We recorded changes in right inferior cardiac and either left inferior cardiac or left vertebral sympathetic nerve discharge (SND) produced by unilateral microinjections of γ-aminobutyric acid (GABA-A) and excitatory amino acid (EAA) receptor antagonists into the ventrolateral medulla (VLM) of urethane-anesthetized, baroreceptor-denervated cats. Unilateral microinjections of GABA-A receptor antagonists, SR-95531 or bicuculline, into single tracks in VLM anywhere between 1 and 5 mm rostral to the obex eliminated or markedly reduced 10-Hz power in SND on both sides of the body. Low frequency components (6 Hz) of SND were unaffected. Complete blockade of the 10-Hz rhythm occurred with a dose of SR-95531 as low as 6.25 pmol in a 50-nl volume. Unilateral microinjections of the non-selective EAA receptor antagonist, kynurenate (KYN, 7.5 nmol), into the caudal or rostral VLM significantly reduced, but did not eliminate, 10-Hz SND ipsilateral to the injection sites, while 10-Hz SND contralateral to the injection sites was not significantly changed. These observations suggest that: 1) GABAergic transmission in VLM is critical for generation of the 10-Hz rhythm, 2) the caudal and rostral portions of VLM act together to generate the 10-Hz rhythm, and 3) 10-Hz rhythm generation depends, at least in part, on tonic or phasic excitatory drive to GABAergic interneurons in caudal VLM and presympathetic neurons in rostral VLM. The data also suggest that pathways interconnecting the two halves of the brain stem play an important role in promoting 10-Hz rhythm generation.