Abstract: Neurogenesis in the avian brain produces new neurons (HVC NNs) into pre-existing neural circuits related to song production. Of the adult born NNs in HVC, 50% can be backfilled from RA while the other 50% cannot. The 50% that do not project to RA express DARPP-32, a phosphoprotein downstream of dopamine receptors. DARPP-32+ NNs do not project to Area X or Nucleus Avalanche, the two other projection targets of HVC, and they are not inhibitory interneurons. These DARPP-32+ HVC NNs cannot be retrogradely labeled to RA at 28 days post mitosis, leading us to hypothesize they may constitute a unique subtype of HVC NN distinct from HVC RA NNs. Our lab previously showed that the DARPP-32+ NNs have transiently larger nuclei than DARPP-32-neg NNs. DARPP-32+ NNs also receive putative tyrosine hydroxylase synapses, showing these neurons could receive dopamine, norepinephrine or epinephrine synaptic contacts. Male zebra finches sing two song types of song, directed and undirected, depending on the social context. Directed song occurs when a female is present and the male sings directly at a potential mate. In this context, each rendition of the song is highly acoustically similar to the previous one. Undirected song is sung in the absence of a female, and has higher rendition to rendition variability, less motifs and introductory notes per bout, and is sung more slowly than directed song. In terms of dopamine, a study showed that when a female is present but dopamine is blocked from binding in HVC, directed song from the male is eliminated but not undirected song, which remains functional. This demonstrates that dopamine in HVC plays a key role in the social aspects of directed versus undirected song. This led us to hypothesize that DARPP-32+ HVC NNs may be preferentially activated during directed song, when there is increased dopamine in HVC. We can test this using an immediate early gene called ZENK, which is expressed following high levels of neuronal activity. HVC RA NNs are sparsely active and will only fire when the birds sing, so we can be confident that the amount of ZENK seen is related to song production. We injected adult male zebra finches with BrdU to label dividing cells. 3, 5, or 8-weeks later we injected fluorogold into RA to backfill HVC-RA neurons. We then placed the birds in sound attenuated recording chambers in one of three conditions: directed song (female present) undirected (no female present), and non-singing control. We are testing whether DARPP-32+ NNs are more likely to be active during directed or undirected song. We expect to see a higher proportion of DARPP-32+ NNs active during directed song, showing that more dopamine is present in HVC during directed song. This could suggest that DARPP-32+ NNs serve as an internal switch between directed and undirected song.
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