Populations experiencing sudden environmental change must be capable of rapidly evolving to survive. Here we explore changes in gene transcription as a mechanism for rapid adaptation at four osmoregulatory genes (CFTR I, NaK ATPase1αa, NaK ATPase1αb and GHRII) in anadromous steelhead trout versus a derived land-locked population after 14 generations. Transcription was measured before and after a 24-h saltwater challenge in pure and reciprocal hybrid offspring of fish from both populations reared in a common environment for two generations. Significant differences between the landlocked and migratory populations were observed, particularly in fresh water at the NaK ATPase1αa and GHRII genes, indicating rapid evolutionary change, possibly associated with reduced energy expenditure in the landlocked lake system. Phenotypic divergence analysis (Q (ST)) shows that the observed transcriptional differences deviate from neutral expectations. Some reciprocal crosses exhibited anomalous transcription consistent with sex-linked epistatic or genetic imprinting effects. Our results highlight unpredictable phenotypic outcomes of hybridization among locally adapted populations and the need to exercise caution when interbreeding populations for conservation purposes.