high-throughput-assay-to-examine-egg-laying-preferences-individual

High Throughput Assay to Examine Egg-Laying Preferences of Individual <em>Drosophila melanogaster</em>

Recently, egg-laying preference of Drosophila has emerged as a genetically tractable model to study the neural basis of simple decision-making processes. ...

Duke University

RFamide-related peptide (RFRP), the mammalian homolog of avian gonadotropin-inhibitory hormone, has a pronounced suppressive action on the reproductive axis across species. In mammals, RFRP acts directly on GnRH neurons, and likely at the level of the pituitary, to inhibit gonadotropin secretion. In the present study, we examined whether RFRP might act outside of mammalian brain on reproductive tissues directly. Using RT-PCR and in situ hybridization, we found that both RFRP and its receptors [G protein-coupled receptor (GPR) 147 and GPR74] are expressed in the testis of Syrian hamster. These results were confirmed and extended using double- and triple-label immunohistochemistry. RFRP expression was observed in spermatocytes and in round to early elongated spermatids. Significant expression of RFRP was not seen in Leydig cells. GPR147 protein was observed in myoid cells in all stages of spermatogenesis, pachytene spermatocytes, maturation division spermatocytes, and in round and late elongated spermatids. GPR74 proteins only appeared in late elongated spermatids. Additionally, we found that RFRP and its receptor mRNA are markedly altered by day length and reproductive condition. These findings highlight a possible novel autocrine and/or paracrine role for RFRP in Syrian hamster testis, potentially contributing to the differentiation of spermatids during spermiogenesis.

RFamide-related peptide and messenger ribonucleic acid expression in mammalian testis: association with the spermatogenic cycle.

Drosophila melanogaster females are highly selective about the chemosensory quality of their egg-laying sites, an important trait that promotes the survival and fitness of their offspring. How egg-laying females respond to UV light is not known, however. UV is a well-documented phototactic cue for adult Drosophila, but it is an aversive cue for larvae. Here, we show that female flies exhibit UV aversion in response to their egg-laying demand. First, females exhibit egg-laying aversion of UV: they prefer to lay eggs on dark sites when choosing between UV-illuminated and dark sites. Second, they also exhibit movement aversion of UV: positional tracking of single females suggests that egg-laying demand increases their tendency to turn away from UV. Genetic manipulations of the retina suggest that egg-laying and movement aversion of UV are both mediated by the inner (R7) and not the outer (R1-R6) photoreceptors. Finally, we show that the Dm8 amacrine neurons, a synaptic target of R7 photoreceptors and a mediator of UV spectral preference, are dispensable for egg-laying aversion but essential for movement aversion of UV. This study suggests that egg-laying demand can temporarily convert UV into an aversive cue for female Drosophila and that R7 photoreceptors recruit different downstream targets to control different egg-laying-induced behavioral modifications.

Egg-laying demand induces aversion of UV light in Drosophila females.

High-throughput analysis of animal behavior requires software to analyze videos. Such software typically depends on the experiments' being performed in good lighting conditions, but this ideal is difficult or impossible to achieve for certain classes of experiments. Here, we describe techniques that allow long-duration positional tracking in difficult lighting conditions with strong shadows or recurring "on"/"off" changes in lighting. The latter condition will likely become increasingly common, e.g., for Drosophila due to the advent of red-shifted channel rhodopsins. The techniques enabled tracking with good accuracy in three types of experiments with difficult lighting conditions in our lab. Our technique handling shadows relies on single-animal tracking and on shadows' and flies' being accurately distinguishable by distance to the center of the arena (or a similar geometric rule); the other techniques should be broadly applicable. We implemented the techniques as extensions of the widely-used tracking software Ctrax; however, they are relatively simple, not specific to Drosophila, and could be added to other trackers as well.

Article	Year
RFamide-related peptide and messenger ribonucleic acid expression in mammalian testis: association with the spermatogenic cycle. Endocrinology\| PubMed ID: 20051487	2010
Egg-laying demand induces aversion of UV light in Drosophila females. Current biology : CB\| PubMed ID: 25455037	2014
Long-duration animal tracking in difficult lighting conditions. Scientific reports\| PubMed ID: 26130571	2015

Edward Zhu

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