Non-invasive large-scale imaging of concurrent neuronal, astrocytic, and hemodynamic activity with hybrid multiplexed fluorescence and magnetic resonance imaging (HyFMRI)

A critical gap currently exists in systematic understanding and experimental validation of the role of astrocytes in neurovascular coupling and their functional links with other brain cells. Despite a broad selection of functional neuroimaging tools for multi-scale brain interrogations, no methodology currently exists that can discern responses from neural and glial cells while simultaneously mapping the associated hemodynamic activity on a large scale. We present a hybrid multiplexed fluorescence and magnetic resonance imaging (HyFMRI) platform for measuring neuronal and astrocytic activity registered to concurrently recorded brain-wide hemodynamic responses. It features a fiberscope-based imaging system for multichannel fluorescence and optical intrinsic signal recordings and a custom surface radiofrequency coil, which are incorporated into the bore of a preclinical magnetic resonance imaging (MRI) scanner. We used HyFMRI to study peripheral-stimulus-evoked brain responses in mice differentially labeled with RCaMP and GCaMP genetically-encoded calcium indicators. Stimulation-evoked neuronal responses displayed the fastest kinetics and highest activation amplitude followed by astrocytic signals and the hemodynamic responses simultaneously recorded with functional MRI. In addition, the activation traces from neurons and astrocytes exhibited high linear correlation, thus providing direct evidence of astrocytic mediation in neurovascular coupling. This newly developed capacity to capture cell-type-specific calcium signaling alongside whole-brain hemodynamics enables the simultaneous investigation of neuro-glial-vascular interactions in health and disease. HyFMRI thus expands the current neuroimaging toolbox for a wide range of studies into synaptic plasticity, neural circuitry, brain function and disorders.