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

Beeldvorming van twee fotonen en breedveldcalcium in de superieure colliculus

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.

Auteur in de schijnwerpers: onthulling van neurale codering en mechanismen van visuele verwerking in de superieure colliculus 

Calciumbeeldvorming bij superieure colliculus bij muizen

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  

Twee-foton calciumbeeldvorming van superieure colliculusneuronen van muizen

Nadat u de kopplaat en plug hebt geïmplanteerd, zet u het dier vast op een roterende loopband met zijn kopplaat. Plaats de loopband onder de twee-fotonenmicroscoop en stel de hoogte in op de juiste positie. Plaats een zwarte aluminium kegel tussen het objectief en de kopplaat om lichtvervuiling van de monitor voor visuele stimulatie te voorkomen.Om beeldvorming met twee fotonen uit te voeren, stelt u het laservermogen op het monstervlak in tussen 20 en 80 milliwatt. Scan een gezichtsveld van 600 bij 600 micrometer met een snelheid van 4,8 hertz met een ruimtelijke resolutie van 2,4 micrometer per pixel en breng neurale activiteit tot 350 micrometer diep in beeld. Gebruik een roterende encoder om de voortbeweging van de muis op de loopband vast te leggen.Gebruik een camera met een lens van 50 millimeter om de pupilgrootte en -positie vast te leggen. Synchroniseer ten slotte deze opnames en beeldacquisities met visuele stimulatie door triggers op te nemen die door de stimulatiecomputer worden verzonden. De calciumresponsen van SC-neuronen van een wildtype muis en de standaarddeviatieprojectie van de fluorescentie over beelden worden gepresenteerd.

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 ...