Insights into the Neural and Genetic Basis of Vocal Communication.

The use of vocalizations to communicate information and elaborate social bonds is an adaptation seen in many vertebrate species. Human speech is an extreme version of this pervasive form of communication. Unlike the vocalizations exhibited by the majority of land vertebrates, speech is a learned behavior requiring early sensory exposure and auditory feedback for its development and maintenance. Studies in humans and a small number of other species have provided insights into the neural and genetic basis for learned vocal communication and are helping to delineate the roles of brain circuits across the cortex, basal ganglia, and cerebellum in generating vocal behaviors. This Review provides an outline of the current knowledge about these circuits and the genes implicated in vocal communication, as well as a perspective on future research directions in this field.

The hidden fitness of the male zebra finch courtship song.

Vocal learning in songbirds is thought to have evolved through sexual selection, with female preference driving males to develop large and varied song repertoires1-3. However, many songbird species learn only a single song in their lifetime4. How sexual selection drives the evolution of single-song repertoires is not known. Here, by applying dimensionality-reduction techniques to the singing behaviour of zebra finches (Taeniopygia guttata), we show that syllable spread in low-dimensional feature space explains how single songs function as honest indicators of fitness. We find that this Gestalt measure of behaviour captures the spectrotemporal distinctiveness of song syllables in zebra finches; that females strongly prefer songs that occupy more latent space; and that matching path lengths in low-dimensional space is difficult for young males. Our findings clarify how simple vocal repertoires may have evolved in songbirds and indicate divergent strategies for how sexual selection can shape vocal learning.

Anterograde transneuronal tracing and genetic control with engineered yellow fever vaccine YFV-17D.

Transneuronal viruses are powerful tools for tracing neuronal circuits or delivering genes to specific neurons in the brain. While there are multiple retrograde viruses, few anterograde viruses are available. Further, available anterograde viruses often have limitations such as retrograde transport, high neuronal toxicity or weak signals. We developed an anterograde viral system based on a live attenuated vaccine for yellow fever-YFV-17D. Replication- or packaging-deficient mutants of YFV-17D can be reconstituted in the brain, leading to efficient synapse-specific and anterograde-only transneuronal spreading, which can be controlled to achieve either monosynaptic or polysynaptic tracing. Moreover, inducible transient replication of YFV-17D mutant is sufficient to induce permanent transneuronal genetic modifications without causing neuronal toxicity. The engineered YFV-17D systems can be used to express fluorescent markers, sensors or effectors in downstream neurons, thus providing versatile tools for mapping and functionally controlling neuronal circuits.

Imaging auditory representations of song and syllables in populations of sensorimotor neurons essential to vocal communication.

Vocal communication depends on the coordinated activity of sensorimotor neurons important to vocal perception and production. How vocalizations are represented by spatiotemporal activity patterns in these neuronal populations remains poorly understood. Here we combined intracellular recordings and two-photon calcium imaging in anesthetized adult zebra finches (Taeniopygia guttata) to examine how learned birdsong and its component syllables are represented in identified projection neurons (PNs) within HVC, a sensorimotor region important for song perception and production. These experiments show that neighboring HVC PNs can respond at markedly different times to song playback and that different syllables activate spatially intermingled PNs within a local (~100 µm) region of HVC. Moreover, noise correlations were stronger between PNs that responded most strongly to the same syllable and were spatially graded within and between classes of PNs. These findings support a model in which syllabic and temporal features of song are represented by spatially intermingled PNs functionally organized into cell- and syllable-type networks within local spatial scales in HVC.

Rapid spine stabilization and synaptic enhancement at the onset of behavioural learning.

Behavioural learning depends on the brain's capacity to respond to instructive experience and is often enhanced during a juvenile sensitive period. How instructive experience acts on the juvenile brain to trigger behavioural learning remains unknown. In vitro studies show that forms of synaptic strengthening thought to underlie learning are accompanied by an increase in the stability, number and size of dendritic spines, which are the major sites of excitatory synaptic transmission in the vertebrate brain. In vivo imaging studies in sensory cortical regions reveal that these structural features can be affected by disrupting sensory experience and that spine turnover increases during sensitive periods for sensory map formation. These observations support two hypotheses: first, the increased capacity for behavioural learning during a sensitive period is associated with enhanced spine dynamics on sensorimotor neurons important for the learned behaviour; second, instructive experience rapidly stabilizes and strengthens these dynamic spines. Here we report a test of these hypotheses using two-photon in vivo imaging to measure spine dynamics in zebra finches, which learn to sing by imitating a tutor song during a juvenile sensitive period. Spine dynamics were measured in the forebrain nucleus HVC, the proximal site where auditory information merges with an explicit song motor representation, immediately before and after juvenile finches first experienced tutor song. Higher levels of spine turnover before tutoring correlated with a greater capacity for subsequent song imitation. In juveniles with high levels of spine turnover, hearing a tutor song led to the rapid ( approximately 24-h) stabilization, accumulation and enlargement of dendritic spines in HVC. Moreover, in vivo intracellular recordings made immediately before and after the first day of tutoring revealed robust enhancement of synaptic activity in HVC. These findings suggest that behavioural learning results when instructive experience is able to rapidly stabilize and strengthen synapses on sensorimotor neurons important for the control of the learned behaviour.

Improving the Success of Strategic Management Using Big Data.

Strategic management involves determining organizational goals, implementing a strategic plan, and properly allocating resources. Poor access to pertinent and timely data misidentifies clinical goals, prevents effective resource allocation, and generates waste from inaccurate forecasting. Loss of operational efficiency diminishes the value stream, adversely impacts the quality of patient care, and hampers effective strategic management. We have pioneered an approach using big data to create competitive advantage by identifying trends in clinical practice, accurately anticipating future needs, and strategically allocating resources for maximum impact.

Neural circuits: How the songbird brain orchestrates courtship displays.

Volitional production of complex behaviors can be motivated by intrinsic rewards and also by extrinsic cues, like social engagement. A new study has revealed the neural circuit permitting social motivation to release multi-component courtship behaviors in a songbird, specifically the zebra finch.

Bidirectional synaptic changes in deep and superficial hippocampal neurons following in vivo activity.

Neuronal activity during experience is thought to induce plastic changes within the hippocampal network that underlie memory formation, although the extent and details of such changes in vivo remain unclear. Here, we employed a temporally precise marker of neuronal activity, CaMPARI2, to label active CA1 hippocampal neurons in vivo, followed by immediate acute slice preparation and electrophysiological quantification of synaptic properties. Recently active neurons in the superficial sublayer of stratum pyramidale displayed larger post-synaptic responses at excitatory synapses from area CA3, with no change in pre-synaptic release probability. In contrast, in vivo activity correlated with weaker pre- and post-synaptic excitatory weights onto pyramidal cells in the deep sublayer. In vivo activity of deep and superficial neurons within sharp-wave/ripples was bidirectionally changed across experience, consistent with the observed changes in synaptic weights. These findings reveal novel, fundamental mechanisms through which the hippocampal network is modified by experience to store information.

What Is the Role of Thalamostriatal Circuits in Learning Vocal Sequences?

Basal ganglia (BG) circuits integrate sensory and motor-related information from the cortex, thalamus, and midbrain to guide learning and production of motor sequences. Birdsong, like speech, is comprised of precisely sequenced vocal elements. Learning song sequences during development relies on Area X, a vocalization related region in the medial striatum of the songbird BG. Area X receives inputs from cortical-like pallial song circuits and midbrain dopaminergic circuits and sends projections to the thalamus. It has recently been shown that thalamic circuits also send substantial projections back to Area X. Here, we outline a gated-reinforcement learning model for how Area X may use signals conveyed by thalamostriatal inputs to direct song learning. Integrating conceptual advances from recent mammalian and songbird literature, we hypothesize that thalamostriatal pathways convey signals linked to song syllable onsets and offsets and influence striatal circuit plasticity via regulation of cholinergic interneurons (ChIs). We suggest that syllable sequence associated vocal-motor information from the thalamus drive precisely timed pauses in ChIs activity in Area X. When integrated with concurrent corticostriatal and dopaminergic input, this circuit helps regulate plasticity on medium spiny neurons (MSNs) and the learning of syllable sequences. We discuss new approaches that can be applied to test core ideas of this model and how associated insights may provide a framework for understanding the function of BG circuits in learning motor sequences.

T-cell Acute Lymphoblastic Leukemia in a Patient with Pre-existing Essential Thrombocythemia: A Case Report and Literature Review.

The occurrence of T-cell acute lymphoblastic leukemia (T-ALL), on a background of preexisting Philadelphia-negative Myeloproliferative neoplasm is rare. Among the few reported cases where no deep molecular sequencing was performed, it was difficult to ascertain whether these leukemia's occurred de-novo or were due to the clonal progression of underlying MPN. We present a case of a 49-year-old man with a history of essential thrombocythemia who subsequently developed T-ALL. By utilizing next generation sequencing we were able to determine that these two entities originated from two distinct clones and were likely random events. We report the outcome and review the literature.

Cellular transcriptomics reveals evolutionary identities of songbird vocal circuits.

Birds display advanced behaviors, including vocal learning and problem-solving, yet lack a layered neocortex, a structure associated with complex behavior in mammals. To determine whether these behavioral similarities result from shared or distinct neural circuits, we used single-cell RNA sequencing to characterize the neuronal repertoire of the songbird song motor pathway. Glutamatergic vocal neurons had considerable transcriptional similarity to neocortical projection neurons; however, they displayed regulatory gene expression patterns more closely related to neurons in the ventral pallium. Moreover, while γ-aminobutyric acid-releasing neurons in this pathway appeared homologous to those in mammals and other amniotes, the most abundant avian class is largely absent in the neocortex. These data suggest that songbird vocal circuits and the mammalian neocortex have distinct developmental origins yet contain transcriptionally similar neurons.

Expression of FoxP2 in the basal ganglia regulates vocal motor sequences in the adult songbird.

Disruption of the transcription factor FoxP2, which is enriched in the basal ganglia, impairs vocal development in humans and songbirds. The basal ganglia are important for the selection and sequencing of motor actions, but the circuit mechanisms governing accurate sequencing of learned vocalizations are unknown. Here, we show that expression of FoxP2 in the basal ganglia is vital for the fluent initiation and termination of birdsong, as well as the maintenance of song syllable sequencing in adulthood. Knockdown of FoxP2 imbalances dopamine receptor expression across striatal direct-like and indirect-like pathways, suggesting a role of dopaminergic signaling in regulating vocal motor sequencing. Confirming this prediction, we show that phasic dopamine activation, and not inhibition, during singing drives repetition of song syllables, thus also impairing fluent initiation and termination of birdsong. These findings demonstrate discrete circuit origins for the dysfluent repetition of vocal elements in songbirds, with implications for speech disorders.

Memory circuits for vocal imitation.

Many complex behaviors exhibited by social species are first learned by imitating the behavior of other more experienced individuals. Speech and language are the most widely appreciated behaviors learned in this way. Vocal imitation in songbirds is perhaps the best studied socially transmitted behavior, and research over the past few years has begun to crack the circuit mechanisms for how songbirds learn from vocal models. Studies in zebra finches are revealing an unexpected and essential role for premotor cortical circuits in forming the behavioral-goal memories used to guide song imitation, challenging the view that song memories used for imitation are stored in auditory circuits. Here, we provide a summary of this recent progress focusing on the What, Where, and How of tutor song memory, and propose a circuit hypothesis for song learning based on these recent findings.

Tocilizumab for severe COVID-19 related illness - A community academic medical center experience.

The SARS-CoV-2 virus responsible for the COVID-19 pandemic can result in severe or fatal disease in a subset of infected patients. While the pathogenesis of severe COVID-19 disease has yet to be fully elucidated, an overexuberant and harmful immune response to the SARS-CoV-2 virus may be a pivotal aspect of critical illness in this patient population. The inflammatory cytokine, IL-6, has been found to be consistently elevated in severely ill COVID-19 patients, prompting speculation that IL-6 is an important driver of the pathologic process. The inappropriately elevated levels of inflammatory cytokines in COVID-19 patients is similar to cytokine release syndrome (CRS) observed in cell therapy patients. We sought to describe outcomes in a series of severely ill patients with COVID-19 CRS following treatment with anti-IL-6/IL-6-Receptor (anti-IL-6/IL-6-R) therapy, including tocilizumab or siltuximab. At our academic community medical center, we formed a multi-disciplinary committee for selecting severely ill COVID-19 patients for therapy with anti-IL-6 or IL-6-R agents. Key selection criteria included evidence of hyperinflammation, most notably elevated levels of C-reactive protein (CRP) and ferritin, and an increasing oxygen requirement. By the data cutoff point, we treated 31 patients with anti-IL-6/IL-6-R agents including 12 who had already been intubated. Overall, 27 (87%) patients are alive and 24 (77%) have been discharged from the hospital. Clinical responses to anti-IL-6/IL-6-R therapy were accompanied by significant decreases in temperature, oxygen requirement, CRP, IL-6, and IL-10 levels. Based on these data, we believe anti-IL-6/IL-6-R therapy can be effective in managing early CRS related to COVID-19 disease. Further study of anti-IL-6/IL-6-R therapy alone and in combination with other classes of therapeutics is warranted and trials are underway.

Telencephalic neurons monosynaptically link brainstem and forebrain premotor networks necessary for song.

Birdsong, like human speech, is a series of learned vocal gestures resulting from the coordination of vocal and respiratory brainstem networks under the control of the telencephalon. The song motor circuit includes premotor and motor cortical analogs, known as HVC (used as a proper name) and RA (the robust nucleus of the arcopallium), respectively. Previous studies showed that HVC projects to RA and that RA projection neurons (PNs) topographically innervate brainstem vocal-motor and respiratory networks. The idea that singing-related activity flows between HVC and RA in a strictly feedforward manner is a central component of all models of song production. In contrast to this prevailing view of song motor circuit organization, we show that RA sends a reciprocal projection directly to HVC. Lentiviral labeling of RA PN axons and transgene tagging of RA PN synaptic terminals reveal a direct projection from RA to HVC. Retrograde tracing from HVC demonstrates that this projection originates exclusively from neurons in dorsocaudal regions of RA. Using dual retrograde tracer injections, we further show that many of these RA(HVC) neurons also innervate the brainstem nucleus retroambigualis, which is premotor to expiratory motoneurons, thereby identifying a population of RA PNs positioned to coordinate activity at higher and lower levels of the song motor circuit. In combination, our findings identify a previously unknown pathway that may enable a subset of RA neurons to provide song-related signals to the respiratory brainstem but also transmit a copy of this information to song patterning networks in HVC.

A synaptic basis for auditory-vocal integration in the songbird.

Songbirds learn to sing by memorizing a tutor song that they then vocally mimic using auditory feedback. This developmental sequence suggests that brain areas that encode auditory memories communicate with brain areas for learned vocal control. In the songbird, the secondary auditory telencephalic region caudal mesopallium (CM) contains neurons that encode aspects of auditory experience. We investigated whether CM is an important source of auditory input to two sensorimotor structures implicated in singing, the telencephalic song nucleus interface (NIf) and HVC. We used reversible inactivation methods to show that activity in CM is necessary for much of the auditory-evoked activity that can be detected in NIf and HVC of anesthetized adult male zebra finches. Furthermore, extracellular and intracellular recordings along with spike-triggered averaging methods indicate that auditory selectivity for the bird's own song is enhanced between CM and NIf. We used lentiviral-mediated tracing methods to confirm that CM neurons directly innervate NIf. To our surprise, these tracing studies also revealed a direct projection from CM to HVC. We combined irreversible lesions of NIf with reversible inactivation of CM to establish that CM supplies a direct source of auditory drive to HVC. Finally, using chronic recording methods, we found that CM neurons are active in response to song playback and during singing, indicating their potential importance to song perception and processing of auditory feedback. These results establish the functional synaptic linkage between sites of auditory and vocal learning and may identify an important substrate for learned vocal communication.

Inception of memories that guide vocal learning in the songbird.

Animals learn many complex behaviors by emulating the behavior of more experienced individuals. This essential, yet still poorly understood, form of learning relies on the ability to encode lasting memories of observed behaviors. We identified a vocal-motor pathway in the zebra finch where memories that guide learning of song-element durations can be implanted. Activation of synapses in this pathway seeds memories that guide learning of song-element duration and can override learning from social interactions with other individuals. Genetic lesions of this circuit after memory formation, however, do not disrupt subsequent song imitation, which suggests that these memories are stored at downstream synapses. Thus, activity at these sensorimotor synapses can bypass learning from auditory and social experience and embed memories that guide learning of song timing.

Transitioning between preparatory and precisely sequenced neuronal activity in production of a skilled behavior.

Precise neural sequences are associated with the production of well-learned skilled behaviors. Yet, how neural sequences arise in the brain remains unclear. In songbirds, premotor projection neurons in the cortical song nucleus HVC are necessary for producing learned song and exhibit precise sequential activity during singing. Using cell-type specific calcium imaging we identify populations of HVC premotor neurons associated with the beginning and ending of singing-related neural sequences. We characterize neurons that bookend singing-related sequences and neuronal populations that transition from sparse preparatory activity prior to song to precise neural sequences during singing. Recordings from downstream premotor neurons or the respiratory system suggest that pre-song activity may be involved in motor preparation to sing. These findings reveal population mechanisms associated with moving from non-vocal to vocal behavioral states and suggest that precise neural sequences begin and end as part of orchestrated activity across functionally diverse populations of cortical premotor neurons.

Corrigendum to "High on Cannabis and Calcineurin Inhibitors: A Word of Warning in an Era of Legalized Marijuana".

[This corrects the article DOI: 10.1155/2016/4028492.].

Thalamostriatal and cerebellothalamic pathways in a songbird, the Bengalese finch.

The thalamostriatal system is a major network in the mammalian brain, originating principally from the intralaminar nuclei of thalamus. Its functions remain unclear, but a subset of these projections provides a pathway through which the cerebellum communicates with the basal ganglia. Both the cerebellum and basal ganglia play crucial roles in motor control. Although songbirds have yielded key insights into the neural basis of vocal learning, it is unknown whether a thalamostriatal system exists in the songbird brain. Thalamic nucleus DLM is an important part of the song system, the network of nuclei required for learning and producing song. DLM receives output from song system basal ganglia nucleus Area X and sits within dorsal thalamus, the proposed avian homolog of the mammalian intralaminar nuclei that also receives projections from the cerebellar nuclei. Using a viral vector that specifically labels presynaptic axon segments, we show in Bengalese finches that dorsal thalamus projects to Area X, the basal ganglia nucleus of the song system, and to surrounding medial striatum. To identify the sources of thalamic input to Area X, we map DLM and cerebellar-recipient dorsal thalamus (DTCbN ). Surprisingly, we find both DLM and dorsal anterior DTCbN adjacent to DLM project to Area X. In contrast, the ventral medial subregion of DTCbN projects to medial striatum outside Area X. Our results suggest the basal ganglia in the song system, like the mammalian basal ganglia, integrate feedback from the thalamic region to which they project as well as thalamic regions that receive cerebellar output.