Name: 냄새 기둥을 탐색하는 생쥐의 해마에서 칼슘 신호를 기록하는 미니스코프 (동영상) | JoVE Journal
Uploaded: 2024-09-20T13:30:00.000+00:00
Duration: 6 min 21 s
Description: 학습하세요 생쥐 해마 CA1에서 미니스코프로 칼슘 신호를 기록하여 냄새 기둥 탐색 행동을 분석하는 방법. 이 기술은 알츠하이머병 모델에서 신경 회로와 공간 인지의 관계를 이해하는 데 활용됩니다.

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This research was supported by the US National Institutes of Health (NIH UF1 NS116241 and NIH R01 DC000566), and the National Science Foundation (NSF BCS-1926676). The authors thank Andrew Scallon for helping setting up the Odor Arena chamber.

Recording Neuronal Activity During Odor Plume Navigation with Miniscope Imaging

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</ol>

The authors declare no conflict of interest.

This protocol meticulously outlines the steps to record place-cells and odor-responsive cells in the dCA1 area of the hippocampus of mice navigating an odor plume. The critical steps in the protocol include stereotaxic surgery, placement of the miniscope baseplate, construction of the odor area, checking the plume in the odor arena, behavioral training, miniscope recording of the freely moving mouse, data preprocessing, and data analysis. Additionally, the protocol explains the process of decoding the mouse trajectory fr...

Although significant progress has been made in understanding neural circuits involved in olfactory navigation in head-fixed mice1,2,3 and navigation strategies in freely moving mice4,5,6,7,8, the role of neural circuits in ethologically relevant freely moving navigation of turbulent odor plumes is still unknown. This article describes monitoring neural activity by imaging Ca2+ transients in cells expressing the genetically encoded calcium sensor GCaMP6f in Thy1-GCaMP6f mice9 to study whether sequential neural dynamics of dorsal CA1 stratum pyramidale neurons in the hippocampus (dCA1) plays a role in odorant plume navigation. The methods provide information on imaged GCaMP6f fluorescence through a miniature epifluorescence microscope focused through a GRIN lens on dCA110,11,12. The methods explain how to monitor simultaneously spatial navigation and dCA1 neuron GCaMP6f calcium transients in mice performing an odor-plume navigation task where they received a water reward when they reached the spout delivering an odorant into an odor arena with a background laminar air flow13,14. This article describes the methods required to achieve this task (Figure 1), including the stereotaxic surgery for the implantation of gradient-index (GRIN) lenses, the placement of a baseplate to secure the miniscope to the skull in a freely moving mouse, imaging with the miniature microscope and monitoring mouse movement with a high-speed digital camera, data preprocessing for removing motion artifacts and finding the regions of interest (ROIs), and preparation of datasets and artificial neural network training and prediction for decoding the X and Y positions of the mouse in the odor arena from changes in fluorescence in dCA1 ROIs7.
Miniscope recording of calcium signals in the CA1 region of the hippocampus of mice navigating an odor plume is relevant for understanding the computation of neural circuits involved with olfaction and spatial information in the complex behavior task of odor-plume navigation2,14,15,16. The CA1 region of the hippocampus plays a role in spatial navigation and is crucial for creating a cognitive map of the environment for efficient navigation17,18. Recording calcium signals with a miniscope is a valuable way to investigate the CA1 neurons that encode spatial information during odor plume navigation.
This technique combines the advantages of miniscope technology for recording GCaMP calcium signals with the well-established role of the CA1 hippocampus in spatial navigation to understand better how neural circuits drive complex behaviors19. Alternatively, approaches using 2-photon microscopy can record CA1 neurons9,20, which requires a head-fixed mouse and restrains the possibility of freely moving to navigate an odor plume21. Local-field electrophysiological recordings of CA1 neurons allow the investigation of freely-moving mice navigating odor plumes22. Still, local field electrical signals impose limitations to estimating intracellular firing by isolating single-unit signals through spike sorting techniques. Miniscope signals allow the identification of ROIs associated directly with intracellular calcium signals in a reliable way10,11&#160;to investigate neural computations at single-cell resolution precisely. Miniscope technology provides a unique opportunity to better understand how the CA1 region encodes spatial information based on odor cues.
Furthermore, this technique investigates how specific neuronal populations process odor information for navigation and the relationship between neuronal activity patterns and decision-making during odor plume tracking. This method can contribute to a better understanding of how the brain processes odor and spatial information. While miniscopes offer a single-cell resolution for recording a freely moving mouse&#39;s brain, they require specialized surgery and data analysis expertise. In this paper, we provide a comprehensive protocol for helping researchers go through each step to investigate the neural mechanisms of odor-plume navigation.
The odor navigating task is a promising framework for studying neural coding and spatial odor cue memory in mice. The article&#39;s findings indicate that it is possible to decode the trajectory of the mouse navigating an odor plume based on neuronal ensemble calcium signals in dCA1. Understanding the role of dCA1 calcium signals in odor plume navigation is a crucial step to crack the neural circuit basis for odor-guided navigation in realistic environments13,14.

<table><tbody><tr><td>Arduino Micro</td><td>Arduino</td><td>Micro</td><td></td></tr><tr><td>Biocompatible Methacrylate Resin</td><td>Parkell</td><td>S380</td><td>C&amp;amp;B-Metabond Adhesive Luting Cement</td></tr><tr><td>Decoding Brain Signals Software</td><td>CU Anschutz</td><td></td><td>https://github.com/restrepd/drgMiniscope</td></tr><tr><td>Dental Drill</td><td>Osada</td><td>LHP-6</td><td>AZ210015</td></tr><tr><td>Dental Drill Box</td><td>Osada</td><td>XL-230</td><td>30000 rotations per minute</td></tr><tr><td>Digital stereotaxic instrument</td><td>Stoelting</td><td>51730D</td><td>Mouse Stereotaxic Instument, #51904 Digital Manipulator Arm, 3-Axes, Add-On, LEFT&amp;nbsp;</td></tr><tr><td>Drill Bit</td><td>FST Fine Science Tools</td><td>19007-05</td><td>Tip diameter 0.5 mm</td></tr><tr><td>Fast Digital Camera</td><td>Edmund Optics</td><td>BFS-U3-63S4C</td><td>FLIR Blackfly S</td></tr><tr><td>Focal Lens</td><td>Edmund Optics</td><td>C-Series</td><td>3.5 mm</td></tr><tr><td>GRIN lens</td><td>Inscopix</td><td>1050-004595</td><td>1 mm diameter and 4 mm length</td></tr><tr><td>GRIN lens Holder</td><td>UCLA</td><td></td><td>http://miniscope.org/index.php/Surgery_Protocol</td></tr><tr><td>Liquid Tissue Adhesive</td><td>3M</td><td>1469C</td><td>Vetbond Tissue Adhesive</td></tr><tr><td>Low-Flow Anesthesia System for Mice</td><td>Kent Scientific Corporation</td><td>SomnoSuite</td><td>https://www.kentscientific.com/products/somnosuite/</td></tr><tr><td>Low Toxicity Silicone Adhesive</td><td>WPI &amp;ndash; World Precision Instruments</td><td>Kwik-sil</td><td></td></tr><tr><td>miniPID Controller</td><td>ASI &amp;ndash; Aurora Scientific Inc.</td><td>Model 200B</td><td>Fast-Response Miniature Photo-Ionization Detector</td></tr><tr><td>Miniscope V4 Holder</td><td>UCLA</td><td>NA</td><td>https://github.com/Aharoni-Lab/Miniscope-v4/tree/master/Miniscope-v4-Holder</td></tr><tr><td>Miniscope V4</td><td>Labmaker</td><td>NA</td><td>https://www.labmaker.org/products/miniscope-v4</td></tr><tr><td>Miniscope Base Plate V2</td><td>Labmaker</td><td>NA</td><td>https://www.labmaker.org/products/miniscope-v4-base-plates-variant-2-pack-of-10</td></tr><tr><td>Miniscope DAQ-QT software</td><td>UCLA</td><td></td><td>https://github.com/Aharoni-Lab/Miniscope-DAQ-QT-Software/wiki</td></tr><tr><td>Motion Correction Software</td><td>CU Anschutz</td><td></td><td>https://github.com/restrepd/drgMiniscope</td></tr><tr><td>Odor Arena Hardware</td><td>Custom Made</td><td>3D Model</td><td>https://www.dropbox.com/scl/fo/lwtpqysnpzis32mhrx3cd/ADomsxyhxu42sqDmTBl2O6k?rlkey=b3l4809eradundt5l3iz0gq74&amp;amp;&lt;br/&gt;dl=0</td></tr><tr><td>Odor Arena Software</td><td>CUAnschutz</td><td></td><td>https://github.com/wryanw/odorarena</td></tr><tr><td>Odorant Isoamyl Acetate</td><td>Aldrich Chemical Co</td><td>06422AX</td><td>Diluted at 1% in odorless mineral oil</td></tr><tr><td>RHD USB Interface Board</td><td>Intan Technologies</td><td>C3100</td><td>Product discontinued. Alternatively use another equivalent board.</td></tr><tr><td>ROI Extraction Software</td><td>CU Anschutz</td><td></td><td>https://github.com/restrepd/drgMiniscope</td></tr><tr><td>Sutter Micromanipulator</td><td>Sutter Instrument Company</td><td>MP-285</td><td></td></tr><tr><td>Synchronization Software</td><td>CU Anschutz</td><td></td><td>https://github.com/fsimoesdesouza/Synchronization</td></tr><tr><td>Thy1-GCaMP6f mice</td><td>Jackson Laboratory</td><td>IMSR_JAX 028281</td><td>C57BL/6J-Tg(Thy1-GCaMP6f)GP5.12Dkim/J)</td></tr></tbody></table>

miniscope recording calcium signals at hippocampus of mice navigating an odor plume

Miniscope Recording Calcium Signals at Hippocampus of Mice Navigating an Odor Plume

Mice navigate an odor plume with a complex spatiotemporal structure in the dark to find the source of odorants. This article describes a protocol to monitor behavior and record Ca2+ transients in dorsal CA1 stratum pyramidale neurons in the hippocampus (dCA1) in mice navigating an odor plume in a 50 cm x 50 cm x 25 cm odor arena. An epifluorescence miniscope focused through a gradient-index (GRIN) lens imaged Ca2+ transients in dCA1 neurons expressing the calcium sensor GCaMP6f in Thy1-GCaMP6f mice. The paper describes the behavioral protocol to train the mice to perform this odor plume navigation task in an automated odor arena. The methods include a step-by-step procedure for the surgery for GRIN lens implantation and baseplate placement for imaging GCaMP6f in CA1. The article provides information on real-time tracking of the mouse position to automate the start of the trials and delivery of a water reward. In addition, the protocol includes information on using an interface board to synchronize metadata describing the automation of the odor navigation task and frame times for the miniscope and a digital camera tracking mouse position. Moreover, the methods delineate the pipeline used to process GCaMP6f fluorescence movies by motion correction using the NorMCorre algorithm followed by identification of regions of interest with EXTRACT. Finally, the paper describes an artificial neural network approach to decode spatial paths from CA1 neural ensemble activity to predict mouse navigation of the odor plume.

Using this procedure allows for visualizing and recording dCA1 GCaMP6f fluorescence transients in mice navigating the odor arena to find the source of odorants (Figure 6A,B, Supplementary Movie 1, and Supplementary Movie 2). The fluorescence images are motion-corrected with NoRMCorre, and EXTRACT is used to extract the ROIs. In addition, recording with an interface board allows for synchronization of the δF/F0 signals from the ROIs with odor...

This protocol investigates the brain-behavior relationship in hippocampal CA1 in mice navigating an odor plume. We provide a step-by-step protocol, including surgery to access imaging of the hippocampus, behavioral training, miniscope GCaMP6f recording and processing of the brain, and behavioral data to decode the mouse position from ROI neural activity.

Mice navigate an odor plume with a complex spatiotemporal structure in the dark to find the source of odorants. This article describes a protocol to ...

My research aims to understand how the brain processes olfactory and spatial information. And we're trying to understand the role of hippocampal CA1 neurons in odor-plume navigation. Currently, the technology of freely-moving recordings of neurons with miniature microscopes has been used to advance research in the field.We found that's possible to decode the trajectory of the mouse navigating odor-plume based on neurons and calcium signals in dorsal CA1. This technique combines the advantages of mini scope technology for recording GCaMP calcium signals with the well-established row of CA1 hippocampus spatial navigation. to understand better how neural circuits drive complex behaviors, We will investigate how odor-plume navigation is impaired in a mouse model of Alzheimer's disease with abnormal CA1 hippocampus function.To begin, construct a chamber with two acrylic walls, and acrylic ceiling, and a wide-expanded polyvinyl chloride floor. The other two unique walls at the front and rear must facilitate airflow. Position four sets of odor sources paired with water delivery spouts 10 centimeters apart along the X-axis.Install a fast digital camera above the arena to monitor animal beh

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Research

JoVE Journal

Neuroscience

This video describes in vivo calcium imaging in Drosophila neurons using GCaMP. The featured protocol clip shows how to record changes in GCaMP fluorescence from mushroom body neurons during an olfactory conditioning experiment.

Drosophila In Vivo Calcium Imaging: A Method for Functional Imaging of Neuronal Activity

This video demonstrates a method for in vivo calcium imaging using synaptically localized calcium sensors in flies. It outlines the steps for preparing the flies for microscopy, conducting olfactory conditioning, and visualizing odor-induced calcium influx.

In Vivo Calcium Imaging Using Synaptically Localized Calcium Sensors in Drosophila melanogaster

Source: Lee, H. et. al., Successful In vivo Calcium Imaging with a Head-Mount Miniaturized Microscope in the Amygdala of Freely Behaving Mouse. J. Vis. Exp. (2020)This video demonstrates in vivo calcium imaging in a mouse using a gradient refractive index, or GRIN lens, and a miniaturized microscope. The setup combines head fixation within a frictionless mobile cage system, stimulating neurons and triggering calcium ion influx. The GRIN lens captures calcium signaling in the brain, enabling...

Real-Time Microendoscopic Calcium Imaging in a Freely Moving Mouse

Source: Thapa, R., et. al., Stereotaxic Viral Injection and Gradient-Index Lens Implantation for Deep Brain In Vivo Calcium Imaging. J. Vis. Exp. (2021)The video demonstrates real-time calcium imaging using a miniscope in a transduced mouse, correlating neuronal activity with behavior.

Real-Time Calcium Imaging of a Mouse Medial Prefrontal Cortex Using a Miniscope

Here we describe a protocol for measuring and analyzing temperature responses in the olfactory bulb of Xenopus laevis. Olfactory receptor neurons and mitral cells are differentially stained, after which calcium changes are recorded, reflecting a sensitivity of some neural networks in the bulb to temperature drops induced at the nose.

Recording Temperature-induced Neuronal Activity through Monitoring Calcium Changes in the Olfactory Bulb of Xenopus laevis

This protocol describes the recording of local field potentials with multi-shank linear silicon probes. Conversion of the signals using current source density analysis allows the reconstruction of local electrical activity in the mouse hippocampus. With this technique, spatially restricted brain oscillations can be studied in freely moving mice.

Recording Spatially Restricted Oscillations in the Hippocampus of Behaving Mice

Here we present a protocol with which pre- and/or postsynaptic calcium can be visualized in the context of Drosophila learning and memory. In vivo calcium imaging using synaptically localized calcium sensors is combined with a classical olfactory conditioning paradigm such that the synaptic plasticity underlying this type of associative learning may be determined.

In Vivo Optical Calcium Imaging of Learning-Induced Synaptic Plasticity in Drosophila melanogaster

This article demonstrates the preparation of a custom-made imaging window supplemented with infusion cannula and its implantation onto the CA1 region of the hippocampus in mice.

Craniotomy Procedure for Visualizing Neuronal Activities in Hippocampus of Behaving Mice

The dentate gyrus of the hippocampus carries out essential and distinct functions in learning and memory. This protocol describes a set of robust and efficient procedures for in vivo calcium imaging of granule cells in the dentate gyrus in freely moving mice.

In Vivo Calcium Imaging of Granule Cells in the Dentate Gyrus of Hippocampus in Mice

This protocol details in vivo calcium imaging of hypothalamic neurons using a miniaturized microscope (miniscope) in freely behaving mice during social state manipulation (social isolation and social reunion). This method enables the imaging at single-neuron resolution of the hypothalamic activity that represents social need and social satiation, enabling mechanistic studies of social homeostasis.

In Vivo Calcium Imaging with a Miniaturized Microscope in the Hypothalamus for Understanding Social Behaviors in Mice

학습하세요 생쥐 해마 CA1에서 미니스코프로 칼슘 신호를 기록하여 냄새 기둥 탐색 행동을 분석하는 방법. 이 기술은 알츠하이머병 모델에서 신경 회로와 공간 인지의 관계를 이해하는 데 활용됩니다.

Miniscope Recording Calcium Signals at Hippocampus of Mice Navigating an Odor Plume

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