visualizing zebrafish brain structures using napari

생체 내 제브라피시 유충의 전뇌 영상

Zebrafish (Danio rerio) has been widely adopted as a model vertebrate animal in neuroscience, owing to its optical transparency at the larval stage, its fast development, its low maintenance cost, and the availability of diverse genetic tools1,2,3,4. In particular, the optical transparency of larvae combined with genetically encoded fluorescent reporters of biological events5,6,7,8,9 provides a unique opportunity to image both neuronal activity and structure at the whole-brain level10,11,12,13,14. However, even with a microscope that supports cellular resolution, the acquired images do not necessarily retain the information at the single-cell level; the optical image quality may be degraded due to the aberration introduced by the agarose gel used for sample mounting, the fish may be mounted at an angle, thus the regions of interest are not fully contained within the field of view of the microscope, and the fish may move during the recording, causing motion artifacts in the images or impeding accurate signal extraction from the images.
Thus, an effective and reproducible protocol is needed to acquire high-quality image data with minimal noise and motion. Unfortunately, publicly available protocols for imaging a whole brain of larval zebrafish in vivo15,16,17,18,19 only briefly describe the procedure, leaving substantial parts of the details, such as agarose solidification, precise mounting techniques, and sample positioning using forceps, up to each experimentalist. In addition, inconsistencies in agarose concentration and immobilization methods10,11,14,15,16,17,18,19 can lead to challenges arising from fish movement during the imaging process.
Here, a detailed protocol for whole-brain imaging of larval zebrafish using three-dimensional fluorescence microscopy is provided. Figure 1 provides a graphical overview of the protocol: sample preparation and immobilization, sample embedding, image acquisition, and visualization after imaging. The structural and functional imaging of the larval zebrafish brain in vivo is demonstrated using a commercial confocal microscope and a custom-designed fluorescence microscope. This protocol can be tailored by practitioners for brain imaging with certain sensory stimuli or behavior contexts depending on the experimental needs and design.

저자 스포트라이트: 3차원 형광 현미경을 사용한 제브라피시 유충의 In Vivo 전뇌 이미징  

생체 내 3차원 형광 현미경을 사용한 제브라피쉬 유충의 전뇌 이미징

Watch this Scientific Journal Video about Visualizing Zebrafish Brain Structures Using Napari at JoVE.com

Visualizing Zebrafish Brain Structures Using Napari  

Napari를 사용한 Zebrafish 뇌 구조 시각화

Napari 및 Napari Animation을 설치한 후 새 Jupyter Notebook 파일을 만듭니다. 렌더링된 제브라피쉬 브레인의 이미지를 시각화하고 동영상을 만들려면 3D 이미지를 로드하고 Napari 창을 엽니다. 그런 다음 Napari 애니메이션 플러그인을 연결합니다.다음으로, 레이어 컨트롤에서 복셀 크기, 색상 맵 및 대비 제한과 같은 매개 변수를 설정합니다. 캔버스에서 뷰어 설정을 조정합니다. 그런 다음 렌더링된 이미지를 캡처하려면 애니메이션 마법사에서 캡처 버튼을 누릅니다.렌더링된 볼륨의 동영상을 생성하려면 뷰어 설정을 수정하고 키 프레임을 추가합니다. 키 프레임을 추가한 후 애니메이션 마법사에서 키 프레임 사이의 프레임 수를 설정합니다. 마지막으로 렌더링된 애니메이션을 저장합니다.범 뉴런 GCaMP 7A를 발현하는 유충 제브라피쉬 뇌의 체적 시야는 전뇌, 중뇌 및 후뇌의 뇌 영역을 덮었습니다. 수질 oblongata, 소뇌 판, 시신경, hebenula 및 등쪽 telencephalon의 신경 세포체는 애벌레 제브라 피쉬의 전체 뇌에서 명확하게 볼 수 있습니다. 2D 기능적 이미징에서 뉴런 세포체는 배경과 중첩된 뉴런 활동 모두에서 명확하게 볼 수 있었습니다.

visualizing-zebrafish-brain-structures-using-napari

Research

JoVE Journal

Neuroscience

Visualizing Zebrafish Brain Structures Using Napari

484 조회수. Napari 및 Napari Animation을 설치한 후 새 Jupyter Notebook 파일을 만듭니다. 렌더링된 제브라피쉬 브레인의 이미지를 시각화하고 동영상을 만들려면 3D 이미지를 로드하고 Napari 창을 엽니다. 그런 다음 Napari 애니메이션 플러그인을 연결합니다.다음으로, 레이어 컨트롤에서 복셀 크기, 색상 맵 및 대비 제한과 같은 매개 변수를 설정합니다. 캔버스에서 뷰어 설정을 조정합니다. ?...

비디오: Visualizing Zebrafish Brain Structures Using Napari  

In Vivo Whole-Brain Imaging of Zebrafish Larvae Using Three-Dimensional Fluorescence Microscopy

As a vertebrate model animal, larval zebrafish are widely used in neuroscience and provide a unique opportunity to monitor whole-brain activity at the cellular resolution. Here, we provide an optimized protocol for performing whole-brain imaging of larval zebrafish using three-dimensional fluorescence microscopy, including sample preparation and immobilization, sample embedding, image acquisition, and visualization after imaging. The current protocol enables in vivo imaging of the structure and neuronal activity of a larval zebrafish brain at a cellular resolution for over 1 h using confocal microscopy and custom-designed fluorescence microscopy. The critical steps in the protocol are also discussed, including sample mounting and positioning, preventing bubble formation and dust in the agarose gel, and avoiding motion in images caused by incomplete solidification of the agarose gel and paralyzation of the fish. The protocol has been validated and confirmed in multiple settings. This protocol can be easily adapted for imaging other organs of a larval zebrafish.

Presented here is a protocol for in vivo whole-brain imaging of larval zebrafish using three-dimensional fluorescence microscopy. The experimental procedure includes sample preparation, image acquisition, and visualization.

As a vertebrate model animal, larval zebrafish are widely used in neuroscience and provide a unique opportunity to monitor whole-brain activity at the ...