Name: Two-Photon Calcium Imaging of Mouse Superior Colliculus Neurons
Uploaded: 2023-04-21T15:00:00
Description: Watch this Scientific Journal Video about Two-Photon Calcium Imaging of Mouse Superior Colliculus Neurons at JoVE.com

two-photon calcium imaging of mouse superior colliculus neurons

Superior Colliculus의 Two-photon 및 Wide-field Calcium 이미징

The superior colliculus (SC) is an important visual center in all vertebrates. In mammals, it receives direct inputs from the retina and the visual cortex1. While optical recording has been widely applied to the cortex2,3,4,5, its application in the SC is hindered by poor optical accesss6,7,8,9,10,11,12,13,14,15,16,17,18,19. The goal of this protocol is to provide details about two complementary methods for optical recording of the neural activity in the SC.
The SC is located beneath the cortex and transverse sinus, which limits optical access to the collicular neurons. One approach to overcome this limitation is to aspirate the overlaying cortex and expose the anterior-lateral SC7,9,10,13,14,19. However, because the SC receives cortical inputs, such an operation might affect how the SC neurons respond to visual stimuli. To overcome this limitation, we detail here an alternative protocol to image the superficial layer of the posterior-medial SC with a silicon plug, while leaving the cortex intact8,11. Specifically, to achieve single-cell resolution, we applied two-photon microscopy to image calcium responses in the posterior-medial SC of wild-type mice. In addition, to achieve broad coverage, we applied wide-field microscopy to image the entire SC of a mutant mouse whose posterior cortex has not developed20.
The two methods described in this protocol are complementary to each other. The two-photon calcium imaging without ablating the cortex is appropriate for recording neural activity at&#160;single-cell resolution with intact cortical inputs. The wide-field calcium imaging is appropriate for recording neural activity in the entire SC while sacrificing spatial resolution.

저자 스포트라이트: Unveiling Neural Coding and Mechanisms of Visual Processing in the Superior Colliculus 

Mouse Superior Colliculus의 칼슘 이미징

My research focuses on neural coding and mechanisms for visual processing. We are trying to answer how various information is encoded in the brain, especially in the superior colliculus under the underlying neuro-mechanisms. To understand visual processing, scientists combine various approaches, including neuronal morphology, antegrade and retrograde tracing, IL sequencing, neuro modeling, machine learning, is understood well.The challenge is to remove the skull over the SSA, cutting and left in the bone in one larger piece, is likely to fail because the bone is attached to the dura. By combining two photo on a wide field calcium image, our recent work revealed the function architecture of motion direction in the mouse SC at both single cell resolution and a global scale. We found that neurons with similar preference form patches up to 500 micrometer under global to the upward on the nasal motion.With our protocol, we are addressing two research gaps. First, researchers can perform long-term cast imaging in mouse SC at a single cell resolution without breaking the cortex. Secondly, researchers can record the neuroactivity across the entire SC using widefield microscoping.Our protocol provides a measure to image the posterior medial SC at single cell resolution with a intact cortex in mice. Also, we use a biocompatible plug to expose the SC, which reduces infection for chronic imaging. Our results provide a way to study the neural coding of visual information across a large visual field.Combined with optogenetics, one can study half inputs from different brain regions modulating the neuroactivity in SC.

Watch this Scientific Journal Video about Two-Photon Calcium Imaging of Mouse Superior Colliculus Neurons at JoVE.com

Two-Photon Calcium Imaging of Mouse Superior Colliculus Neurons  

Mouse Superior Colliculus Neuron의 Two-Photon Calcium 이미징

헤드 플레이트와 플러그를 이식한 후 헤드 플레이트가 있는 회전하는 트레드밀에 동물을 고정합니다. 트레드밀을 이광자 현미경 아래에 놓고 높이를 적절한 위치로 조정합니다. 대물렌즈와 헤드 플레이트 사이에 검은색 알루미늄 콘을 놓아 시각적 자극을 위해 모니터의 빛이 오염되는 것을 방지합니다.2광자 이미징을 수행하려면 샘플 평면에서 레이저 출력을 20에서 80밀리와트 사이로 조정합니다. 픽셀당 2.4마이크로미터의 공간 해상도와 최대 350마이크로미터 깊이의 이미지 신경 활동으로 4.8Hz에서 600 x 600마이크로미터의 시야를 스캔할 수 있습니다. 로터리 인코더를 사용하여 트레드밀에서 마우스의 움직임을 기록합니다.50mm 렌즈가 장착된 카메라를 사용하여 동공 크기와 위치를 기록합니다. 마지막으로, 이러한 기록과 이미지 획득을 자극 컴퓨터에서 보낸 트리거를 기록하여 시각적 자극에 동기화합니다. 야생형 마우스의 SC 뉴런의 칼슘 반응과 이미지 전반에 걸친 형광의 표준 편차 투영이 표시됩니다.

Research

JoVE Journal

Neuroscience

Two-Photon Calcium Imaging of Mouse Superior Colliculus Neurons

Calcium Imaging in Mouse Superior Colliculus

The superior colliculus (SC), an evolutionarily conserved midbrain structure in all vertebrates, is the most sophisticated visual center before the ...

연구

신경과학

Preparation of a Suction Cup and Plugs for Calcium Imaging in Mouse

마우스의 칼슘 이미징을 위한 흡입 컵 및 플러그 준비

Viral Construct (AAV) Injection in Mouse Superior Colliculus for Calcium Imaging

칼슘 이미징을 위한 Mouse Superior Colliculus의 Viral Construct (AAV) 주입