David Perkel - US grants

This project will investigate the cellular basis of vocal learning in songbirds. Behavioral studies have established that male songbirds learn their songs from their fathers, and investigations using anatomical and electrophysiological approaches have elucidated the brain circuitry necessary for vocal learning. Work in a variety of systems has pointed toward a critical role for synaptic plasticity in learning. It is now possible to make and test specific cellular hypotheses concerning how changes in synaptic strength could give rise to vocal learning in songbirds, but little is known of the cellular electrophysiological properties of the neurons in the songbird brain. The proposed experiments will investigate neurons in nucleus RA of the songbird brain, an important site of convergence of auditory and motor information which has been implicated in song learning. Voltage-clamp recordings will be obtained from neurons in in vitro brain slices to test the hypothesis that activity-driven synaptic plasticity, resembling long- term potentiation (LTP) or depression (LTD) as studied in other preparations, could contribute to song learning. Previous findings indicate that the auditory and motor pathways projecting to RA neurons use different combinations of postsynaptic neurotransmitter receptors. The existence of synaptic plasticity in one or both of these pathways will be tested by applying, individually and in combination, various types of stimuli that are effective in other systems exhibiting plasticity. If a form of synaptic plasticity is found, it will be necessary to test whether it might play a role in behavioral plasticity. Although it is difficult to design experiments to test this hypothesis directly, a number of indirect tests are possible. The developmental profile of synaptic plasticity will be investigated to determine whether changes in synaptic strength become less pronounced or more difficult to induce at times when the bird's song is not plastic. Secondly, by shifting the critical period for song learning using hormonal manipulations, it should be possible to shift the critical periods for cellular changes. These studies will be among the first to examine, in a vertebrate system, the link between changes in synaptic strength and learning of a complex behavior.

DESCRIPTION (Adapted from applicant's abstract): Vocal learning in songbirds is a unique, experimentally accessible model of human vocal learning that exemplifies the general process of motor learning. A male songbird learns his song by first memorizing his father's song, and later using auditory feedback to match his own song to the memory of his father's song. When a good match is achieved, the song becomes highly stereotyped and less dependent on auditory feedback. Forebrain nucleus RA is implicated as a site of plasticity for song learning because: (i) it receives convergent motor input from nucleus HVc and auditory input from nucleus L-MAN; (ii) the HVcRA connection is first formed at the onset of song practicing; (iii) and input for L-MAN, which is that nucleus' sole output to the motor system, is crucial for learning, but not for adult song production. Numerous collaterals within RA also contribute the majority of excitatory connections to RA neurons. My working hypothesis is that vocalization driven auditory activity in L-MAN guides selection of motor connections in RA through synaptic enhancement or depression and that this plasticity disappears or becomes substantially reduced after song crystallization. This proposal will (1) measure developmental changes in basic properties of synaptic connections in the L-MAN RA, HVc RA and RA RA pathways using whole cell patch clamp recording in brain slices; (2) determine activity patterns in the three pathways that elicit changes in synaptic strength; and (3) compare plasticity in slices prepared from juveniles and from adults. These studies will be the first to examine a role for changes in synaptic strength in learning of a complex vocal behavior. Progress in illuminating mechanisms underlying plasticity in this system will further our long-term goal of understanding more general mechanisms of learning in the central nervous system.

LAY ABSTRACT
Proposal #: IBN-9817889
PI Name: David Perkel

Humans, cetaceans (whales and dolphins) and some birds are the only animals known to learn their vocalizations. Songbirds thus provide a uniquely accessible model system for studying how the brain learns vocal behaviors. The brain structures involved in avian song production and learning are distinct and well defined, but their functional connections are not well understood. The present proposal will examine an avian brain circuit, the anterior forebrain pathway, which (1) is necessary for vocal learning but not song production and (2) resembles a circuit known in
mammals to be involved in a variety of motor and cognitive functions.

These experiments will use immunological and electrophysiological techniques to determine whether the AFP, as in its mammalian counterpart, has two sequential stages of inhibitory projections, i.e. whether the same functional wiring occurs in the songbird. Two primary benefits are: (1) clarification of how the circuit works in songbirds to mediate vocal learning and (2) testing for evolutionarily conserved circuitry, which would suggest more general rules for motor
learning. Such similarities would provide essential guidance for further experiments aimed at understanding the detailed cellular mechanisms of vocal learning and of motor learning in general.

DESCRIPTION (Adapted from applicant's abstract): Vocal learning in songbirds is a unique, experimentally accessible model of human vocal learning that exemplifies the general process of motor learning. A male songbird learns his song by first memorizing his father's song, and later using auditory feedback to match his own song to the memory of his father's song. When a good match is achieved, the song becomes highly stereotyped and less dependent on auditory feedback. Forebrain nucleus RA is implicated as a site of plasticity for song learning because: (i) it receives convergent motor input from nucleus HVc and auditory input from nucleus L-MAN; (ii) the HVcRA connection is first formed at the onset of song practicing; (iii) and input for L-MAN, which is that nucleus' sole output to the motor system, is crucial for learning, but not for adult song production. Numerous collaterals within RA also contribute the majority of excitatory connections to RA neurons. My working hypothesis is that vocalization driven auditory activity in L-MAN guides selection of motor connections in RA through synaptic enhancement or depression and that this plasticity disappears or becomes substantially reduced after song crystallization. This proposal will (1) measure developmental changes in basic properties of synaptic connections in the L-MAN RA, HVc RA and RA RA pathways using whole cell patch clamp recording in brain slices; (2) determine activity patterns in the three pathways that elicit changes in synaptic strength; and (3) compare plasticity in slices prepared from juveniles and from adults. These studies will be the first to examine a role for changes in synaptic strength in learning of a complex vocal behavior. Progress in illuminating mechanisms underlying plasticity in this system will further our long-term goal of understanding more general mechanisms of learning in the central nervous system.

This project addresses a fundamental question of brain evolution: how well conserved is the main brain circuit diagram across the vertebrates? Specifically, this project aims to resolve contradicting predictions arising from two recent lines of research into a structure called the basal ganglia, which is important in initiating and learning complex behaviors. One set of work, primarily on traditionally studied bird species such as pigeons and chickens, has shown close similarities between birds and mammals. Another line of work, using the specific brain structures of songbirds, which learn their vocalizations, has found some key differences in how the basal ganglia circuits are organized. Work proposed here will determine how portions of the chicken basal ganglia are wired, and how individual nerve cells in those structures work. Emphasis will be on how similar or different the structure and function of these brain regions are between birds and mammals. Data gathered through this sort of comparative research has two major beneficial effects: they shed light on how brains have evolved; and they provide information on how different species have solved problems in similar or different ways.

[unreadable] DESCRIPTION (provided by applicant): Vocal learning in songbirds is a unique, experimentally accessible model of human vocal learning that also exemplifies the general process of motor learning using sensory feedback. A male songbird learns his courtship song by first memorizing his father's song, and later using auditory feedback to match his own song to his memory of his father's song. One major advantage to this model system is the existence of separate forebrain circuits involved in producing the song and in learning it. The so-called "anterior forebrain pathway", which is essential for vocal learning, has recently been found to bear gross similarities to the mammalian basal ganglia pathway, which is known to be involved in motor control and motor learning. Specifically, the general circuit connectivity, neurotransmitters used, and neuron classes present, are consistent with the hypothesis that the same basic circuitry underlies information processing in avian and mammalian basal ganglia. The experiments proposed here will use electrophysiological and morphological approaches to test whether key microcircuitry of the anterior forebrain pathway supports this view. In addition, the experiments will test whether neuromodulatory actions of dopamine in the anterior forebrain pathway are similar to those in mammals. These experiments will provide necessary information about how the avian anterior forebrain pathway works. If the hypotheses are supported, two major benefits will ensue. First, work on the avian song system will accelerate because it can be guided by the wealth of information already available for mammals. Moreover, the song system, with its advantages of a well-studied, naturally-learned behavior, will be validated as a more directly applicable model system of mammalian basal ganglia function than heretofore realized. Such complementary approaches to understanding the basal ganglia are valuable because approaches to disorders of the basal ganglia such as Parkinson's and Huntington's diseases will likely require fundamental understanding of information processing in the basal ganglia. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Birdsong is an excellent model for understanding the neural basis of complex vocal behavior, like human speech. Songbirds naturally learn and produce their song, and like human speech, the learning process depends on hearing. Our long-term goal is to understand the neural mechanisms of song production. As a beginning, this proposal considers the role played by the telencephalic nucleus HVc in zebra finches, which produce one song with high stereotypy throughout life. HVc is likely to participate in driving song production: it is required for singing, its neurons display premotor and patterned activity during song production, and it is poised anatomically to influence the output of the syrinx, the avian vocal organ. Furthermore, studies of HVc in vivo have implicated it as a pattern generator for song. The experiments described here propose to develop a slice preparation of HVc that can be induced to produce activity in vitro that mimics the rhythmic activity observed during song production in vivo. Recent indirect evidence shows that patterned activity can be elicited in brain slice preparations of HVc; the goal here is to directly observe the spiking activity, and to induce it reliably. Specifically, extracellular recordings will be used to monitor the spiking patterns of HVc cells in vitro in response to brief high frequency stimuli, application of neuromodulators, or both. If patterned activity results, we will analyze it for stereotypy and similarity to the song of the bird from which the brain slice was made. The proposal is highly exploratory because it aims to discover the conditions under which rhythmic, singing-related activity can be induced in HVc in vitro. This approach will at least clarify the extent to which HVc is capable of generating rhythmic activity in isolation. If these experiments yield reliable rhythmic activity related to song, then this will be a strong candidate for a preparation of "fictive singing." Such a preparation would provide one of the few vertebrate examples of pattern generation in vitro, claim the first instance of in vitro pattern generation for vocal behavior within the telencephalon, and it would form a basis for future mechanistic investigations of rhythmic activity in HVc, song production, and more generally the motor control of sequenced and patterned motor outputs. This knowledge would be applicable to understanding the mechanisms governing speech production in humans, and enhance the understanding of speech, how it is learned, and its pathologies.

The overall goal of the Computer Resources Core is to establish and maintain computer support for a core group of scientists. This support will increase the productivity of our research through better use of computers. Users and support personnel (Computer Specialists) will work together to generate and implement ideas to use software or hardware to enable and facilitate research. The goal is to provide computer expertise within a broad enough scientific context that any one solution would have potential for more than one application. The specific aims are: 1) Communicate and establish common goals. The first aim is to identify problems and solutions that would benefit the maximum number of investigators in the group. Through regular meetings and electronic communications, we will discuss and describe the scientific issues and this will help us focus on potential solutions to improving experimental data collection or analysis using information technology. This discourse will help us identify computational methods that will increase our productivity. 2) Software and hardware solutions. Once a problem or approach is identified, the task will be delegated to the Computer Specialists. The Computer Specialists will examine the possible solutions among the choices of commercial or custom software/hardware. The Computer Specialists will interact with members of the Core to choose the best solution. 3) Software and hardware evaluation and development. In consultation with the members of the Core, the Computer Specialists will select a software package or develop custom software to fit the goals of the experiment or analysis desired. The Computer Specialists will be responsible for testing and debugging software applications and training and assisting core users to apply the software in their research. Likewise, hardware will be implemented in a similar manner. They will also help to disseminate information about solutions to particular problems to other Core investigators who might be able to incorporate such solutions into their own research.

The Computer Resources Core (Core B) provides a spectrum of services to support and enhance the research endeavors of a core group of scientists investigating questions related to hearing, communication and balance at the University of Washington. The overall goal is to increase the productivity of our research through better use of computers. Specifically, we aim to: enable completely new types of research; accelerate development and spread of technologies new to the University of Washington; facilitate collaboration or interaction among users of the Core; help reduce redundant work by providing consolidated computer expertise accessible by all users of the core; maintain a high level of basic computer support for all supported research groups. We endeavor to provide computer expertise within a broad enough scientific context that any one solution would have potential for more than one application. In consultation with the members of the Core, the Computer Specialists supported by the Core will select a software package or develop custom software to fit the goals of the experiment or analysis desired. The Computer Specialists are responsible for testing and debugging software applications and training and assisting core users to apply the software in their research. Likewise, hardware solutions will be implemented in a similar manner. The Computer Specialists will also help to disseminate information about solutions to particular problems to other Core investigators who might be able to incorporate such solutions into their.own research.

DESCRIPTION (provided by applicant): The University of Washington (UW) has a long-standing history of commitment to research in the areas of normal and disordered hearing, speech, language and communication. In addition, it has an outstanding interdisciplinary community of investigators in all of the subdisciplines of basic neuroscience. At the intersection of these communities lies a diverse and highly productive group of investigators who study the fundamental neural mechanisms that underlie hearing and communication. One important mission of the auditory neuroscience community at UW is to mentor the trainees who will carry on this line of research and advance our knowledge of the field in the future. The Auditory Neuroscience Training Program, established in 2002, helps train the basic neuroscience researchers whose work will form the foundation for research in the clinical disciplines. It therefore complements the UW clinical training programs in Otolaryngology and in Speech and Hearing Sciences. The training experience at UW currently includes six predoctoral training slots since this is the area in which strong support during the early stages of training is most crucial, and three postdoctoral training positions for those transitioning to auditory neuroscience from other disciplines. Trainees participate in active research programs in neuroanatomy, development, genetics, cell and molecular biology, neuropharmacology, and electrophysiology of the peripheral and central auditory system as well as psychoacoustics, language perception and processing, and communication behavior. They also have the opportunity to combine research in more than one area through collaborative efforts. Through courses, journal clubs and retreats, program trainees are exposed to a wide range of research techniques, enabling them to conduct conceptually and technologically sophisticated research programs. Importantly, continued support through the training program should greatly enhance the ability of the UW to attract and retain high-caliber trainees from underrepresented groups and prepare them for future research careers in auditory neuroscience.

Project Summary Understanding speech depends on the capacity of the auditory system to accurately represent salient sounds. This representation may be altered by a variety of factors, including disorders involving neuromodulatory systems. For example, patients with Parkinson's disease have speech processing problems suggesting that dopamine alters normal representation of these salient signals. The proposed studies focus on the role of dopamine in altering representation of salient sounds in the inferior colliculus (IC). The IC is a prime location for modulating auditory processing of salient signals because it receives input from multiple auditory and non-auditory source, it contains dopamine receptors and fibers, and preliminary data from this proposal indicate that dopamine modulates IC auditory responses. The objective of this proposal is to determine the mechanisms by which dopamine alters the representation of vocalizations in IC. The first Aim will use in vivo single unit recordings with application of pharmacological agents in the IC of awake mice to determine the effects of dopamine receptor activation on responses to vocalizations. The second Aim will use whole-cell recordings in mouse IC brain slices to determine the effects of dopamine on intrinsic and synaptic properties of different neuron types. The third Aim will use in vivo whole-cell recordings to identify how intrinsic properties of different neuron types shape selectivity to vocalizations. Aims 1-3 will thus provide an integrated understanding of the cellular and synaptic mechanisms underlying auditory responses to complex sounds. The fourth Aim will determine the sources of dopaminergic input to the IC, an important step towards understanding the behavioral contexts that elicit dopamine release into the IC. The significance of this proposal is that it is the first integrated study of the effects of dopamine on the cellular, synaptic and circuit properties underlying IC responses to salient sounds. The results will increase our mechanistic understanding of auditory processing of meaningful sounds and how this encoding changes with different social contexts, physiological states and communication disorders. These studies using mice with normal hearing will facilitate future studies of genetically engineered mice to further probe the mechanisms underlying specific communication and neurological disorders.

Hummingbirds, songbirds and parrots learn their vocalizations from adults, just like human infants learn to speak by imitating their parents' speech. This capacity provides the basis for how humans acquire speech and language, yet how the brain achieves this goal is unknown. Using an array of powerful techniques, this collaborative project will examine the anatomical, electrical, and molecular properties of brain circuits that control vocalizations in hummingbirds and songbirds, comparing them with each other and with prior human studies. Understanding how these different organisms evolved brain circuits to accomplish similar goals will reveal insights into fundamental properties of vocal learning systems. Traditional lab animals cannot be used as they lack vocal learning; hence, the use of vocal learner birds is critical. The project will provide training in multiple research techniques to high school, undergraduate, and graduate students, emphasizing underrepresented groups. It will also promote broad dissemination of findings and outreach activities, related to both scientific and conservation efforts. The project is also integrated with an International Consortium (funded by the Brazilian Government) for cataloguing and characterizing the diversity of tropical birds, including integration of Museum collections, generation of genome sequences, and examination of brain specimens relevant to the evolution of vocal learning. These activities will enable interactions between US and Brazilian faculty and students, while promoting training in molecular and histological methods through site visits, field trips, and workshops.

The vocal control system of songbirds is critical for song production and learning, and is well characterized anatomically, electrophysiologically, and molecularly. However, knowledge of the analogous areas in other avian vocal learners is limited. Recent phylogenomics efforts reveal that hummingbirds evolved vocal learning independently of songbirds; thus, comparing their vocal control systems will reveal convergently evolved features that may be fundamentally required for this trait. The investigators will use tract-tracing to determine how vocal control areas are connected in hummingbirds, in vitro electrophysiological recordings to determine intrinsic neuronal properties of vocal areas in hummingbirds and songbirds, and in situ hybridization to identify molecular markers of vocal nuclei. Evidence of shared anatomical, physiological and molecular specializations will point to convergent features representing possible universal properties of vocal learning systems that may also be shared with humans. Alternatively, differences would suggest that multiple circuit and cellular/molecular architectures can subserve vocal learning. Outcomes will provide novel clues as to evolutionary origins and constraints of vocal learning and associated pathways, leading to insights into fundamental requirements of vocal learning.

Project Summary The University of Washington (UW) has a long-standing history of commitment to research in the areas of normal and disordered hearing, balance, speech, language and communication. In addition, it has an outstanding interdisciplinary community of investigators in all of the subdisciplines of basic neuroscience. At the intersection of these communities lies a diverse and highly productive group of investigators who study the fundamental neural mechanisms that underlie hearing and communication. One important mission of the auditory neuroscience community at UW is to mentor the trainees who will carry on this line of research and advance our knowledge of the field in the future. The Auditory Neuroscience Training Program, established in 2002, helps train the basic neuroscience researchers whose work will form the foundation for research in the clinical disciplines. It therefore complements the existing UW clinical training program in Otolaryngology and one proposed in Speech and Hearing Sciences. The training experience at UW currently includes five predoctoral training slots since this is the area in which strong support during the early stages of training is most crucial, and three postdoctoral training positions for those transitioning to auditory neuroscience from other disciplines. Trainees participate in active research programs in neuroanatomy, development, genetics, cell and molecular biology, neuropharmacology, and electrophysiology of the peripheral and central auditory system as well as psychoacoustics, language perception and processing, and communication behavior. They also have the opportunity to combine research in more than one area through collaborative efforts. Through courses, journal clubs and retreats, program trainees are exposed to a wide range of research techniques, enabling them to conduct conceptually and technologically sophisticated research programs. Importantly, continued support through the training program should greatly enhance the ability of the UW to attract and retain high-caliber trainees from underrepresented groups and prepare them for future research careers in auditory neuroscience.

Project Summary: The neural circuit that regulates birdsong, a highly precise, learned sensorimotor behavior, excels for study of fundamental mechanisms of adult circuit plasticity. The song system is a unique model of naturally occurring degeneration and compensatory regeneration in a behaviorally relevant neural circuit in adult brains. This circuit shows exaggerated seasonal degeneration and reconstruction via neurogenesis, in response to changes in circulating steroid hormone levels. Our long-term goal is to understand the fundamental mechanisms by which steroid hormones and neurotrophins interact to regulate plasticity of neural circuits and behavior. On a translational level, our goal is to understand how forebrain circuits can regenerate to support performance of complex learned motor skills. The central hypothesis of the proposed aims is that seasonal changes in hormones trigger changes in anterograde and retrograde trophic signaling that lead to remodeling of the HVC-RA circuit and changes in song behavior in adult birds.The goal of this application is to identify the trophic signaling pathways (molecular and electrophysiological) that regulate the the incorporation of newborn neurons to regenerate this circuit. This research will advance the field by elucidating fundamental issues of adult circuit plasticity. This topic is of translational relevance for exploiting endogenous or exogenous stem cells for therapeutic repair of injured or dysfunctional circuits in humans. These fundamental issues include whether new neurons added to adult circuits establish functional connections with efferent nuclei and restore behavior (Aim 1), the role of activity regulated genes in mediating retrograde trophic effects of neuronal activity on presynaptic adult neurogenesis (Aim 2), the role of calcium channels in mediating the transsynaptic neurotrophic regulation of postsynaptic activity (Aim 3), and the role of pre- and/or postsynaptic neuronal activity in maintaining a regenerated adult circuit (Aim 4).