Licking Microstructure Behavior Classifies a Spectrum of Emotional States in Mice

Randa Salalha^1,2Federica Cruciani²Micky Holzman²Gil Ben David²Yam Amir²Firas Mawase^3,4Kobi Rosenblum^1,2*

• ¹University of Haifa, Haifa, Israel
• ²Sagol Department of Neuroscience, center for gene manipulation in the brain, Haifa, Israel
• ³Faculty of Biomedical Engineering, Technion–Israel Institute of Technology, Haifa, Israel. Faculty of Biomedical Engineering, Technion–Israel Institute of Technology, Haifa, Israel
• ⁴Faculty of Biomedical Engineering, Technion–Israel Institute of Technology, Haifa, Israel

Measuring precise emotional tagging for taste information, with or without the use of words, is challenging. While affective taste valence and salience are core components of emotional experiences, traditional behavioral assays for taste preference, which often rely on cumulative consumption, lack the resolution to distinguish between different affective states, such as innate versus learned aversion, which are known to be mediated by distinct neural circuits. To overcome this limitation, we developed an open-source system for high-resolution microstructural analysis of licking behavior in freely moving mice. Our approach integrates traditional lick burst analysis with a proprietary software pipeline that utilizes interlick interval (ILI) distributions and principal component analysis (PCA) to create a multidimensional behavioral profile of the animal. Using this system, we characterized the licking patterns associated with innate appetitive, aversive, and neutral tastants. While conventional burst analysis failed to differentiate between two palatable stimuli (water and saccharin), our multidimensional approach revealed distinct and quantifiable behavioral signatures for each.Critically, this approach successfully dissociates innate and learned aversive taste valences, a distinction that cannot be achieved using standard metrics. By providing the designs for our custom-built setup and analysis software under an open-source license, this study offers a comprehensive and accessible methodology for examining hedonic responses in future studies. This powerful toolkit enhances our understanding of sensory valence processing and provides a robust platform for future investigations of the neurobiology of ingestive behavior.