Ronald Dubner - US grants

DESCRIPTION (Adapted from the Investigator's Abstract): The major hypothesis of this study is that following induction of inflammation, ascending tonic nociceptive inputs induce prolonged functional changes in the neural circuitry at brainstem sites leading ultimately to a net inhibitory modulation of spinal cord activity. This input to the brainstem triggers a cascade of molecular, biochemical and electrophysiological events that result in enhanced excitability and inducible changes in neurotransmitters and receptor populations. Specific Aim 1 will test the hypothesis that persistent inflammation leads to time-dependent, dynamic changes in the neuronal excitability of medullary brain stem nuclei. Specific Aim 2 will test the hypothesis that endogenous mechanisms of inflammation-induced changes in the excitability of medullary brain stem neurons involve excitatory and inhibitory amino acids, opioid peptides and their receptors. Specific Aim 3 will use molecular and immunohistochemical probes to test the hypothesis that alterations in EAA receptor activation play a key role in enkephalinergic transmission and that reduced GABAergic transmission in the RVM is mediated via postsynaptic mu opioid receptors (MORs) after inflammation. Specific Aim 4 will test the hypothesis that changes in brain stem excitability after inflammation lead to alterations in the response profile of several different classes of neurons in the RVM. These events will be explored in a lightly anesthetized rat preparation in which both behavioral and electrophysiological responses to hindpaw inflammation can be ascertained sequentially over time. Hindpaw inflammation will be induced with complete Freund's adjuvant (CFA) and the studies done over specific, but relatively short, time frames during the development phase of this persistent inflammatory state. Fixed brainstem tissues from other animals with CFA inflamed hindpaws will be compared to control tissues to determine specific inducible neurotransmitter and receptor changes subsequent to inflammation. The screening process will include anatomical studies identifying excitatory amino acid (EAA) and preproenkephalin (PPE) receptor mRNA expression increases, as well as immunocytochemical localization of changes in Fos, GABA, mu opioid receptor, and glutamate receptor subunit NMDA NR1 expression with specific antibodies. The findings of this study will improve understanding of supraspinally initiated descending mechanisms that modulate the pain experience through alteration of the nociceptive information relayed from the spinal cord.

The purpose of this program is to develop the organizational structure that will support the activities of a multidisciplinary regional Center devoted to the study of the mechanisms, diagnosis and treatment of chronic or persistent pain conditions of deep tissues and those associated with nerve injury. A primary focus will be on orofacial pain conditions of such origin and their comparison with similar pain conditions existing in other regions of the body. The program will be diverse and span the basic and clinical sciences and clinical management from molecular studies to clinical trial studies to outreach and demonstration projects in the community. We are fortunate that major expertise in these areas exists in the Baltimore area at the University of Maryland at Baltimore and at Johns Hopkins University. We have also sought out colleagues in surrounding regions with unique qualifications to join us in this exciting endeavor. Our working model is that research on pain has reached a level of understanding that facilitates the transfer of basic knowledge to the study of persistent pain in humans, and ultimately to new methods of diagnosing and treating these conditions in the general population. Further advances will require multidisciplinary collaborations among basic and clinical scientists and clinicians. A multidisciplinary Center in the thematic area of persistent pain will facilitate this collaboration and interaction and hopefully lead to improved management of orofacial pain and oral health. The specific aims of the Center will be: 1) To elucidate the neural basis, psychosocial influences, and proper treatment of temporomandibular disorders (TMD) and other orofacial pains. 2) To elucidate biological and psychosocial factors that predict the occurrence of chronic or persistent pain following acute back injury. 3) To elucidate the mechanisms of visceral pain and the differences and interaction between visceral and cutaneous pain. 4) To further elucidate the underlying basic peripheral and central nervous system mechanisms associated with persistent pain of nerve injury in animal models and to translate these findings rapidly into the development of new treatment approaches. 5) To transfer these research findings: a) to health professionals and the public with an emphasis on research methods for educating health professionals including demonstration projects and b) to community outreach programs with a focus on disadvantaged populations. A sub-aim will be to transfer technology advances from the research arena to clinical management and elsewhere.

Pain is the major patient complaint with systemic illness, traumatic injuries and surgery. The last decade has resulted in new knowledge that is leading to significant improvements in the management of persistent pain. Nevertheless, important gaps in our understanding remain. Many difficult to treat persistent pains involve deep visceral and muscle tissue. Research on the development of nociceptive systems is minimal as the effect of early infant persistent injury is limited above the level of the brain stem, yet multiple forebrain structures participate in the perception of pain. The major theme of this program project is the study of modulatory influences in nociceptive circuitry in response to persistent visceral and cutaneous tissue injury and the effect of neonatal exposure to pain on such plasticity. The individual projects all take advantage of the development of new animal models of inflammation and examine the changes that take place in primary afferent neurons, dorsal horn circuitry and descending modulatory pathways in adults after injury and the impact of neonatal injury on these changes. Project #1 will study the changes in dorsal horn excitability that take place after cholonic inflammation in the rat and how descending modulatory mechanisms contribute to the sensitization process. Project #2 will focus on the role of early postnatal pain experience as a risk factor for the development of nociceptive systems and the neurochemical reorganization of dorsal root ganglia neurons and spinal dorsal horn neurons. Project #3 will study the modulation of primary afferent excitability and their underlying biophysical properties after inflammation in the adult and the effect of neonatal injury on these changes. Project #4 will examine the influence of forebrain structures on descending modulatory effects in a rat model of cutaneous inflammation and the effects of neonatal cutaneous and visceral pain experience on these descending circuits. Project #5 will use a model of inflammatory hyperalgesia systems in these effects, and the neuroprotective role of gustatory stimulation in preventing developmental changes in nociceptive circuitry produced by neonatal injury. In summary, this program project grant brings together a group of experts in the field of pain to study the role of neuronal modulation in animal models of persistent pain.

DESCRIPTION (provided by applicant): Previously, we analyzed the effects of prenatal cocaine exposure during the neocortical neuronogenesis (which in primates corresponds to the second trimester of gestation) on the development of the neocortex in rhesus monkeys. We found that the neocortex in such prenatally-cocaine-exposed monkeys is characterized by: (i) a loss of the normal lamination, (ii) an inappropriate positioning of the cortical neurons, and (iii) a reduction in the density and number of these neurons. We have also demonstrated that cocaine can induce these long-term effects only if it is administered during the period of neocortical neuronogenesis, with no such abnormalities being observed in the animals exposed to cocaine either prior to or after that developmental period. It is reasonable to hypothesize that these structural neocortical abnormalities in monkeys exposed to cocaine in utero are accompanied by behavioral deficits. The demonstration such deficits is important because it would not only validate the functional significance of the detected morphological changes, but also suggest which specific aspects of behavior may be compromised in children of drug-abusing human mothers. It is well documented that postnatal development of the primate brain is very prolonged with new functional capabilities appearing throughout neonatal, infant, juvenile and adolescent periods. The brain is also known for its remarkable adaptability. Consequently various behavioral deficits in our animals may not necessarily be continuously present throughout life; some of them may be detected only transiently as developmental retardations at certain stages of brain maturation, while others may appear later in life with maturation of their adult neural substrates. Therefore, an evaluation of behavior in prenatally cocaine-exposed animals should be done in a format of a longitudinal study. The goal of the present study is to employ longitudinal behavioral tests and observations to address the hypothesis that the significant prenatal cocaine-induced cerebral cortical structural disorganization is associated with identifiable behavioral deficits: I) We will determine whether neonatal monkeys in our model of prenatal cocaine exposure display deficits on items of the Neonatal Behavioral Assessment Scale; 2) We will assess whether infant, juvenile and adolescent monkeys with significant central cortical structural abnormalities, which are generated in our model of prenatal cocaine exposure, display deficits in specific cognitive, motor, and perceptual functions which rely on cerebral cortex as one of their major neural substrates; and 3) We will evaluate whether prenatal cocaine exposure producing cerebral cortical neuropathology in our animals affects normal development of non-social and social behaviors.

DESCRIPTION (provided by applicant): Brain stem descending pathways constitute a major mechanism in the modulation of pain transmission by which attentional, motivational and cognitive variables filter ascending information. Recent studies indicate that hyperalgesia in animal models of persistent pain is linked to enhanced activation of descending inhibitory and facilitatory modulatory circuits. Such mechanisms may play an important role in persistent pain conditions such as fibromyalgia, temporomandibular joint disorders, and irritable bowel syndrome by increasing endogenous facilitation leading to an enhancement and spread of the pain. In earlier studies we focused mainly on the activity-dependent plasticity and dynamic temporal changes occurring in the rostral ventromedial medulla (RVM) within the first 24h after complete Freund's adjuvant (CFA)-induced inflammation. The cellular mechanisms mainly responsible for descending inhibition included excitatory amino acid receptors. Less is known about the effects of chemical mediators that contribute to the initiation and maintenance of descending facilitation and behavioral hyperalgesia after inflammation. Therefore, a major objective of our proposal is to determine the chemical signature of RVM circuits involved in the initiation and maintenance of descending facilitation of inflammatory hyperalgesia. Our major hypothesis is that inflammation leads to an increase in the release of multiple chemical mediators whose chemical signature involves activation of a signal transduction cascade in RVM neurons that couples their activity to subunits of the N-methyl-D-aspartate (NMDA) receptor and a descending serotoninergic pathway. We will test these hypotheses utilizing behavioral, molecular, immunohistochemical and pharmacological approaches in the aims presented below. Aim 1 will test the hypothesis that the balance between descending facilitation and inhibition and their contribution to inflammatory hyperalgesia is dependent, in part, upon the activation of multiple receptors for glutamate, proinflammatory cytokines, brain-derived neurotrophic factor (BDNF) and Substance P. Aim 2 will test the hypothesis that activation of the receptors for these chemical mediators in the RVM involves changes in receptor expression levels and their activation of signal transduction cascades that lead to phosphorylation of NMDA receptor subunits. Aim 3 will test the hypothesis that the time course of effect of these chemical mediators is differentially dependent upon primary afferent drive during the initiation and maintenance phases of behavioral hyperalgesia. Aim 4 will test the hypothesis that descending facilitation produced by BDNF, SP and proinflammatory cytokines is mediated through serotonin-containing descending pathways and spinal 5-HT3 receptors. Finally, Aim 5 will test the hypothesis that these chemical mediators in the RVM contribute to the aversive component of behavioral hyperalgesia via descending circuits originating in the RVM.

DESCRIPTION (provided by applicant): The development of animal models of persistent pain has advanced our knowledge of the initiation of pain hypersensitivity due to sensitization of peripheral nociceptors and neurons in the medullary and spinal dorsal horns as well as other sites in the central nervous system (CNS). It is now commonly proposed that the initiation of this plasticity and resultant pain amplification is dependent upon activation of peripheral nociceptors. It is assumed that peripheral mechanisms are also responsible for the maintenance of the pain hypersensitivity, though the findings in this area are less compelling. Recent findings suggest that behavioral hyperalgesia following injury persists despite a reduction in peripheral neuronal activity suggesting that peripheral drive may be necessary but not sufficient for the maintenance of pain hypersensitivity after injury. The primary aim of this proposal is to study the mechanisms that underlie the maintenance of pain amplification after injury and to determine whether there are CNS mechanisms that participate in pain chronicity and how persistent pain emerges from the more acute stage after injury. Current animal models are not entirely adequate for the study of the chronicity and maintenance of pain hypersensitivity. We have developed two new models in the trigeminal system: a tendomyositis model of the masseter muscle and a neuropathic pain model of the infraorbital nerve. Both models exhibit long-term hyperalgesia/allodynia that is constant and lasts for months, and the inflammation due to the surgical procedure can be separated with the use of long-duration local anesthetics. Our major hypothesis is that tissue and nerve injury in the orofacial region can lead to the maintenance of secondary hyperalgesia that involves the activation CNS descending mechanisms and is less dependent on the peripheral drive associated with the injured target. The following Specific Aims will test these hypotheses using multidisciplinary approaches. #1: Test the hypothesis that the maintenance of long-term hyperalgesia after injury involves an attenuation of peripheral afferent drive and a transition to central mechanisms. #2: Test the hypothesis that the maintenance of long-term hyperalgesia is dependent upon an enhancement of descending facilitatory or a reduction of descending inhibitory inputs. #3: Test the hypothesis that the maintenance of long-term hyperalgesia is dependent upon an enhancement of descending facilitatory drive involving activation of trigeminal non-neural glial cells and their release of cytokines. #4: Test the hypothesis that the maintenance of long-term hyperalgesia is dependent upon an enhancement of descending facilitatory drive that involves shifts in the anionic reversal potential of trigeminal GABA-responsive neurons and a reduction in GABAA-induced inhibitory tone. #5: Test the hypothesis that the maintenance of long-term mechanical hyperalgesia is dependent upon rostral ventromedial medulla activation of a trigeminal brain stem interactive signaling cascade of 5-HT3, GABAA and NMDA receptors as well as glia and cytokines. PUBLIC HEALTH RELEVANCE: The major goal of the proposed research is to study the central nervous system mechanisms that account for the chronicity of persistent orofacial pain after tissue or nerve injury. Although it is now commonly proposed that the initiation of injury-induced neural plasticity and resultant pain amplification is dependent upon activation of peripheral nociceptors, it is unclear whether peripheral mechanisms are also responsible for the maintenance and chronicity of pain hypersensitivity and how persistent pain emerges from the more acute stage after injury. Our findings will lead to a transformative shift in the search for unique treatment approaches for persistent pain.