Miniscope Recording Calcium Signals at Hippocampus of Mice Navigating an Odor Plume

Although significant progress has been made in understanding neural circuits involved in olfactory navigation in head-fixed mice^1,2,3 and navigation strategies in freely moving mice^4,5,6,7,8, the role of neural circuits in ethologically relevant freely moving navigation of turbulent odor plumes is still unknown. This article describes monitoring neural activity by imaging Ca²⁺ transients in cells expressing the genetically encoded calcium sensor GCaMP6f in Thy1-GCaMP6f mice⁹ to study whether sequential neural dynamics of dorsal CA1 stratum pyramidale neurons in the hippocampus (dCA1) plays a role in odorant plume navigation. The methods provide information on imaged GCaMP6f fluorescence through a miniature epifluorescence microscope focused through a GRIN lens on dCA1^10,11,12. The methods explain how to monitor simultaneously spatial navigation and dCA1 neuron GCaMP6f calcium transients in mice performing an odor-plume navigation task where they received a water reward when they reached the spout delivering an odorant into an odor arena with a background laminar air flow^13,14. This article describes the methods required to achieve this task (Figure 1), including the stereotaxic surgery for the implantation of gradient-index (GRIN) lenses, the placement of a baseplate to secure the miniscope to the skull in a freely moving mouse, imaging with the miniature microscope and monitoring mouse movement with a high-speed digital camera, data preprocessing for removing motion artifacts and finding the regions of interest (ROIs), and preparation of datasets and artificial neural network training and prediction for decoding the X and Y positions of the mouse in the odor arena from changes in fluorescence in dCA1 ROIs⁷.

Miniscope recording of calcium signals in the CA1 region of the hippocampus of mice navigating an odor plume is relevant for understanding the computation of neural circuits involved with olfaction and spatial information in the complex behavior task of odor-plume navigation^2,14,15,16. The CA1 region of the hippocampus plays a role in spatial navigation and is crucial for creating a cognitive map of the environment for efficient navigation^17,18. Recording calcium signals with a miniscope is a valuable way to investigate the CA1 neurons that encode spatial information during odor plume navigation.

This technique combines the advantages of miniscope technology for recording GCaMP calcium signals with the well-established role of the CA1 hippocampus in spatial navigation to understand better how neural circuits drive complex behaviors¹⁹. Alternatively, approaches using 2-photon microscopy can record CA1 neurons^9,20, which requires a head-fixed mouse and restrains the possibility of freely moving to navigate an odor plume²¹. Local-field electrophysiological recordings of CA1 neurons allow the investigation of freely-moving mice navigating odor plumes²². Still, local field electrical signals impose limitations to estimating intracellular firing by isolating single-unit signals through spike sorting techniques. Miniscope signals allow the identification of ROIs associated directly with intracellular calcium signals in a reliable way^10,11 to investigate neural computations at single-cell resolution precisely. Miniscope technology provides a unique opportunity to better understand how the CA1 region encodes spatial information based on odor cues.

Furthermore, this technique investigates how specific neuronal populations process odor information for navigation and the relationship between neuronal activity patterns and decision-making during odor plume tracking. This method can contribute to a better understanding of how the brain processes odor and spatial information. While miniscopes offer a single-cell resolution for recording a freely moving mouse's brain, they require specialized surgery and data analysis expertise. In this paper, we provide a comprehensive protocol for helping researchers go through each step to investigate the neural mechanisms of odor-plume navigation.

The odor navigating task is a promising framework for studying neural coding and spatial odor cue memory in mice. The article's findings indicate that it is possible to decode the trajectory of the mouse navigating an odor plume based on neuronal ensemble calcium signals in dCA1. Understanding the role of dCA1 calcium signals in odor plume navigation is a crucial step to crack the neural circuit basis for odor-guided navigation in realistic environments^13,14.