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

Imágenes de calcio de dos fotones y de campo amplio en el colículo superior

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.

Autor destacado: Desvelando la codificación neuronal y los mecanismos de procesamiento visual en el colículo superior 

Imágenes de calcio en el colículo superior de ratón

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  

Obtención de imágenes de calcio de dos fotones de neuronas del colículo superior de ratón

Después de implantar la placa de la cabeza y el tapón, asegure al animal en una cinta de correr giratoria con su placa de la cabeza. Coloque la cinta de correr debajo del microscopio de dos fotones y ajuste la altura a una posición adecuada. Coloque un cono de aluminio negro entre el objetivo y la placa de la cabeza para evitar la contaminación lumínica del monitor para la estimulación visual.Para realizar imágenes de dos fotones, ajuste la potencia del láser en el plano de muestra entre 20 y 80 milivatios. Escanee un campo de visión de 600 x 600 micrómetros a 4,8 hercios con una resolución espacial de 2,4 micrómetros por píxel y una actividad neuronal de imagen de hasta 350 micrómetros de profundidad. Con un codificador rotatorio, registre la locomoción del ratón en la cinta de correr.Utiliza una cámara con una lente de 50 milímetros para registrar el tamaño y la posición de la pupila. Por último, sincronice estas grabaciones y las adquisiciones de imágenes con la estimulación visual mediante el registro de los disparadores enviados por la computadora de estimulación. Se presentan las respuestas de calcio de las neuronas SC de un ratón de tipo salvaje y la proyección de la desviación estándar de la fluorescencia a través de las imágenes.

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 ...

Investigación

Neurociencias

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

Preparación de una ventosa y tapones para la obtención de imágenes de calcio en ratones

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

Inyección de construcción viral (AAV) en el colículo superior de ratón para obtener imágenes de calcio

Implantation of the Headplate in Mouse for Calcium Imaging 

Implantación de la placa de la cabeza en ratón para la obtención de imágenes de calcio 

Implanting the Plug in the Mouse for Calcium Imaging

Implantar el enchufe en el ratón para obtener imágenes de calcio

Wide-Field Calcium Imaging of Mouse Superior Colliculus Neurons

Obtención de imágenes de calcio de campo amplio de neuronas del colículo superior de ratón