Neural circuits in zebrafish

Principal Investigator: Florian Engert
Program in Neuroscience @Harvard

Title: “Neural circuits in zebrafish: form, function and plasticity”
BRAIN CategoryUnderstanding Neural Circuits (RFA NS-14-009)

Dr. Engert’s team will combine a wide array of cutting-edge neuroscience techniques to watch the entire brain activity of a see-through fish while it swims, and to make detailed maps of its brain circuitry.

NIH Webpages

In order to examine response properties of specific neuronal subpopulation in freely swimming larvae we have developed an imaging technique based on the bioluminescence of Aquorin-GFP that does not require excitation light and can therefore operate on a zero background signal 17. This optical signal can be collected with large angle optics and detected as a one-dimensional temporal signal by a photon multiplier tube. Figure 3 shows data from a first series of experiments in which Aquorin is expressed in the hypocretin system, a small nucleus consisting of less than 20 neurons that are known to regulate the sleep-wake cycle.The left side shows an in-vivo two-photon image of the transgenic fish and a diagram of the set-up. On the right side bioluminescence photon counts from the same fish are shown in green below traces describing the simultaneously monitored behavior of the fish in black. These data illustrate the feasibility of recording neuronal activity in a small set of identified neurons in a freely behaving animal with high temporal resolution and over many hours.

In order to examine response properties of specific neuronal subpopulation in freely swimming larvae we have developed an imaging technique based on the bioluminescence of Aquorin-GFP that does not require excitation light and can therefore operate on a zero background signal 17. This optical signal can be collected with large angle optics and detected as a one-dimensional temporal signal by a photon multiplier tube. Figure 3 shows data from a first series of experiments in which Aquorin is expressed in the hypocretin system, a small nucleus consisting of less than 20 neurons that are known to regulate the sleep-wake cycle.The left side shows an in-vivo two-photon image of the transgenic fish and a diagram of the set-up. On the right side bioluminescence photon counts from the same fish are shown in green below traces describing the simultaneously monitored behavior of the fish in black. These data illustrate the feasibility of recording neuronal activity in a small set of identified neurons in a freely behaving animal with high temporal resolution and over many hours.

Project Description

We propose to combine whole brain 2-photon imaging of neural activity in behaving larval zebrafish with detailed anatomical and connectivity information extracted from the same animals. The final goal is to generate quantitative models of brain wide neural circuits that explain the dynamic processing of sensory information as well as the generation of motor output by these circuits. Anatomical data will be generated by two complementary technologies: 1) whole brain EM data sets will be prepared from the same fish that were used for calcium imaging. Respective data sets will be registered to each other, functionally relevant neuronal ensembles will then be identified in the EM stacks and connectivity will be analyzed in these sub-networks via sparse reconstruction. 2) EM based connectivity information will be supplemented by trans-synaptic viral tracing technology. These two technologies for identifying synaptic connections have complementary strengths and weaknesses and are thus ideally suited for combination with in-vivo 2-photon calcium imaging studies. The specific power of this approach is that all three techniques, whole brain calcium imaging, viral tracing and EM reconstruction, can be done in the same animal. Functional, anatomical and behavioral data can then be analyzed in the context of the specific stimuli and quantified behavioral output and subsequently synthesized into a theoretical framework. To that end we will start with quantitative models of simple reflex behaviors, like the optomotor and optokinetic reflex, where the transformation of sensory input to motor output is relatively straightforward and well defined. These elementary models will serve as a scaffold that can be refined and complemented by additional data from structure function studies from fish performing in more sophisticated behavioral assays that involve more complex stimuli, different modalities and plastic changes. As such the process of building such a “virtual fish” will be an iterative, open ended process that requires continuous and bidirectional exchange of information between the theoretical and experimental groups of the research team.

Public Health Relevance Statement

Physical neural circuits and their patterned activation give rise to all thought and behavior, and abnormal circuits and activity patterns underlie neurodevelopmental and psychiatric disorders. To make focused progress, we will undertake a comprehensive integration of experiments and computational models of the structure and function of whole-brain circuits in transparent fish experiencing virtual reality.

NIH Spending Category

Basic Behavioral and Social Science; Behavioral and Social Science; Bioengineering; Mental Health; Neurosciences

Project Terms

Animals; Area; base; Behavior; Behavioral; Behavioral Assay; Behavioral Paradigm; Brain; Calcium; calcium indicator; cell type; Cells; Code; Complement; Complex; Computer Simulation; computerized tools; Data; Data Set; design; Development; Electron Microscopy; Engineering; experience; Fishes; Functional Imaging; Future; Generations; Goals; Image; in vivo; information processing; insight; Label; Larva; learned behavior; light microscopy; Machine Learning; Maps; Mathematics; Measures; Mental disorders; Modality; Modeling; Mono-S; Motion; Motor; Motor output; Nature; neural circuit; Neurodevelopmental Disorder; Neurons; Neurosciences; Noise; Output; Pattern; Plastics; Process; public health relevance; reconstruction; Reflex action; relating to nervous system; Research; research study; Resolution; response; scaffold; Sensory; Sensory Process; statistics; Stimulus; Structure; Subgroup; Synapses; Techniques; Technology; theories; tool; Transgenic Organisms; two-photon; Viral; virtual; virtual reality; Virus; Virus Receptors; visual stimulus; Zebrafish

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