Stephen Nowicki - US grants

This award provides funds to the Undergraduate Neurosciences Program at Duke University to establish an NSF-REU site which will support a Summer Undergraduate Research initiative. The program emphasizes integrative, systems, and behavioral neuroscience, reflecting the breadth of neuroscience research at Duke University. Research, and training experiences will be organized around the theme of "mechanisms of behavior", capitalizing on the strengths of the present academic Undergraduate program and targeting laboratories in which supportive and stimulating research environments are found. Duke University has a history of successful summer research programs, especially those that provide training experiences for women and minorities. The mentors have drawn form this experience in developing the present program, and are able to make use of existing administrative structures both for recruiting candidates to the program and for providing the logistical support necessary for an engaging and stimulating research experience.

The Undergraduate Neuroscience Program at Duke University plans a summer research program for undergraduates who will have the opportunity to become involved in neuroscience research at a major research university. Research and training experiences will be organized around the theme of "Mechanisms of Behavior". The emphasis on integrative, systems, and behavioral neuroscience that unifies faculty research interests within this interdisciplinary field at Duke serves as the common theme for the individual projects available to student participants in the program. The focus of the ten-week summer program for each student will be a research mentorship with a participating faculty member. Two weekly seminar series - "Research in the Neuroscience" and "Science Basics" - will be included in the summer schedule. "Research in the Neuroscience" will focus on ongoing faculty research so those students in the program gain exposure to the field of neuroscience research beyond their mentor's specific interests. "Science Basics" will address practical matters pertaining to science careers. The summer program will begin with a two-day familiarization retreat and will close with a research forum. Applications to participate in "Mechanisms in Behavior" will be accepted from sophomore undergraduates. The Program is especially interested in receiving applications from women and minorities as well as undergraduates at liberal arts colleges.

NONTECHNICAL SUMMARY Nowicki & Searcy, IBN 9408360, Perception, Function and Development of Complex Vocal Signals Animal communication plays an important role in many biologically critical arenas, particularly social behavior, including territorial defense and the attraction and choice of a mate. Bird song is one of the most spectacular cases on this type of behavior, and much of what has been learned about the development and evolution of animal communication has been the result of studies of bird song. In this research project, Dr. Searcy will investigate several aspects of a common characteristic of bird song, different song types. In many songbirds, the males sing variable songs which can be classified into a small number of song types. For example, male song sparrows sing five to thirteen different song types each. This pattern is found in many species but little is known about either how such variability in song develops in the individual or about how it functions in an evolutionary sense. A series of experiments conducted in both the laboratory and the field will investigate: (1) the abilities of song sparrows to discriminate variation both within and between song types; (2) the ways in which such variation contributes toward either attracting a mate or discouraging other territorial males: (3) the extent to which song sparrows differ in the amount of variability in their songs in different regions of the country; and (4) the role of learning and cultural transmission in the development of song type variability. Taken together, the results of these studies will advance the understanding of the evolution and development of animal communication.

Drs. Nowicki and Searcy propose to test whether early nutrition affects the
development of song in birds. Song development and production are controlled in
songbirds by a series of well defined brain areas, which develop to a large
extent during the first two months after hatching. In many species, the details
of song are learned during this same two month interval. Nowicki and Searcy
hypothesize that nutrition during this period affects the development of the
song control regions in the brain and consequently affects song learning. As a
test, they will manipulate level of nutrition in hand-reared sparrows during the
first three weeks post-hatching. The sparrows will be tutored with
tape-recorded songs, and the songs produced by these birds as adults will be
recorded. The stressed and unstressed groups will be compared for the quality
of learning, in terms of how faithfully they have copied the notes of the tutor
songs, and for the quantity of learning, in terms of how many songs are learned
per individual. The brains of the two groups will also be compared, for the
size of the song control regions and the density of neurons in those regions.
Finally, the songs of stressed and unstressed birds will be compared in terms of
how well they function on ecologically relevant tasks.

The idea that stress early in life affects brain development and hence learning
ability is applicable to a wide variety of organisms, from birds to humans.
Song in songbirds provides an excellent model system for studying such effects,
in that the neural control, development, and ecological function of the behavior
are all relatively well known. The study will use this system to enhance our
understanding of the effects of nutrition on the development of complex
behaviors.

Organisms communicate using a diversity of signals, but there is relatively little known of the factors that contribute to this diversity. One approach to addressing this question is to examine how the morphology involved in signal production may influence the way these signals can change over evolutionary time. Limits on morphological variation (morphological constraints) during the evolution of a communication system are rarely addressed because signal-producing morphology is often too complex to be analyzed. Spiny lobsters (Palinuridae) offer an important model system because both morphology and signal features can be easily quantified and because there is significant variation across this clade of organisms. This proposed research addresses two specific aims: (1) to construct a combined morphological and molecular phylogeny of the spiny lobsters and (2) to use comparative methods to test for correlations between sound-producing morphology, the surrounding antennal morphology and signal features. Results from these studies will be used to test hypotheses for how constraints on morphological variation influence signal diversity. Through an understanding of how constraints on morphological variation can lead to patterns of signal diversity, this research will offer novel insights into variation in the structure and function of communication systems.

The function of multiple signals in avian vocal communication
Stephen Nowicki
Martin D. Beebee

The puzzling occurrence of highly conspicuous traits in the males of many species of animals is thought to be explained by the fact that these traits act as signals to potential mates, indicating something about phenotypic or genotypic quality. This apparent resolution, however, does not easily explain cases in which individuals exhibit more than one conspicuous trait. For example, many songbirds use a combination of bright coloration and complex vocal displays in courtship, or multiple kinds of courtship vocalizations. If an individual trait can act as an indicator of male quality, what is the added function, if any, of possessing two or more such traits? The goal of this project is to study the function of multiple vocal traits in the yellow warbler (Dendroica petechia), and test a hypothesis for how different traits of an individual might function as signals. As with several groups of North American and European songbirds, yellow warblers have two distinct modes of singing: one relatively simple and one more complex. Results from previous work suggest that these singing modes are specialized for communicating different messages to males and females. This hypothesis will be evaluated using a combination of field and laboratory studies to examine three important aspects of signal function: how the singing modes are used by males, what kinds of information the singing modes convey, and how males and females respond to the different singing modes.

Collaborative research: Developmental and receiver-dependent costs of avian signals.

Stephen Nowicki & William A. Searcy

A central question in the study of animal communication is why signals provide reliable information, especially in cases where the signaler has different interests from the receiver. Theory suggests that signals may be reliable if they are in some way costly. The proposed research applies this theory to a well studied signaling system, birdsong.
Female songbirds use male song to judge the quality of prospective mates. The "nutritional stress hypothesis" proposes that the costs of song development produce a reliable link between a male's song and his quality as a mate. Because song development is costly, any male experiencing an early stress such as food shortage will produce both poor song and a poor overall phenotype, making song quality a reliable measure of overall quality. Previous work has shown that early nutritional stress has a negative impact on the development of the brain areas that control song, on the ability of males to learn songs, and on other aspects of adult phenotype such as body size. The proposed research will extend this work in three ways. First, it will test whether early nutritional stress affects a male's vocal performance, that is his ability to sing songs that are physically challenging to produce. Second, the research will test whether early stress affects the quality of the immune system of adult birds, a trait that should be of special interest to females. Third, the research will ask whether early stress also affects the development of song preferences in females.
Song also serves as an aggressive signal between competing males. In this context, signal reliability may be maintained by costs imposed by the aggressive response of receivers. Signals can reliably predict escalation if those signals also elicit the most aggressive response by receivers. Previous work shows that "soft songs" (songs produced very quietly) reliably predict aggression on the part of the signaler. The proposed research will test whether soft song also is likely to provoke an aggressive response from receivers, as theory suggests. Second, research will test whether soft song reduces the ability of third party individuals to gain information from an interaction. Third, the proposed research will investigate how patterns of signal matching contribute to signaling in aggressive interactions.
The proposed research integrates theory and data concerning animal signaling systems and mate choice with testable hypotheses about the development of brain and behavior. This area has proven to be an effective platform for recruiting and training women and minority scientists, and for disseminating the process of how science is done to the general public. An interactive website will be developed to allow students in grades 6-12 to simulate playback and learning experiments with songbirds.

DISSERTATION RESEARCH:
FITNESS COST OF NEST DEFENSE IN A PASSERINE BIRD:
A TRADE-OFF WITH OFFSPRING CARE?

PI: Stephen Nowicki & Renee A. Duckworth

In many organisms, the ability of young to grow and survive depends on the care they receive from their parents. Despite the importance of parental care for offspring survival, adults often vary widely in the quantity and quality of care they provide. One of the reasons why this variation is observed may be because individuals face a trade-off in the use of their time and resources for parental care and for other activities. A fitness cost to the expression of nest defense behavior was recently documented in western bluebirds (Sialia mexicana). Bluebirds benefit from defending their nest cavities from competitors because suitable breeding cavities are a limited resource. However, male western bluebirds that defend their nests more aggressively produce fewer offspring on average each year. The proposed research examines three alternative hypotheses that could account for this observed relationship between nest defense behavior and reproductive success. Specifically, it tests whether there is a time/energy trade-off between offspring care and defense, whether aggressive males are inherently poorer parents, or whether a correlation between elevation and nest defense best explains the relationship between nest defense and reproductive success. Identifying the mechanisms that produce variation in parents' ability to raise young successfully is essential for our understanding of the evolution of parental investment strategies.

A primary function of male bird song is to attract females as mates. However, not all songs are equally effective at achieving this goal. For example, females are more likely to mate with males that produce longer songs or more kinds of songs, presumably because a male's ability to do so corresponds to some aspect of his quality as a potential mate. This project investigates a new mechanism by which females use song to assess males as potential mates: the ability of males to produce songs that are hard to sing. When birds sing they produce pure tone sounds by opening their beaks to produce high frequency tones and by closing their beaks to produce low frequency tones making it difficult to produce trilled songs that are both broadband and rapid. "Vocal performance" refers to the ability of males to cope with this difficulty and may reflect male quality. This project proposes to test the hypothesis that females use vocal performance to choose mates because it reflects male quality, to investigate a mechanism by which females are able to discriminate such subtle features of song, and to test a key prediction of how beak movements affect vocal performance. The broader impacts of this project are to foster the professional development of women in science, disseminate science to the public and provide biological data important for conservation of an endangered habitat.

Animals use signals to identify potential mates of their own species, and also to assess the quality of potential mates. Thus, the processes by which individuals choose mates may play a role in speciation. Bird song is an important model system for studying the production, perception, development, and function of animal signals, including human speech. The proposed research aims to identify how the production, perception and function of bird song influence the way bird song changes over time. An interesting pattern of change in the trilled portion of songs over a 30-year period was demonstrated recently in five different populations of white-crowned sparrows (Zonotrichia leucophrys). This pattern suggests that trills may be changing due to selection by female choice or through correlated change with morphology or with habitat structure. The proposed research uses experiments and observations to test the relative importance of female choice, morphological change, and habitat change in explaining patterns of trill evolution. This study is the first to assess in a single species whether these multiple forces influence changes in song over time and the potential balance between selection and constraints on song in multiple populations. The intellectual merit of this project is to increase understanding of how and why animal signals change over time and more generally to increase scientific understanding of processes driving speciation.

Broader impacts: This project will support the training of one female graduate student and increase her chances of achieving her professional goals as a woman in science. Preliminary work on this project has already additionally contributed to the research education of several undergraduate women in science, including those from under represented groups. This project also will support collaboration between Duke University and the Point Reyes Bird Observatory (PRBO). Part of this collaboration will include an exchange of knowledge on long-term trends in white-crowned sparrow breeding habitat and song behavior, which can play a valuable role in understanding the factors that affect the viability of songbird populations and thus contribute to their conservation in the future. Finally, work on this project will lead to long-term preservation and availability of an important set of field audio recordings of white-crowned sparrow populations over a 30-year period and currently archived at the California Academy of Science.

A long-held assumption in animal behavior is that females and males differ fundamentally in their mating strategies, with females carefully choosing their mates, and males competing among themselves for access to females and resources. This assumption is challenged by the increasing realization that females also compete aggressively. It remains unclear, however, whether females benefit directly from their aggressiveness, or whether aggressive behavior in females is a byproduct of selection for aggressiveness in males. The proposed research aims to understand the selective pressures shaping aggressive behavior in females, using free-living tree swallows (Tachycineta bicolor) as a model. Preliminary work has demonstrated that more aggressive females benefit by gaining access to limited nest sites. Offspring of more aggressive females develop more slowly, however, and as a result, may have lower fitness. Thus, females also experience a cost associated with their aggressiveness. The proposed studies use experimental and observational approaches (1) to test mechanisms by which offspring of more aggressive females develop more slowly, and (2) to determine the extent to which the behavior of males may mitigate the cost of female aggressiveness. The focus on aggressiveness as a sexually selected trait in females provides a much needed parallel to the wealth of information already known about the selective pressures shaping sexually selected traits in males. The broader impacts of this work include public education about animal behavior and basic biology, conservation of cavity-nesting birds, and advancement of the scientific education and training of undergraduates and women in science.

Individual differences in the ability to compete for mates can drive evolution through a process known as sexual selection. While examples of sexual selection acting on behavior or morphology are well known, much less established is the understanding of how sexual selection acts at the molecular level to affect individual reproductive success.
In species where females mate with multiple males, sperm compete for access to a female?s eggs. Sperm competition can lead to selection on seminal fluid proteins (Sfps), a class of proteins transferred by a male during mating that aid in successful fertilization by that male while reducing success of subsequent matings by other males. Because of their link to sperm competitiveness, Sfps may evolve differently in species with polygamous mating systems (with high sperm competition) compared to monogamous species (with low sperm competition).
This project investigates the relationship between mating behavior and selection on Sfps in New World blackbirds (genus Agelaius), a group whose diverse behavior enables the study of how sexual selection affects DNA sequence evolution. Genes coding for Sfps will be isolated to test the hypothesis that such genes evolve faster in polygamous than in monogamous species. These genes will be archived on the NIH sequence database GenBank, providing researchers with tools for future comparative studies. Ultimately, results will clarify how selection links traits across a wide range of biological scales, from the molecular to organismal level.
Mentoring and outreach efforts are integral to this project. Undergraduate students will be introduced to the basic research process by learning fundamental lab techniques and writing original papers. Results will be shared at Caribbean field sites, where a primary challenge facing natural resources agencies is reconciling the need for preservation with those of a tourist-driven economy. Work will also be communicated broadly at the university, community and national levels.

This research will examine associations between communication signals and cognitive ability in two species of songbirds. Song in these species is learned: males memorize external models early in life, and then use these memories to shape their own production when they begin to sing many months later. Because song development involves learning and memory, it is logical to hypothesize an association between song and other aspects of cognition. To test that hypothesis, cognitive ability will be measured in a series of laboratory tasks, for example a novel motor task, in which subjects learn to remove lids from the wells of a foraging grid to obtain food, a color association task, in which subjects learn to associate food with lids of a particular color, and a spatial task, in which subjects learn that food can be found only in certain locations. These cognitive measures will then be related to aspects of song that are the outcome of learning, such as the accuracy with which song models are copied during development and the frequency of sharing of song types with others in the local population. Vocal learning in songbirds shows a series of striking parallels with speech development in humans, and therefore results on the tie between vocal development and cognition in songbirds will have implications for the relationship between cognition and speech in humans. Results from the study will be disseminated not only in scientific outlets but also in workshops for high school biology teachers designed to demonstrate how the study of animal behavior can be used to teach the scientific method.

Some of the most remarkable behaviors, including speech, musical, and athletic performance, depend on the brain's ability to precisely represent and control the timing and order of elementary movements. How the brain accomplishes this feat is largely unknown. This project will identify how the brain encodes the vocal elements and sequences used in learned vocal communication. To accomplish this goal, state of the art high-resolution optical imaging and electrical recording methods will be used to measure neural activity in animals as they engage in vocal learning and communication. This project will determine how populations of interconnected neurons that are necessary to vocal communication encode vocal elements and sequences. Accomplishing this goal is essential to understanding the neural basis of communication and also has other important potential benefits, including the ability to diagnose and ultimately repair brain pathologies that impair perception or movement, and to design machines that emulate these processes. This project will advance our understanding of neural mechanisms that enable the production and perception of complex, sequential behaviors, train professional scientists at the doctoral and postdoctoral levels, and enrich the science curriculum of high school students both locally and nationally.

The intellectual merit of this project is that it will shed light into fundamental cellular processes by which the brain encodes complex learned behaviors. This project's goal is to understand how the brain encodes syllables and syllable sequences in birdsong, a complex learned vocal behavior. The study will focus on how syllables are represented by spatiotemporal patterns of activity in populations of sensorimotor neurons important to vocal communication. Resolving this problem is an essential step to understand the neural basis of vocal communication. This project blends leading edge methods, including multiphoton calcium imaging of neuronal population activity and intracellular recordings and genetic manipulations in singing birds, to explore the synaptic, cellular, and circuit mechanisms that encode learned vocalizations. Until now, it has been impractical to interrogate how the brain encodes sensory and motor representations of such complex behaviors with cellular and synaptic resolution.