We previously found different effects on behavior, serotonin (5-HT) concentrations, 5-HT uptake sites, and 5-HT1A binding sites of neonatal 5,7-dihydroxytryptamine (5,7-DHT) lesions depending on the route of 5,7-DHT injection. To study the impact of early lesions on 5-HT1B sites as putative 5-HT terminal autoreceptors, we labelled them autoradiographically with [3H]5-HT 4 months after intraperitoneal (i.p.) or intracisternal (i.c.) 5,7-DHT injection during the first postnatal week and quantitated specific binding in 22 brain regions. Changes were confined to the subiculum and substantia nigra, regions with the most 5-HT1B-specific binding and projection areas of structures with high mRNA expression. Both routes of 5,7-DHT injection were associated with increases in specific binding in subiculum (24% for i.p. and 47% for i.c. route). In contrast, there was a 32% increase in specific binding in the substantia nigra in rats with lesions made i.c. but not i.p. No significant differences were found in nucleus accumbens, caudate-putamen or other brain areas. In saturation homogenate binding studies of 5-HT1B sites using [125I]iodocyanopindolol 1 month after i.p. injections, neonatal 5,7-DHT lesions did not significantly alter Bmax or Kd in the neocortex, striatum, diencephalon or brainstem. These data indicate the differential effects of the route of neonatal 5,7-DHT injections on plasticity of 5-HT1B receptor recognition sites and suggest the presence of a subpopulation of post-synaptically located 5-HT1B sites which increases in response to denervation. The data also suggest that sprouting of 5-HT neurons after neonatal 5,7-DHT lesions does not involve 5-HT1B sites.