<p>ARP2/3 is a seven-subunit protein complex involved in the formation of actin filament branching, which is essential for the formation of membrane protrusions, cell migration, and establishment of cell polarity. Among these functions, ARP2/3 has been implicated in myoblast fusion. Since myogenesis is a complex multistep process, here we decided to explore deeper the distribution patterns and functions of ARP2/3 during the initial steps of embryonic chicken skeletal myogenesis. The chosen biological experimental model was the cell culture of pectoralis muscle obtained from 11-d-old chick embryos, which is composed of myoblasts, multinucleated myotubes, and muscle fibroblasts. Our results show that ARP2/3 was found in myoblasts, myotubes, and muscle fibroblasts in four main distributions: the perinuclear region, in small puncta in the cytoplasm, along F-actin structures in the cytoplasm, and in circular structures in myotubes. Inhibition of ARP2/3 function by CK-666 led to a significant reduction in several parameters of skeletal myogenesis, including the area of muscle cells (desmin-positive cells), myotube thickness, the number of myoblasts, the number of nuclei within myotubes, the number of fibroblasts, the total number of nuclei (including nuclei in myoblasts, myotubes, and fibroblasts), and the myoblast fusion index. Interestingly, CK-666 reduced myotube formation and induced the formation of spindle-shaped myoblasts. Live cell video microscopy showed that inhibition of ARP2/3 induced a decrease in myoblast cell migration and the formation of blebs in the membranes of cells. The collection of our results shows that ARP2/3 is essential for the initial steps of embryonic chick skeletal myogenesis, and its inhibition leads to a major reduction in myoblast proliferation, migration, fusion, and muscle fiber formation.</p>