Recently developed in vitro preparations of the brainstem-spinal cord from neonatal rat suitable for investigation of motor control systems for mammalian locomotion and respiration are described. The preparations remain viable for extended periods under standard in vitro conditions and generate rhythmic motor patterns for locomotion and respiration. The methodology of the preparations and characteristics of the motor output patterns are described. The preparations retain functional circuitry for major components of the motor control systems, including brainstem respiratory and spinal locomotor pattern generating networks, brainstem locomotor command regions, descending bulbospinal and ascending spinal pathways, and mechanosensory afferent input systems. They therefore offer potential for investigation of diverse aspects of the mammalian respiratory and locomotor control systems.