Properties of a potent LHRH antagonist (Org 30850) in female and male rats.

Org 30850 (Ac-D-pClPhe1,2,D-Bal3,D-Lys6,D-Ala10-LHRH) is a novel LHRH antagonist, which is being developed for the treatment of hormone-dependent disorders. The activities of this compound with respect to its endocrinological properties and side-effects were tested in rats and the results were compared with one of the first LHRH antagonists: Ac-D-pClPhe1,2,D-Trp3,D-Arg6,D-Ala10-LHRH (Org 30276). A single subcutaneous (s.c.) dose of 0.3 micrograms/kg Org 30850 administered to rats in pro-estrus gave inhibition of ovulation in approx. 50% of the rats, whereas Org 30276 was approx. 4 times less potent. The effect of a single s.c. injection of Org 30850 on testosterone levels in young adult male rats was also studied. The administration of 250 micrograms/kg or higher of Org 30850 induced a significant decrease in testosterone levels after 3 h, this effect lasted for at least 48 h. Treatment of female rats for 14 days with a daily dose of 12 micrograms/kg Org 30850 decreased statistically significantly uterine and ovarian weights. At a daily dose of 50 micrograms/kg Org 30850 completely suppressed estrous cycles and significantly decreased estradiol and FSH serum levels. The LH levels were below the detection level in both control and treated animals on the (expected) second day of di-estrus. Treatment of male rats for 14 days (25-200 micrograms/kg) resulted in a dose-dependent reduction of the gonads, accessory sex organs, testosterone levels and gonadotrophins. The decrease in gonadal function in both sexes was reversible since the females proved to be as fertile as the controls 6 weeks after the last treatment and an almost complete recovery of the weight of testes, seminal vesicles and ventral prostate was observed in the males 4 weeks after cessation of treatment. In contrast to Org 30276, Org 30850 exerted very slight irritation at the site of injection and no edematous reactions in the extremities at a daily dose of up to 8 mg/kg in male rats. It is concluded that Org 30850 is a very potent LHRH antagonist without edematous reactions and with a more favourable therapeutic index than Org 30276.     

Identification of a large family of genes for putative chemoreceptor proteins in an ordered library of the Desulfovibrio vulgaris Hildenborough genome.

A library of 879 recombinant lambda phages, constructed for the genome of Desulfovibrio vulgaris Hildenborough, has been ordered by restriction fingerprinting. Restriction endonuclease HinfI digestion patterns were entered into a data base and sorted into 87 overlapping groups (contigs), with 19 clones remaining unattached. Eight of ten cloned genes of D. vulgaris, including dcrA, which encodes a transmembrane methyl-accepting protein, were assigned to contigs. Probing of a filter containing the lambda DNAs of the library with the labeled, conserved 3' end of the dcrA gene indicated hybridization to 54 clones distributed over multiple contigs. The presence of 11 additional dcr genes (dcrB to dcrL) was confirmed by direct cycled dideoxy sequencing of positive lambda clones. Since the ordered library provides only partial coverage of the D. vulgaris Hildenborough genome, we estimate that the dcr gene family has 16 members spread throughout the genome, making it the second largest gene family found in prokaryotes.     

How Long Depends on How Fast—Perceived Flicker Dilates Subjective Duration

How do humans perceive the passage of time and the duration of events without a dedicated sensory system for timing? Previous studies have demonstrated that when a stimulus changes over time, its duration is subjectively dilated, indicating that duration judgments are based on the number of changes within an interval. In this study, we tested predictions derived from three different accounts describing the relation between a changing stimulus and its subjective duration as either based on (1) the objective rate of changes of the stimulus, (2) the perceived saliency of the changes, or (3) the neural energy expended in processing the stimulus. We used visual stimuli flickering at different frequencies (4–166 Hz) to study how the number of changes affects subjective duration. To this end, we assessed the subjective duration of these stimuli and measured participants'' behavioral flicker fusion threshold (the highest frequency perceived as flicker), as well as their threshold for a frequency-specific neural response to the flicker using EEG. We found that only consciously perceived flicker dilated perceived duration, such that a 2 s long stimulus flickering at 4 Hz was perceived as lasting as long as a 2.7 s steady stimulus. This effect was most pronounced at the slowest flicker frequencies, at which participants reported the most consistent flicker perception. Flicker frequencies higher than the flicker fusion threshold did not affect perceived duration at all, even if they evoked a significant frequency-specific neural response. In sum, our findings indicate that time perception in the peri-second range is driven by the subjective saliency of the stimulus'' temporal features rather than the objective rate of stimulus changes or the neural response to the changes.     

Paradoxical Sensitivity to an Integrated Stress Response Blocking Mutation in Vanishing White Matter Cells

The eukaryotic translation initiation factor eIF2B promotes mRNA translation as a guanine nucleotide exchange factor (GEF) for translation initiation factor 2 (eIF2). Endoplasmic reticulum (ER) stress-mediated activation of the kinase PERK and the resultant phosphorylation of eIF2’s alpha subunit (eIF2α) attenuates eIF2B GEF activity thereby inducing an integrated stress response (ISR) that defends against protein misfolding in the ER. Mutations in all five subunits of human eIF2B cause an inherited leukoencephalopathy with vanishing white matter (VWM), but the role of the ISR in its pathogenesis remains unclear. Using CRISPR-Cas9 genome editing we introduced the most severe known VWM mutation, EIF2B4^A391D, into CHO cells. Compared to isogenic wildtype cells, GEF activity of cells with the VWM mutation was impaired and the mutant cells experienced modest enhancement of the ISR. However, despite their enhanced ISR, imposed by the intrinsic defect in eIF2B, disrupting the inhibitory effect of phosphorylated eIF2α on GEF by a contravening EIF2S1/eIF2α^S51A mutation that functions upstream of eIF2B, selectively enfeebled both EIF2B4^A391D and the related severe VWM EIF2B4^R483W cells. The basis for paradoxical dependence of cells with the VWM mutations on an intact eIF2α genotype remains unclear, as both translation rates and survival from stressors that normally activate the ISR were not reproducibly affected by the VWM mutations. Nonetheless, our findings support an additional layer of complexity in the development of VWM, beyond a hyperactive ISR.     

Using simulations to evaluate Mantel‐based methods for assessing landscape resistance to gene flow

Mantel‐based tests have been the primary analytical methods for understanding how landscape features influence observed spatial genetic structure. Simulation studies examining Mantel‐based approaches have highlighted major challenges associated with the use of such tests and fueled debate on when the Mantel test is appropriate for landscape genetics studies. We aim to provide some clarity in this debate using spatially explicit, individual‐based, genetic simulations to examine the effects of the following on the performance of Mantel‐based methods: (1) landscape configuration, (2) spatial genetic nonequilibrium, (3) nonlinear relationships between genetic and cost distances, and (4) correlation among cost distances derived from competing resistance models. Under most conditions, Mantel‐based methods performed poorly. Causal modeling identified the true model only 22% of the time. Using relative support and simple Mantel r values boosted performance to approximately 50%. Across all methods, performance increased when landscapes were more fragmented, spatial genetic equilibrium was reached, and the relationship between cost distance and genetic distance was linearized. Performance depended on cost distance correlations among resistance models rather than cell‐wise resistance correlations. Given these results, we suggest that the use of Mantel tests with linearized relationships is appropriate for discriminating among resistance models that have cost distance correlations <0.85 with each other for causal modeling, or <0.95 for relative support or simple Mantel r. Because most alternative parameterizations of resistance for the same landscape variable will result in highly correlated cost distances, the use of Mantel test‐based methods to fine‐tune resistance values will often not be effective.