Anterior forebrain pathway in parrots is necessary for producing learned vocalizations with individual signatures.

Parrots have enormous vocal imitation capacities and produce individually unique vocal signatures. Like songbirds, parrots have a nucleated neural song system with distinct anterior (AFP) and posterior forebrain pathways (PFP). To test if song systems of parrots and songbirds, which diverged over 50 million years ago, have a similar functional organization, we first established a neuroscience-compatible call-and-response behavioral paradigm to elicit learned contact calls in budgerigars (Melopsittacus undulatus). Using variational autoencoder-based machine learning methods, we show that contact calls within affiliated groups converge but that individuals maintain unique acoustic features, or vocal signatures, even after call convergence. Next, we transiently inactivated the outputs of AFP to test if learned vocalizations can be produced by the PFP alone. As in songbirds, AFP inactivation had an immediate effect on vocalizations, consistent with a premotor role. But in contrast to songbirds, where the isolated PFP is sufficient to produce stereotyped and acoustically normal vocalizations, isolation of the budgerigar PFP caused a degradation of call acoustic structure, stereotypy, and individual uniqueness. Thus, the contribution of AFP and the capacity of isolated PFP to produce learned vocalizations have diverged substantially between songbirds and parrots, likely driven by their distinct behavioral ecology and neural connectivity.

Songbird mesostriatal dopamine pathways are spatially segregated before the onset of vocal learning.

Diverse dopamine (DA) pathways send distinct reinforcement signals to different striatal regions. In adult songbirds, a DA pathway from the ventral tegmental area (VTA) to Area X, the striatal nucleus of the song system, carries singing-related performance error signals important for learning. Meanwhile, a parallel DA pathway to a medial striatal area (MST) arises from a distinct group of neighboring DA neurons that lack connectivity to song circuits and do not encode song error. To test if the structural and functional segregation of these two pathways depends on singing experience, we carried out anatomical studies early in development before the onset of song learning. We find that distinct VTA neurons project to either Area X or MST in juvenile birds before the onset of substantial vocal practice. Quantitative comparisons of early juveniles (30-35 days post hatch), late juveniles (60-65 dph), and adult (>90 dph) brains revealed an outsized expansion of Area X-projecting neurons relative to MST-projecting neurons in VTA over development. These results show that a mesostriatal DA system dedicated to social communication can exist and be spatially segregated before the onset of vocal practice and associated sensorimotor experience.

Dopaminergic error signals retune to social feedback during courtship.

Hunger, thirst, loneliness and ambition determine the reward value of food, water, social interaction and performance outcome1. Dopamine neurons respond to rewards meeting these diverse needs2-8, but it remains unclear how behaviour and dopamine signals change as priorities change with new opportunities in the environment. One possibility is that dopamine signals for distinct drives are routed to distinct dopamine pathways9,10. Another possibility is that dopamine signals in a given pathway are dynamically tuned to rewards set by the current priority. Here we used electrophysiology and fibre photometry to test how dopamine signals associated with quenching thirst, singing a good song and courting a mate change as male zebra finches (Taeniopygia guttata) were provided with opportunities to retrieve water, evaluate song performance or court a female. When alone, water reward signals were observed in two mesostriatal pathways but singing-related performance error signals were routed to Area X, a striatal nucleus specialized for singing. When courting a female, water seeking was reduced and dopamine responses to both water and song performance outcomes diminished. Instead, dopamine signals in Area X were driven by female calls timed with the courtship song. Thus the dopamine system handled coexisting drives by routing vocal performance and social feedback signals to a striatal area for communication and by flexibly re-tuning to rewards set by the prioritized drive.

Dopaminergic signals for improved parental behavior.

In this issue of Neuron, Xie et al.1 record and manipulate dopaminergic activity as mice engage in parental care. Dopaminergic prediction error signals previously implicated in food rewards were associated with retrieving isolated pups to the nest, showing that neural mechanisms long associated with reinforcement learning can be repurposed for aspects of parenting.

Dopamine neurons evaluate natural fluctuations in performance quality.

Many motor skills are learned by comparing ongoing behavior to internal performance benchmarks. Dopamine neurons encode performance error in behavioral paradigms where error is externally induced, but it remains unknown whether dopamine also signals the quality of natural performance fluctuations. Here, we record dopamine neurons in singing birds and examine how spontaneous dopamine spiking activity correlates with natural fluctuations in ongoing song. Antidromically identified basal ganglia-projecting dopamine neurons correlate with recent, and not future, song variations, consistent with a role in evaluation, not production. Furthermore, maximal dopamine spiking occurs at a single vocal target, consistent with either actively maintaining the existing song or shifting the song to a nearby form. These data show that spontaneous dopamine spiking can evaluate natural behavioral fluctuations unperturbed by experimental events such as cues or rewards.

Songbird subthalamic neurons project to dopaminergic midbrain and exhibit singing-related activity.

Skill learning requires motor output to be evaluated against internal performance benchmarks. In songbirds, ventral tegmental area (VTA) dopamine neurons (DA) signal performance errors important for learning, but it remains unclear which brain regions project to VTA and how these inputs may contribute to DA error signaling. Here, we find that the songbird subthalamic nucleus (STN) projects to VTA and that STN microstimulation can excite VTA neurons. We also discover that STN receives inputs from motor cortical, auditory cortical, and ventral pallidal brain regions previously implicated in song evaluation. In the first neural recordings from songbird STN, we discover that the activity of most STN neurons is associated with body movements and not singing, but a small fraction of neurons exhibits precise song timing and performance error signals. Our results place the STN in a pathway important for song learning, but not song production, and expand the territories of songbird brain potentially associated with song learning.NEW & NOTEWORTHY Songbird subthalamic (STN) neurons exhibit singing-related signals and are interconnected with the motor cortical nucleus, auditory pallium, ventral pallidum, and ventral tegmental area, areas important for song generation and learning.

Movement signaling in ventral pallidum and dopaminergic midbrain is gated by behavioral state in singing birds.

Movement-related neuronal discharge in ventral tegmental area (VTA) and ventral pallidum (VP) is inconsistently observed across studies. One possibility is that some neurons are movement related and others are not. Another possibility is that the precise behavioral conditions matter-that a single neuron can be movement related under certain behavioral states but not others. We recorded single VTA and VP neurons in birds transitioning between singing and nonsinging states while monitoring body movement with microdrive-mounted accelerometers. Many VP and VTA neurons exhibited body movement-locked activity exclusively when the bird was not singing. During singing, VP and VTA neurons could switch off their tuning to body movement and become instead precisely time-locked to specific song syllables. These changes in neuronal tuning occurred rapidly at state boundaries. Our findings show that movement-related activity in limbic circuits can be gated by behavioral context.NEW & NOTEWORTHY Neural signals in the limbic system have long been known to represent body movements as well as reward. Here, we show that single neurons dramatically change their tuning from movement to song timing when a bird starts to sing.

Cortex-dependent corrections as the tongue reaches for and misses targets.

Precise tongue control is necessary for drinking, eating and vocalizing1-3. However, because tongue movements are fast and difficult to resolve, neural control of lingual kinematics remains poorly understood. Here we combine kilohertz-frame-rate imaging and a deep-learning-based neural network to resolve 3D tongue kinematics in mice drinking from a water spout. Successful licks required corrective submovements that-similar to online corrections during primate reaches4-11-occurred after the tongue missed unseen, distant or displaced targets. Photoinhibition of anterolateral motor cortex impaired corrections, which resulted in hypometric licks that missed the spout. Neural activity in anterolateral motor cortex reflected upcoming, ongoing and past corrective submovements, as well as errors in predicted spout contact. Although less than a tenth of a second in duration, a single mouse lick exhibits the hallmarks of online motor control associated with a primate reach, including cortex-dependent corrections after misses.

Fabrication of Injectable Micro-Scale Opto-Electronically Transduced Electrodes (MOTEs) for Physiological Monitoring.

In vivo, chronic neural recording is critical to understand the nervous system, while a tetherless, miniaturized recording unit can render such recording minimally invasive. We present a tetherless, injectable micro-scale opto-electronically transduced electrode (MOTE) that is ~60µm × 30µm × 330µm, the smallest neural recording unit to date. The MOTE consists of an AlGaAs micro-scale light emitting diode (µLED) heterogeneously integrated on top of conventional 180nm complementary metal-oxide-semiconductor (CMOS) circuit. The MOTE combines the merits of optics (AlGaAs µLED for power and data uplink), and of electronics (CMOS for signal amplification and encoding). The optical powering and communication enable the extreme scaling while the electrical circuits provide a high temporal resolution (<100µs). This paper elaborates on the heterogeneous integration in MOTEs, a topic that has been touted without much demonstration on feasibility or scalability. Based on photolithography, we demonstrate how to build heterogenous systems that are scalable as well as biologically stable - the MOTEs can function in saline water for more than six months, and in a mouse brain for two months (and counting). We also present handling/insertion techniques for users (i.e. biologists) to deploy MOTEs with little or no extra training.

Actor-critic reinforcement learning in the songbird.

It feels rewarding to ace your opponent on match point. Here, we propose common mechanisms underlie reward and performance learning. First, when a singing bird unexpectedly hits the right note, its dopamine (DA) neurons are activated as when a thirsty monkey receives an unexpected juice reward. Second, these DA signals reinforce vocal variations much as they reinforce stimulus-response associations. Third, limbic inputs to DA neurons signal the predicted quality of song syllables much like they signal the predicted reward value of a place or a stimulus during foraging. Finally, songbirds may solve difficult problems in reinforcement learning - such as credit assignment and catastrophic forgetting - with node perturbation and consolidation of reinforced vocal patterns in motor cortical circuits. Consolidation occurs downstream of a canonical 'actor-critic' circuit motif that learns to maximize performance quality in essentially the same way it learns to maximize reward: by computing and learning from prediction errors.

Songbird Ventral Pallidum Sends Diverse Performance Error Signals to Dopaminergic Midbrain.

Motor skills improve with practice, requiring outcomes to be evaluated against ever-changing performance benchmarks, yet it remains unclear how performance error signals are computed. Here, we show that the songbird ventral pallidum (VP) is required for song learning and sends diverse song timing and performance error signals to the ventral tegmental area (VTA). Viral tracing revealed inputs to VP from auditory and vocal motor thalamus, auditory and vocal motor cortex, and VTA. Our findings show that VP circuits, commonly associated with hedonic functions, signal performance error during motor sequence learning.

Automated home cage training of mice in a hold-still center-out reach task.

An obstacle to understanding neural mechanisms of movement is the complex, distributed nature of the mammalian motor system. Here we present a novel behavioral paradigm for high-throughput dissection of neural circuits underlying mouse forelimb control. Custom touch-sensing joysticks were used to quantify mouse forelimb trajectories with micron-millisecond spatiotemporal resolution. Joysticks were integrated into computer-controlled, rack-mountable home cages, enabling batches of mice to be trained in parallel. Closed loop behavioral analysis enabled online control of reward delivery for automated training. We used this system to show that mice can learn, with no human handling, a direction-specific hold-still center-out reach task in which a mouse first held its right forepaw still before reaching out to learned spatial targets. Stabilogram diffusion analysis of submillimeter-scale micromovements produced during the hold demonstrate that an active control process, akin to upright balance, was implemented to maintain forepaw stability. Trajectory decomposition methods, previously used in primates, were used to segment hundreds of thousands of forelimb trajectories into millions of constituent kinematic primitives. This system enables rapid dissection of neural circuits for controlling motion primitives from which forelimb sequences are built. NEW & NOTEWORTHY A novel joystick design resolves mouse forelimb kinematics with micron-millisecond precision. Home cage training is used to train mice in a hold-still center-out reach task. Analytical methods, previously used in primates, are used to decompose mouse forelimb trajectories into kinematic primitives.

Vocal learning in songbirds requires cholinergic signaling in a motor cortex-like nucleus.

Cholinergic inputs to cortex modulate plasticity and sensory processing, yet little is known about their role in motor control. Here, we show that cholinergic signaling in a songbird vocal motor cortical area, the robust nucleus of the arcopallium (RA), is required for song learning. Reverse microdialysis of nicotinic and muscarinic receptor antagonists into RA in juvenile birds did not significantly affect syllable timing or acoustic structure during vocal babbling. However, chronic blockade over weeks reduced singing quantity and impaired learning, resulting in an impoverished song with excess variability, abnormal acoustic features, and reduced similarity to tutor song. The demonstration that cholinergic signaling in a motor cortical area is required for song learning motivates the songbird as a tractable model system to identify roles of the basal forebrain cholinergic system in motor control. NEW & NOTEWORTHY Cholinergic inputs to cortex are evolutionarily conserved and implicated in sensory processing and synaptic plasticity. However, functions of cholinergic signals in motor areas are understudied and poorly understood. Here, we show that cholinergic signaling in a songbird vocal motor cortical area is not required for normal vocal variability during babbling but is essential for developmental song learning. Cholinergic modulation of motor cortex is thus required for learning but not for the ability to sing.

A Stable Neural Code for Birdsong.

Proteins, synapses, and neural connections are in constant flux, yet motor behaviors somehow remain stable. In this issue of Neuron, Katlowitz et al. (2018) show that temporally precise neural activity driving birdsong production is stable for weeks.

Place preference and vocal learning rely on distinct reinforcers in songbirds.

In reinforcement learning (RL) agents are typically tasked with maximizing a single objective function such as reward. But it remains poorly understood how agents might pursue distinct objectives at once. In machines, multiobjective RL can be achieved by dividing a single agent into multiple sub-agents, each of which is shaped by agent-specific reinforcement, but it remains unknown if animals adopt this strategy. Here we use songbirds to test if navigation and singing, two behaviors with distinct objectives, can be differentially reinforced. We demonstrate that strobe flashes aversively condition place preference but not song syllables. Brief noise bursts aversively condition song syllables but positively reinforce place preference. Thus distinct behavior-generating systems, or agencies, within a single animal can be shaped by correspondingly distinct reinforcement signals. Our findings suggest that spatially segregated vocal circuits can solve a credit assignment problem associated with multiobjective learning.

Dopamine neurons encode performance error in singing birds.

Many behaviors are learned through trial and error by matching performance to internal goals. Yet neural mechanisms of performance evaluation remain poorly understood. We recorded basal ganglia-projecting dopamine neurons in singing zebra finches as we controlled perceived song quality with distorted auditory feedback. Dopamine activity was phasically suppressed after distorted syllables, consistent with a worse-than-predicted outcome, and was phasically activated at the precise moment of the song when a predicted distortion did not occur, consistent with a better-than-predicted outcome. Error response magnitude depended on distortion probability. Thus, dopaminergic error signals can evaluate behaviors that are not learned for reward and are instead learned by matching performance outcomes to internal goals.

Origins of basal ganglia output signals in singing juvenile birds.

Across species, complex circuits inside the basal ganglia (BG) converge on pallidal output neurons that exhibit movement-locked firing patterns. Yet the origins of these firing patterns remain poorly understood. In songbirds during vocal babbling, BG output neurons homologous to those found in the primate internal pallidal segment are uniformly activated in the tens of milliseconds prior to syllable onsets. To test the origins of this remarkably homogenous BG output signal, we recorded from diverse upstream BG cell types during babbling. Prior to syllable onsets, at the same time that internal pallidal segment-like neurons were activated, putative medium spiny neurons, fast spiking and tonically active interneurons also exhibited transient rate increases. In contrast, pallidal neurons homologous to those found in primate external pallidal segment exhibited transient rate decreases. To test origins of these signals, we performed recordings following lesion of corticostriatal inputs from premotor nucleus HVC. HVC lesions largely abolished these syllable-locked signals. Altogether, these findings indicate a striking homogeneity of syllable timing signals in the songbird BG during babbling and are consistent with a role for the indirect and hyperdirect pathways in transforming cortical inputs into BG outputs during an exploratory behavior.

A variability-generating circuit goes awry in a songbird model of the FOXP2 speech disorder.

FOXP2 mutations cause a monogenic speech disorder in humans. In this issue of Neuron, Murugan et al. (2013) show that knockdown of FoxP2 in the songbird basal ganglia causes abnormal vocal variability and excess bursting in a frontal cortical nucleus.

Basal ganglia output to the thalamus: still a paradox.

The basal ganglia (BG)-recipient thalamus controls motor output but it remains unclear how its activity is regulated. Several studies report that thalamic activation occurs via disinhibition during pauses in the firing of inhibitory pallidal inputs from the BG. Other studies indicate that thalamic spiking is triggered by pallidal inputs via post-inhibitory 'rebound' calcium spikes. Finally excitatory cortical inputs can drive thalamic activity, which becomes entrained, or time-locked, to pallidal spikes. We present a unifying framework where these seemingly distinct results arise from a continuum of thalamic firing 'modes' controlled by excitatory inputs. We provide a mechanistic explanation for paradoxical pallidothalamic coactivations observed during behavior that raises new questions about what information is integrated in the thalamus to control behavior.

Integration of cortical and pallidal inputs in the basal ganglia-recipient thalamus of singing birds.

The basal ganglia-recipient thalamus receives inhibitory inputs from the pallidum and excitatory inputs from cortex, but it is unclear how these inputs interact during behavior. We recorded simultaneously from thalamic neurons and their putative synaptically connected pallidal inputs in singing zebra finches. We find, first, that each pallidal spike produces an extremely brief (∼5 ms) pulse of inhibition that completely suppresses thalamic spiking. As a result, thalamic spikes are entrained to pallidal spikes with submillisecond precision. Second, we find that the number of thalamic spikes that discharge within a single pallidal interspike interval (ISI) depends linearly on the duration of that interval but does not depend on pallidal activity prior to the interval. In a detailed biophysical model, our results were not easily explained by the postinhibitory "rebound" mechanism previously observed in anesthetized birds and in brain slices, nor could most of our data be characterized as "gating" of excitatory transmission by inhibitory pallidal input. Instead, we propose a novel "entrainment" mechanism of pallidothalamic transmission that highlights the importance of an excitatory conductance that drives spiking, interacting with brief pulses of pallidal inhibition. Building on our recent finding that cortical inputs can drive syllable-locked rate modulations in thalamic neurons during singing, we report here that excitatory inputs affect thalamic spiking in two ways: by shortening the latency of a thalamic spike after a pallidal spike and by increasing thalamic firing rates within individual pallidal ISIs. We present a unifying biophysical model that can reproduce all known modes of pallidothalamic transmission--rebound, gating, and entrainment--depending on the amount of excitation the thalamic neuron receives.