The relationship between the anatomy and neurochemistry of neurons in the ventral medulla oblongata in regions that is responsible for cardiovascular, airways, and respiratory regulation was investigated. Microinjections of excitant amino acid (glutamate, 100 nl, 100 mM) were made throughout the ventral medulla in anesthetized rats. Arterial blood pressure, sympathetic nerve activity and phrenic nerve discharge was recorded. Injection sites were marked with albumin-colloidal gold or methylene blue. At the end of recording, brains were removed, fixed, sectioned and stained for gold spots, dopamine-beta-hydroxylase (DBH), phenylethanolamine-N-methyltransferase (PNMT) and serotonin. In the ventrolateral medulla (VLM), several interesting features emerged; first, the entire region corresponding to the pressor and depressor regions of the rostral and caudal VLM (RVLM and CVLM) lay in the area containing PNMT-immunoreactive neurons (C1 cell group). Caudal to the CVLM pressor sites were identified as far caudally as the spino-medullary junction. Activation of the B?tzinger region consistently inhibited phrenic amplitude and frequency whereas activation of the pre B?tzinger increased the basal level of phrenic nerve discharge as well as its frequency and amplitude. The PreBotzinger is found at the same level as the most rostral part of the CVLM. Medial sites in CVLM tended to be excitatory while lateral sites evoked inhibition. In the midline, few sites were found where excitation elicited large (±10%) changes in blood pressure or sympathetic nerve activity. However at one site (11.3 mm caudal to bregma) injection of glutamate elicited phrenic apnea in both paralyzed and non-paralyzed animals. The ventral medulla contains many areas from which responses that affect the control of airways, breathing and circulation exist; these responsive sites are coextensive with populations of neurons that are immunoreactive for many neurochemicals including amines and peptides. Functional studies are required to determine the individual roles played by these different neurotransmitters.