Invasive sea lampreys have had substantial negative ecological and economic impacts on the Laurentian Great Lakes region. Control efforts, such as lampricide application and barriers, have resulted in a reduction in number of sea lampreys in the Great Lakes. Due to environmental and non-target impacts of existing control mechanisms, coupled with the fact that no non-physical barrier is completely effective at stopping fish movement, there is a critical need to develop novel control technologies to assist with the control and suppression of sea lamprey populations. Recent work has indicated that carbon dioxide gas (CO2) applied to water will influence the movement and behavior of fishes, providing the potential for CO2 to act as a non-physical barrier that can exclude fish from a target area. To date, the effectiveness of CO2 at influencing the movement and behavior of sea lampreys has not been explored. The current study showed that CO2 applied to water will result in behavioral agitation for both adult and transformer sea lampreys, and will eventually result in equilibrium loss. More importantly, both adult and transformer sea lampreys will ‘choose’ to avoid water with CO2 concentrations of 85 and 160 mg/L (respectively). Together, results from this study suggest that CO2 applied to water has the potential to act as a non-physical barrier to deter the movement of free-swimming lamprey in the wild. Carbon dioxide gas can be integrated with existing control technologies to act as a novel barrier technology and augment existing control strategies for sea lampreys.