AbstractThe aim of this study was to characterize further the transmitter content and the location of the parent cells of tyrosine hydroxylase‐immunoreactive boutons terminating on luteinizing hormone‐releasing hormone‐ and glutamic acid decarboxylase‐immunoreactive neurons in the rat medial preoptic area. Electron microscopic immunostaining for luteinizing hormone‐releasing hormone, tyrosine hydroxylase or glutamic acid decarboxylase was performed on desipramine‐pretreated (to protect norepinephrine and epinephrine axons) rats which received a stereotaxic injection of 6‐hydroxydopamine into the medial preoptic area anteroventral periventricular nucleus 48 h prior to sacrifice. This treatment induced acute degeneration of dopamine axon terminals characterized by the development of autophagous cytolysosomes, an early morphological sign of catecholamine axon degeneration. To further define the cells of origin of these dopamine boutons, the anterograde markerPhaseolus vulgarisleucoagglutinin was iontophoretically applied to the zona incerta. Six days later, rats received a 6‐hydroxydopamine injection into the zona incerta or the lateral ventricle, and 48 h later, double immunostaining was performed forPhaseolus vulgarisleucoagglutinin and tyrosine hydroxylase, luteinizing hormone‐releasing hormone, or glutamic acid decarboxylase on preoptic area vibratome sections.Following the 6‐hydroxydopamine injection into the anteroventral periventricular nucleus, autophagous cytolysosome‐containing degenerated axons were found in synaptic contact with both luteinizing hormone‐releasing hormone and GABA neurons in the medial preoptic area, confirming that these are dopaminergic connections. Following the double injection treatment, 6‐hydroxydop‐amine‐induced degenerated,Phaseolus vulgarisleucoagglutinin‐labeled dopamine axons originating in the zona incerta were not found to contact luteinizing hormone‐releasing hormone‐containing or GABA cells. Instead, many degenerated,Phaseolus vulgarisleucoagglutinin‐immunopositive boutons were observed in the dorsomedial and paraventricular nuclei of the hypothalamus. These observations indicate that tyrosine hydroxylase‐immunoreactive fibers terminating on the medial preoptic area luteinizing hormone‐releasing hormone and GABA cells are dopaminergic and most probably originate from dopamine neurons located in anterior