Rhanor Gillette - US grants

In order to optimize interdisciplinary approaches that capitalize upon new technical and theoretical developments, a Center for the Neurobiology of Learning and Memory is being established at the Beckman Institute (currently under construction) of the University of Illinois at Urbana-Champaign . This center will serve as 1) a Resource Center, providing advanced facilities for the study of the learning and memory process, including optical imaging (for histological studies), multi-electrode array recording (to allow functional patterns of interactions among neurons to be examined), and rapid tissue freezing (for assessment of sub-cellular dynamics); 2) a Research Center that fosters communication and collaboration among scientists pursuing common and related problems of memory and neural plasticity; 3) a Training Center which prepares graduate and postdoctoral investigators for research careers in learning and memory, and 4) a Recruiting Center that to attract outstanding young people to scientific careers. This program of scientific development and interaction is taking advantage of the unusual resources of the Beckman Institute and the University of Illinois Urbana-Champaign campus in neurobiology, interdisciplinary collaboration and cooperation, and strengths of the component disciplines of neural and behavioral sciences. Technical foci of the Center include large array neurophysiological recording facilities, with which the interactions among brain regions during learning are studied; rapid freezing facilities for examining brain slices in vitro, (with which the nature of plasticity at the level of the synapse is studied), and neuroanatomical imaging and analysis facilities (where memory processes are studied at levels ranging from the molecular to the morphological). Several types of learning are being studied, including discriminative conditioning, acquisition of motor skill, acquisition of acoustic discriminative ability, and the traditional psychological animal learning tasks such as mazes. In addition, current "models" of memory (such as long-term potentiation and kindling) are being examined. The function of the Center for the Neurobiology of Learning and Memory is to advance our knowledge of brain substrates of learning and memory from the cellular and molecular to the integrative brain system levels.

This project addresses the regulation of a cyclic AMP-dependent Na+ current, one of a class widely distributed among tissues and phyla. In olfactory epithelium the current is mediated by direct nucleotide binding, not phosphorylation. Evidence suggests that the current in molluskan neurons, like that of vertebrate olfactory epithelium, is mediated by direct binding. Despite the lack of kinase regulation, the current is richly modulated by multiple factors: both extracellular and intracellular a Ca2+, intracellular pH and a factor sensitive to calmodulin blockers. We have developed a model for regulation of the current at the channel level by interactions of Ca and cyclic AMP based on observations of the whole cell current. The character of the model and the kinetics of the current have allowed formulation of a relatively non-invasive and quantitative electrophysiological assay for adenylate cyclase and phosphodiesterase activities, and for levels of cyclic AMP in vivo. We propose 1) to test the independence of I(Na)(cAMP) from phosphorylation by single channel recording; 2) to assay the effects of intracellular and extracellular Ca2+ in regulating cyclic AMP activation of I(Na)(cAMP) and the conductance in single channels; 3) to compare the regulatory mechanisms of I(Na)(cAMP)in neurons where its voltage dependence differs; 4) to complete development of the in vivo assay and test it against radioimmunoassay; 5) to assess the role of cyclic AMP phosphodiesterase in mediating some effects of Ca 2+, pH(i), and calmodulin blockers. The likely broad distribution of the type of current under study suggests that the mechanisms regulating it are also widely distributed and participate in diverse cell functions and pathological states. The assay method under development may be also broadly applicable.

Neuronal pattern analysis (NPA) documents the dynamic brain processes of sensation, perception, learning and cognition by recording the electrical activity of brain neurons. Recent advances in multi-array recording technologies have greatly expanded the rate at which NPA data can be obtained, and these technologies have fostered means not previously available to study the intercorrelations of dynamic activities in neuronal networks. Computational modeling of brain dynamic activity has fostered major increment in the requirements of data processing due to the need to analyze simulated neuronal spike trains and to compare real and simulated neuronal data. These developments call for parallel development of adequate database systems for organization, rapid access, and sharing of NPA data. This project will establish a database system (DBS) for time series neurophysiological data recorded in experiments of members of the University of Illinois Beckman Institute Neuronal Pattern Analysis Group. System design and implementation will be carried out with consultation and guidance of the National center for Supercomputer Applications. This proposed system will foster community-wide sharing of times series and other forms of neural data, proved a model DBS that can generalize to other neuroscience groups, and enhance the research in the involved laboratories.

LAY ABSTRACT IBN-9808400 Neural Mechanisms of Nutritional Homeostasis Motivational processes, in particular hunger, are driving influences in behavioral decision-making; however, it is not known how neural networks mediating feeding or other motivated behaviors are organized by the physiological mediators of hunger. One critically involved neurochemical is the biogenic amine serotonin (5-HT), which modulates affect and arousal in vertebrates and invertebrates alike. In a simple and accessible model system, 5-HT stimulates readiness to feed acting in the feeding motor network to enhance specific aspects of neuron excitability. Evidence suggests that 5-HT stimulates production of nitric oxide (NO), a critical modulator of neural activity in many animals. This proposal outlines plans to elucidate the role and mechanisms of action of 5-HT in regulating the enzyme that makes NO, and thereby hunger/satiation state. These studies combine behavioral and electrical measures with analysis of 5-HT and NO-related chemical compounds in single cells. These studies approach the basic organization of the feedback loops regulating nutritional homeostasis in all motile animals and can lead to a fuller understanding of mechanisms of hunger/satiation. They relate directly to health issues of weight-control in anorexia and obesity. Knowledge of these factors is likely to be significant to the development of autonomous robots capable of making least- probable-error decisions in a noisy environment, and to the evolution of artificial intelligence for whose optimal function motivation-based processes can provide the critical regulation of goal related activity, just as they do in real intelligence systems.

The potent neuromodulatory roles of NO emerge from both regulation of its synthetic enzyme and its own diverse effects on neural function. Major aspects of the regulation, actions and integrative function of this critical cellular modulator continue to be uncovered. In a simple model system the regulation and modulatory role of NO production are approachable at levels that relate behavior, cell signalling pathways and ion channels. NO synthase (NOS) is found in identified and well characterized neurons of the feeding motor network, an is up-regulated by serotonin (5-HT), itself a physiological arousal factor. NO regulates excitability of the feeding neurons and network arousal state by potentiating a prominent cAMP-gated cation current (INacAMP). NOS activity varies with satiation state, acting like a homeostatic regulator of feeding behavior. This proposal has three specific and integrated aims: 1) exploring the dynamic regulation of NOS by Ca2+, calmodulin, pHi and arginine substrate in single, physiologically defined cells, 2) probing the signalling pathways through which 5-HT affects the activity and expression of NOS in single identified neurons, and 3) elucidating cellular and kinetic mechanisms by which NO regulates neuron excitability via cAMP-gated Na2 current. A major procedural strength is that these studies will take place in single identified and living neurons of well defined functional roles, for which new and potent methods for assessing NOS activity in single neurons are combined with conventional biochemical and pharmacological assays. In elucidating the neural substrates of cellular plasticity in which NO action is embedded, this proposal has fundamental relevance to understanding mechanisms of learning and memory and their disorders, hyperexcitability and depression, and motor and sensory dysfunction. It has applied relevance to understanding physiological consequences of altered 5-HT levels in integrated neural network functions that result in altered mood and affective behavior.

Successful animals tend to make efficient decisions in their daily activities in terms of effort spent, risk taken and benefit gained. How they do this at the level of the dynamical neuronal circuit has broad implications for the understanding of both normal and abnormal animal and human behavior. This project outlines a plan to describe how the nervous system integrates sensation, internal state and learning.

In preliminary work on a simple animal model system, a simple and robust neural network model has been derived for cost-benefit decision-making. Now, investigations are aimed at explaining the neural bases for switching between orienting and avoidance motor acts, how this switch is determined by sensation, appetite and learning, and how avoidance decision may suppress feeding behavior.

Studies are to be carried out with conventional electrophysiological recording methods. This project contributes to laboratory training of undergraduate and graduate students, and through its content to field classroom education in neuroethology. In these venues an effective synthetic approach, linked by content to problems in animal decision, to neuronal systems and behavior in field and lecture is important in training the next generation of systems and computational neurobiologists. The expected results will elucidate the interactions of goal-directed neuronal network circuitries to show how animals can make successful behavioral decisions that balance perception of available resources and risk against their own needs. They will provide a simple model for approaching the neural circuitry for similar value-based decision-making in more complex animals. The further potential impact of the work is to enable improved ecological and economic modeling using biologically based decision making mechanisms.

[unreadable] DESCRIPTION (provided by applicant): This project combines two fundamentally new approaches toward a model for changing the characteristic behavior of drug abuse. It exploits the decision-making neural circuitry of foraging behavior, which is subverted in addictive behavior. The experimental targets are specific neural elements that switch expression of approach and avoidance behaviors. The first approach, by locating and describing neural switches for appetence vs. avoidance in a simple system, provides descriptors to facilitate discovery of similar elements in mammalian brain. The second tests utility of focal switch control for regulating behavioral expression that could markedly alter addiction treatment strategies. Applied to human drug abusers, such methods could lessen the influence of hedonic impulse and enhance effectiveness of conventional treatment. We will utilize the predatory gastropod Pleurobranchaea californica, a simple model system for which neural circuitry of decision is well studied by microelectrode and computational simulation. Our initial specific aim is to characterize the switch neurons that toggle the motor network for directional turning between orienting and avoidance, under the control of appetitive state. We will elucidate effects of altering appetitive state on the switch neurons and consequent motor output of the turns stimulated by appetent and noxious stimuli. Our second specific aim is to achieve an exemplar paradigm in which appetitive stimuli induce involuntary avoidance behavior. We will elucidate biasing of the switch via external intervention. Regulation of the switch will be probed by direct electrical control at the elements themselves via intracellular current injection, by directly modifying the state of the feeding network, and by focal application of neuromodulators to the switch elements. Towards developing and testing a model of neural intervention in drug abuse, we will assess effects of gel beads impregnated with neuromodulators to focally affect switch neuron activity and promote bias in behavioral choice. This project seeks to develop a model for treating addictive behavior through characterizing and regulating neural elements that switch behavior between approach and avoidance. [unreadable] [unreadable] [unreadable]

This research will illuminate one of the most challenging aspects of the evolution of neuronal circuits: genomic mechanisms underlying cell-specific adaptive modifications and the origin of novel behaviors. The evolutionary approach is less developed in modern neuroscience. However, it is crucial to understand how complex networks and brains are formed or to answer "why" questions (e.g. why different subsets of signal molecules were selected in distinct neuronal circuits). The evolution of centralized complex brains occurs in parallel, where distinct neural patterning might emerge independently in different lineages but use similar molecular building blocks or toolkits.

This project proposes to identify and characterize cellular homologs within defined neural circuitries across opisthobranch species (e.g. Pleurobranchaea and Tritonia) to understand how changes in the genomic organization of homologous neurons lead to adaptive modifications of networks underlying escape and other behaviors. As a result, it will lead to conceptually new approaches when nervous system evolution can be portrayed on an entire genomic scale with single-cell resolution. The hypothesis about whether divergent evolution of neural circuits resulted in the appearance of novel signaling systems and other neuron-specific markers will be tested. Alternatively, novel network properties and connections might emerge as modular rearrangements of preexisting molecular components.

Training opportunities for interdisciplinary students will arise during the development of a nation-wide comparative genomic database that will be searchable for neuronal markers and signal transduction pathways. The extensive collection of transcripts will also allow testing evolutionary relationships across neurobiological models with different levels of centralization of their nervous systems. The proposed approaches and methodologies can be generalized to any system and thus will dramatically increase both the information and education opportunities that can be gained from studying classical electrophysiological preparations. The research will provide a long desired marriage of neuroscience and comparative genomics to understand of how specific neuronal networks are organized and evolved.

The research objective of this award is to understand how neurons develop mechanical force (tension), and how this tension influences their functionality. It is known that memory, learning and locomotion in animals are mediated by neurotransmitters that are released from vesicles clustered at the synapse (junction between two neurons, or between a neuron and muscle). Recent experiments on the embryonic Drosophila (fruit fly) nervous system show that vesicle clustering at the neuromuscular synapse requires mechanical tension within the axons. Vesicle clustering vanishes upon severing the axon from the cell body, but is restored simply by applying mechanical tension on the severed axon. Using micro mechanical force sensors, it has been shown that embryonic axons that form neuromuscular junctions maintain a rest tension of about 1 nN. These results suggest that neuromuscular synapses employ mechanical tension as a signal to modulate vesicle accumulation. In this project, the mechanism of force generation in axons, as well as the mechanism by which tension modulates vesicle clustering, transport and release will be explored using a combination of nano scale force sensors, advanced imaging and mechanics models.

Understanding the force paradigm of synaptic function may lead to new treatments for neurological diseases. The findings of this research will be disseminated to the broader audience through the web and journal publications. The instruments to be developed will be made available to others for mechanobiological studies. The knowledge gained through this project will be integrated with education through development of a new course on neuro-mechanics that incorporates neuroscience and engineering, as well as through student seminars, involvement of undergraduate students from minorities in research, and hands-on teaching modules at the local Children's Science Museum.