Long-Term Two-Photon Calcium Imaging of Neuronal Populations with Subcellular Resolution in Adult Non-human Primates

Abstract

Two-photon imaging with genetically encoded calcium indicators (GECIs) enables long-term observation of neuronal activity in vivo. However, there are very few studies of GECIs in primates. Here, we report a method for long-term imaging of a GECI, GCaMP6f, expressed from adeno-associated virus vectors in cortical neurons of the adult common marmoset (Callithrix jacchus), a small New World primate. We used a tetracycline-inducible expression system to robustly amplify neuronal GCaMP6f expression and up- and downregulate it for more than 100 days. We succeeded in monitoring spontaneous activity not only from hundreds of neurons three-dimensionally distributed in layers 2 and 3 but also from single dendrites and axons in layer 1. Furthermore, we detected selective activities from somata, dendrites, and axons in the somatosensory cortex responding to specific tactile stimuli. Our results provide a way to investigate the organization and plasticity of cortical microcircuits at subcellular resolution in non-human primates.

Super-wide-field two-photon imaging with a micro-optical device moving in post-objective space

“…If a GRIN lens can be inserted between the mirror pair as a relay optical system, the mirror-to-mirror distance d m–m could be extended further, regardless of the WD of the objective lens. These extensions may be useful when observing the broad (>1 cm) neocortex of non-human primates 56–58 . The post-objective space is a new frontier in microscope design.…”

Two-photon imaging of cerebral hemodynamics and neural activity in awake and anesthetized marmosets

“…Additionally, marmosets can be trained to rest comfortably with restraint, allowing researchers to bypass anesthesia and its confounding influences on brain physiology (Liu et al, 2013). Recently, 2PM was used to image neuronal populations in anesthetized marmosets (Sadakane et al, 2015). In the present study, we aim to establish a longitudinal two-photon imaging technique in awake marmosets, providing a critical tool to visualize cortical vasculature, neurons, and their dynamics at the cellular level.…”

Chronic multiscale imaging of neuronal activity in the awake common marmoset

“…For future experiments, it will be useful to enhance the expression level of GCaMP with the tetracycline-inducible AAV system that has been used with mice1718 and was recently reported with marmosets8.…”

“…However, to gain cellular- and circuit- level insights into the plasticity of cortical networks underlying normal behaviors and disease states, an experimental technique to chronically track neuronal ensemble activity is required. Recently, in vivo Ca 2+ imaging was applied to marmosets8, but the experiments were performed exclusively under anesthesia, which not only prevents the recording of behaviorally relevant neuronal activity, but also artificially modifies brain activity at the macroscopic9 and cellular levels1011. Here we established, for the first time, multiscale macroscopic and cellular chronic imaging of brain activity in the awake marmoset, by combining a novel body fixation device, systematic acclimation training and subject screening based on their behavior.…”