Background. The main function of extender solutions for trout semen is to preserve viability while keeping the sperm motility inactivated. The longer the sperm remain viable and inactivated the better, so that later they can be intentionally activated when the sperm must travel to reach the egg micropyle for fertilization. In rainbow trout (Oncorhynchus mykiss), considerable advances have been made concerning the production of sperm extenders, quality control protocols, and the handling of gametes. However, the protocols for using an extender can differ considerably, especially regarding the extender composition. Additionally, the precise impact of pH on sperm motility, and its effect in the design and selection of a sperm extender remain unclear. Goals. The aim of this study was to determine the effect of pH on rainbow trout sperm motility and activation during storage at 5 °C and using 5 different extenders. Methods. Three simple composition sperm extenders (306, 512, and Mounib) and two complex composition sperm extenders (Erdahl & Graham, and Hank's) were tested at different pHs (7, 7.2, 7.4, 7.6, 7.8, 8.0, and 8.2), as well as 3 activation solutions (DIA 532, saline solution, and pond water). Results. The extenders showed a high buffer capacity. The highest percentages of motility were obtained with DIA 532. Extenders 306, 512, and Hanks exhibited a positive relationship between pH and % motility. Erdahl & Graham and Mounib extender did not induce motility in rainbow trout sperm at different pHs. Conclusions. A positive relationship was found between the pH and motility of trout sperm, as well as a negative effect regarding the activation solution DIA 532 with the 306, 512, and Hanks extenders.