Michael P. Stryker - US grants

Description (provided by applicant): This proposal seeks to discover the intercellular signaling and specific neural circuit substrates of the loss and recovery of binocular responses in the visual cortex during the critical period of susceptibility to the effects of monocular visual deprivation. It also seeks to understand the differences between the high degree neural plasticity at the height of the critical period and reduced plasticity in the adult visual cortex. The methods to be used are efficient means of measuring visual responses and neural circuits longitudinally in individual mice, including transcranial intrinsic signal optical imaging, extracellular microelectrode recording, 2-photon laser scanning microscopy for longitudinal anatomical studies in vivo, and 2-photon laser scanning of multiple neurons bulk-filled with calcium indicators. The time course of visual cortical plasticity in response to monocular visual deprivation and recovery at the peak of the critical period consists of distinct temporal phases. Our preliminary results show these phases to be distinct in the molecular signals that operate in each phase, so that mice mutant in specific signaling pathways are deficient in only one of the three phases. We will determine in which cells of the upper cortical layers these mechanisms influence plasticity, and how these signaling pathways and neural plasticity in response to them change in adulthood. PUBLIC HEALTH RELEVANCE The long term goal of the proposed research is to understand the cellular mechanisms and changes in the circuitry of the visual cortex that are responsible for the loss of influence of the deprived eye in experimental models of amblyopia, and to determine what is the potential for recovery. Much of the basic science in this field has concentrated on animal models of deprivation amblyopia, of which there are many, in the hope that an understanding of the neural signaling and circuit bases of plasticity would enlighten further attempts at prevention and treatment in human patients. This proposal, by identifying mechanisms that operate in different phases of plasticity and by identifying the origins of the difference between the repair potential of juvenile and visual adult cortex, should be valuable in guiding future attempts at therapy for visual cortical abnormalities.

Neural plasticity refers to the ability of central nervous system connections to be modified or rearranged, particularly during a critical period early in development. The overall goal of Dr. Stryker's research over the past several years has been to understand the role of neural activity in mammalian visual system plasticity and development. Much recent evidence from studies of long-term potentiation (LTP) in the hippocampus suggests that the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor plays a crucial role in this neural activity-dependent synaptic plasticity. LTP plasticity is found in both young and adult nervous systems. In this project, Dr. Stryker and his colleagues will conduct several studies to elucidate the role of NMDA receptors in mediating the effects of activity on the plasticity of the cat visual cortex. Plasticity in the visual cortex is only found during an early stage of development. The investigators will infuse NMDA-antagonists into the visual cortex; measure electrophysiologically the specificity of the receptor blockade so produced; record the effects of NMDA- receptor blockade on patterns of neural activity; and study physiologically and anatomically effects on plasticity in development. Dr. Stryker has a long history of making significant contributions to our knowledge of nervous system development. This new work promises to reveal whether the cellular and molecular details of plasticity in the adult central nervous system (LTP), now receiving enormous research attention, apply also to the rearrangement of neuronal connections in brain development.

This application proposes neurophysiological studies to elucidate the mechanisms by which patterns of neural activity may guide or affect the development and plasticity of the mammalian visual cortex. Our previous research has shown that spontaneous neural activity in the visual system is require for the development of ocular dominance columns in the visual cortex, an event that begins in utero in higher primates. Our more recent experiments have demonstrated a similar requirement for neural activity for the development of eye-specific laminae in the lateral geniculate nucleus; this process is essentially complete before the time of birth even in our feline model. The findings suggest that abnormalities in the spontaneous patterns of neural activity in utero may be a hitherto unsuspected cause of birth defects. In addition, and understanding in detail of the mechanisms of plasticity in the developing visual system should provide a rational basis to therapy for ambloyopia, a clinical disorder affecting as many as 2% of all children. The specific aim of this proposal is to explore in detail the synaptic plasticity in the developing visual cortex produced by pharmacologically inhibiting the postsynaptic cortical cells. Our recent work has demonstrated that infusion of the GABA-A agonist muscimol into kitten visual cortex causes plasticity in favor of the less-active input when one eye is deprived of vision. The implications of this unprecedented phenomenon will be explored by determining the necessary does of a variety of inhibitory Agents, be delineating the period of susceptibility to the effects of these agents on cortical plasticity, by making quantitative measurements of visual responses following such plasticity, by conducting longer-term physiological experiments and anatomical studies of transneuronally labelled ocular dominance columns, and by pharmacological studies of effects on various receptor types.

This award provides funds to aid in the purchase of optical microscopy equipment. The equipment will be located in a central facility where it will be available to a group of neuroscientists in the Keck Center for Integrative Neuroscience. The scientists are all interested in various aspects of local circuit analysis of the mammalian central nervous system (CNS). The goal of their studies is an improved understanding of how small groups of neurons generate behavior. The instruments requested will be used for combined anatomical and physiological studies of brain function that include the use of double or triple labeling techniques. The development of new instrumentation for optical microscopy and of new methods that permit specific labeling of certain cells or certain cell constituents has been key to progress in our understanding of many aspects of cellular and developmental biology. Neuroscience has benefited greatly from these developments, since they permit the rapid and reliable identification of particular neurons or groups of neurons in the brain or at other sites that typically contain many nerve cells or similar morphology.

The orientation column, and orientation-selective cortical neurons, have long been appreciated as the most distinctive emergent features of the visual cortex. Experiments of the past few years have disclosed two anatomical phenomena related to orientation columns: (1) the discovery in our laboratory of the alignment of the collection of receptive fields of geniculocortical afferents that arborize within an orientation column, and (2) the discoveries from Singer's and Katz's laboratories of the development of the patchy intrinsic corticocortical connections that appear from the findings of Gilbert and Wiesel to connect columns of similar orientation preference in adult animals. These two phenomena mirror two contrasting views of the mechanism that produces orientation- selective responses in adult animals: the Hubel and Wiesel (1962) model of simple cells, and the specification or refinement of orientation selectivity by local corticocortical connections. These two aspects of afferent and corticocortical organization might be either causes or consequences of the development of orientation selectivity and orientation columns. We propose to find out which if either of these phenomena is responsible for the development of orientation selectivity and the formation of orientation columns by first determining the times in normal development when each of these kinds of organization emerge in relation to the orientation-selectivity of single neurons. We will then interfere with the development of orientation selectivity by changing afferent and cortical activity by a variety of means and will assess effects on the development of afferent receptive- field alignment and the formation of patchy corticocortical connections. The results of these experiments will be essential for a comprehensive understanding of the mechanisms that organize the visual cortex in development. This understanding should yield insight into the mechanisms responsible for amblyopia, a visual disorder that affects up to 2% of children, as well as possible approaches to therapy.

This project proposes to develop further a transgenic mouse model of neocortical plasticity in order to investigate the roles of neurotrophins and their receptors, protein kinase A and downstream pathways and serotonin systems in the form of activity-dependent neural plasticity that refines thalamocortical and connections during normal development. In preliminary work, we have already established that many aspects of the physiological plasticity of the developing mouse visual cortex are similar to those in the cat, monkey, ferret, and human that we and others have studied earlier. In all these species, similar competitive interactions between inputs from the two eyes regulate, in an activity-dependent fashion, the relative strength of those inputs (referred to as ocular dominance), and this plasticity is observed only during a well-defined critical period in early life. In the mouse, as in the other species, we have delineated the critical period and have now shown that a brief period of monocular visual occlusion during the critical period results in the loss of responsiveness to the deprived eye and a complementary increase in cortical responses to the open eye; while a similar period of bilateral occlusion has no effect. The present proposal is to build upon this preliminary work to determine whether deletions of genes needed for models of adult plasticity in vitro also alter this form of developmental plasticity thought to be responsible for organizing neuronal connections to and within the developing visual cortex.

The visual system of the mouse offers the opportunity for the use of genetic approaches to an understanding of central nervous system development, function, and pathology. The mouse visual system has many features in common with that of the human and other higher mammals. The present proposal in response to RFA MH-99-006 is for the development, verification, and dissemination of procedures for a quantitative, rapid and reliable assessment of visual function in the mouse that would be sensitive to abnormalities from the level of the optic media and retina up to and including the level of the primary visual cortex. Swept visual evoked potentials recorded over the visual cortex will be used to define the visibility of grating stimuli of a range of contrasts and spatial frequencies. Signal-to-noise ratios in normal mice are sufficiently large to allow reliable measurement of vision through one eye in 240 sec of data collection, consistent with a goal of screening vision through both eyes at a rate of 4 mice/hour. This proposal represents a collaboration between a laboratory that has over the past several years delineated mechanisms of activity-dependent plasticity in the mouse primary visual cortex, along with genetic disturbances in those mechanisms, and one that perfected the swept VEP and distributed it widely for uses including the screening of visual development in human infants. These assays should permit the investigation of the genetic bases of visual system development and function at levels from the eye to the cortex.

The long-term objective of our research is to understand the computations performed by cerebral cortical circuit. Toward this end, we focus on understanding the circuitry of cat primary visual cortex (VI) and the manner in which it produces the functional response properties of cat VI neurons. We focus on can VI because it is by far the best-studied piece of cerebral cortex, and because it is part of the process of visual perception which is the sensory modality that is best understood at the cortical level. In particular, we aim to understand the functional connectivity of cat V1, that is, which neurons are connected to one another by excitatory or inhibitory synapses, as a function of the visual response properties, cortical layers of origin, and excitatory or inhibitory nature of the neurons. This will be accomplished by simultaneously recording from multiple nearby neurons using the tetrode method of recording, which allows the simultaneous isolation of multiple neurons at single recording sites. We will use cross-correlation analysis to infer which of the simultaneously recorded neurons make monosynaptic connections to one another and the sign of the connection when it exists. Evidence of an inhibitory connection simultaneously provides evidence of the excitatory of inhibitory nature, respectively, of the presynaptic neuron. We will functionally characterize the recorded neurons using a combination of traditional grating stimuli and noise stimuli. We will determine the cortical layers in which recorded neurons are located through histological analysis after the conclusion of the experiment. By studying large numbers of pairs, and determining who is connected to whom, vs. cell layers functional response properties, and excitatory or inhibitory nature of the connection, we will build up a statistical picture of the functional connectivity of cat V1. This information, in interaction with modeling of the cortical circuit, provides the basis for understanding how the function of cat V1 is created from its circuit structure. Understanding of cortical function in turn provides the basis for understanding visual disorders such as amblyopia and strabismus and neurological disorders due to stroke, and more generally for understanding normal function and its disorders such as learning disabilities.

To identify genes required specifically for neocortical plasticity, we will focus on the activity-dependent[unreadable] plasticity of responses to the two eyes in the primary visual cortex during the critical period. We will screen[unreadable] memory mutants selected as likely to be defective in cortical plasticity in Project 1 for deficits in this form of[unreadable] visual cortical plasticity. We will use monocular visual deprivation during the critical period to induce[unreadable] plasticity, and test the mice with microelectrode recordings and intrinsic signal optical imaging methods[unreadable] recently developed in our laboratory. We will then study in detail by the methods noted below mutants that[unreadable] affect plasticity in both somatosensory (Project 2) and visual (this project, Project 3) cortex. Therefore, we[unreadable] will coordinate the sequence of mutant studies in projects 2 and 3 so that mutants that do not affect either[unreadable] visual or somatosensory plasticity can be set aside. The next series of experiments on selected mutant[unreadable] genes will focus on the development of cortical maps and the receptive field properties of individual neurons.[unreadable] Mutations that do not perturb the early development of the cortex and visual responses but do alter cortical[unreadable] plasticity will be the subject of detailed analysis. We will search for the basis of of plasticity defects in the[unreadable] selected mutants by measuring their the effects on the turnover of dendritic spines and synaptic boutons in[unreadable] vivo and the change in this turnover elicited by stimuli that would normally cause plasticity of visual[unreadable] responses. Because the appropriate regulation of intracortical inhibition is known to be a crucial regulator of[unreadable] visual cortical plasticity, we will also determine the effects of selected mutations on inhibitory[unreadable] neurotranmission by making whole cell patch recordings in vitro.[unreadable] These studies will identify novel genetic pathways that are specifically involved in neocortical plasticity.[unreadable] Nearly all of earlier research on genes involved in forebrain plasticity has begun from defects in plasticity in[unreadable] the hippocampus. We are selecting mutants in which hippocampal function should be normal. Defects in[unreadable] neocortical plasticity are likely to underly defects in cognition and awareness that underly many forms of[unreadable] mental illness and neurobehavioral developmental disorders. Identification of genetic pathways in[unreadable] experimental animals will be a first step toward understanding them and formulating rational therapy for[unreadable] these afflictions.

DESCRIPTION (provided by applicant): Most forebrain interneurons originate in the developing medial ganglionic eminence (MGE), from where they migrate into cortex, hippocampus, striatum, and amygdala to form local inhibitory circuits. When transplanted into the juvenile or adult mouse cortex, MGE cells retain the ability for migration, functional integration, and differentiation primarily into parvalbumin (PV) and somatostatin (SOM) expressing GABAergic cells. Previous work has shown that GABAergic inhibition is required for the induction of cortical plasticity and brain repair. Recent work from our laboratories has shown that transplantation of MGE cells into the neonatal or juvenile mouse visual cortex can induce a new period of ocular dominance plasticity (ODP). The transplantation of MGE cells can also enhance the brain's capacity for functional recovery and is being developed as a possible cell therapy for several brain disorders. The ability of these cells to induce plasticity de novo also offers a powerful tool to study the mechanisms and limits of cortical plasticity. The present proposal has three Aims. Aim (1) will determine which type of cortical interneuron is responsible for the induction of cortical plasticity. We have developed and validated genetic tools to ablate PV, SOM, or both cell types from the MGE grafts. Previous research suggests that PV cells may be responsible for the induction of ODP, but this hypothesis has not been formally tested. Our preliminary studies suggest that ODP can still be rekindled, even when most PV cells are eliminated from MGE grafts. We will determine if SOM interneuron depletion is sufficient for the elimination of ODP, or whether both these populations have the capacity to induce ODP. This ODP is induced in young animals weeks after the end of the critical period, but it remains unknown if the adult brain is similarly receptive to interneuron-induced plasticity. Aim (2) will determine if the transplantation of cortical interneurons in the adult brain can induce ODP and contribute to recovery of function. We have developed and validated optical recording techniques to study ODP induction in adult mice. We also have preliminary evidence that MGE cells grafted into the adult mouse cortex migrate and integrate, suggesting that they could also modify cortical circuits and possibly induce ODP. Aim (3) will determine the normal pattern of interneuron maturation in the human visual cortex. If cortical interneurons are key to the induction of critical period plasticity in humans, what is normal pattern of interneuron maturation in infants? Studies from our labs suggest that interneurons continue to be recruited into some regions in the infant brain and this could underlie extended periods of plasticity. Using a collection of brain samples from our developmental tissue bank, we have validated staining and stereological methods to quantify and map PV, SOM, and other markers of immature and mature interneurons. Identification of cortical interneurons responsible for the induction of plasticity, the age range when plasticity can be induced, and the normal patterns of human interneuron maturation will provide new information for the use of MGE cells in brain repair.

The visual system of the mouse is now widely studied as a model for developmental neurobiology, as well as for the understanding of human disease, because it can be studied with the most powerful modern genetic and optical tools. This project aims to discover how neurons in the visual cortex of the mouse allow it to see well by measuring how the cortex represents ecologically-relevant properties of the visual world. Quantitative studies of neurons in the mouse's primary visual cortex to date reveal only very poor vision, but their behavior indicates that mice can see much better than that -- they avoid predators and catch crickets in the wild. To understand mouse vision, the investigators will study responses to novel, mathematically tractable stimuli resembling the flow of images across the retina as the mouse moves through a field of grass. Studies based on these new stimuli indicate that most V1 neurons respond reliably to fine details of the visual scene. A mathematical understanding of how the brain takes in the visual world should have real implications for how we see, and should have great benefits for artificial vision by computers and robots. Bringing these ideas into the classroom will provide the foundation for new technologies, and will expose students to both real and artificial vision systems.

Analyses of the brain's visual function are limited by the stimuli used to probe them. Conventional quantitative approaches to understanding biological vision have been based on models with linear kernels in which only the output might be subject to a nonlinearity, all derived from responses of neurons in the brain to gratings of a range of spatial frequencies. This analysis fails to capture relevant features of natural images, which can not be constrained to linearity. The goal of this project is to probe the mouse visual system beyond the linear range but below the barrier posed by the complexity of arbitrary natural images. The investigators have identified an intermediate stimulus class--visual flow patterns--that formally approximate important features of natural visual scenes, resembling what an animal would see when running through grass. Flow patterns have a rich geometry that is mathematically tractable. This project will develop such stimuli and test them on awake-behaving mice, while recording the resultant neural activity in the visual cortex. Studying the mouse opens up the possibility of applying the entire range of powerful modern neuroscience tools-- genetic, optical, and electrophysiological. Visual responses will be analyzed using a novel variety of machine learning algorithms, which will allow the investigators to model the possible neural circuits and then test predictions from those model circuits. Such an understanding of the brain will inform both primate vision and the next generation of artificially-intelligent algorithms which, as a result, should benefit from being more "brain-like."

This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

PROJECT SUMMARY The major goal of the Rapid-Prototyping and Design Core (RPDC) remains to support the user's needs for the design of new unique research equipment. Additional goals for the RPDC is the education and training of users, empowering them to participate in the manufacturing, construction, maintenance, and upgrading of their unique research equipment designs.

Project Summary The use of stimuli with increasingly naturalistic properties has become critical to advance our understanding of vision. Many studies demonstrate that simple artificial stimuli (e.g. sinusoidal gratings and white noise) fail to engage nonlinearities that profoundly alter responses in the retina, lateral geniculate nucleus (LGN), and primary visual cortex (V1). A recent and striking example comes from the use of naturalistic ?flow? stimuli, which engage robust responses in V1 that are not predicted from responses to gratings. This gap in understanding motivates the development of a stimulus ensemble and analysis framework that produces a quantitative understanding of visual processing to increasingly naturalistic stimuli and the nonlinearities that they engage. Our objective is to understand how flow stimuli are processed from retina through visual cortex. To meet this goal, we will make neural population recordings in retina (Aims 1 & 3), LGN (Aims 1 & 3) and V1 (Aim 3) using matched experimental conditions and a unified theoretical/modeling framework to map the transformations that occur across these stages of visual processing. Our central hypothesis is that V1 transforms a discrete and heavily light-level-de- pendent retinal representation of natural stimuli into a continuous (uniform) representation that is relatively in- variant to changes in the mean luminance. This invariance places a strong constraint on the class of nonlineari- ties that transform retinal responses to those observed in LGN and V1. We test this hypothesis in three aims: (1) determine early visual processing (retina & LGN) of naturalistic flow stimuli; (2) develop an encoding manifold to capture the population activity at each processing stage and transforms from one stage to the next; (3) test the ability of the manifold description to predict the impact of light adaptation on processing flow stimuli from retina to V1. Aim 1 will yield a matched experimental dataset to an interesting and novel class of ecologically-relevant stimuli. Aim 2 will yield a quantitative framework by which to understand the transformations that occur between retina, LGN, and V1. Aim 3 will provide a platform for globally perturbing the output of the retina by switching from photopic to mesopic and scotopic conditions, and thereby compare predictions of our model to measured changes in LGN and V1 activity. The primary significance of this research is that it will provide a computationally and experimentally unified framework for understanding the transformations that occur in the processing of stim- uli across multiple stages of visual processing. The major innovations are (1) presenting visual stimuli for retinal recordings that are matched to eye movements and pupil dynamics in alert animals; (2) creating a novel analysis framework that captures the responses of neurons at all three levels and the inter-level transformations to in- creasingly complex stimuli; (3) utilizing light adaptation as a method of perturbing retinal output to test our model and the stability (invariance) of LGN and V1 responses to adapting retinal signals. The expected outcome is a data-driven model of the processing from retina to LGN and V1 that generalizes from starlight to sunlight.

Adult; Affect; Apoptosis; Brain; Bypass; Calcium; Cell Adhesion Molecules; Cell Death; Cells; Cerebral cortex; Code; Cognition; combinatorial; conditional knockout; critical period; Data; Development; Electrophysiology (science); Elements; Esthesia; Excision; Family; Functional disorder; Ganglia; Gene Cluster; Goals; GTP-Binding Protein alpha Subunits, Gs; hippocampal pyramidal neuron; Homologous Gene; Image; Impairment; In Vitro; in vivo; Interneurons; Life; Link; Medial; Mediating; Mental disorders; Microelectrodes; migration; Morphology; Mus; mutant; Mutant Strains Mice; Mutation; Neocortex; Nervous system structure; neural circuit; neuronal survival; Neurons; Pattern; postnatal; postnatal development; Process; Prosencephalon; Protein Isoforms; Pyramidal Cells; Regulation; Resolution; Retina; Role; Signal Transduction; Slice; Spinal Cord; Structure; System; Testing; time use; Transfection; Transplantation; two-photon; Visual Cortex; visual information;