Lesions to Caudomedial Nidopallium Impair Individual Vocal Recognition in the Zebra Finch.

Many social animals can recognize other individuals by their vocalizations. This requires a memory system capable of mapping incoming acoustic signals to one of many known individuals. Using the zebra finch, a social songbird that uses songs and distance calls to communicate individual identity (Elie and Theunissen, 2018), we tested the role of two cortical-like brain regions in a vocal recognition task. We found that the rostral region of the Cadomedial Nidopallium (NCM), a secondary auditory region of the avian pallium, was necessary for maintaining auditory memories for conspecific vocalizations in both male and female birds, whereas HVC (used as a proper name), a premotor areas that gates auditory input into the vocal motor and song learning pathways in male birds (Roberts and Mooney, 2013), was not. Both NCM and HVC have previously been implicated for processing the tutor song in the context of song learning (Sakata and Yazaki-Sugiyama, 2020). Our results suggest that NCM might not only store songs as templates for future vocal imitation but also songs and calls for perceptual discrimination of vocalizers in both male and female birds. NCM could therefore operate as a site for auditory memories for vocalizations used in various facets of communication. We also observed that new auditory memories could be acquired without intact HVC or NCM but that for these new memories NCM lesions caused deficits in either memory capacity or auditory discrimination. These results suggest that the high-capacity memory functions of the avian pallial auditory system depend on NCM.SIGNIFICANCE STATEMENT Many aspects of vocal communication require the formation of auditory memories. Voice recognition, for example, requires a memory for vocalizers to identify acoustical features. In both birds and primates, the locus and neural correlates of these high-level memories remain poorly described. Previous work suggests that this memory formation is mediated by high-level sensory areas, not traditional memory areas such as the hippocampus. Using lesion experiments, we show that one secondary auditory brain region in songbirds that had previously been implicated in storing song memories for vocal imitation is also implicated in storing vocal memories for individual recognition. The role of the neural circuits in this region in interpreting the meaning of communication calls should be investigated in the future.

Brain-Derived Neurotrophic Factor Has a Transsynaptic Trophic Effect on Neural Activity in an Adult Forebrain Circuit.

While hormone-driven plasticity in the adult brain is well studied, the underlying cellular and molecular mechanisms are less well understood. One example of this is seasonal plasticity in the avian brain, where song nuclei exhibit hormonally driven changes in response to changing photoperiod and circulating sex steroid hormones. Hormone receptor activation in song nucleus HVC (proper name) elicits a robust change in activity in target nucleus RA (robust nucleus of the arcopallium), but the molecular signal responsible for this is unknown. This study addressed whether brain-derived neurotrophic factor (BDNF) mediates a transsynaptic effect from HVC to RA in male Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii). In situ hybridization confirmed an increase in BDNF expression in HVC neurons of birds switched to a long-day (LD) photoperiod plus systemically elevated testosterone (T) levels, compared with short-day (SD) conditions. BDNF expression was virtually absent in RA neurons of SD birds, increasing to barely detectable levels in a small subset of cells in LD+T birds. Infusion of BDNF protein adjacent to the RA of SD birds caused an increase in the spontaneous neuron firing rate. Conversely, the infusion of ANA12, a specific antagonist of the tyrosine-related kinase B (TrkB) for BDNF, prevented the increase in RA neuron firing rate in LD+T birds. These results indicate that BDNF is sufficient, and TrkB receptor activation is necessary, for the transsynaptic trophic effect exerted by HVC on RA. The dramatic change in the activity of RA neurons during the breeding season provides a clear example of transsynaptic BDNF effects in the adult brain in a functionally relevant circuit.SIGNIFICANCE STATEMENT Sex steroid hormones drive changes in brain circuits in all vertebrates, both within specific neurons and on their synaptic targets. Such changes can lead to profound changes in behavior, but little is known about the precise molecular mechanisms that underlie this process. We addressed this question in a seasonally breeding songbird and found that the trophic effects of one forebrain song nucleus on its target are mediated transsynaptically by the neurotrophin BDNF. This suggests that, in addition to their role in development, neurotrophins have critical roles in adult brain plasticity.

Network analysis of microRNA and mRNA seasonal dynamics in a highly plastic sensorimotor neural circuit.

BACKGROUND: Adult neurogenesis and the incorporation of adult-born neurons into functional circuits requires precise spatiotemporal coordination across molecular networks regulating a wide array of processes, including cell proliferation, apoptosis, neurotrophin signaling, and electrical activity. MicroRNAs (miRs) - short, non-coding RNA sequences that alter gene expression by post-transcriptional inhibition or degradation of mRNA sequences - may be involved in the global coordination of such diverse biological processes. To test the hypothesis that miRs related to adult neurogenesis and related cellular processes are functionally regulated in the nuclei of the avian song control circuit, we used microarray analyses to quantify changes in expression of miRs and predicted target mRNAs in the telencephalic nuclei HVC, the robust nucleus of arcopallium (RA), and the basal ganglia homologue Area X in breeding and nonbreeding Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelli). RESULTS: We identified 46 different miRs that were differentially expressed across seasons in the song nuclei. miR-132 and miR-210 showed the highest differential expression in HVC and Area X, respectively. Analyzing predicted mRNA targets of miR-132 identified 33 candidate target genes that regulate processes including cell cycle control, calcium signaling, and neuregulin signaling in HVC. Likewise, miR-210 was predicted to target 14 mRNAs differentially expressed across seasons that regulate serotonin, GABA, and dopamine receptor signaling and inflammation. CONCLUSIONS: Our results identify potential miR-mRNA regulatory networks related to adult neurogenesis and provide opportunities to discover novel genetic control of the diverse biological processes and factors related to the functional incorporation of new neurons to the adult brain.

HTR2 receptors in a songbird premotor cortical-like area modulate spectral characteristics of zebra finch song.

Serotonin [5-hydroxytryptamine (5-HT)] is involved in modulating an array of complex behaviors including learning, depression, and circadian rhythms. Additionally, HTR2 receptors on layer V pyramidal neurons are thought to mediate the actions of psychedelic drugs; the native function of these receptors at this site, however, remains unknown. Previously, we found that activation of HTR2 receptors in the zebra finch forebrain song premotor structure the robust nucleus of the arcopallium (RA) led to increased excitation, and that endogenous 5-HT could roughly double spontaneous firing rate. Here, using in vivo single-unit recordings, we found that direct application of 5-HT to these same RA projection neurons, which are analogous to layer V cortical pyramidal neurons, caused a significant increase in the number of action potentials per song-related burst, and a dramatic decrease in signal-to-noise ratio. Injection of the serotonergic neurotoxin 5,7-dihydroxytryptamine into the third ventricle greatly reduced telencephalic 5-HT and resulted in decreased fundamental frequency of harmonic syllables as well as increased goodness of pitch. Both of these results can be explained by the observed actions of 5-HT on RA projection neurons, and both effects recovered to baseline within 2 weeks following the toxin injection. These results show that 5-HT is involved in modulating spectral properties of song, likely via effects on RA projection neurons, but that adult zebra finches can partially compensate for this deficit within 7 d.

Serotonin, via HTR2 receptors, excites neurons in a cortical-like premotor nucleus necessary for song learning and production.

Serotonin (5-HT) is a neuromodulator that is important for neural development, learning and memory, mood, and perception. Dysfunction of the serotonin system is central to depression and other clinically important mood disorders and has been linked with learning deficits. In mammals, 5-HT release from the raphe nuclei in the brainstem can modulate the functional properties of cortical neurons, influencing sensory and motor processing. Birds also have serotonergic neurons in the dorsal raphe, suggesting that 5-HT plays similar roles in sensory and motor processing, perhaps modulating brain circuitry underlying birdsong. To investigate this possibility, we measured the effects of 5-HT on spontaneous firing of projection neurons in the premotor robust nucleus of the arcopallium in brain slices from male zebra finches. These neurons are thought be akin to cortical layer V pyramidal neurons. 5-HT dramatically and reversibly enhanced the endogenous firing of RA neurons. Using pharmacological agonists and antagonists in vitro, we determined this action is mediated via HTR2 receptors, which we verified are expressed by in situ hybridization. Finally, focal administration of the serotonin selective reuptake inhibitor fluvoxamine revealed that endogenous 5-HT is sufficient to mediate this effect in vivo. These findings reveal a modulatory action of serotonin on the physiology of the song system circuitry and suggest a novel role of serotonin in regulating song production and/or learning; further understanding of the role of 5-HT in this system may help illuminate the complex role of this neuromodulator in social interactions and motor plasticity in humans.

A canonical neural mechanism for behavioral variability.

The ability to generate variable movements is essential for learning and adjusting complex behaviours. This variability has been linked to the temporal irregularity of neuronal activity in the central nervous system. However, how neuronal irregularity actually translates into behavioural variability is unclear. Here we combine modelling, electrophysiological and behavioural studies to address this issue. We demonstrate that a model circuit comprising topographically organized and strongly recurrent neural networks can autonomously generate irregular motor behaviours. Simultaneous recordings of neurons in singing finches reveal that neural correlations increase across the circuit driving song variability, in agreement with the model predictions. Analysing behavioural data, we find remarkable similarities in the babbling statistics of 5-6-month-old human infants and juveniles from three songbird species and show that our model naturally accounts for these 'universal' statistics.

A daily oscillation in the fundamental frequency and amplitude of harmonic syllables of zebra finch song.

Complex motor skills are more difficult to perform at certain points in the day (for example, shortly after waking), but the daily trajectory of motor-skill error is more difficult to predict. By undertaking a quantitative analysis of the fundamental frequency (FF) and amplitude of hundreds of zebra finch syllables per animal per day, we find that zebra finch song follows a previously undescribed daily oscillation. The FF and amplitude of harmonic syllables rises across the morning, reaching a peak near mid-day, and then falls again in the late afternoon until sleep. This oscillation, although somewhat variable, is consistent across days and across animals and does not require serotonin, as animals with serotonergic lesions maintained daily oscillations. We hypothesize that this oscillation is driven by underlying physiological factors which could be shared with other taxa. Song production in zebra finches is a model system for studying complex learned behavior because of the ease of gathering comprehensive behavioral data and the tractability of the underlying neural circuitry. The daily oscillation that we describe promises to reveal new insights into how time of day affects the ability to accomplish a variety of complex learned motor skills.

Dietary retinoic acid affects song maturation and gene expression in the song system of the zebra finch.

Vitamin A, an essential nutrient, is required in its acidic form (retinoic acid) for normal embryogenesis and neuronal development, typically within well-defined concentration ranges. In zebra finches, a songbird species, localized retinoic acid synthesis in the brain is important for the development of song, a learned behavior sharing significant commonalities with speech acquisition in humans. We tested how dietary retinoic acid affects the development of song behavior and the brain's system for song control. Supplemental doses of retinoic acid given to juveniles during the critical period for song learning resulted in more variable or plastic-like songs when the birds reached adulthood, compared to the normal songs of vehicle-fed controls. We also observed that several genes (brinp1, nrgn, rxr-alpha, and sdr2/scdr9) had altered levels of expression in specific nuclei of the song system when comparing the experimental and control diet groups. Interestingly, we found significant correlations between gene expression levels in nuclei of the anterior forebrain pathway (lMAN and area X) and the degree of variability in the recorded songs. We observed, however, no major morphological effects such as changes in the volumes of song nuclei. Overall, our results lend further support to a fundamental role of retinoic acid in song maturation and point to possible molecular pathways associated with this action. The data also demonstrate that dietary content of Vitamin A can affect the maturation of a naturally learned complex behavior.