Fluorescent In Situ Hybridization Chain Reaction for RNA in the Embryonic and Larval Central Nervous System.

In the nerve cord, much is known about the generation of neurons from neuronal stem cells. Over the lifetime of a neuron, the cumulative expression of genes within that neuron determines its fate. Furthermore, gene expression in mature neurons determines their functional characteristics.

Studying Larval Behavior in Agarose Channels.

Larvae of the fruit fly are a popular and tractable model system for studying the development and function of sensorimotor circuits, thanks to the relative numerical simplicity of their nervous system and the wealth of available genetic tools to manipulate the anatomy, activity, and function of specific cell types. Researchers studying the role of a particular gene or cell type in sensorimotor circuit activity or function may wish to observe the effects of an experimental manipulation during behavior in the intact animal. Observing these effects, which may include changes in the intracellular calcium concentration or movement of small numbers of neurons, muscles, etc.

The Study of Sensorimotor Circuit Assembly in Embryos and Larvae.

In animals, movement is generated by the activity of motor circuits housed in the vertebrate spinal cord or the arthropod nerve cord. How motor circuits form is a fundamental question, with wide-ranging impacts on the fields of development, neurobiology, medicine, evolution, and beyond. Until recently, studying circuit assembly had been experimentally difficult, with a paucity of suitable models.

Single-Neuron Labeling in Using Multicolor FLP-Out.

Neurons exhibit some of the most striking examples of morphological diversity of any cell type. Thus, when studying neurons, the morphology of each neuron must be considered individually. However, neurons densely populate the central nervous system (CNS), making it difficult to ascertain fine morphological features due to a lack of spatial resolution.

Imaging Neural Activity in Intact, Semirestrained Larvae.

The larval nerve cord, which is the equivalent of the vertebrate spinal cord, houses the circuits required to process somatosensory stimuli (e.g., tactile, temperature, vibration, and self-movement) and generate the patterned muscle contractions underlying movement and behavior.

CRASH2p: Closed-loop Two Photon Imaging in a Freely Moving Animal.

Direct measurement of neural activity in freely moving animals is essential for understanding how the brain controls and represents behaviors. Genetically encoded calcium indicators report neural activity as changes in fluorescence intensity, but brain motion confounds quantitative measurement of fluorescence. Translation, rotation, and deformation of the brain and the movements of intervening scattering or auto-fluorescent tissue all alter the amount of fluorescent light captured by a microscope.

Distinctive features of the central synaptic organization of larval proprioceptors.

Proprioceptive feedback is critically needed for locomotor control, but how this information is incorporated into central proprioceptive processing circuits remains poorly understood. Circuit organization emerges from the spatial distribution of synaptic connections between neurons. This distribution is difficult to discern in model systems where only a few cells can be probed simultaneously.

The Role of Even-Skipped in Larval Somatosensory Circuit Assembly.

Proper somatosensory circuit assembly is critical for processing somatosensory stimuli and for responding accordingly. In comparison to other sensory circuits (e.g.

Development of motor circuits: From neuronal stem cells and neuronal diversity to motor circuit assembly.

In this review, we discuss motor circuit assembly starting from neuronal stem cells. Until recently, studies of neuronal stem cells focused on how a relatively small pool of stem cells could give rise to a large diversity of different neuronal identities. Historically, neuronal identity has been assayed in embryos by gene expression, gross anatomical features, neurotransmitter expression, and physiological properties.

RNA-binding protein syncrip regulates starvation-induced hyperactivity in adult Drosophila.

How to respond to starvation determines fitness. One prominent behavioral response is increased locomotor activities upon starvation, also known as Starvation-Induced Hyperactivity (SIH). SIH is paradoxical as it promotes food seeking but also increases energy expenditure.

Temporal transcription factors determine circuit membership by permanently altering motor neuron-to-muscle synaptic partnerships.

How circuit wiring is specified is a key question in developmental neurobiology. Previously, using the motor system as a model, we found the classic temporal transcription factor Hunchback acts in NB7-1 neuronal stem cells to control the number of NB7-1 neuronal progeny form functional synapses on dorsal muscles (Meng et al., 2019).

How prolonged expression of Hunchback, a temporal transcription factor, re-wires locomotor circuits.

How circuits assemble starting from stem cells is a fundamental question in developmental neurobiology. We test the hypothesis that, in neuronal stem cells, temporal transcription factors predictably control neuronal terminal features and circuit assembly. Using the Drosophila motor system, we manipulate expression of the classic temporal transcription factor Hunchback (Hb) specifically in the NB7-1 stem cell, which produces U motor neurons (MNs), and then we monitor dendrite morphology and neuromuscular synaptic partnerships.

Direction Selectivity in Drosophila Proprioceptors Requires the Mechanosensory Channel Tmc.

Drosophila Transmembrane channel-like (Tmc) is a protein that functions in larval proprioception. The closely related TMC1 protein is required for mammalian hearing and is a pore-forming subunit of the hair cell mechanotransduction channel. In hair cells, TMC1 is gated by small deflections of microvilli that produce tension on extracellular tip-links that connect adjacent villi.

Temporal Cohorts of Lineage-Related Neurons Perform Analogous Functions in Distinct Sensorimotor Circuits.

Neuronal stem cell lineages are the fundamental developmental units of the brain, and neuronal circuits are the fundamental functional units of the brain. Determining lineage-circuitry relationships is essential for deciphering the developmental logic of circuit assembly. While the spatial distribution of lineage-related neurons has been investigated in a few brain regions [1-9], an important, but unaddressed question is whether temporal information that diversifies neuronal progeny within a single lineage also impacts circuit assembly.

The Hunchback temporal transcription factor establishes, but is not required to maintain, early-born neuronal identity.

Background: Drosophila and mammalian neural progenitors typically generate a diverse family of neurons in a stereotyped order. Neuronal diversity can be generated by the sequential expression of temporal transcription factors. In Drosophila, neural progenitors (neuroblasts) sequentially express the temporal transcription factors Hunchback (Hb), Kruppel, Pdm, and Castor.

Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior.

Drosophila larval crawling is emerging as a powerful model to study neural control of sensorimotor behavior. However, larval crawling behavior on flat open surfaces is complex, including: pausing, turning, and meandering. This complexity in the repertoire of movement hinders detailed analysis of the events occurring during a single crawl stride cycle.

Functional Genetic Screen to Identify Interneurons Governing Behaviorally Distinct Aspects of Drosophila Larval Motor Programs.

Drosophila larval crawling is an attractive system to study rhythmic motor output at the level of animal behavior. Larval crawling consists of waves of muscle contractions generating forward or reverse locomotion. In addition, larvae undergo additional behaviors, including head casts, turning, and feeding.

Even-Skipped(+) Interneurons Are Core Components of a Sensorimotor Circuit that Maintains Left-Right Symmetric Muscle Contraction Amplitude.

Bilaterally symmetric motor patterns--those in which left-right pairs of muscles contract synchronously and with equal amplitude (such as breathing, smiling, whisking, and locomotion)--are widespread throughout the animal kingdom. Yet, surprisingly little is known about the underlying neural circuits. We performed a thermogenetic screen to identify neurons required for bilaterally symmetric locomotion in Drosophila larvae and identified the evolutionarily conserved Even-skipped(+) interneurons (Eve/Evx).

Atlas-builder software and the eNeuro atlas: resources for developmental biology and neuroscience.

A major limitation in understanding embryonic development is the lack of cell type-specific markers. Existing gene expression and marker atlases provide valuable tools, but they typically have one or more limitations: a lack of single-cell resolution; an inability to register multiple expression patterns to determine their precise relationship; an inability to be upgraded by users; an inability to compare novel patterns with the database patterns; and a lack of three-dimensional images. Here, we develop new 'atlas-builder' software that overcomes each of these limitations.

A resource for manipulating gene expression and analyzing cis-regulatory modules in the Drosophila CNS.

Here, we describe the embryonic central nervous system expression of 5,000 GAL4 lines made using molecularly defined cis-regulatory DNA inserted into a single attP genomic location. We document and annotate the patterns in early embryos when neurogenesis is at its peak, and in older embryos where there is maximal neuronal diversity and the first neural circuits are established. We note expression in other tissues, such as the lateral body wall (muscle, sensory neurons, and trachea) and viscera.

Characterization of Drosophila larval crawling at the level of organism, segment, and somatic body wall musculature.

Understanding rhythmic behavior at the developmental and genetic levels has important implications for neurobiology, medicine, evolution, and robotics. We studied rhythmic behavior--larval crawling--in the genetically and developmentally tractable organism, Drosophila melanogaster. We used narrow-diameter channels to constrain behavior to simple, rhythmic crawling.

An image-free opto-mechanical system for creating virtual environments and imaging neuronal activity in freely moving Caenorhabditis elegans.

Non-invasive recording in untethered animals is arguably the ultimate step in the analysis of neuronal function, but such recordings remain elusive. To address this problem, we devised a system that tracks neuron-sized fluorescent targets in real time. The system can be used to create virtual environments by optogenetic activation of sensory neurons, or to image activity in identified neurons at high magnification.