Abstract: Fish muscle, as an important source of high-quality animal protein for humans, has long been a core scientific issue and industrial demand in aquaculture in terms of improving both yield and quality. Muscle growth mainly depends on the processes of muscle fiber hyperplasia and hypertrophy, and the elucidation of key regulatory factors is of great significance for genetic improvement. Myostatin (Mstn), a member of the transforming growth factor-β superfamily, is a key negative regulator of skeletal muscle growth. This review systematically summarizes the structural characteristics and evolutionary divergence patterns of fish mstn genes, and outlines their tissue-specific expression profiles as well as regulatory roles in myoblast proliferation, differentiation, and muscle fiber type formation. On this basis, this review focuses on the molecular mechanisms by which Mstn-mediated Smad and non-Smad signaling pathways regulate muscle growth, cellular metabolism, and homeostasis maintenance, and further analyzes its multidimensional functions in glucose metabolism regulation and lipid deposition. In addition, this review summarizes recent phenotypic studies in which gene-editing technologies have been employed to generate targeted mstn knockouts in various fish species, highlighting alterations in muscle fiber number, intramuscular fat deposition, and skeletal morphology. Finally, with mstn as a key molecular target, its application prospects and implementation pathways in the breeding of new fish varieties with rapid growth and superior flesh quality are discussed, with the aim of providing theoretical support and technical reference for the precise genetic improvement of fish muscle traits.