Abstract: To investigate effective methods for restoring the natural spawning of the Yangtze sturgeon (Acipenser dabryanus), a nature-like river channel with diverse topographies and pebble substrates was constructed in a semi-enclosed side channel of the middle Chishui River in April 2025. Through the implementation of flow regulation, captive-bred Yangtze sturgeon were successfully induced to achieve natural reproduction. Our findings indicate that the nature-like channel offers a broad selection window for spawning, with depth variations of 0.2—1.1 m and flow velocities ranging from 0.27 to 1.44 m/s. The highest spawning density was recorded at water depth of 0.65 m and a flow velocity of 0.38 m/s, and substrates of large-sized cobbles (mean long axis ≥0.15 m). Furthermore, the sturgeons exhibited a significant preference for micro-topographies on stoss slopes (average downstream slope of 6.0%,slope angle of–3.45°) and substrates composed of large-sized cobbles. This study confirms the feasibility of using nature-like channels as functional spawning grounds in field environments, providing a critical scientific basis and technical paradigm for the functional restoration of Yangtze sturgeon habitats.