Genetic dissection of a regionally differentiated network for exploratory behavior in Drosophila larvae.

An efficient strategy to explore the environment for available resources involves the execution of random walks where straight line locomotion alternates with changes of direction. This strategy is highly conserved in the animal kingdom, from zooplankton to human hunter-gatherers. Drosophila larvae execute a routine of this kind, performing straight line crawling interrupted at intervals by pause turns that halt crawling and redirect the trajectory of movement. The execution of this routine depends solely on the activity of networks located in the thoracic and abdominal segments of the nervous system, while descending input from the brain serves to modify it in a context-dependent fashion. I used a genetic method to investigate the location and function of the circuitry required for the different elements of exploratory crawling. By using the Slit-Robo axon guidance pathway to target neuronal midline crossing defects selectively to particular regions of the thoracic and abdominal networks, it has been possible to define at least three functions required for the performance of the exploratory routine: (1) symmetrical outputs in thoracic and abdominal segments that generate the crawls; (2) asymmetrical output that is uniquely initiated in the thoracic segments and generates the turns; and (3) an intermittent interruption to crawling that determines the time-dependent transition between crawls and turns.J.B. was funded by EMBO long-term fellowship, by the Wellcome Trust Institutional Strategic Support Fund, by the Department of Zoology at University of Cambridge, and by a Sir Henry Dale Fellowship (Wellcome Trust and the Royal Society) Grant 105568/Z/14/Z. The work benefited from facilities supported by Wellcome Trust Equipment Grant WT079204.This is the final published version. It first appeared at http://www.sciencedirect.com/science/article/pii/S0960982215003383

Imaging fictive locomotor patterns in larval Drosophila.

We have established a preparation in larval Drosophila to monitor fictive locomotion simultaneously across abdominal and thoracic segments of the isolated CNS with genetically encoded Ca(2+) indicators. The Ca(2+) signals closely followed spiking activity measured electrophysiologically in nerve roots. Three motor patterns are analyzed. Two comprise waves of Ca(2+) signals that progress along the longitudinal body axis in a posterior-to-anterior or anterior-to-posterior direction. These waves had statistically indistinguishable intersegmental phase delays compared with segmental contractions during forward and backward crawling behavior, despite being ∼10 times slower. During these waves, motor neurons of the dorsal longitudinal and transverse muscles were active in the same order as the muscle groups are recruited during crawling behavior. A third fictive motor pattern exhibits a left-right asymmetry across segments and bears similarities with turning behavior in intact larvae, occurring equally frequently and involving asymmetry in the same segments. Ablation of the segments in which forward and backward waves of Ca(2+) signals were normally initiated did not eliminate production of Ca(2+) waves. When the brain and subesophageal ganglion (SOG) were removed, the remaining ganglia retained the ability to produce both forward and backward waves of motor activity, although the speed and frequency of waves changed. Bilateral asymmetry of activity was reduced when the brain was removed and abolished when the SOG was removed. This work paves the way to studying the neural and genetic underpinnings of segmentally coordinated motor pattern generation in Drosophila with imaging techniques.S.R.P. was supported by a Newton International Fellowship (Royal Society) and a Junior Fellowship (Janelia Research Campus, Howard Hughes Medical Institute). T.G.B. was supported by a Medical Research Council (UK) PhD grant. J.B. was supported by a Henry Dale Fellowship (Royal Society and Wellcome Trust). M.B. was supported by the Isaac Newton Trust.This is the final version of the article. It first appeared from the American Physiological Society via http://dx.doi.org/10.1152/jn.00731.201

Drosophila para bss Flies as a Screening Model for Traditional Medicine: Anticonvulsant Effects of Annona senegalensis.

Epilepsy is among the most common serious neurological disorders and affects around 50 million people worldwide, 80% of which live in developing countries. Despite the introduction of several new Anti-Epileptic Drugs (AEDs) in the last two decades, one third of treated patients have seizures refractory to pharmacotherapy. This highlights the need to develop new treatments with drugs targeting alternative seizure-induction mechanisms. Traditional medicine (TM) is used for the treatment of epilepsy in many developing countries and could constitute an affordable and accessible alternative to AEDs, but a lack of pre-clinical and clinical testing has so far prevented its wider acceptance worldwide. In this study we used Drosophila melanogaster paralytic bangsensitive (para bss ) mutants as a model for epileptic seizure screening and tested, for the first time, the anti-seizure effect of a non-commercial AED. We evaluated the effect of the African custard-apple, Annona senegalensis, which is commonly used as a TM for the treatment of epilepsy in rural Africa, and compared it with the classical AED phenytoin. Our results showed that a stem bark extract from A. senegalensis was significantly more effective than a leaf extract and similar to phenytoin in the prevention and control of seizure-like behavior. These results support that Drosophila constitutes a robust animal model for the screening of TM with potential value for the treatment of intractable epilepsy

Purriato is a conserved small open reading frame gene that interacts with the CASA pathway to regulate muscle homeostasis and epithelial tissue growth in Drosophila

Recent advances in proteogenomic techniques and bioinformatic pipelines have permitted the detection of thousands of translated small Open Reading Frames (smORFs), which contain less than 100 codons, in eukaryotic genomes. Hundreds of these actively translated smORFs display conserved sequence, structure and evolutionary signatures indicating that the translated peptides could fulfil important biological roles. Despite their abundance, only tens of smORF genes have been fully characterised; these act mainly as regulators of canonical proteins involved in essential cellular processes. Importantly, some of these smORFs display conserved functions with their mutations being associated with pathogenesis. Thus, investigating smORF roles in Drosophila will not only expand our understanding of their functions but it may have an impact in human health. Here we describe the function of a novel and essential Drosophila smORF gene named purriato (prto). prto belongs to an ancient gene family whose members have expanded throughout the Protostomia clade. prto encodes a transmembrane peptide which is localized in endo-lysosomes and perinuclear and plasma membranes. prto is dynamically expressed in mesodermal tissues and imaginal discs. Targeted prto knockdown (KD) in these organs results in changes in nuclear morphology and endo-lysosomal distributions correlating with the loss of sarcomeric homeostasis in muscles and reduction of mitosis in wing discs. Consequently, prto KD mutants display severe reduction of motility, and shorter wings. Finally, our genetic interaction experiments show that prto function is closely associated to the CASA pathway, a conserved mechanism involved in turnover of mis-folded proteins and linked to muscle dystrophies and neurodegenerative diseases. Thus, this study shows the relevance of smORFs in regulating important cellular functions and supports the systematic characterisation of this class of genes to understand their functions and evolution

Circadian Remodeling of Neuronal Circuits Involved in Rhythmic Behavior

Clock output pathways are central to convey timing information from the circadian clock to a diversity of physiological systems, ranging from cell-autonomous processes to behavior. While the molecular mechanisms that generate and sustain rhythmicity at the cellular level are well understood, it is unclear how this information is further structured to control specific behavioral outputs. Rhythmic release of pigment dispersing factor (PDF) has been proposed to propagate the time of day information from core pacemaker cells to downstream targets underlying rhythmic locomotor activity. Indeed, such circadian changes in PDF intensity represent the only known mechanism through which the PDF circuit could communicate with its output. Here we describe a novel circadian phenomenon involving extensive remodeling in the axonal terminals of the PDF circuit, which display higher complexity during the day and significantly lower complexity at nighttime, both under daily cycles and constant conditions. In support to its circadian nature, cycling is lost in bona fide clockless mutants. We propose this clock-controlled structural plasticity as a candidate mechanism contributing to the transmission of the information downstream of pacemaker cells

A Functional Misexpression Screen Uncovers a Role for Enabled in Progressive Neurodegeneration

Drosophila is a well-established model to study the molecular basis of neurodegenerative diseases. We carried out a misexpression screen to identify genes involved in neurodegeneration examining locomotor behavior in young and aged flies. We hypothesized that a progressive loss of rhythmic activity could reveal novel genes involved in neurodegenerative mechanisms. One of the interesting candidates showing progressive arrhythmicity has reduced enabled (ena) levels. ena down-regulation gave rise to progressive vacuolization in specific regions of the adult brain. Abnormal staining of pre-synaptic markers such as cystein string protein (CSP) suggest that axonal transport could underlie the neurodegeneration observed in the mutant. Reduced ena levels correlated with increased apoptosis, which could be rescued in the presence of p35, a general Caspase inhibitor. Thus, this mutant recapitulates two important features of human neurodegenerative diseases, i.e., vulnerability of certain neuronal populations and progressive degeneration, offering a unique scenario in which to unravel the specific mechanisms in an easily tractable organism

PRRG4 function reveals that Robo trafficking is evolutionarily conserved

Achieving a correct set of neuronal connections during development is key for a healthy functioningnervous system. Autism, which is characterised by impairments in social interaction,language, and range of interests, has been hypothesised to originate from defective synapticfunction and abnormal brain connectivity [1,2]. Moreover, genetic alterations such as the deficiencyin proline-rich carboxyglutamic acid protein 4 (PRRG4) have been associated withautistic features present in WAGR syndrome (Wilm´s tumour, aniridia, genitourinary anomaliesand ªmental retardationº). Therefore, understanding the genetic mechanisms underlyingthe assembly of brain circuits is likely to be essential for the design of new diagnostic tools andtherapeutic strategies for Autistic Spectrum Disorders (ASD). In this issue of PLOS Genetics,Justice et al. link genetic alterations and neural circuitry development, revealing a novel rolefor the PRRG4 as a regulator of Roundabout (Robo) receptor subcellular localization in thenervous system [3].Fil: Berni, Jimena. University of Cambridge; Reino Unido. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

How larvae feel the world around them

A complete map of the external sense organs shows how fruit fly larvae detect different aspects of their environment

The fundamentals of flying:simple and inexpensive strategies for employing Drosophila genetics in neuroscience teaching laboratories

Drosophila researchers have developed a powerful suite of genetic techniques for studying the neural basis of animal behavior. Many of these tools can be exported to neuroscience teaching laboratories (Berni et al., 2010; Pulver et al., 2011a,b), but often neuroscience educators lack the basic knowledge and resources to obtain, generate and rear transgenic fruit flies on their own. Fly researchers in turn may take for granted resources that are readily available in research laboratories, but out of reach for educators. Our goal is to provide a primer for neuroscience educators who want to incorporate Drosophila genetics into their teaching, but have limited knowledge of fruit fly genetics, and/or small budgets. First we review the available methods for manipulating gene expression in Drosophila. Then we provide educators with blueprints for obtaining transgenic animals tailored for specific types of teaching modules. We outline simple techniques for rearing transgenic Drosophila, performing genetic crosses, and preparing a teaching laboratory without the use of expensive animal-care facilities. Overall, we try to break down the practical barriers educators may face when integrating modern neurogenetic experiments into teaching laboratories