Young, active and well-connected: adult-born neurons in the zebra finch are activated during singing

Neuronal replacement in the pallial song control nucleus HVC of adult zebra finches constitutes an interesting case of homeostatic plasticity; in spite of continuous addition and attrition of neurons in ensembles that code song elements, adult song remains remarkably invariant. New neurons migrate into HVC and later synapse with their target, arcopallial song nucleus RA (HVCRA). New HVCRA neurons respond to auditory stimuli (in anaesthetised animals), but whether and when they become functionally active during singing is unknown. We studied this, using 5-bromo-2'-deoxyuridine to birth-date neurons, combined with immunohistochemical detection of immediate-early gene (IEG) expression and retrograde tracer injections into RA to track connectivity. Interestingly, singing was followed by IEG expression in a substantial fraction of new neurons that were not retrogradely labelled from RA, suggesting a possible role in HVC-intrinsic network function. As new HVC neurons matured, the proportion of HVCRA neurons that expressed IEGs after singing increased significantly. Since it was previously shown that singing induces IEG expression in HVC also in deaf birds and that hearing song does not induce IEG expression in HVC, our data provide the first direct evidence that new HVC neurons are engaged in song motor behaviour.