Genetic Manipulation of Genes and Cells in the Nervous System of the Fruit Fly

Venken, Koen J. T.; Simpson, Jeffrey H.; Bellen, Hugo J.

doi:10.1016/j.neuron.2011.09.021

Cited by 402 publications

(358 citation statements)

References 366 publications

Supporting

Mentioning

340

Contrasting

Unclassified

Order By: Relevance

“…Many genetic approaches in Drosophila rely on the FLP/FRT system, including MARCM, recombinase-mediated genetic intersections, and lineage tracing (38,39). To facilitate combining such methods with MCFO, for example, to reveal individual cells in a MARCM clone, we generated stop-cassette reporters in which the FRT sites were replaced by recognition sites for two Flp-related recombinases with orthogonal site specificity, B3 and KD (28) (SI Appendix, Table S1).…”

Section: Mcfo Facilitates Visualization Of Complex Arrangements Of Nementioning

confidence: 99%

Optimized tools for multicolor stochastic labeling reveal diverse stereotyped cell arrangements in the fly visual system

Nern

Pfeiffer

Rubin

2015

Proc. Natl. Acad. Sci. U.S.A.

522

691

View full text Add to dashboard Cite

We describe the development and application of methods for high-throughput neuroanatomy in Drosophila using light microscopy. These tools enable efficient multicolor stochastic labeling of neurons at both low and high densities. Expression of multiple membrane-targeted and distinct epitope-tagged proteins is controlled both by a transcriptional driver and by stochastic, recombinase-mediated excision of transcription-terminating cassettes. This MultiColor FlpOut (MCFO) approach can be used to reveal cell shapes and relative cell positions and to track the progeny of precursor cells through development. Using two different recombinases, the number of cells labeled and the number of color combinations observed in those cells can be controlled separately. We demonstrate the utility of MCFO in a detailed study of diversity and variability of Distal medulla (Dm) neurons, multicolumnar local interneurons in the adult visual system. Similar to many brain regions, the medulla has a repetitive columnar structure that supports parallel information processing together with orthogonal layers of cell processes that enable communication between columns. We find that, within a medulla layer, processes of the cells of a given Dm neuron type form distinct patterns that reflect both the morphology of individual cells and the relative positions of their arbors. These stereotyped cell arrangements differ between cell types and can even differ for the processes of the same cell type in different medulla layers. This unexpected diversity of coverage patterns provides multiple independent ways of integrating visual information across the retinotopic columns and implies the existence of multiple developmental mechanisms that generate these distinct patterns.neuroanatomy | Drosophila | interneuron diversity | light microscopy | recombinase N ervous systems contain numerous and diverse cells displaying complex anatomical relationships. The specification and patterning of these cells must be generated by the execution of a much smaller set of instructions encoded in the genome. How many different genetic algorithms are needed? How precise are their outcomes? What types of rules do they follow? Answering such questions requires knowledge of the anatomy of neuronal processes for many different cell types, for numerous cells of the same type, and in multiple individuals. We describe here the development of a set of methods for collecting such data by light microscopy and their application in the adult visual system of Drosophila.Neuronal morphology is often sufficiently stereotyped to identify cell types. For example, many cell populations in vertebrate and invertebrate visual systems can be reliably distinguished on the basis of cell shape (1-3). In at least some cases, these anatomical cell types have been shown to correlate well with classifications based on genetic marker expression or functional properties. Important anatomical features are not limited to cell shape, but also include the spatial relationships between cells. One critical ...

show abstract

Section: Mcfo Facilitates Visualization Of Complex Arrangements Of Nementioning

confidence: 99%

Optimized tools for multicolor stochastic labeling reveal diverse stereotyped cell arrangements in the fly visual system

Nern

Pfeiffer

Rubin

2015

Proc. Natl. Acad. Sci. U.S.A.

522

691

View full text Add to dashboard Cite

show abstract

“…The fruit fly Drosophila melanogaster offers sophisticated tools for genetic manipulation of the nervous system and has proven to be a robust and relevant system for elucidating neuronal function at the levels of genes, cellular physiology, and neuronal circuits (Venken et al, 2011). More recently, flies have become a valuable experimental system for identifying glial genes and signaling pathways that mediate neuron-glia interactions (Egger et al, 2002;Freeman et al, 2003;Altenhein et al, 2006;Coutinho-Budd and Freeman, 2013).…”

Section: Introductionmentioning

confidence: 99%

Astrocytic glutamate transport regulates a Drosophila CNS synapse that lacks astrocyte ensheathment

MacNamee

Liu

Gerhard

et al. 2016

J of Comparative Neurology

View full text Add to dashboard Cite

Anatomical, molecular, and physiological interactions between astrocytes and neuronal synapses regulate information processing in the brain. The fruit fly Drosophila melanogaster has become a valuable experimental system for genetic manipulation of the nervous system and has enormous potential for elucidating mechanisms that mediate neuron glia interactions. Here, we show the first electrophysiological recordings from Drosophila astrocytes and characterize their spatial and physiological relationship with particular synapses. Astrocyte intrinsic properties were found to be strongly analogous to those of vertebrate astrocytes, including a passive current-voltage relationship, low membrane resistance, high capacitance, and dye-coupling to local astrocytes. Responses to optogenetic stimulation of glutamatergic pre-motor neurons were correlated directly with anatomy using serial electron microscopy reconstructions of homologous identified neurons and surrounding astrocytic processes. Robust bidirectional communication was present: neuronal activation triggered astrocytic glutamate transport via Eaat1, and blocking Eaat1 extended glutamatergic interneuron-evoked inhibitory post-synaptic currents in motor neurons. The neuronal synapses were always located within a micron of an astrocytic process, but none were ensheathed by those processes. Thus, fly astrocytes can modulate fast synaptic transmission via neurotransmitter transport within these anatomical parameters. Graphical AbstractCorresponding Author: Sarah E. MacNamee, University of Arizona Dept. of Neuroscience, 1040 E 4 th St GS Rm 611, Tucson AZ 85721, (520) 621 6671, Smac3@email.arizona.edu. Role of AuthorsAll authors had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: SEM, LAO. Acquisition of data: SEM, KEL, SG, CTT, RDF, AC. Analysis and interpretation of data: SEM, KEL, CTT. Drafting of the manuscript: SEM. Critical revision of the manuscript for important intellectual content: LAO, LPT, AC. Statistical analysis: SEM. Obtained funding: LAO, LPT, AC. Administrative, technical, and material support: none. Study supervision: LAO, LPT. Conflict of Interest StatementThe authors have no conflict of interest to disclose. Data AccessibilityData generated in the lab and the subsequent analyses will be made available upon request to the Oland/Tolbert laboratory by qualified individuals as long as doing so would not compromise intellectual property interests or interfere with publication. Any shared data would include notations, standards, and the like that are necessary for interpretation of the data. HHMI Author Manuscript HHMI Author Manuscript HHMI Author ManuscriptThe authors show the first recordings from Drosophila astrocytes and find that their electrophysiological properties are strongly analogous to those of vertebrate astrocytes. Using correlative electron microscopy and physiology, they found no glial ensheathment of glutamatergic synap...

show abstract

“…These optogenetic techniques use the vast repertoire of the different promoters available in Drosophila 14,16 . These promoters are used to express genetically encoded calcium and cAMP reporters in the memory neurons 16,27 to study the effect of specific gene mutations on memory traces.…”

Section: Discussionmentioning

confidence: 99%

“…3. For experiments using Shibire temperature sensitive to block synaptic output 11,14,24 : Rear the flies at 18 °C and conduct testing at 30 °C. Note: This storage allows the flies to acclimatize to the subsequent learning test performed in the environmentally controlled room, which had the optimal conditions for Drosophila learning and, importantly, removes any daily environmental variations that might have affected the behavioral phenotype.…”

Section: Introductionmentioning

confidence: 99%