A Dual-Color Fiber Photometry Method for Recording Astrocyte-Neuron Activity Across Multiple Brain Regions During Learning and Memory Behaviors.

Learning and memory depend critically on the coordination between neurons and astrocytes across distributed brain regions. Astrocytes modulate synaptic transmission and neuronal excitability, thereby contributing to the encoding and consolidation of information within memory-related circuits. The intra- and inter-regional correlations between astrocytes and neurons are crucial for understanding memory and cognitive disorders. However, simultaneous monitoring of astrocytic and neuronal activity across multiple regions during memory behavior remains technically challenging. Here, a method is presented that facilitates the concurrent recording of these calcium signals from astrocytes and neurons in several memory-related regions. This strategy combines cell-type-specific dual-color genetically encoded calcium indicators (GECIs), a multi-channel fiber photometry system, and synchronized behavioral tracking to achieve real-time monitoring of neuronal and astrocytic activity in the medial prefrontal cortex (mPFC), hippocampal CA1 region, and medial entorhinal cortex (MEC). Representative results demonstrate coordinated calcium dynamics across regions and cell types during the object recognition task. This method provides a reproducible and efficient experimental framework for investigating astrocyte-neuron interactions underlying learning and memory processes.