Zebrafish Reproduction: Revisiting In Vitro Fertilization to Increase Sperm Cryopreservation Success

Although conventional cryopreservation is a proven method for long-term, safe storage of genetic material, protocols used by the zebrafish community are not standardized and yield inconsistent results, thereby putting the security of many genotypes in individual laboratories and stock centers at risk. An important challenge for a successful zebrafish sperm cryopreservation program is the large variability in the post-thaw in vitro fertilization success (0 to 80%). But how much of this variability was due to the reproductive traits of the in vitro fertilization process, and not due to the cryopreservation process? These experiments only assessed the in vitro process with fresh sperm, but yielded the basic metrics needed for successful in vitro fertilization using cryopreserved sperm, as well. We analyzed the reproductive traits for zebrafish males with a strict body condition range. It did not correlate with sperm volume, or motility (P>0.05), but it did correlate with sperm concentration. Younger males produced more concentrated sperm (P<0.05). To minimize the wastage of sperm during the in vitro fertilization process, 106 cells/ml was the minimum sperm concentration needed to achieve an in vitro fertilization success of ≥ 70%. During the in vitro process, pooling sperm did not reduce fertilization success (P>0.05), but pooling eggs reduced it by approximately 30 to 50% (P<0.05). This reduction in fertilization success was due not to the pooling of the females' eggs, but to the type of tools used to handle the eggs. Recommendations to enhance the in vitro process for zebrafish include: 1) using males of a body condition closer to 1.5 for maximal sperm concentration; 2) minimizing sperm wastage by using a working sperm concentration of 106 motile cells/ml for in vitro fertilization; and 3) never using metal or sharp-edged tools to handle eggs prior to fertilization

Human and Chimpanzee Gene Expression Differences Replicated in Mice Fed Different Diets

Although the human diet is markedly different from the diets of closely related primate species, the influence of diet on phenotypic and genetic differences between humans and other primates is unknown. In this study, we analyzed gene expression in laboratory mice fed diets typical of humans and of chimpanzees. The effects of human diets were found to be significantly different from that of a chimpanzee diet in the mouse liver, but not in the brain. Importantly, 10% of the genes that differ in their expression between humans and chimpanzee livers differed also between the livers of mice fed the human and chimpanzee diets. Furthermore, both the promoter sequences and the amino acid sequences of these diet-related genes carry more differences between humans and chimpanzees than random genes. Our results suggest that the mouse can be used to study at least some aspects of human-specific traits

The reference frame for encoding and retention of motion depends on stimulus set size

YesThe goal of this study was to investigate the reference frames used in perceptual encoding and storage of visual motion information. In our experiments, observers viewed multiple moving objects and reported the direction of motion of a randomly selected item. Using a vector-decomposition technique, we computed performance during smooth pursuit with respect to a spatiotopic (nonretinotopic) and to a retinotopic component and compared them with performance during fixation, which served as the baseline. For the stimulus encoding stage, which precedes memory, we found that the reference frame depends on the stimulus set size. For a single moving target, the spatiotopic reference frame had the most significant contribution with some additional contribution from the retinotopic reference frame. When the number of items increased (Set Sizes 3 to 7), the spatiotopic reference frame was able to account for the performance. Finally, when the number of items became larger than 7, the distinction between reference frames vanished. We interpret this finding as a switch to a more abstract nonmetric encoding of motion direction. We found that the retinotopic reference frame was not used in memory. Taken together with other studies, our results suggest that, whereas a retinotopic reference frame may be employed for controlling eye movements, perception and memory use primarily nonretinotopic reference frames. Furthermore, the use of nonretinotopic reference frames appears to be capacity limited. In the case of complex stimuli, the visual system may use perceptual grouping in order to simplify the complexity of stimuli or resort to a nonmetric abstract coding of motion information

Transcriptomic Characterization of Temperature Stress Responses in Larval Zebrafish

Temperature influences nearly all biochemical, physiological and life history activities of fish, but the molecular mechanisms underlying the temperature acclimation remains largely unknown. Previous studies have identified many temperature-regulated genes in adult tissues; however, the transcriptional responses of fish larvae to temperature stress are not well understood. In this study, we characterized the transcriptional responses in larval zebrafish exposed to cold or heat stress using microarray analysis. In comparison with genes expressed in the control at 28°C, a total of 2680 genes were found to be affected in 96 hpf larvae exposed to cold (16°C) or heat (34°C) for 2 and 48h and most of these genes were expressed in a temperature-specific and temporally regulated manner. Bioinformatic analysis identified multiple temperature-regulated biological processes and pathways. Biological processes overrepresented among the earliest genes induced by temperature stress include regulation of transcription, nucleosome assembly, chromatin organization and protein folding. However, processes such as RNA processing, cellular metal ion homeostasis and protein transport and were enriched in genes up-regulated under cold exposure for 48 h. Pathways such as mTOR signalling, p53 signalling and circadian rhythm were enriched among cold-induced genes, while adipocytokine signalling, protein export and arginine and praline metabolism were enriched among heat-induced genes. Although most of these biological processes and pathways were specifically regulated by cold or heat, common responses to both cold and heat stresses were also found. Thus, these findings provide new interesting clues for elucidation of mechanisms underlying the temperature acclimation in fish

Looming sounds enhance orientation sensitivity for visual stimuli on the same side as such sounds

Several recent multisensory studies show that sounds can influence visual processing. Some visual judgments can be enhanced for visual stimuli near a sound occurring around the same time. A recent TMS study (Romei et al. 2009) indicates looming sounds might influence visual cortex particularly strongly. But unlike most previous behavioral studies of possible audio-visual exogenous effects, TMS phosphene thresholds rather than judgments of external visual stimuli were measured. Moreover, the visual hemifield assessed relative to the hemifield of the sound was not varied. Here, we compared the impact of looming sounds to receding or "static" sounds, using auditory stimuli adapted from Romei et al. (2009), but now assessing any influence on visual orientation discrimination for Gabor patches (well-known to involve early visual cortex) when appearing in the same hemifield as the sound or on the opposite side. The looming sounds that were effective in Romei et al. (2009) enhanced visual orientation sensitivity (d0) here on the side of the sound, but not for the opposite hemifield. This crossmodal, spatially specific effect was stronger for looming than receding or static sounds. Similarly to Romei et al. (2009), the differential effect for looming sounds was eliminated when using white noise rather than structured sounds. Our new results show that looming structured sounds can specifically benefit visual orientation sensitivity in the hemifield of the sound, even when the sound provides no information about visual orientation itself. © The Author(s) 2011