David C. Yeomans - US grants

Perhaps the most basic division in the pain system is between sensory receptors with myelinated (Asigma) and those with unmyelinated (C) afferent fibers. Pain evoked by the activation of these two nociceptor types feels different and the two nociceptor types have been implicated in different kinds of clinical pain. Previous work demonstrated that these two nociceptor types are differentially activated by using different rates of noxious radiant heating of the hindpaw of the rat. Using a behavioral analgesiometric model based on this distinction, the previous supported work also demonstrated clear differences in the effects of analgesic treatments, such as opiates, on nociceptive responses evoked by these two nociceptor types. Work proposed in this application would extend this study by examining and comparing the neurochemistry and neuropharmacology of the two nociceptor types at the first synapse as they convey information to the central nervous system. This will be accomplished by measuring the release of putative neurotransmitters from presynaptic terminals and subsequent activation of neurotransmitter receptors on postsynaptic cells as evoked by selective activation of either Asigma or C fiber nociceptors. In this way, the differential use of neurotransmitters by different nociceptor types can be determined. In addition, serotonergic, noradrenergic, and opioid agents will be applied to determine whether part of the nociceptor-selective analgesic effects of these agents may be mediated through inhibition of the release of neurotransmitters from presynaptic spinal terminals of the nociceptors. These studies should provide important information as the mechanisms underlying the differential modulation of Asigma and C fiber mediated nociception. Since these two types of nociceptors probably mediate different kinds of clinical pain, a better understanding of their basic properties should lead to better targeting of analgesic drugs for specific types of pain, reducing the need for broad analgesic approaches, such as systemic application of opiates.

Over the last several years, the scientific literature has been replete with numerous new approaches to the treatment of pain. Unfortunately, approaches that were promising in the laboratory do not always translate to better clinical treatments. Similarly, the underlying physiology and clinical relevance of some animal pain models used to evaluate these new treatments have not been clearly demonstrated. One of the reasons for these disparities may have to do with the fact that some analgesic treatments appear to differentially attenuate only some kinds of nociception. Some standard tests then, may miss the potential utility of a treatment for a pain type that is not modeled by those tests. One distinction that we have found to be important in determining antinociceptive capacity of treatments is whether, and to what extent a nociceptive response is mediated by the activation of unmyelinated or myelinated primary afferent nociceptors. Another reason that results of rodent tests do not always predict clinical outcomes and vice versa may be the phylogenetic distance between humans and rodents. It is likely that testing potential treatments using non-human primates models might be more predictive of clincial utility in some cases. Thus, there is a need for the development of a primate assay which allows for the differential assessement of nociception mediated by myelinated or unmyelinated nociceptors. The purpose of the work outlined in this proposal is to provide preliminary evidence that the principles determined in rodents with regards to differential testing of Adelta and C fiber mediated nociception are also applicable to the testing of non-human primates. The experiments described here make use of demonstrated differences in the sensitivity of nociception mediated by myelinated or unmyelinated nociceptors to different rates of skin heating, topical application of the neurotoxin capsaicin, and the systemic application of the opiate analgesic morphine. The goal of this work is to develop a primate nociceptive assay which will allow for better predictions of the clinical utility of novel approaches to the treatment of pain.

DESCRIPTION (provided by applicant): Despite many years of research, the mainstays of clinical treatment of pain remain highly problematic. High dose opiate analgesics have numerous side effect, induce tolerance and dependence and have significant abuse potential. New knowledge and technologies, however, may allow for entirely new approaches to the management of chronic pain. It has recently been established that there are 12 or more different isoforms of neuronal sodium channels, which are critical to these neurons' ability to convey information. One isoform in particular, Nav1.7, appears to be selectively present in neurons that carry pain information from the periphery to the central nervous system. Furthermore, levels of these channels, as well as their location on these neurons, change dramatically during chronic pain states, implying that they are important in establishing and/or maintaining these pain states. Thus, an examination of Nav1.7 channel function may be significant in designing new approaches to the treatment of chronic pain. In order to establish the importance of this sodium channel in pain, virally directed gene transfer will be used to decrease or eliminate expression of the gene for Nav1.7 selectively in those neurons that carry pain information. Herpes viral vectors have been previously used to insert novel transgenes into these neurons, and to decrease synthesis of endogenous proteins, but this approach has never been applied to sodium channels. Thus, a herpes virus encoding an antisense sequence for the Nav1.7 channel gene will be used to knock-down production of this channel, so that immunochemical, behavioral, and electrophysiological experiments can be conducted in order to examine the contribution of Nav1.7 in pain biology. In this way, it should become clear whether or not this channel represents a potential target for the development of new pharmaceutical tools. Another implication of this work, however, is that this approach may in itself provide a means of therapy, and it may be possible to use herpes vectors to inhibit production of endogenous Nav1.7 channels in chronic pain patients, providing a long-term genetic therapy for their pain.

DESCRIPTION (provided by applicant): Head cancer pain, trigeminal neuralgia, migraine headache, dental pain and temporomandibular joint pain are all examples of pain syndromes that are unique to the trigeminal system. In many instances, these pain types are hard to treat clinically, with last-line opiates working only marginally well in some instances, and not at all in others. In addition, the tolerance and addiction potential of strong, systemic opioids sometimes limit the duration over which they can be used effectively. Thus, there is a need for novel approaches to the treatment of trigeminal pain. We have previously demonstrated the potential of using replication-defective herpes viral constructs to alter the function of pain-sensing nerve cells, such that we have been able to produce robust, highly localized analgesia for months after a single application. In doing this we have applied the virus locally to targeted tissues, such as skin. Doing so, we have observed a very long-lasting (> 20 weeks) attenuation of pain responses limited to those areas treated with the virus. In many trigeminal syndromes however, pain is relatively diffuse or multicentered. Applications of vectors to peripheral tissues may be of limited utility in these cases, as a more widely distributed analgesic effect is desirable. One method that has not yet been investigated would be to inject vectors directly into the trigeminal ganglia, the grouping of neurons that make up the cell bodies of the sensory nerves of the trigeminal. In doing this, we would expect to introduce recombinant vectors over a wide distribution of trigeminal neurons, and thus, potentially, producing a widespread analgesic effect. The experiments described here will provide evidence as to whether direct trigeminal injection of recombinant herpes vectors, encoding genes for analgesic peptides, will alter the sensitivity to nociceptive stimulation of tissue innervated by the trigeminal. In so doing, we hope to provide initial support for what may be a new long-lasting treatment for trigeminal nerve-related pain.

DESCRIPTION (provided by applicant): One of the main determinants of the temperature at which C or Adelta thermonociceptors are excited appears to be the rate at which the peripheral terminals of nociceptors are heated. The relationship between rate and threshold temperature is not simple however, and is likely modulated by other direct factors, such as intensity and duration of stimulation, and surface area stimulated, as well as numerous indirect factors, including body, skin and air temperature, depth of the receptor in the skin, density of the skin, and specific characteristics of the heat source (thermode, normal radiant, various kinds of lasers). Perhaps the greatest advance in investigating these issues has been the development of standardized laser stimulation. However, there are great differences in the types of laser used, which can complicate comparisons between laboratories. We have chosen to investigate the use of a diode infrared laser, which should give optimal heating characteristics, in terms of direct heating of nociceptors in the skin, and which should inherently produce an extremely stable output. In addition, we will use a thermal camera to precisely measure temperature changes in the skin with an extremely precise temporal accuracy, so that the very rapid changes that can occur with lasers can be measured. Finally, we will combine these technologies with single unit peripheral nerve recordings in rats. In this way, we hope to determine optimal laser heating protocol for selective activation of either C or Adelta thermonociceptors. Based on our preliminary work, as well as our examination of the literature, we hypothesize that very rapid rise times with this laser will allow for selective activation of Adelta nociceptors, whereas lower rise time stimuli will selectively excite C nociceptors. Thus, we hope to show that temperatures of activation are not absolute, but rather are modulated by heating rate (dictated by laser power) and duration, and size of area stimulated. We also intend to use the basic data gleaned from these studies to generate predictive modeling of temperature response functions at different depths of skin In this way, we hope, with this R21 early stage project, to develop a method which will allow future studies in terms of nociceptimetric testing in rats, examining the biomolecular mechanisms underlying selective nociceptor activation, and finally in the examination of central nervous system consequences, in humans, of specific activation of these different nociceptor types.

DESCRIPTION (provided by applicant): Trigeminal neuralgia (tic douloureux) is a devistatingly painful craniofacial disease, characterized by intense stabbing pains of the face which may be elicited by light touch. In the majority of cases, the pain can be traced to chronic compression of the trigeminal nerve roots by a blood vessel, leading to demyelination of the compressed roots. These compression and myelination changes may then alter Na+ channel density along the nerve root, possibly leading to ectopic discharges, and abnormal fiber-to-fiber transmission of impulses. Although some Na+ channel blocking anti-seizure drugs are helpful in the short-term, many patients end up having invasive neurosurgery to decompress the nerve root. Even surgery however, typically only provides temporary relief. A primary issue limiting the development of new, more effective therapies, has been the lack of an animal model which accurately simulates trigeminal neuralgia. One of the two main goals of RFA-DE-07-006 "New Models of Pain Relevant to the Trigeminal System" is to "stimulate the development and utilization of novel animal models of chronic orofacial pain conditions. As an adjunct to these two goals, this Initiative also encourages the development of novel measures of pain in patients and animals that are non-invasive and objective and that permit a behavioral or functional assessment of pain." The goal of the proposed research fits closely with this goal, in that we plan to develop an accurate, predictive animal model of trigeminal neuralgia. Our preliminary work has shown that transcranial placement of a biocompatible superabsorbant polymer next to the trigeminal nerve root pushes the root against the bone, providing a means of non-inflammatory chronic compression, simulating human pathology. The technique is relatively non-invasive and simple to perform. Furthermore, these experiments have shown that these animals demonstrate behavioral and electrophysiological changes that are highly consistent with the symptoms and signs of trigeminal neuralgia. We propose to further characterize this model, providing clear endpoints to test potential therapies and a means to examine mechanisms underlying trigeminal neuralgia. These mechanistic studies, in turn, can lead to the determination of new targets for pharmacdceutical development, as well as for gene therapy approaches.