Delay eyeblink conditioning performance and brain-wide c-Fos expression in male and female mice

Delay eyeblink conditioning has been extensively used to study associative learning and the cerebellar circuits underlying this task have been largely identified. However, there is a little knowledge on how factors such as strain, sex and innate behaviour influence performance during this type of learning. In this study, we used male and female mice of C57BL/6J (B6) and B6CBAF1 strains to investigate the effect of sex, strain and locomotion in delay eyeblink conditioning. We performed a short and a long delay eyeblink conditioning paradigm and used a c-Fos immunostaining approach to explore the involvement of different brain areas in this task. We found that both B6 and B6CBAF1 females reach higher learning scores compared to males in the initial stages of learning. This sex-dependent difference was no longer present as the learning progressed. Moreover, we found a strong positive correlation between learning scores and voluntary locomotion irrespective of the training duration. c-Fos immunostainings after the short paradigm showed positive correlations between c-Fos expression and learning scores in the cerebellar cortex and brainstem, as well as previously unreported areas. By contrast, after the long paradigm, c-Fos expression was only significantly elevated in the brainstem. Taken together, we show that differences in voluntary locomotion and activity across brain areas correlate with performance in delay eyeblink conditioning across strains and sexes.