Effects of device color and sex on mice in von Frey test

Pain-related behavioral research in animals is profoundly influenced by a constellation of interacting factors. This study explored the effect of device color and sex on exploratory and anxiety-related behavior of C57BL/6 mice in the von Frey test. Mice in 4 groups based on device color (red vs clear) and sex (male and female), behavioral observations were made during a 15 min adaptation period following the mice’s entry into the test device, and mechanical pain thresholds were assessed using the von Frey test before and after inducing neuropathic pain through spared nerve injury surgery. Results showed mice in the red device exhibited significantly reduced exploratory and anxiety-related behavior, entered a resting state more rapidly and stabilized across the three 5 min periods compared to those in the clear one. On the 7th and 14th day after surgery, the mechanical pain threshold decreased in all groups. However, there were no significant differences between male and female. Device color could affect behavioral responses in the adaptation of von Frey test, and red device potentially improves test efficiency by reducing the necessary adaptation period. These findings emphasize the importance of considering environmental factors in pain research with von Frey test, and the red device can simulate red visible light environments that facilitate faster behavioral stabilization, which enhances the experiment efficiency and respects rodent nocturnal habits.