Mapping Neurotransmitter Identity in the Whole-Mount Drosophila Brain Using Multiplex High-Throughput Fluorescence in Situ Hybridization

Genetics: A Periodical Record of Investigations Bearing on Heredity and Variation

Robert H. Singer^2¹  Aljoscha Nern^1²  Geoffrey W. Meissner^1³ 

[1] Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461^2;Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia 20147;Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia 20147^1

关键词: neurotransmitter;    Drosophila;    fluorescence in situ hybridization;    gene expression;    mRNA;   

DOI  :  10.1534/genetics.118.301749

学科分类：医学（综合）

来源: Genetics Society of America

【 摘 要 】

Identifying the neurotransmitters used by specific neurons is a critical step in understanding the function of neural circuits. However, methods for the consistent and efficient detection of neurotransmitter markers remain limited. Fluorescence in situ hybridization (FISH) enables direct labeling of type-specific mRNA in neurons. Recent advances in FISH allow this technique to be carried out in intact tissue samples such as whole-mount Drosophila melanogaster brains. Here, we present a FISH platform for high-throughput detection of eight common neurotransmitter phenotypes in Drosophila brains. We greatly increase FISH throughput by processing samples mounted on coverslips and optimizing fluorophore choice for each probe to facilitate multiplexing. As application examples, we demonstrate cases of neurotransmitter coexpression, reveal neurotransmitter phenotypes of specific cell types, and explore the onset of neurotransmitter expression in the developing optic lobe. Beyond neurotransmitter markers, our protocols can in principle be used for large-scale FISH detection of any mRNA in whole-mount fly brains.

【 授权许可】

CC BY