1990 — 1997 |
Julius, David J |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Molecular Analysis of Serotonin Receptor Function @ University of California San Francisco
The overall objective of the work proposed here is to understand how neurotransmitters modulate cellular and physiological processes by interacting with specific cell surface receptors. The focus will be on serotonin [5-hydroxytryptamine, 5HT], a biogenic amine that is involved in a wide array of physiological responses in the central and peripheral nervous system. Serotonin exerts its physiological effects by binding to a family of structurally- and functionally-related cell surface receptors, each having distinct pharmacological properties. In addition, these subtypes couple to different intracellular second messenger signaling pathways. In the brain, serotonin receptors are believed to play a key role in modulating affective and perceptual states, and are sites of action of numerous psychotropic drugs, including LSD. In the spinal cord, serotonin is involved in the central regulation of pain, while in the periphery serotonin modulates enteric reflexes and the contraction of smooth muscle. As such, these receptors are potential targets for the pharmaceutical treatment of affective disorders (obsessive-compulsive behavior, depression and schizophrenia), migraine headaches and pain. Genes encoding three 5HT receptor subtypes (5HT1a, 5HT1c and 5HT2) have now been cloned, permitting a molecular analysis of receptor structure and function. When expressed in the unnatural environment of a fibroblasts, the 5HT1c and 5HT2 receptor subtypes bind ligands and activate intracellular second messenger signaling systems. The first aim of this proposal is to extend the molecular characterization of 5HT receptor subtypes by isolating other members of this gene family using standard recombinant DNA methodologies. The second objective is to use the fibroblast expression system as a means for identifying mutations in these receptors that alter their ligand binding or signal transduction properties. The third aim is to biochemically characterize the intracellular second messenger signaling pathways to which these receptors couple in fibroblasts and neuroblastoma cells in culture. Using these simple in vitro systems as models, it should be possible to further our understanding of how these receptors operate in neurons.
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0.958 |
1990 — 1996 |
Julius, David |
N/AActivity Code Description: No activity code was retrieved: click on the grant title for more information |
Presidental Young Investigator Award @ University of California-San Francisco
Transmitter receptors exist as multiple receptor subtypes with different binding affinities and different cellular and physiological responses to the same transmitter. The expression and sensitivity of these receptor subtypes can be independently regulated. Such multiplicity of receptor subtypes enables a single transmitter to play a variety of physiological roles in the same tissue. The investigator is employing molecular genetic, biochemical and electrophysiological methods to analyze and manipulate cloned serotonin receptor genes to determine how these receptors bind specific ligands and interact with other cellular proteins. This award will enable the investigator to isolate novel serotonin receptors and to characterize the cellular responses they elicit as well as to determine which cells in the rat brain express them. In addition, the regions of serotonin receptors responsible for ligand binding, ligand.induced conformational changes in receptor structure, and G protein interaction will be determined using mutational techniques. Differences in the second messenger systems activated by different serotonin receptor subtypes will also be explored.
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1 |
1996 — 2000 |
Julius, David J |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Substances That Mediate Signaling in Nociceptive Pathways @ University of California San Francisco
The identification of endogenous substances that mediate sensory and synaptic signaling nociceptive pathways has resulted from pharmacological and physiological studies. For a number of these ligand-receptor systems, selective and potent pharmacological probes have not been developed that can discriminate among closely related receptor subtypes. This lack of pharmacological reagents has hampered progress in understanding signaling mechanisms in nociceptive pathways and in the development of novel drug- based therapies for the treatment of acute and chronic pain. The application of molecular biological methods to neuropharmacology has greatly facilitated the identification and functional characterization of distinct receptor subtypes for most neurotransmitters. Gene cloning has also provided molecular probes for determining the location of specific receptor subtypes within functional neural pathways, and their regulation under physiological and pathological conditions. This proposal is focused on the molecular biology of two receptors that are important in sensory transduction and neural transmission in nociceptive pathways. One class of receptors is activated by extracellular adenosine triphosphate (ATP), and the other by vanilloid compounds (e.g., capsaicin and resiniferatoxin). We have recently isolated genes encoding ATP-gated ion channels (termed P/2X receptors) from the mammalian nervous system. We propose to use these molecular probes to determine where these receptors are expressed in the nociceptive pathway. In addition, we plan to elucidate physiological roles for these receptors in sensory transduction and synaptic transmission by generating mutant mice through gene targeting that lack specific ATP receptor subtypes. We are also initiating efforts to isolate a functional cDNA clone encoding the capsaicin receptor, which serves as an important marker for sensory afferents in the pain pathway and which represents a potential target for novel therapeutic agents in the treatment of pain.
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0.958 |
1997 — 2000 |
Julius, David J |
P50Activity Code Description: To support any part of the full range of research and development from very basic to clinical; may involve ancillary supportive activities such as protracted patient care necessary to the primary research or R&D effort. The spectrum of activities comprises a multidisciplinary attack on a specific disease entity or biomedical problem area. These grants differ from program project grants in that they are usually developed in response to an announcement of the programmatic needs of an Institute or Division and subsequently receive continuous attention from its staff. Centers may also serve as regional or national resources for special research purposes. |
Receptor Gene Expression in Hippocampal Neurons @ University of California San Francisco
Serotonin (5-hydroxytryptamine; 5-HT) is a prominent neurotransmitter that modulates a wide variety of sensory, motor, and behavioral responses in the mammalian nervous system. Serotonergic synapses are major targets for the action of psychotropic and antidepressant drugs like LSD, psilocybin, and Prozac and it is therefore believed that transmission at these synapses plays an important role in the regulation of mood, behavior, and perceptual states. Indeed, drugs that modify transmission at synapses are presently used in the management of depression, obsessive-compulsive behavior, eating disorder, anxiety, migraine headache, and chemotherapy-induced nausea. In addition to its well established role as a mediator of synaptic transmission in the adult nervous system, a number of independent observations suggest that serotonin may also function during embryogenesis to modulate development and morphogenesis in neural and non-neural tissues. The diverse responses to serotonin are mediated through its interaction with at least fourteen distinct and surface receptor subtypes. The complexity of this signaling system and the paucity of selective drugs have made it difficult to define specific functions for individual 5-HT receptor subtypes, or to determine how serotonergic drugs modulate mood and behavior. The long-term goal is to address these issues by using genetic methods to define roles for serotonin receptor subtypes in the regulation of developmental, physiological, or behavioral states. The primary focus of this research proposal is to elucidate functional roles for the 5-HT-3 receptor, a serotonin-gated ion channel that mediates rapid excitatory responses in central and peripheral neurons. The first specific aim is to generate transgenic mice that lack functional 5-HT3 receptors. This will be achieved by targeted disruption of the endogenous 5-HT3 receptor gene through homologous recombination, or by ectopic expression of a truncated amino-terminal fragment of the 5-HT3 receptor subunit that can inactivate native ion channel complexes via a dominant-negative effect. 5-HT3 receptor deficient mice will be examined for developmental, physiological, or behavioral abnormalities compared to wildtype siblings. The second specific aim is to identify cis-acting DNA elements and trans-acting nuclear factors that regulate 5-HT3 receptor gene transcription in neurons or neuroendocrine cells. This will be accomplished by using gene transfer and transgenic methods to pinpoint promoter elements upstream of the 5-HT3 receptor transcript that confer regulated expression of this gene in vitro and in vivo. Candidate transcription factors will be examined for their potential to interact with these promoter elements. The goal is to use the 5-HT3 receptor as a model system for studying mechanisms that regulate gene expression in the nervous system, and to identify specific promoter elements that may be used to direct the ectopic expression of heterologous proteins in the nervous system.
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0.958 |
1999 — 2016 |
Julius, David |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Molecular Mechanisms of Receptor and Channel Function @ University of California, San Francisco
DESCRIPTION (provided by applicant): Nociception is the process whereby primary afferent somatosensory neurons recognize and respond to noxious stimuli, resulting in pain and neurogenic inflammation. Members of the TRP ion channel family play important roles in nociception and pain by functioning as sensors for a variety of noxious stimuli, including heat, cold, and inflammatory agents. More broadly, genetic studies have highlighted the importance of these and other TRP channel subtypes in processes ranging from calcium adsorption to neuronal growth cone guidance, keratinocyte development, and numerous aspects of sensory transduction. Thus, understanding how these channels respond to physiological stimuli and drugs is of direct clinical and therapeutic relevance to disorders that affect virtually every majo organ system in the body. This proposal is focused primarily on understanding the structure and biophysical properties of the capsaicin- and heat-activated receptor, TRPV1 - perhaps the best-characterized member of the mammalian TRP channel family. Its widely validated role in pain physiology, together with the availability of well characterized pharmacological agents (natural and synthetic), make it a 'poster child' for elucidating basic principles underlying TRP channel pharmacology, structure, and regulation. The studies proposed here are aimed at broadening our understanding of the structural and biophysical principles whereby TRPV1 and related channels are activated or modulated by chemical or physical stimuli. The specific aims are to (i) analyze intrinsic sensitivity of TRPV1 to heat, phospholipids, and other agents in a defined environment consisting of purified channel protein and synthetic lipids; (ii) exploit purified, functional TRPV1 protein for in vitro spectroscopic studies to examine stimulus-evoked conformational movements, and (iii) use voltage-clamp fluorometry to assess the dynamics of stimulus-evoked conformational rearrangements of TRPV1 in cells. Together, these aims will address unresolved issues concerning TRP channel function and structure while laying important groundwork for the long-term goal of obtaining three-dimensional structures of TRPV1 or other TRP channels - which represents a logical and essential next step for the field. Such information is key to the rational development of therapeutic agents that target chronic inflammatory pain syndromes (e.g. arthritis, irritable bowel syndrome, and asthma) and other disorders involving TRP channels. PUBLIC HEALTH RELEVANCE: This project is focused on elucidating structural and biophysical mechanisms that regulate ion channels involved in temperature and pain sensation. Results from these studies will expand our understanding of an important class of ion channels that contribute to numerous physiological processes. In doing so, this work will aid in the development of therapeutic strategies (such as novel drugs) for treating chronic pain and other clinical disorders.
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2001 — 2005 |
Julius, David J |
P01Activity Code Description: For the support of a broadly based, multidisciplinary, often long-term research program which has a specific major objective or a basic theme. A program project generally involves the organized efforts of relatively large groups, members of which are conducting research projects designed to elucidate the various aspects or components of this objective. Each research project is usually under the leadership of an established investigator. The grant can provide support for certain basic resources used by these groups in the program, including clinical components, the sharing of which facilitates the total research effort. A program project is directed toward a range of problems having a central research focus, in contrast to the usually narrower thrust of the traditional research project. Each project supported through this mechanism should contribute or be directly related to the common theme of the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence, i.e., a system of research activities and projects directed toward a well-defined research program goal. |
Genetic Analysis of Nociceptor Function @ University of California San Francisco
The long-term objective of this proposal is to understand how primary afferent neurons of the "pain" pathway detection noxious and transduce this information this information into action potentials, a process referred to as nociception. Identification of the molecular components involved in nociception should improve our understanding of how tissue injury produces both acute and persistent pain. In the past few years, a number of candidate molecules have been proposed to mediate responses of nociceptors to noxious thermal or chemical stimuli. These include the receptor for capsaicin and other vanilloid compounds (VR1), a vallinoid-insensitive, heat-sensitive VR1 homologue (VRL1), and members of the acid-sensing ion channel family (ASICs). In the proposed study, a combination of genetic, neuroanatomical, and electrophysiological methods will be used to assess the contribution of these molecules to nociception and pain in vivo. The specific aims of the proposal are to: (i) ascertain the relative contributions of VR1 and VRL1 to nociception by assessing the ability of mice bearing deletions in these genes to detect noxious physical and chemical stimulant; (ii) determine the involvement of VR1 and VRL1 in neurotrophin-mediated hyperalgesia by examining heat and chemical sensitivity of VR1 and VRL1 mutant mice in which nerve growth factor is transgenically over-produced in the skin; (iii) assess the relative contributions of VR1 and ASICs to acid sensitivity by generating VR1/ASIC double mutant mice and examining the sensitivity to noxious physical and chemical stimuli and the cellular and behavioral level. In addition to providing new information about the basic cellular mechanisms underlying nociception and pain, the proposed studies will highlight potential targets for the development of novel analgesic agents.
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0.958 |
2004 — 2010 |
Julius, David J |
R37Activity Code Description: To provide long-term grant support to investigators whose research competence and productivity are distinctly superior and who are highly likely to continue to perform in an outstanding manner. Investigators may not apply for a MERIT award. Program staff and/or members of the cognizant National Advisory Council/Board will identify candidates for the MERIT award during the course of review of competing research grant applications prepared and submitted in accordance with regular PHS requirements. |
Molecular Mechanism of Receptor and Channel Function @ University of California San Francisco
DESCRIPTION (provided by applicant): The long-term objective of this proposal is to understand how cell surface receptors and ion channels detect extracellular signals and transduce this information into physiological changes at the cellular and organismal level. This project will focus on two members of the TRP channel family that are expressed on primary afferent neurons of the pain pathway and mediate thermosensation in the mammalian peripheral nervous system. TRPV1 is an excitatory ion channel that is activated by noxious heat or capsaicin, the pungent ingredient in chili peppers. Electrophysiological and genetic studies have shown that TRPV1 contributes to the detection of noxious heat in vivo and is modulated by a variety of inflammatory agents (e.g. extracellular protons, bioactive lipids, nerve growth factor, and bradykinin), making it an essential component of the signaling pathway through which injury increases sensitivity to heat. TRPM8 is a cold-activated channel that also responds to menthol and other cooling compounds. Determining how these channels detect thermal and chemical stimuli will provide important insight into the basic molecular processes that underlie nociception and pain sensation under normal and pathological conditions. This information will also stimulate the design and development of novel analgesic agents for treating peripheral pain syndromes, such as those associated with rheumatoid arthritis, viral and diabetic neuropathies, or peri-operative wound healing. A combination of molecular genetic, biochemical, and electrophysiological methods will be used to probe the mechanisms whereby TRPV1 and TRPM8 detect and respond to chemical and physical stimuli that produce or exacerbate pain. The specific aims of the proposal are to: (i) delineate regions of TRPV1 that are required for modulation by phospholipase C and phospholipid interaction; (ii) pinpoint domains of TRPV1 that interact with TrkA, the receptor for nerve growth factor; (iii) delineate regions of TRPM8 that are required for detection of menthol and cold; (iv) determine whether and how TRPM8 is regulated by inflammatory agents or prolonged exposure to cold (i.e. adaptation).
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2005 — 2006 |
Julius, David J |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Molecular Basis of Mechanoreception in Sensory Neurons @ University of California San Francisco
[unreadable] DESCRIPTION (provided by applicant): The long-term goal of this research is to elucidate the molecular basis of mechanotransduction by mammalian somatosensory neurons. Somatosensory mechanoreceptors mediate the senses of pain, touch and proprioception. The importance of these senses to human health is underscored by diseases that cause peripheral neuropathy, such as rheumatoid arthritis, diabetes and acquired immunodeficiency syndrome. Because patients with peripheral neuropathy cannot feel injuries, even minor insults can lead to irreversible tissue damage and chronic pain. The objective of this exploratory study is to understand how cell surface receptors and ion channels detect mechanical stimuli and transduce this information into physiological changes at the cellular level. A combination of molecular genetic, histological, and electrophysiological, and live-cell imaging methods will be used to probe the mechanisms whereby stretch, changes in osmolarity or direct pressure lead to depolarization of the primary afferent neuron. One of the major goals of this application is to develop in vitro systems that can be used to simultaneously detect and characterize mechanosensory responses in large numbers of cultured sensory neurons from rat dorsal root or trigeminal ganglia. Histological and pharmacological analyses will then be used to profile mechanosensitive cells with respect to the expression of a variety of molecular markers that delineate subsets of primary afferent neurons. Furthermore, we will use the high-throughput stimulus paradigms and detection methods developed here to carry out a screen to identify molecules that may be involved in mechanosensory transduction. Once identified, candidate molecules will be evaluated using electrophysiological, histological and genetic methods. [unreadable] [unreadable]
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0.958 |
2006 — 2016 |
Julius, David |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Cellular Physiology of Sensory Ion Channels @ University of California, San Francisco
DESCRIPTION (provided by applicant): Chemo-nociception is the process whereby primary afferent sensory neurons recognize and respond to noxious substances, resulting in pain, discomfort, and/or neurogenic inflammation. Such substances include exogenous environmental irritants, as well as endogenous pro-algesic agents that are produced or released in response to injury or disease. The ion channel TRPA1 also known as the 'wasabi receptor' functions as a major detector of chemical irritants and contributes significantly to mechanisms underlying acute pain and chronic inflammatory pain syndromes, such as arthritis, inflammatory bowel disease, hemorrhagic cystitis, asthma and other constrictive airway disorders. Thus, understanding basic mechanisms whereby TRPA1 detects noxious chemical stimuli and responds to cellular regulatory mechanisms is relevant to both basic sensory physiology and pain therapeutics. Recent studies have shown that chemically reactive (electrophilic) irritants activate TRPA1 through an unusual process involving covalent modification of cysteine residues within the so-called 'linker region' located in the cytoplasmic amino-terminus of the channel protein. An ankyrin-repeat rich domain (ARD) adjacent to the linker region also plays a significant role in specifying stimulus sensitivity, suggesting that it engages in a functional, and possibly structura interaction with the linker region to mediate irritant-evoked channel gating. Intracellular calcium also serves as a key modulator of TRPA1 function, promoting both potentiation and inactivation. Recent structure-function studies further suggest that calcium-dependent modulation is also specified by a site(s) within the ARD, although the cellular and biochemical mechanisms underlying this important regulatory process remain obscure. G protein beta-gamma subunits may also serve as TRPA1 modulators. The goal of this proposal is to elucidate biochemical and structural mechanisms whereby the TRPA1 amino terminus functions as an integrator of physiological stimuli that regulate sensory neuron excitation. The specific aims are to: (i) define the key structural elements of the linker region - such as length, proximity to the ARD domain, and relative location of modifiable cysteine residues that specify irritant detection and efficienc of channel gating; (ii) elucidate cellular signaling mechanisms underlying G protein and calcium-dependent channel modulation using electrophysiological and biochemical methods; and (iii) develop a structural model of the TRPA1 amino-terminus using biochemical and crystallographic methods. Together, these studies will provide a rational basis for the development and design of novel analgesic agents for controlling chronic pain and neurogenic inflammatory syndromes.
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2009 — 2017 |
Julius, David |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. R37Activity Code Description: To provide long-term grant support to investigators whose research competence and productivity are distinctly superior and who are highly likely to continue to perform in an outstanding manner. Investigators may not apply for a MERIT award. Program staff and/or members of the cognizant National Advisory Council/Board will identify candidates for the MERIT award during the course of review of competing research grant applications prepared and submitted in accordance with regular PHS requirements. |
Exploiting Toxins to Probe Sensory Signaling @ University of California, San Francisco
DESCRIPTION (provided by applicant): Natural plant products have served as tremendously valuable tools for deciphering cellular and molecular mechanisms contributing to somatosensation, nociception, and pain. Notable examples include the use of natural analgesics, such as morphine (from the opium poppy) and salicylate (from willow bark) to discover opioid receptors and cyclooxgenases, respectively. Other important examples include the use of natural irritants, such as capsaicin (from chili peppers) and menthol (from mint leaves) to identify ion channels that detect heat and cold, respectively. Indeed, each of these proteins represents a validated or potential target for pharmacological management of acute or chronic pain. Plants are not unique in their capacity to produce chemical agents that target sensory neurons or other excitable cells. Indeed, venoms from animals (ranging from arachnids to mammals) represent a vast pharmacopoeia that has great potential to yield novel agents with which to identify or characterize receptors, ion channels, or other signaling molecules that contribute to sensory transduction. Indeed, in the previous funding period we identified two such toxins - one from spider and another from snake - that serve as novel, potent, and highly selective agonists for TRPV1 and ASIC1 channels, respectively. In each case, these toxins enabled elucidation of the activated, fully open state of the channel at atomic resolution, providing unprecedented insights into structural mechanisms underlying channel gating and modulation. This proposal builds on our success and expertise in toxin discovery and characterization, with the goal of expanding the repertoire of pharmacological agents with which to study known or novel somatosensory receptors. The first aim is focused on characterizing two spider toxins that we identified in a sensory neuron- based screening assay, and which target a specific voltage-gated sodium channel (Nav) subtype expressed by these cells. We propose to identify the Nav channel domain(s) that specifies toxin sensitivity and accounts for its subtype selectivity. Furthermore, we shall test toxin selectivity in vivo using mouse genetics, and identify the subpopulation of sensory neurons that mediate the excitatory and algogenic actions of these toxins in cellular and behavioral paradigms. The second aim is focused on characterizing two novel toxins - one from centipede and the other from snake - that we also identified by functional screening, and which activate primary afferent sensory neurons to elicit nocifensive responses in mice. We propose to identify the molecular targets of these toxins and determine the signaling mechanisms through which they activate sensory neurons of the pain pathway. These studies will uncover novel sensory transduction molecules and/or provide powerful new tools for determining how known transducers work to modulate nociceptor excitability. Information gleaned from this work will provide important pharmacologic leads and insights pertinent to the development of analgesic agents.
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2009 — 2011 |
Julius, David |
P41Activity Code Description: Undocumented code - click on the grant title for more information. |
Toxin Interactions With Trpv1 @ University of California, San Francisco
This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. One of the most powerful tools for analyzing ion channel structure/function has been the identification of natural toxins agonists and antagonists. Several organisms produce small protein toxins that act on ion channels and aid in prey capture or defense. We propose to study toxin interaction with the ion channel TRPV1. We will use MALDI mass spectrometry to assess the molecular composition of toxins isolated by reverse phase chromatography. The UCSF Mass Spectrometry Facility will assist in the collection to determine the molecular weight of isolated toxins.
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2009 — 2011 |
Julius, David |
P41Activity Code Description: Undocumented code - click on the grant title for more information. |
Trp Channel Modulation @ University of California, San Francisco
This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The purpose of this project is to perform biochemical characterization of proteins that modulate the activity of channels important for pain sensation. The targets of study will include channel cytoplasmic domains, small molecule modulators, peptide modulators, and associated membrane or cytoplasmic proteins. The objective is to purify proteins, determine molecular interaction sites, and perform structural studies.The UCSF mass spectrometry facility will aid us in determining the accurate mass and of purified products, as well as confirming the identity of purified products. The aid of the facility will also allow us to interpret results from chemical crosslinking studies in the future.
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2012 — 2016 |
Julius, David |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Asic Channels and Pain @ University of California, San Francisco
DESCRIPTION (provided by applicant): Nociception is the process whereby primary afferent somatosensory neurons recognize and respond to noxious stimuli. In addition to initiating acute pain responses, nociceptor activation can produce local inflammation leading to pain hypersensitivity. Tissue acidosis (i.e. reduction in local pH) is an important hallmark of this response, and is associated with a range of physiological insults, such as infection, ischemia, tumor growth, and arthritis. Indeed, extracellular protons enhance excitability of primary afferent nociceptors, thereby producing acute pain and/or pain hypersensitivity. Members of the acid sensing ion channel (ASIC) family are believed to play important roles in nociception and pain by functioning as sensors for extracellular protons. For example, the ASIC3 subtype likely accounts for ischemic pain associated with large, rapidly inactivating proton-evoked currents in neurons that innervate skeletal or cardiac muscle. However, additional roles for ASIC channels in nociception have remained enigmatic for a variety of reasons. First, a dearth of pharmacological tools has made it difficult to manipulate these channels in vivo. Second, mice lacking specific ASIC channel subtypes have failed to revealed clear or robust phenotypes in regard to acid-evoked pain or other aspects of nociception. Third, a comprehensive analysis of ASIC channel expression and localization - which is critical to deciphering physiological roles for these channels in nociception and pain - remains incomplete. Finally, some ASIC subtypes (e.g. ASIC2a) respond only to extreme extracellular acidosis (pH < 5), suggesting the existence of other endogenous modulators for these channels that may be produced under pathophysiological conditions of tissue injury and/or chronic inflammation. The goal of this proposal is to address these and other questions by applying genetic, physiologic, and biochemical methods to develop a comprehensive view of ASIC subtype function, pharmacology, and expression in the somatosensory system. The specific aims are to (i) develop a comprehensive map of ASIC channel expression using gene targeted reporter mice to visualize ASIC-positive nerve fibers with exquisite sensitivity and fidelity; (ii) characterize functional properties of ASIC-expressing sensory neurons and nerve fibers from these genetically-labeled mice using a range of electrophysiological and live-cell imaging methods; (iii) screen for novel endogenous ASIC modulators in extracts of normal and injured tissues using a range of biochemical, functional, and behavioral assays. Together, these aims will address important unresolved questions concerning ASIC function as key steps toward the rational development of novel therapeutic agents that target a range of chronic inflammatory pain syndromes.
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2018 — 2021 |
Julius, David |
R35Activity Code Description: To provide long term support to an experienced investigator with an outstanding record of research productivity. This support is intended to encourage investigators to embark on long-term projects of unusual potential. |
Natural Products as Probes of the Pain Pathway @ University of California, San Francisco
Nociception is the process whereby a subset of somatosensory nerve fibers (called nociceptors) detect noxious stimuli and transmit this information to the spinal cord and brain, ultimately producing a percept of discomfort or pain. Nociceptors are faced with the complex task of detecting disparate environmental and endogenous signals of both a physical and chemical nature; these include temperature, pressure, irritants, pruritogens, and inflammatory agents. Consequently, nociceptor activation elicits acute pain as well as injury-evoked pain hypersensitivity and can contribute to so-called ?maladaptive? processes underlying persistent pain syndromes. Our goal is to understand how nociceptors detect and integrate these signals in response to changing environmental or physiological conditions. Natural products from plants or venomous creatures have served as tremendously valuable tools for deciphering cellular and molecular mechanisms contributing to nociception and pain. Notable examples include the use of natural analgesics, such as morphine (from the opium poppy) and salicylate (from willow bark) to discover opioid receptors and cyclooxgenases, respectively. Other important examples include the use of natural irritants, such as capsaicin (from chili peppers) and menthol (from mint leaves) to identify ion channels that detect heat and cold, respectively. Indeed, each of these natural product receptors represents a validated or potential target for pharmacological management of acute or chronic pain. This proposal is aimed at elucidating molecules, cells, and mechanisms that contribute to nociception in the context of acute (protective) or pathological (chronic) pain states. We shall continue to exploit the vast chemical ?space? of natural product pharmacology to identify and characterize ion channels and sensory neuron subtypes that contribute to distinct nociceptive modalities. At the most reductionist level, we will use biophysical, biochemical, and pharmacological tools to elucidate structural mechanisms underlying ion channel function, including stimulus detection, drug binding, and gating. Here, the main emphasis will be on members of the TRP channel family that are targeted by natural pungent agents and play major roles in thermo- or chemo-nociception. At a more integrative level, we will use molecular and pharmacological probes to characterize distinct nociceptor subtypes, whose functionalities will be examined in mice using genetic, anatomical, and in vivo imaging methods. As one such example, we are interested in characterizing a population of presumptive A? nociceptors and asking how they contribute to mechanical pain. Together, these studies will provide important mechanistic insights for the development of novel analgesic therapies.
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2020 — 2021 |
Ingraham, Holly A. (co-PI) [⬀] Julius, David |
U01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Mapping Gut-Spinal Cord Connections in Visceral Pain @ University of California, San Francisco
Project Summary/Abstract Our current understanding of mechanisms underlying visceral pain, including that associated with irritable bowel syndrome, remains rudimentary. Importantly, opiates are ineffective at treating visceral pain syndromes, and only exacerbate discomfort by producing constipation, reflecting a clear need for alternative treatment options. The goal of this proposal is to bring greater mechanistic insight to this underserved area of pain research, and to approach the problem in a multifaceted strategy designed to maximize the relevance of our basic research discoveries to future pain treatments. Here, we will ask how enterochromaffin (EC) cells transmit noxious signals from the gut lumen to the spinal cord. EC cells are key sensory cells in the intestinal epithelium that release serotonin onto primary sensory nerve fibers, thereby evoking a sensation of discomfort and pain in response to luminal irritants, such as bacterial metabolites, inflammatory agents, or ingested chemicals. The goals of this collaborative effort are to use activating and silencing approaches to examine functional connections between EC cells and sensory nerve fibers. We will couple these methods with transcriptome profiling, viral tracing, and electrophysiological methods to gain insights into the molecular and functional identity of these fibers. Another key goal is to determine whether EC cell signaling pathways exhibit sex-specific differences, an important question that may relate to the higher prevalence of GI visceral pain syndromes experienced by women. Our team brings an unusually wide ranging and innovative approach to this area of pain research that includes expertise in the neurophysiology, pharmacology, and anatomy of nociceptive and pain circuits, visceral tissue anatomy and development, and relevant clinical experience. This knowledge base is supported by complementary technological approaches that will enable us to connect molecular and mechanistic insights to physiology, visceral nociception, and disease. Our focus on the epithelial-nociceptor connectome highlights EC and other enteroendocrine cell types as potentially powerful control points for neuromodulation of visceral discomfort and pain. A comprehensive functional, pharmacological, genetic and anatomical characterization of EC-primary afferent-spinal circuits is an essential first step toward achieving this important goal. As such, our research program fits squarely within the SPARC mandate to transform our understanding of peripheral nerve-organ interactions and advance strategies for controlling organ system function.
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