Isolated Murine Brain Model for Large-Scale Optoacoustic Calcium Imaging

Abstract

Real-time visualization of large-scale neural dynamics in whole mammalian brains is hindered with existing neuroimaging methods having limited capacity when it comes to imaging large tissue volumes at high speeds. Optoacoustic imaging has been shown to be capable of real-time three-dimensional imaging of multiple cerebral hemodynamic parameters in rodents. However, optoacoustic imaging of calcium activity deep within the mammalian brain is hampered by strong blood absorption in the visible light spectrum as well as a lack of activity labels excitable in the near-infrared window. We have developed and validated an isolated whole mouse brain preparation labeled with genetically encoded calcium indicator GCaMP6f, which can closely resemble in vivo conditions. An optoacoustic imaging system coupled to a superfusion system was further designed and used for rapid volumetric monitoring of stimulus-evoked calcium dynamics in the brain. These new imaging setup and isolated preparation’s protocols and characteristics are described here in detail. Our new technique captures calcium fluxes as true three-dimensional information across the entire brain with temporal resolution of 10 ms and spatial resolution of 150 μm, thus enabling large-scale neural recording at penetration depths and spatio-temporal resolution scales not covered with any existing neuroimaging techniques.

Photoacoustic Neuroimaging - Perspectives on a Maturing Imaging Technique and its Applications in Neuroscience

“…In proof-of-principle experiments, simultaneous fluorescence, and photoacoustic recordings were conducted in GCaMP-expressing zebrafish larvae and ex vivo adult zebrafish brains using chemical calcium triggers. These results have been corroborated by an ex vivo murine brain preparation showing signal changes from GCaMP6f in response to an activating chemoconvulsant (Gottschalk et al, 2019a).…”

Optoacoustic Calcium Imaging of Deep Brain Activity in an Intracardially Perfused Mouse Brain Model

“…Furthermore, it is now possible to observe and evaluate the transition between an in-vivo brain model (prior to perfusion) and a blood free in-situ preparation (after perfusion), providing a platform to further explore the relation between epifluorescence and optoacoustic signals for GCaMP6 indicators. A simpler, quicker surgical procedure in comparison to the isolated brain model [40] increased the success rate of performed experiments, thus reducing the number of animals required for the study. By utilizing the vascular system we were able to fully replace the blood with ACSF, and thereby double the maximum depth from which GCaMP6f signals could be collected.…”

Advanced high resolution three-dimensional imaging to visualize the cerebral neurovascular network in stroke

“…Neuroimaging of PA can also be based on hemoglobin gradient and oxygen saturation changes, but the resolution is limited to several hundred microns [172]. GCaMP is primarily used, while it cannot be used to image neurons deep in the brains of living mammals [173,174].…”