day old chick sympathetic nerve chain dorsal root ganglia were cultured with hSlit2 protein, which induced significant neurite branching and outgrowth. Using kidney explants as a model system, metanephric development in the presence of hSlit2 protein was examined. Addition of hSlit2 up to a final concentration of 1 mug/ml had no detectable effect on the formation of nephrons or on branching morphogenesis of the ureteric tree after 2 or 4 days in culture, as assessed via immunofluorescence for the markers WT1 and calbindin 28K respectively. Similarly, maturation of the nephrogenic mesenchyme occurred in a phenotypically normal fashion. In situ analysis of the Slit receptors, Robot and Robot, the vasculogenic markers VEGFA and Flk-1, and the stromal cell marker BF2 displayed no difference in comparison to controls.