1. The properties of the excitatory synapse made by callosal inputs onto layer V and layer VI cells in the anterior cingulate cortex were studied in an in vitro slice preparation with intracellular recording. 2. In the presence of picrotoxin, the excitatory postsynaptic potential (EPSP) had two components, a fast component blocked by the non-N-methyl-D-aspartate (NMDA) receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and a slow component blocked by the NMDA receptor antagonist DL-2-amino-5-phosphonovalerate (APV). 3. Delivery of a brief tetanus to the afferent fibres led to a long-term potentiation (LTP) of the initial slope of the monosynaptic EPSP. The LTP displayed the property of co-operativity and could be blocked by APV or by buffering intracellular calcium. 4. Pairing of low frequency presynaptic activity or weak tetanic stimulation with postsynaptic depolarization failed to potentiate the EPSP. This suggests that postsynaptic depolarization alone is unable to explain the co-operativity. 5. It is concluded that the transmitter mediating the excitatory input between callosal afferents and layer V and layer VI pyramidal neurones is glutamate. Tetanic stimulation of these afferents leads to LTP which shares many but not all the properties of LTP seen in the CA1 region of the hippocampus.