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The innervation of the adult abdominal heart of Drosophila melanogaster was studied by neuronal staining with green fluorescent protein and immunocytochemical techniques. The investigation was undertaken to determine whether the adult heart receives neuronal input or whether its complex activity must be considered independent from the nervous system. The larval heart lacks innervation, suggesting that the cardiac impulse is totally myogenic. At metamorphosis, segmental neural processes grow onto the myocardium. A pair of transverse nerves innervates bilaterally each cardiac chamber and its alary muscles. These nerve terminals are immunoreactive to glutamate and form unique synaptic structures on the ventral layer of longitudinal cardiac muscles of the conical chamber. This characteristic cardiac synapse may represent part of the neural mechanism controlling the retrograde heartbeat, and, thus, the cardiac reversal that is characteristic of adults. In addition, crustacean cardioactive peptide–immunoreactive fibers originating from peripheral, bipolar neurons (BpNs) fasciculate with the transverse nerve projections and terminate segmentally throughout the abdominal heart. An additional cluster composed of four large, CCAP-positive neurons innervates the terminal chamber. The cardioacceleratory effect of CCAP release at this location may modulate the properties of a pacemaker producing the anterograde heartbeat. J. Comp. Neurol. 465:560–578, 2003. © 2003 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]