Paired box mutations in familial and sporadic aniridia predicts truncated aniridia proteins.

Aniridia, an autosomal dominant ocular disorder characterized by iris hypoplasia, results from mutations in the PAX6 gene, which encodes paired box and homeobox motifs. In this report we describe five new mutations in the paired box region of the human PAX6 gene that are associated with aniridia. The paired box mutations that we detected were in both familial (three) and sporadic (two) cases. All five mutations predict truncated PAX6 proteins. Our study indicates that early premature translational termination mutations in the PAX6 gene result in haploinsufficiency and generate the aniridia phenotype.     

Discharge patterns of chicken cochlear ganglion neurons following kanamycin-induced hair cell loss and regeneration

Hair cells in the basal, high frequency region (>1100 Hz) of the chicken cochlea were destroyed with kanamycin (400 mg/kg/d × 10 d) and allowed to regenerate. Afterwards, single unit recordings were made from cochlear ganglion neurons at various times post-treatment. During the first few weeks post-treatment, only neurons with low characteristic frequencies (<1100 Hz) responded to sound. Despite the fact that the low frequency region of the cochlea was not destroyed, neurons with low characteristic frequencies had elevated thresholds, abnormally broad U-shaped or W-shaped tuning curves and low spontaneous discharge rates. At 2 days post-treatment, the spontaneous discharge rates of some acoustically unresponsive units fluctuated in a rhythmical manner. As recovery time increased, thresholds decreased, tuning curves narrowed and developed a symmetrical V-shape, spontaneous rate increased and neurons with higher characteristic frequencies began to respond to sound. In addition, the proportion of interspike interval histograms with regularly spaced peaks increased. These improvements progressed along a low-to-high characteristic frequency gradient. By 10–20 weeks post-treatment, the thresholds and tuning curves of neurons with characteristic frequencies below 2000 Hz were within normal limits; however, the spontaneous discharge rates of the neurons were still significantly lower than those from normal animals.Abbreviations KM kanamycin - BrdU bromodeoxyuridine - CF characteristic frequency - CAP compound action potential - ISI interspike interval     

Middle-ear development

Laser interferometry was used to measure umbo velocity in the developing BALB/c mouse middle ear at 133 pure-tone frequencies between 2.0 kHz and 40.0 kHz, all at a constant 100 dB sound pressure level. Umbo velocities increased with age across the entire frequency range, and reached adult-like levels by about 19 days between 2.0 and 22.0 kHz. Velocities at 28.0 and 34.0 kHz took 27 and 52 days respectively to reach adult-like levels.A simple middle-ear model utilizing compliance, resistance, and inertia elements matched the general trends of our velocity results and provided an indication of the anatomical basis for the growth in umbo velocity. The model suggested that velocity development at the lowest frequencies may be attributed to increases in tympanic membrane compliance. The model also indicated that both the frictional resistance of the middle ear and the inertia of the tympanic membrane and ossicles decreased during the growth period.At frequencies below 20.0 kHz, age-related increases in umbo velocity coincided with improvements in Nj₁ thresholds recorded from the round window and evoked potential thresholds obtained from the cochlear nucleus. These results indicated that the functional development of the middle-ear plays a major role in the development of hearing in the mouse.Portions of this work were presented at the Fifteenth Meeting of the Association for Research in Otolaryngology     

Phase shifts of the adult locomotor activity rhythm in Calliphora vicina induced by non-steroidal ecdysteroid agonist RH 5849

RH 5849, a non-steroidal ecdysteroid mimic, was found to cause consistent phase shifts in the circadian rhythm of locomotor activity of the blowfly, Calliphora vicina. This compound causes phase advances in the early subjective night and phase delays in the late subjective night. This effect is the opposite, but not the mirror image of the phase response curve obtained for 1 h light pulses. This suggests that ecdysteroids might act as entraining agents via the output pathway by feedback to clock neurons in the brain. A computer model based on 12 pacemaker neurons with circadian periods ( values) from short to long without simulated feedback from the ecdysteroid system becomes arrhythmic; with feedback, the oscillators become synchronized to a common period. The possible role of ecdysteroids as endogenous synchronizing agents in the insect circadian system is discussed.     

Direct comparison of bromodeoxyuridine and Ki-67 labelling indices in human tumours

Direct comparison of bromodeoxyuridine (BrdUrd) and Ki-67 labelling indices was achieved by selecting similar areas from serial sections of human tumours. Fifteen patients were selected who had been administered BrdUrd in vivo and both proliferation markers were assessed by immunohistochemistry. The data show a good correlation between both BrdUrd LI and MIB-1 LI and T_pot (calculated using the flow cytometry derived duration of S phase) and MIB-1 LI. The contribution of BrdUrd LI to growth fraction varied as a function of proliferation characteristics. In tumours with a high LI, the number of DNA synthesizing cells represented half the growth fraction, whilst in tumours with lower LI's (<10%) the ratio of DNA precursor labelled cells as a function of growth fraction fell to between 10% and 20%. T_pot showed a linear correlation with MIB-1/BrdUrd ratio with a slope approaching unity. It was apparent that both intra- and interpatient variation in proliferation index was greater for BrdUrd labelling than for MIB-1 expression.     

Mandible Form Relative to the Main Food Type in Ladybirds (Coleoptera: Coccinellidae)

The Coccinellidae is an economically important family within the Coleoptera. Some members are phytophagous pests, but many are beneficial predators and valuable biocontrol agents. This study investigates the morphology of the mandibles of adult Coccinellidae in relation to diet. Using scanning electron microscopy on 86 species of Coccinellidae, it was found that the morphology of the mandibles was dictated by the general feeding method, and could only be used to indicate a phytophagous, mycophagous or carnivorous diet. Phytophagous Coccinellidae of the subfamily Epilachninae had mandibles with denticulate apical teeth and setae for feeding on leaf material. The mandibles of the mycophagous Psylloborini had secondary teeth on the ventral apical tooth for collecting fungal spores. The mandibles of carnivorous Coccinellidae and Scymninae had either a bifid or unidentate apex. The unidentate mandible seemed to be restricted to coccidophagous species. Many species also had a mandibular groove along which prey body juices were conducted. Although mandible morphology could be related to the overall feeding method, there was no relationship between specific diet or food taxon and mandible shape. Mandible shape does not appear to be especially restricting for changes in diet either in the ecological sense or over evolutionary time. Mandible morphology is of limited use in determining diet and host specificity in Coccinellidae that are being selected as potential biocontrol agents.     

P-Element Insertion Alleles of Essential Genes on the Third Chromosome of Drosophila Melanogaster: Correlation of Physical and Cytogenetic Maps in Chromosomal Region 86e-87f

We have established a collection of 2460 lethal or semi-lethal mutant lines using a procedure thought to insert single P elements into vital genes on the third chromosome of Drosophila melanogaster. More than 1200 randomly selected lines were examined by in situ hybridization and 90% found to contain single insertions at sites that mark 89% of all lettered subdivisions of the Bridges' map. A set of chromosomal deficiencies that collectively uncover ~25% of the euchromatin of chromosome 3 reveal lethal mutations in 468 lines corresponding to 145 complementation groups. We undertook a detailed analysis of the cytogenetic interval 86E-87F and identified 87 P-element-induced mutations falling into 38 complementation groups, 16 of which correspond to previously known genes. Twenty-one of these 38 complementation groups have at least one allele that has a P-element insertion at a position consistent with the cytogenetics of the locus. We have rescued P elements and flanking chromosomal sequences from the 86E-87F region in 35 lines with either lethal or genetically silent P insertions, and used these as probes to identify cosmids and P1 clones from the Drosophila genome projects. This has tied together the physical and genetic maps and has linked 44 previously identified cosmid contigs into seven ``supercontigs' that span the interval. STS data for sequences flanking one side of the P-element insertions in 49 lines has identified insertions in the αγ element at 87C, two known transposable elements, and the open reading frames of seven putative single copy genes. These correspond to five known genes in this interval, and two genes identified by the homology of their predicted products to known proteins from other organisms.     

The Drosophila Gene abnormal spindle Encodes a Novel Microtubule-associated Protein That Associates with the Polar Regions of the Mitotic Spindle

abnormal spindle, a gene required for normal spindle structure and function in Drosophila melanogaster, lies immediately adjacent the gene tolloid at 96A/B. It encodes a 220-kD polypeptide with a predicted pI of 10.8. The recessive mutant allele asp¹ directs the synthesis of a COOH terminally truncated or internally deleted peptide of ~124 kD. Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins. The bacterially expressed NH₂-terminal 512-amino acid peptide, which has a number of potential phosphorylation sites for p34^cdc2 and MAP kinases, strongly binds to microtubules. The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins. Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome. These findings are discussed in relation to the known spindle abnormalities in asp mutants.     

Protein Phosphorylation and Calcium Uptake into Rat Forebrain Synaptosomes: Modulation by the σ Ligand, 1,3-Ditolylguanidine

Abstract: The σ ligand 1,3-di- O -tolylguanidine (DTG) increased basal dynamin and decreased depolarization-stimulated phosphorylation of the synaptosomal protein synapsin Ib without having direct effects on protein kinases or protein phosphatases. DTG dose-dependently decreased the basal cytosolic free Ca²⁺ concentration ([Ca²⁺]_i) and blocked the depolarization-dependent increases in [Ca²⁺]_i. These effects were inhibited by the σ antagonists rimcazole and BMY14802. The nitric oxide donors sodium nitroprusside (SNP) and 8-( p -chlorophenylthio)guanosine-3',5'-cyclic monophosphorothioate decreased basal [Ca²⁺]_i and the KCl-evoked rise in [Ca²⁺]_i to an extent similar to DTG. SNP, but not DTG, produced a rise in cyclic GMP levels, suggesting that the effect of DTG on [Ca²⁺]_i was not mediated via downstream regulation of cyclic GMP levels. DTG increased ⁴⁵Ca²⁺ uptake and efflux under basal conditions and inhibited the ⁴⁵Ca²⁺ uptake induced by depolarization with KCl. The KCl-evoked rise in [Ca²⁺]_i was inhibited by ω-conotoxin (ω-CgTx)-GVIA and -MVIIC but not nifedipine and ω-agatoxin-IVA. The effect of DTG on decreasing the KCl-evoked rise in [Ca²⁺]_i was additive with ω-CgTx-MVIIC but not with ω-CgTx-GVIA. These data suggest that DTG was producing some of its effects on synapsin I and dynamin phosphorylation and intrasynaptosomal Ca²⁺ levels via inhibition of N-type Ca²⁺ channels.     

A PHYLOGENETIC ANALYSIS OF THE SYNUROPHYCEAE USING MOLECULAR DATA AND SCALE CASE MORPHOLOGY

The phylogeny of the Synurophyceae was investigated by parsimony analysis of scale case characters and small-sub unit (18S) ribosomal RNA (rRNA) sequence data. Analysis of 1 eustigmatophycean (outgroup), 3 chrysophycean, and 10 synurophycean 18S rRNA sequences corroborated the inference from ultrastructural information that the Synurophyceae is a monophyletic assemblage . Tessellaria vol-vocina, which had been tentatively proposed as a member of the Synurophyceae, was confirmed as the earliest lineage within the Synurophyceae by both the molecular analysis and an evaluation of published ultrastructural data. A second set of analyses investigated the relationships among Tessellaria volvocina, 6 Synura species, and 10 Mallomonas species/varieties, with particular reference to the validity of current classifications of the Synurophyceae and the characters upon which they are based. The molecular and scale case phylogenies were not totally resolved but were largely congruent. The data sets were combined to produce another phylogeny, which showed greater resolution. The combined phylogeny weakly supported our representatives of Synura and Mallomonas as monophyletic groupings and also upheld several of the sections within these genera that are recognized by current classifications. However, some changes to the classification and delineation of these genera are recommended and predicted. Both our 18S rRNA sequence and scale case data sets were more appropriate for examining the branching order among the more closely related text rather than resolving the deeper branching points of the synurophycean phylogeny .