Calcium is a secondary messenger in plant signaling, and its concentration changes spatially and temporally during the course of heat stress. In the present study, potassium antimonate was used to visualize calcium localization in blades of a marine macroalga, the juvenile Laminaria japonica sporophytes under heat stress (25 °C). Result showed that loosely bound calcium was mainly distributed on the cell wall under normal conditions (10 °C), and flowed into the cytoplasm when exposed to heat. The simutaneous assay on the antioxidant system changes was performed. Oxidative damage, as measured by generation of reactive oxygen species (ROS) malondialdehyde (MDA) content, increased significantly during heat stress, and calcium pretreatment alleviated oxidative damage. The assay on the activities of six antioxidant enzymes demonstrated that their enzymatic activities were inhibited when exposed to heat stress, but Ca2+ pretreatment effectively attenuated the inhibition. Results in the present study inferred that calcium homeostasis plays an essential role in L. japonica sporophyte when exposed to heat, and calcium pretreatment could improve its thermo-tolerance. Calcium is a secondary messenger in plant signaling, and its concentration changes spatially and temporally during the course of heat stress. In the present study, potassium antimonate was used to visualize calcium localization in blades of a marine macroalga, the juvenile Laminaria japonica sporophytes under heat stress (25 ° C). Result showed that loosely bound calcium was mainly distributed on the cell wall under normal conditions (10 ° C), and flowed into the cytoplasm when exposed to heat. The simutaneous assay on the antioxidant system changes was performed. Oxidative damage, as measured by generation of reactive oxygen species (ROS) malondialdehyde (MDA) content, increased significantly during heat stress, and calcium pretreatment alleviated oxidative damage. The assay on the activities of six antioxidant enzymes demonstrated that their enzymatic activities were inhibited when exposed to heat stress, but Ca2 + pretreatment effectively attenuated the inhibition. Results in the pre sent study inferred that calcium homeostasis plays an essential role in L. japonica sporophyte when exposed to heat, and calcium pretreatment could improve its thermo-tolerance.