Possible interactions between octopamine-immunoreactive (IR) and serotonergic neurons in the CNS of the medicinal leech were investigated. Simultaneous intracellular recordings of serotonin-containing neurons (either the Retzius neuron or cell 21) and the dorsolateral octopamine-IR (DLO) neuron demonstrated that both sets of neurons are coactive at times. Depolarization of either serotonergic cell 21 or the Retzius neuron did not alter the membrane potential of the DLO. Similarly depolarization of the DLO did not affect the serotonergic neurons examined. Because it was found that the DLO and either the serotonergic cell 21 or Retzius neuron were at times coactive, we looked for possible sources of common excitatory inputs. The centrally located pressure (P)- and touch (T)-sensitive mechanosensory neurons excited the DLOs through a polysynaptic pathway. Stimulation of nociceptive (N) mechanosensory neurons did not cause a measurable depolarization in the membrane potential of the DLO. Through simultaneous recordings of the DLO, cell 21, and a particular identified mechanosensory neuron, it was demonstrated that activity in the T or P cells can excite both serotonergic cell 21 and the octopamine-IR DLO. These findings indicate that, in many instances, both serotonin and octopamine, biogenic amines with neuromodulatory actions in many different invertebrates, may be released simultaneously in the leech.