Nitric oxide serves as an orthograde synaptic cotransmitter between identified neurons in the cerebral ganglion of Aplysia. Nitric oxide synthase, the enzyme that produces nitric oxide, is localized in a few specific neurons in the ganglia, including neuron C2. Guanylyl cyclase the target enzyme of nitric oxide, is found in neurons C4 and MCC, which are synaptic followers of C2. Stimulation of C2 causes a vsEPSP in these neurons that is reduced to 50% of its amplitude by nitric oxide synthase inhibitors and guanylyl cyclase inhibitors. The remaining portion of the vsEPSP is mediated by histamine. Thus, nitric oxide and histamine act as orthograde cotransmitters in producing the vsEPSP. Both cotransmitters cause closure of a background potassium channel, which depolarizes the neuron and enhances its response to synaptic inputs. Exogenous nitric oxide (released by nitric oxide donor molecules) and histamine mimic the vsEPSP's depolarization and decreased membrane conductance. When neurons C4 or MCC are isolated in cell culture they respond just as they do in the ganglion, i.e., the nitric oxide response but not the histamine response is blocked by guanylyl cyclase inhibitors, and the membrane conductance is decreased by both histamine and nitric oxide. Aplysia hemolymph partially suppresses the response to nitric oxide, due to nitric oxide scavenging by hemocyanin, which contains copper and is the equivalent of hemoglobin. Neuron C2 followers that are hyperpolarized by histamine are insensitive to nitric oxide. Thus, only select follower neurons respond to both transmitters.