Ca2+ is a key element in the sperm activation process of Salmo salar. However, the molecular mechanisms by which this ion enters the sperm cell have been poorly studied. In this study, we examined, for the first time, the role of the voltage-gated T-type Ca2+ channel in the activation of sperm motility of Salmo salar. Using an in vitro inhibition assay, a significant decrease in total and progressive motility (P < 0.0001) was observed in Salmo salar sperm when they were treated with NNC-55-0396, a highly selective blocker. The in silico analysis showed that this blocker is docked with a strong affinity for the pore of the voltage-gated T-type calcium channel suggesting the blocking of Ca2+ ions. The results show that the T-type voltage-gated Ca2+ channel is key to sperm motility in Salmo salar.