In the abdominal ganglion of Aplysia californica, seven motoneurons have been described which modulate the myogenic heart beat and vasomotor tone (28). These neurons mediate their motor effects by chemical transmission. In this paper we have attempted to specify the transmitters of six of these motoneurons. We have 1) studied the effects of several common transmitters on the innervated structures and compared these effects with the effects of firing the motoneurons, 2) examined whether blocking agents influence similarly the effect of a putative transmitter applied to the innervated structure and the effect of firing a motoneuron, and 3) tested the capability of the motoneurons to synthesize the putative transmitters from precursors. The positive inotropic and chronotropic effects of firing the excitor motoneuron RB(HE) were mimicked by perfusion of the heart with serotonin at a low concentration. Cinanserin blocked both the effects of motoneuron excitation and serotonin perfusion. RB(HE) was also shown to synthesize [3H]serotonin from L-[3H]tryptophan injected directly into the cell body. The effects of firing the two LD(HI) heart-inhibitory motoneurons were mimicked by perfusion of the heart with acetylcholine. Benzoquinonium blocked the effects of the inhibitory motoneuron and acetylcholine perfusion. Perfusion with arecoline also inhibited the heart beat. Acetylcholine applied to the arteries mimicked the vasoconstriction caused by the LB(VC) motoneurons. Aortic constriction in response to activity in LB(VC) cells or to acetylcholine was blocked by hexamethonium and curare. The heart inhibitor and vasoconstrictor motoneurons synthesized [3H] acetylcholine from [3H] choline injected into their cell bodies. Thus, as in vertebrates, acetylcholine mediates inhibition to the heart. Unlike vertebrates, however, serotonin mediates excitation to the heart and acetylcholine mediates peripheral vasoconstriction.