SignificanceContraction of skeletal muscle is triggered by regulatory structural changes in the thin filaments following calcium binding to troponin. Structural changes in the thick filaments control the availability of myosin motors for actin interaction and the strength and speed of contraction. Here, we elucidate the coupling between thin and thick filament regulatory mechanisms in demembranated fibers from mammalian skeletal muscle in near-physiological conditions, both in the steady state and on the millisecond timescale following a calcium jump. We show that physiological activation of skeletal muscle depends on two positive feedback loops, involving mechano-sensing by the thick filament and myosin-sensing by the thin filament. The rapid activation of skeletal muscle following electrical stimulation depends on the coordinated activation of both filaments.