Spontaneous breathing occurs transiently in spinalized animals suggesting that the cervical spinal cord generates respiratory rhythm. We recorded optical signals from isolated brainstem-spinal cord preparations of neonatal rats using a voltage-sensitive dye, and visualized respiratory-related activity by processing the optical data with a correlation coefficient imaging technique. We identified a novel region that extended from the medullo-spinal junction to the C2 segment, approximately 100-300 mum deep from the ventral surface, and termed the high cervical respiratory group. We recorded inspiratory and preinspiratory-inspiratory neurons in this region. High cervical respiratory group neurons are distinct from motoneurons, because they are small, spindle in shape, and have only two or three long processes, and thus may be interneurons that are involved in respiratory rhythmogenesis in the cervical spinal cord.