We investigated the afferents and intra-cortical synaptic organization of the anterior cingulate cortex (ACC) during noxious electrical stimulation. Extracellular field potentials were recorded simultaneously from sixteen electrodes spanning all layers of the ACC in male Sprague-Dawley rats anesthetized by halothane inhalation. Laminar-specific transmembrane currents were calculated with the current source density analysis method. Two major groups of evoked sink currents were identified: an early group (latency = 54.04 ±2 .12 ms; 0.63±0.07 mv/mm2) in layers V-VI and a more intense late group (latency = 80.07±4.85 ms; 2.16±0.22 mv/mm2) in layer II/III and layer V. Multi-unit activities were evoked mainly in layer V and deep layer II/III with latencies similar to that of the early and late sink groups. The evoked EPSP latencies of pyramidal neurons in layers II/III and V related closely with the sink currents. The sink currents were inhibited by intracortical injection of CNQX (1 mM, 1 µl), a glutaminergic receptor antagonist, and enhanced by intraperitoneal (5 mg/kg) and intracortical (10 µg/µl, 1 µl) injection of morphine, a µ-opioid receptor agonist. Paired pulse depression was observed with inter-pulse intervals of 50 to 1000 ms. High frequency stimulation (100 Hz, 11 pulses) enhanced evoked responses in the ACC and evoked medial thalamic (MT) unit activities. MT lesions blocked evoked responses in the ACC. Our results demonstrated that two distinct synaptic circuits in the ACC were activated by noxious stimuli and that the MT is the major thalamic relay that transmits nociceptive information to the ACC.