Chronic heart failure (CHF) is characterized by increased sympathetic tone. The glutamatergic input in the rostral ventrolateral medulla (RVLM), which is a key region involved in sympathetic outflow, seems not to be involved in the generation of sympathetic tone in the normal state. The aim of this study was to determine the role of the RVLM glutamate receptors in the generation of sympathetic tone in CHF. CHF was produced by coronary artery ligation. Bilateral microinjection of the glutamate receptor antagonist kynurenic acid, the N-methyl-d-aspartate (NMDA) receptor antagonist D-2-amino-5-phosphonopentanoate, or the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione into the RVLM dose-dependently reduced resting blood pressure and renal sympathetic nerve activity in CHF but not in sham rats. Picoinjection of kynurenic acid (100 pmol in 5 nL) significantly decreased the basal discharge by 47% in 25 RVLM presympathetic neurons in CHF rats. In contrast, kynurenic acid had no effect on the discharge in all 22 of the RVLM presympathetic neurons tested in sham rats. These data suggest that upregulated glutamate receptors, including NMDA and non-NMDA, in the RVLM are involved in tonic control of elevated sympathetic tone in CHF.