Effect of bantam knockdown on mRNA levels in the the γ5β′2a/β′2mp/β′2mp_bilateral Mushroom Body Output Neurons (MBONs) in Drosophila Melanogaster
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ABSTRACT:
PROVIDER: PRJNA860542 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR20325739_1.fastq.gz | Fastqsanger.gz | |||
| SRR20325739_2.fastq.gz | Fastqsanger.gz | |||
| SRR20325740.fastq.gz | Fastqsanger.gz | |||
| SRR20325740_1.fastq.gz | Fastqsanger.gz | |||
| SRR20325740_2.fastq.gz | Fastqsanger.gz |
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Project description:In order to knockdown bantam in the γ5β′2a/β′2mp/β′2mp_bilateral Mushroom Body Output Neurons (MBONs), we expressed the bantam microRNA-sponge (or the control scrambled-sponge) under the MB011B-split-Gal4 driver. Additionally, we expressed a membrane localized GFP transgene. At ZT12-13.5, MB011B>ban-spg, GFP and MB011B>scramble-spg, GFP female fly brains were diseccted, homogenized and FACS sorted using GFP flourescence to isolate the MBONs of interest. mRNA was extracted and used to construct Illumina RNAseq libraries
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Project description:Sleep circuitry evolved to have both dedicated and context-dependent modulatory elements. Identifying modulatory subcircuits and understanding their molecular machinery is a major challenge for the sleep field. Previously, we identified 25 sleep-regulating microRNAs in Drosophila melanogaster, including the developmentally important microRNA bantam. Here we show that bantam acts in the adult to promote early nighttime sleep through a population of glutamatergic neurons that is intimately involved in applying contextual information to behaviors, the γ5β'2a/β'2mp/β'2mp_bilateral Mushroom Body Output Neurons (MBONs). Calcium imaging revealed that bantam inhibits the activity of these cells during the early night, but not the day. Blocking synaptic transmission in these MBONs rescued the effect of bantam knockdown. This suggests bantam promotes early night sleep via inhibition of the γ5β'2a/β'2mp/β'2mp_bilateral MBONs. RNAseq identifies Kelch and CCHamide-2 receptor as possible mediators, establishing a new role for bantam as an active regulator of sleep and neural activity in the adult fly.
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Project description:The mushroom body (MB) is a well-characterized associative memory structure within the Drosophila brain. Analyzing MB connectivity using multiple approaches is critical for understanding the functional implications of this structure. Using the genetic anterograde transsynaptic tracing tool, trans-Tango, we identified divergent projections across the brain and convergent downstream targets of the MB output neurons (MBONs). Our analysis revealed at least three separate targets that receive convergent input from MBONs: other MBONs, the fan-shaped body (FSB), and the lateral accessory lobe (LAL). We describe, both anatomically and functionally, a multilayer circuit in which inhibitory and excitatory MBONs converge on the same genetic subset of FSB and LAL neurons. This circuit architecture enables the brain to update and integrate information with previous experience before executing appropriate behavioral responses. Our use of trans-Tango provides a genetically accessible anatomical framework for investigating the functional relevance of components within these complex and interconnected circuits.
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Project description:Memories are stored in the fan-out fan-in neural architectures of the mammalian cerebellum and hippocampus and the insect mushroom bodies. However, whereas key plasticity occurs at glutamatergic synapses in mammals, the neurochemistry of the memory-storing mushroom body Kenyon cell output synapses is unknown. Here we demonstrate a role for acetylcholine (ACh) in Drosophila. Kenyon cells express the ACh-processing proteins ChAT and VAChT, and reducing their expression impairs learned olfactory-driven behavior. Local ACh application, or direct Kenyon cell activation, evokes activity in mushroom body output neurons (MBONs). MBON activation depends on VAChT expression in Kenyon cells and is blocked by ACh receptor antagonism. Furthermore, reducing nicotinic ACh receptor subunit expression in MBONs compromises odor-evoked activation and redirects odor-driven behavior. Lastly, peptidergic corelease enhances ACh-evoked responses in MBONs, suggesting an interaction between the fast- and slow-acting transmitters. Therefore, olfactory memories in Drosophila are likely stored as plasticity of cholinergic synapses.
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Project description:The Drosophila melanogaster mushroom bodies (MBs) are brain structures critical for olfactory memory. The approximately 2000 intrinsic MB neurons are divisible into alpha/beta neurons, alpha'/beta' neurons and gamma neurons by morphology and roles in memory processing. It has also been shown that the different subtypes of MB neurons have different function in the process of memory formation. This difference in function, suggested to us that different types of MB neurons might have different expression profiles. In this study, we used cell-type specific gene expression profiling to gain insight into cellular properties of MB neurons.
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Project description:Active memory forgetting is a well-regulated biological process thought to be adaptive and to allow proper cognitive functions. Recent efforts have elucidated several molecular players involved in the regulation of olfactory forgetting in Drosophila, including the small G protein Rac1, the dopamine receptor DAMB, and the scaffold protein Scribble. Similarly, we recently reported that dopaminergic neurons mediate both learning- and forgetting-induced plasticity in the mushroom body output neuron MBON-γ2α'1. Two open questions remain: how does forgetting affect plasticity in other, highly plastic, mushroom body compartments and how do genes that regulate forgetting affect this cellular plasticity? Here, we show that forgetting reverses short-term synaptic depression induced by aversive conditioning in the highly plastic mushroom body output neuron MBON-γ1pedc>α/β. In addition, our results indicate that genetic tampering with normal forgetting by inhibition of small G protein Rac1 impairs restoration of depressed odor responses to learned odor by intrinsic forgetting through time passing and forgetting induced acutely by shock stimulation after conditioning or reversal learning. These data further indicate that some forms of forgetting truly erase physiological changes generated by memory encoding.
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Project description:Memory-relevant neuronal plasticity is believed to require local translation of new proteins at synapses. Understanding this process requires the visualization of the relevant mRNAs within these neuronal compartments. Here, we used single-molecule fluorescence in situ hybridization to localize mRNAs at subcellular resolution in the adult Drosophila brain. mRNAs for subunits of nicotinic acetylcholine receptors and kinases could be detected within the dendrites of co-labeled mushroom body output neurons (MBONs) and their relative abundance showed cell specificity. Moreover, aversive olfactory learning produced a transient increase in the level of CaMKII mRNA within the dendritic compartments of the γ5β'2a MBONs. Localization of specific mRNAs in MBONs before and after learning represents a critical step towards deciphering the role of dendritic translation in the neuronal plasticity underlying behavioral change in Drosophila.
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Project description:Multisensory integration plays a central role in perception, as all behaviors usually require the input of different sensory signals. For instance, for a foraging honeybee the association of a food source includes the combination of olfactory and visual cues to be categorized as a flower. Moreover, homing after successful foraging using celestial cues and the panoramic scenery may be dominated by visual cues. Hence, dependent on the context, one modality might be leading and influence the processing of other modalities. To unravel the complex neural mechanisms behind this process we studied honeybee mushroom body output neurons (MBON). MBONs represent the first processing level after olfactory-visual convergence in the honeybee brain. This was physiologically confirmed in our previous study by characterizing a subpopulation of multisensory MBONs. These neurons categorize incoming sensory inputs into olfactory, visual, and olfactory-visual information. However, in addition to multisensory units a prominent population of MBONs was sensitive to visual cues only. Therefore, we asked which visual features might be represented at this high-order integration level. Using extracellular, multi-unit recordings in combination with visual and olfactory stimulation, we separated MBONs with multisensory responses from purely visually driven MBONs. Further analysis revealed, for the first time, that visually driven MBONs of both groups encode detailed aspects within this individual modality, such as light intensity and light identity. Moreover, we show that these features are separated by different MBON subpopulations, for example by extracting information about brightness and wavelength. Most interestingly, the latter MBON population was tuned to separate UV-light from other light stimuli, which were only poorly differentiated from each other. A third MBON subpopulation was neither tuned to brightness nor to wavelength and encoded the general presence of light. Taken together, our results support the view that the mushroom body, a high-order sensory integration, learning and memory center in the insect brain, categorizes sensory information by separating different behaviorally relevant aspects of the multisensory scenery and that these categories are channeled into distinct MBON subpopulations.
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