The aim of the present study was to examine the effects of L-glutamate and glycine microinjected into the rostral ventrolateral medulla (RVLM) in conscious unrestrained rats. Microinjection of 2 nmol of L-glutamate increased the mean arterial pressure (MAP) and renal sympathetic nerve activity (RSNA) in the conscious rats. The RSNA responses were significantly larger in the conscious rats than in anesthetized rats, while the magnitude of the pressor responses was similar in conscious and urethane-anesthetized rats. L-Glutamate injection significantly decreased heart rate in the conscious rats, whereas it increased the heart rate slightly but not significantly in the anesthetized rats. Microinjection of 100 nmol of glycine into the RVLM of conscious rat decreased MAP and RSNA. In 2 of the 6 rats examined, the depressor and sympathoinhibitory responses were preceded by a few seconds of a pressor and sympathoexcitatory phase. The decreases of RSNA in response to glycine injection were significantly larger in the conscious rats than in the anesthetized rats, whereas the magnitude of the depressor responses was similar in the two groups of rats. Heart rate decreased in response to glycine injection into the RVLM in the conscious and the anesthetized rats. In conclusion, in conscious unrestrained rats, as well as in urethane-anesthetized rats, L-glutamate acts as a sympathoexcitatory agent and glycine acts as a sympathoinhibitory agent in the RVLM. The sympathetic responses to these amino acids are larger in conscious rats than in anesthetized rats.