In this report we describe the actions of two novel compounds, (RS)-alpha-cyclopropyl-4-phosphonophenylglycine (CPPG) and (S)-alpha-ethylglutamate (EGLU), which are potent antagonists at two types of presynaptic metabotropic glutamate (mGlu) receptors in the neonatal rat spinal cord. Selective activation of these receptors by L-2-amino-4-phosphonobutyrate (L-AP4) or (1S,3S)-1-aminocyclopentane-1,3-dicarboxylic acid ((1S,3S)-ACPD) results in the depression of the monosynaptic component of the dorsal root-evoked ventral root potential (DR-VRP). CPPG produces rightward parallel shifts of the dose-response curves for both L-AP4- and (1S,3S)-ACPD, with Schild slope in each case close to unity, consistent with a competitive mechanism of antagonism. CPPG is the most potent antagonist yet described for both L-AP4- and (1S,3S)-ACPD-sensitive presynaptic mGlu receptors but displays a 30-fold selectivity for the L-AP4-sensitive receptor over the (1S,3S)-ACPD-sensitive receptor (KD values 1.7 microM and 53 microM, respectively). EGLU, on the other hand, is selective for the (1S,3S)-ACPD-sensitive receptor, displaying little or no activity at the L-AP4-sensitive site. EGLU produces a rightward parallel shift of the dose-response curve to (1S,3S)-ACPD, with Schild slope close to unity, again indicative of a competitive mode of antagonism (KD 66 microM). Both CPPG and EGLU displayed only weak or no antagonist activity at postsynaptic metabotropic and ionotropic glutamate receptors.