# Unbiased Segmentation and Multifeature Classification of Cortical Neuronal Activity Reveals Complex Dynamics Under Anesthesia

## TL;DR

This study uses advanced imaging to understand how anesthesia affects brain activity in mice, revealing complex patterns and recovery dynamics.

## Contribution

The study introduces an unbiased method to segment and classify neuronal activity in cortical layer 2/3 under anesthesia.

## Key findings

- Anesthesia causes nonlinear changes in neuronal activity in cortical layer 2/3.
- A rebound effect in neuronal activity is observed during recovery from anesthesia.
- Superficial cortical layer activity shows consistent characteristics under anesthesia.

## Abstract

Cortical layer 2/3 plays a pivotal role in regulating perception and consciousness. However, the effects of anesthetic agents on the dynamic activity patterns in this layer remain poorly understood. This study examined how neuronal activity in cortical layer 2/3 dynamically changes under anesthesia. Using high-resolution wide-field microscopy, we performed whole-brain synchronous imaging of layer 2/3 neuronal activity in mice. Using these recordings, we performed an unbiased segmentation of the awake, anesthesia, and recovery stages and classified neurons into three categories according to their activity features. Our findings revealed the characteristics of cortical dynamics under anesthesia, including a rebound effect during recovery and nonlinear changes in neuronal activity. We also confirmed the consistent and uniform characteristics of superficial cortical layer activity under anesthesia. These results increase the understanding of cortical dynamics and provide a theoretical basis for improving clinical monitoring techniques and protocols.

The online version contains supplementary material available at 10.1007/s12264-025-01527-9.