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

Abstract

Background Marmosets are a powerful, emerging model for human behavior and neurological disorders. However, longitudinal imaging modalities that visualize both cellular structure and function within the cortex are not available in this animal model. Hence, we implemented an approach to quantify vascular topology, hemodynamics, and neural activity in awake marmosets using two-photon microscopy (2PM). New method Marmosets were acclimated to a custom stereotaxic system. AAV1-GCaMP5G was injected into somatosensory cortex to optically indicate neural activity, and a cranial chamber was implanted. Results Longitudinal 2PM revealed vasculature and neurons 500 µm below the cortical surface. Vascular response and neural activity during sensory stimulation were preserved over 5 and 3 months, respectively, before optical quality deteriorated. Vascular remodeling including increased tortuosity and branching was quantified. However, capillary connectivity from arterioles to venules remained unchanged. Further, behavioral assessment before and after surgery demonstrated no impact on cognitive and motor function. Immunohistochemistry confirmed minimal astrocyte activation with no focal damage. Over 6 months, total cortical depth visualized decreased. When under anesthesia, the most prominent isoflurane-induced vasodilation occurred in capillaries and smaller arterioles. Comparison with existing method(s) These results demonstrate the capability to repeatedly observe cortical physiology in awake marmosets over months. Conclusions This work provides a novel and insightful technique to investigate critical mechanisms in neurological disorders in awake marmosets without introducing confounds from anesthesia.

Anatomical and functional neuroimaging in awake, behaving marmosets

“…We used an AAV1-GCaMP5G construct injected into S1, and an intravascular fluorescent dye to visualize individual neurons and blood vessels located up to 500 mm below the cortical surface. We observed task-induced vascular responses and neural activity for over 5 months before optical quality of the cranial window deteriorated (Santisakultarm et al, 2016). The use of optical methods for direct visualization of neural activity in the brain of awake behaving marmosets is a major future direction that significantly boosts the value of this species to neuroscience research.…”

The marmoset as a model system for studying voluntary motor control

“…One of the most promising aspects of the marmoset as a model system for neuroscience derives from its potential for genetic manipulation, since the initial demonstration of transgenic marmosets expressing enhanced green fluorescent protein (Sasaki et al, 2009;Cyranoski, 2014;Sasaki, 2015). More recently there have been demonstrations of using viral strategies to introduce genes for expressing calcium indicators which have allowed for in vivo calcium imaging in anesthetized marmosets (Sadakane et al, 2015;Santisakultarm et al, 2016). Successful optogenetic stimulation has also recently been reported (MacDougall et al, 2016).…”

Two-photon probes for in vivo multicolor microscopy of the structure and signals of brain cells

“…New kinds of detectors like GaAsP photomultipliers or hybrid avalanche photodiodes allow detection of dimmer signals. Also, promising advances in the miniaturization of two-photon microscopes are enabling simultaneous recordings of neural circuit activity in freely moving animals (Yu et al 2015; Zong et al 2017) while avoiding the side effects associated with the use of anesthetics (Tran and Gordon 2015; Santisakultarm et al 2016). …”