DIRECT AND INDIRECT ENKEPHALINERGIC SYNAPTIC INPUTS TO THE RAT ARCUATE NUCLEUS STUDIED BY COMBINATION OF RETROGRADE TRACING AND IMMUNOCYTOCHEMISTRY

The origin of both direct and indirect enkephalinergic innervation pot entially able to influence neurons of the rat arcuate nucleus has been investigated by combining enkephalin immunocytochemistry and retrogra de axonal transport of a wheatgerm agglutinin-Apo horseradish peroxida se-gold complex. Twenty four hours after tissue injections of small vo lumes (20 nl) of the tracer into the arcuate nucleus, rats were treate d with colchicine and killed. In order to localize the enkephalinergic cells which directly innervate the arcuate nucleus, Vibratome section s were first silver-stained for detection of the wheatgerm agglutinin- Apohorseradish peroxidase-gold complex and then processed for enkephal in immunohistochemistry. To study the indirect enkephalinergic input t o the arcuate nucleus, an electron microscope detection of immunoreact ive synapses was carried out in areas rich in retrogradely labeled per ikarya. Perikarya both immunoreactive and retrogradely labeled were ob served ipsilaterally to the injection site in telencephalic structures such as the bed nucleus of the stria terminalis, medial preoptic and adjacent periventricular areas. Hypothalamic ipsilateral doubly labele d cells were localized principally in the dorsomedial nucleus and rost ral arcuate nucleus. The major direct inputs arising from brainstem st ructures concerns the dorsal and ventral parabrachial nuclei. Moreover , at the ultrastructural level, numerous enkephalinergic terminals wer e demonstrated to synapse with retrogradely labeled perikarya and dend rites localized in the medial preoptic area, the hypothalamic paravent ricular nucleus and the parabrachial nuclei providing evidence for an important enkephalinergic input on neurons projecting to the arcuate n ucleus. Taken together, our light and electron microscope studies stro ngly suggest that the arcuate nucleus is the target of an enkephaliner gic control originating from several regions and acting either directl y or indirectly on neurons projecting to the arcuate nucleus.