Paul B. Manis - US grants

DESCRIPTION: (Adapted from applicant's abstract): The dorsal cochlear nucleus (DCN) is a relatively complex neural region thought to provide rapid and early processing of complex acoustic stimuli, and to associate auditory and non-auditory events. The overall goals of this grant are to investigate cellular mechanisms of information processing in the molecular layer of this nucleus. This circuitry consists of an obligatory set of interneurons, the granule cells, which receive input from diverse mossy fiber afferents and in turn form excitatory parallel fibers that innervate two major targets: a set of inhibitory neurons, the cartwheel cells, and the principal projection neurons of the nucleus, the pyramidal cells. The pyramidal cells are the final common pathway through which this intricate neural circuit processes acoustic information, and provide a direct and significant input to the principal auditory midbrain nucleus, the inferior colliculus. The current proposal focuses on the intrinsic integrative mechanisms of pyramidal cells. The first aim is to investigate the hypotheses that the voltage-dependence of pyramidal discharge patterns depends on the voltage-dependence of a rapidly inactivating potassium conductance present in these cells. The modifiability of this conductance and its effects on discharge patterns, as well as sensitivity to peptide toxins, will also be studied. The second aim is to investigate the hypothesis that subthreshold oscillations in the membrane during slow depolarization play a role in regulating the first spike latency and first interspike interval, and to determine the ionic basis of the oscillations. The third aim is to evaluate the hypothesis that conductances operating in different voltage regimes differentially participate in the dynamic integration of synaptic inputs, using dynamic current clamp. The fourth aim is to continue development of single-cell models of pyramidal cells based on experimental results and to use those models to evaluate experimental results and develop new hypotheses. These experiments utilize patch-clamp recordings from cells in brain slices, including current and voltage-clamp studies on outside-out patches, high-speed calcium imaging, and pharmacological manipulations. These experiments will help us to understand key cellular mechanisms involved in neural integration of information by an important class of cells in the cochlear nucleus. The results will have an impact on our understanding of information processing in the DCN and its dynamic characteristics, and may suggest new functions for this primary auditory center. These studies may also lead to new knowledge about the general rules of information processing by neurons throughout the central nervous system.

This project proposes to do an in-depth evaluation of patients with Meniere's disease from the audiological, auditory physiological and vestibular standpoints. Cochlear function will be qualified using routine audiometry, electrocochleography, auditory brain-stem responses, and otoacoustic emissions in a pre-and postglycerol ingestion paradigm. Vestibular function will be quantified using computerized measures of vestibulo-ocular and vestigulospinal reflexes. Patients will be randomized into two groups: 1) endolymphatic sac-mastoid shunting and 2) controls. Long term follow-up will use same measures with occasional glycerol challenges over a two year period. By systematically applying these objective measures of cochlear and vestibular function, we should be able to 1) reliably group patients with Meniere's disease into various categories according to sites of involvement of the end organ, i.e., sensory hair cells vs neural elements, and 2) define the diagnosis and natural course of the disease and test the validity of the endolymphatic sac-mastoid shunt. Ultimately, it should be possible to outline objective indicators that can predict success of various treatments in this disease.

DESCRIPTION (provided by applicant): Neurons and neural circuits in the cochlear nuclei are on the front line of auditory information processing. The CN receives a stereotyped representation of sound in the form of spike trains from the auditory nerve (AN), and produces highly modified parallel representations that drive the ascending auditory pathways. Traditionally, the cochlear nuclei are viewed as consisting of three major regions containing distinct populations of projection neurons that each emphasize different aspects of information about the acoustic environment in a parallel, but largely independent, fashion. However, coordinated spiking between output pathways can aid in the reconstruction of auditory objects and the detection of signals in noise by providing temporal cues that contribute to integration in higher auditory neurons. Our overarching hypothesis is that the relative spike timing between these pathways is coordinated not only by features in the acoustic stimulus, but importantly by shared local excitatory and inhibitory circuits, implying that the pathways are not independent. Coordinated activity may aid in the reconstruction of auditory objects and the detection of signals in noise by providing temporal coherence of activity across cells that can be integrated when these pathways converge onto higher auditory neurons. Yet, how the local circuits contribute to processing and their connectivity with other cells in the cochlear nuclei are only partially understood. In this proposal, we address how the output pathways of the cochlear nuclei can be coordinated through local circuits. The first stage of the work takes place in the context of normal hearing, and examines hypotheses about the functional synaptic connectivity of three cell types in the cochlear nuclei with the principal neurons and with each other, using optogenetic techniques and targeted patch clamp recording in brain slices form mice. The second stage incorporates the spatial structure of these circuits and the temporal dynamics of their synapses into a network model to evaluate how the activity between and within the output pathways is structured by the local circuits. We will then test predictions from the model with in vivo single unit studies. The finals stage considers the effects of a high-frequency noise-induced hearing loss on the functional organization of the CN circuit to determine how excitatory and inhibitory balance is altered. The rationale of the proposed research is that the successful restoration of function with cochlear implants or hearing aids depends on the ability to optimally engage the functional network architecture of the cochlear nucleus, which in turn requires an understanding of how information is integrated in the cochlear nuclei and how the output activity is coordinated by the local networks.

This is a proposal to begin a new research venture in studies of auditory cortex, in a laboratory that has previously been exclusively working in the cochlear nucleus. The aims of our exploration are threefold: (1) We will study synaptic function and connections between pairs of identified cortical neurons in auditory areas of the rat, using simultaneous intracellular recordings. (2) We will study long-term potentiation and depression of synaptic responses in the supragranular layers of primary and non-primary auditory cortex, with a particular focus on the timing relationship between synaptic activity and post-synaptic spiking, between pairs of identified cells. (3) We will examine changes in synaptic function from before the onset of hearing through early adulthood, to see whether synaptic plasticity is expressed differently during critical periods. These aims take advantage of our existing technical expertise and hardware, develop additional experimental capabilities, and begin the exploration of a new class of problems in auditory information processing.

We propose to continue our program of research training for residents in the basic sciences related to Otolaryngology/Head and Neck Surgery at the University of North Carolina at Chapel Hill. The training program consists of two tracks. The first track is for residents in the Otolaryngology program. One resident in each incoming class receives 2 contiguous protected years of research training in the Department?s research laboratories, starting after their surgical internship year, in preparation for careers as academic Otolaryngologists and researchers. The second track is for one non-resident postdoctoral fellow per year, who will be appointed to receive 2 contiguous years of training. All levels of the training program are designed to provide cross-fertilization between the trainees in Otolaryngology with basic and clinical scientists. Didactic elements of the training include a year-long course that covers diverse topics including experimental design, research ethics, publication and grantsmanship, is designed to ensure the production of outstanding researchers. Training progress is monitored with monthly research meetings and trainee presentations. This training program is also extended to the regular non-research track residents, medical students, graduate students, and postdoctoral trainees in the labs affiliated with the department. The training faculty includes basic and clinical faculty of the Department of Otolaryngology/Head and Neck Surgery at the University of North Carolina at Chapel Hill, and selected faculty from other departments whose research interests intersect with Otolaryngology/Head and Neck Surgery. Research areas include auditory neurobiology, cochlear implants, auditory perception, cancer mechanisms, head and neck cancer epidemiology, and sinus and airway function. An External Advisory Committee with expertise related to the research training tracks represented by the program provides guidance regarding the overall program. An Internal Advisory Committee provides additional oversight of the training program and helps guide trainee selection. The long-term goal is to produce academic clinicians and basic researchers in the field of Otolaryngology/Head and Neck Surgery with exceptional research credentials.

DESCRIPTION (provided by applicant): Support is requested to renew and continue a broad, comprehensive and fundamental interdisciplinary predoctoral training program in the neurosciences at the University of North Carolina at Chapel Hill. The training will be administered by the interdepartmental Curriculum in Neurobiology which is in its third decade of existence as a Ph.D. degree granting entity. Training will involve 48 faculty members of the Curriculum representing research laboratories in 14 departments or programs. These research facilities are well equipped and funded for a wide variety of anatomical, molecular, genetic, biochemical, physiological, behavioral, and biophysical investigations. During the past year the Curriculum has become closely integrated with the new UNC Neuroscience Center providing expanded opportunities for training through the development of new research laboratories, the recruitment of new faculty, and occupancy of a new research building in the Fall of 2001. The formal training program is already in place and constitutes a series of required and elective learning activities leading to the Ph.D. in Neurobiology. Learning activities include formal coursework in a recently reorganized curriculum, laboratory apprenticeship research with individual faculty mentors, focused dissertation research, research seminars, techniques seminars, clinical correlation experiences, journal clubs, and discussion groups on topics of career development and research integrity. In addition an annual Carolina Neuroscience Symposium will expose trainees to current thought on specialized topics presented by distinguished scientists from outside UNC. An important central goal is to train individuals to utilize methods from a variety of disciplines (e.g. ultrastructure, molecular biology, molecular genetics, electrophysiology) to probe important problems in neurobiology. The proposed training program will take advantage of several areas of particular strength in neurobiology research at UNC including: (1) molecular and genetic control of neural development (2) molecular correlations of specific sensory neuronal function, (3) glial cell biology, (4) structure, function, and regulation of neurotransmitters, their receptors and transporters, (5) mechanisms of signal transduction and ion channel function, (6) neuroendocrine and neuroimmune interactions, (7) functional imaging of nervous system activity in vitro, in vivo, and in situ, and (8) distribution and regulation of neuropeptides active in various regions of the central nervous system. Support is requested for 10 predoctoral trainees. Qualified minority candidates will be aggressively recruited.

DESCRIPTION (provided by applicant): Sensory systems perform adaptive processing of the sensory environment on a moment-to- moment basis. In the cortex, adaptive processing develops the basic network, optimizes sensory learning for specific perceptual tasks, and supports compensatory responses to long- term changes in sensory input. Cortical plasticity depends on the organization of intracortical circuits as well as the intrinsic plasticity of local microcircuits. In this proposal, we will explore local circuit organization within and orthogonal to the tonotopic axes of the primary auditory cortex, the mechanisms regulating synaptic plasticity in those circuits, and the effects of hearing loss on circuit organization and synaptic plasticity. In the first aim, we will test the hypothesis that the organization of synaptic connections in L2/3 in primary auditory cortex is anisotropic with respect to the tonotopic axes, and we will compare the strength and organization of the supragranular input to L4 neurons with that from layers 5 and 6. We will measure the tonotopic map, then use a thalamocortical brain slice preparation to dissect the responses of morphologically identified neurons in physiologically defined regions to thalamic stimulation and to local intracortical stimulation, using a combination of electrophysiological and optical methods. In the second aim, we will examine cellular mechanisms that regulate a key trigger of synaptic plasticity, action potential back-propagation, in dendrites of L4 and L2/3 neurons. Stimulation of basal forebrain cholinergic systems has been shown to enhance map plasticity in vivo, and we find that activation of cholinergic receptors in auditory cortex affects spike timing-dependent plasticity. We will test the hypotheses that dendritic potassium channels regulate calcium signaling produced by back-propagating action potentials in dendrites, and that these channels are in turn regulated by muscarinic receptor activation. In the third aim we will test the hypothesis that noise-induced hearing loss increases synaptic connectivity between L2/3 pyramidal neurons in the normal-hearing region and the hearing- loss region, and that the hearing loss also decreases synaptic plasticity. Our experiments are aimed at identifying key circuits and cellular mechanisms that support adaptive processing functions at the initial stages of cortical processing, and to understand how those mechanisms respond to hearing loss.

The cochlear nucleus is the gateway for central nervous system processing of auditory information in mammals. It has been proposed that parallel processing channels are set up in the CN, and these form the basis for further computation at higher stations of the auditory system. Despite decades of study, enumeration of CN cell types is incomplete and CN circuitry is described only superficially. In neuroscience generally, classification and naming of neurons has relied primarily upon qualitative approaches based upon human observational capabilities. We have implemented and in some cases developed novel high-throughput and unbiased techniques for labeling, segmenting and classifying neurons in 3D, generated from large-scale electron microscopy image volumes. We propose to deliver a nanoscale map, or connectome, of the mouse CN with enumerated and localized cell types and their synaptic connections. This effort is unbiased because all neurons will be sampled. To achieve this goal, we bring together four parallel modes of tissue analysis for neuron classification: morphology, connectivity, molecular identity and function. We propose that connectivity analysis will define long-proposed parallel processing circuits that will be tested functionally using realistic biophysical models of identified cell types. Notably, the cochlear nucleus contains both amorphous and layered organizations of cells, which serve as templates for all other brain regions. By investigating the fundamental structure of this sensory center, we will establish principles of neural computation and methods for structural and functional phenotyping that will apply to other brain regions regardless of their particular neural architecture.