Optical Imaging-Based Guidance of Viral Microinjections and Insertion of a Laminar Electrophysiology Probe Into a Predetermined Barrel in Mouse Area S1BF.

Victor M Mocanu,Amir Shmuel

doi:10.3389/fncir.2021.541676

Abstract

Wide-field Optical Imaging of Intrinsic Signals (OI-IS; Grinvald et al., 1986) is a method for imaging functional brain hemodynamic responses, mainly used to image activity from the surface of the cerebral cortex. It localizes small functional modules – such as cortical columns – with great spatial resolution and spatial specificity relative to the site of increases in neuronal activity. OI-IS is capable of imaging responses either through an intact or thinned skull or following a craniotomy. Therefore, it is minimally invasive, which makes it ideal for survival experiments. Here we describe OI-IS-based methods for guiding microinjections of optogenetics viral vectors in proximity to small functional modules (S1 barrels) of the cerebral cortex and for guiding the insertion of electrodes for electrophysiological recording into such modules. We validate our proposed methods by tissue processing of the cerebral barrel field area, revealing the track of the electrode in a predetermined barrel. In addition, we demonstrate the use of optical imaging to visualize the spatial extent of the optogenetics photostimulation, making it possible to estimate one of the two variables that conjointly determine which region of the brain is stimulated. Lastly, we demonstrate the use of OI-IS at high-magnification for imaging the upper recording contacts of a laminar probe, making it possible to estimate the insertion depth of all contacts relative to the surface of the cortex. These methods support the precise positioning of microinjections and recording electrodes, thus overcoming the variability in the spatial position of fine-scale functional modules.

Highlights

Optogenetics activates light-sensitive ion channels – or pumps – termed opsins at a physiologically relevant, millisecond-scale on/off kinetics (Zhang et al, 2010; Fenno et al, 2011; Yizhar et al, 2011)
For guiding microinjections which are followed by the recovery of the animal, we propose to perform minimally invasive OI-IS through the thinned skull (Figures 2A, 3A, 4) and gently break the surface of the skull at the selected injection points
In addition to guiding the insertion of the electrode to the pre-determined barrel, the user can use the OI system to estimate the spatial extent of the optogenetics photostimulation, by comparing the image obtained under the fiber-optic illumination to the pial vessels around the optic fiber (Figure 6)

Summary

Introduction

Optogenetics activates light-sensitive ion channels – or pumps – termed opsins at a physiologically relevant, millisecond-scale on/off kinetics (Zhang et al, 2010; Fenno et al, 2011; Yizhar et al, 2011). Selecting the cortical sites for microinjections and for inserting the recording electrodes is commonly done by using stereotaxic coordinates referenced from structural brain atlases (Kirkcaldie et al, 2012; Watson et al, 2012; Knutsen et al, 2016; Yang et al, 2017; Paxinos and Franklin, 2019) This approach has been commonly used and optimized for applying optogenetics in rodents (Cetin et al, 2007). Atlas-based positioning of microinjections and electrodes provides only an approximation of the true locations of functional modules, as there can be significant inter-individual (between-subject) variation (Jellema et al, 2004; Oberlaender et al, 2012; Knutsen et al, 2016; Paxinos and Watson, 2017) This is especially problematic for small functional modules such as cortical columns with diameters as small as 200 – 300 microns. Localizing an insertion with high precision with respect to cortical columns cannot be based solely on stereotactic coordinates

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