The enteric nervous system (ENS) is situated in the intestinal wall and autonomously controls most aspects of digestion, including intestinal motility. The current model of neuronal control of intestinal motility describes reflex circuits responding to locally detected luminal contents, but provides no explanation for long-distance communication and organ-wide responses to stimuli. However, like the central nervous system, the ENS contains a diverse array of neuronal subtypes, and the contributions of many of these subtypes to intestinal function are unknown. The excitatory neurotransmitter glutamate is expressed in a small number of enteric neuron subtypes, the characteristics, circuitry and function of which have been little explored. Dysregulation of glutamatergic signalling is thought to be involved in several pathologies of the digestive tract, including irritable bowel syndrome and ischemia/reperfusion injury. We use optogenetic activation, immunohistochemistry and single neuron tracing of glutamatergic neurons to demonstrate their role in colonic motility. Additionally, we identify a novel neuron subpopulation that expresses glutamate and is present only in the colon. This is the first illustration of glutamatergic neuron function in the gut, and could represent a mechanistic basis for long-distance communication in the ENS.