Superficial skeletal muscle activation is associated with an electric activity. Bidimensional High-Density Surface Electromyography (HD-sEMG) is a non-invasive technique that uses a grid of equally spaced electrodes applied on the skin surface to detect and portray superficial skeletal muscle activation. The goal of the study was to evaluate the feasibility of HD-sEMG to detect electrical activation of skeletal muscle and its application during rehabilitation exercises in horses. To fulfil this aim, activation of the superficial descending pectoral and external abdominal oblique core muscles were measured using HD-sEMG technology during dynamic mobilization exercises to induce lateral bending and flexion/extension tasks of the trunk. Masseter muscle was instrumented during mastication as a control condition. A 64 surface EMG channel wireless system was used with a single 64 electrode grid or a pair of 32 electrode grids. HD-sEMG provided unique information on the muscular activation onset, duration, and offset, along each motor task, and permitting inferences about the motor control strategy actuated by the central nervous system. Signals were further processed to obtain firing frequencies of few motor-neurons. Estimation of electromyographic amplitude and spectral parameters allowed detecting the onset of muscular fatigue during the motor tasks performed. HD-sEMG allows the assessment of muscular activation in horses performing specific motor tasks, supporting its future application in clinical and research settings.