Abstract: Since the industrial revolution, massive amount of anthropogenic carbon dioxide (CO₂) have been generated to elevate the atmospheric CO₂ concentration. Some anthropogenic CO₂ have been absorbed by the ocean to cause “ocean acidification” (OA). Although the negative impacts of OA on sperm motility are increasingly found in various marine invertebrate species, the cellular and molecular mechanisms for these effects are still poorly understood. This study investigated the effect of OA (pH7.8 and 7.4) on sperm motility and energy supplying pathway in blood clam Tegillarca granosa. The results showed that the sperm swimming speed reduced significantly in acidified seawater. Since the adenosine triphosphate (ATP) level of sperm is closely related to its motility, we analyzed the sperm ATP content and activities of key enzymes during ATP synthesis under different OA scenarios. OA treatments significantly reduced ATP content as well as activities of 6-phosphofructokinase and pyruvate kinase in the sperm of T. granosa. The sperm Ca²⁺-ATPase of various animals has been reported to regulate sperm motility. Therefore, we explored the Ca²⁺-ATPase activity of T. granosa sperm under OA treatment. The results found that Ca²⁺-ATPase activities in the sperm of T. granosa were significantly declined under OA scenarios. In conclusion, these results suggested that OA could constrain sperm motility through inhibiting ATP synthesis and disturbing intracellular Ca²⁺ regulation.