The actions of pentobarbital sodium (Pent) on receptors for glutamate (Glu) and related compounds were studied in thin sections of the guinea pig hippocampus. Depolarizations induced by Glu and quisqualate (Quis) in CA3 neurons were reduced in amplitude during iontophoretic administration of Pent. This action of Pent was not accompanied by any noticeable changes in membrane potential or neuron input resistance. Depolarizations induced by N-methyl-D-aspartate were less sensitive to Pent. The fast kainate (KA) response was as susceptible as the Glu response, whereas the slow KA response was unaffected by Pent in three quarters of the neurons examined. Pent suppressed the Glu response at lower concentrations than required to potentiate responses to gamma-amino butyric acid. Excitatory postsynaptic potentials (EPSPs) elicited by stimulation of mossy fibers were suppressed by Pent. The EPSPs were a little more resistant to Pent than were the Glu responses. These results indicate that Pent blocks receptors for excitatory amino acids in the hippocampus. Of the three different populations of the receptors, Quis receptors are the most sensitive to Pent and KA receptors are the least sensitive. The suppression of the EPSPs is in accordance with the notion that Glu is the transmitter released from mossy fibers.