Chronic pain is costly, emotionally and psychologically debilitating and dispiriting to tens of thousands of people. The Strichartz laboratory is investigating three areas of chronic pain: 1. The role of endothelin-1 (ET-1), and endogenous peptide, in abnormal pain from tissue injury, inflammation and cancer, 2. The mechanisms by which prolonged pain and pain sensitization is caused by nerve growth factor, another endogenous molecule that is released when peripheral tissues are injured, and 3. The changes in the peripheral tissues and in the central nervous system that underlie the development and persistence of chronic post-operative pain.
Endothelin receptor dynamics.
The G-protein coupled receptors for ET-1, an endogenous peptide, appear to be involved in several forms of acute and chronic pain. Our work has shown that injection of ET-1 in the paw or application directly on peripheral nerve causes immediate and transient pain in rats, induces spontaneous action potentials selectively in peripheral nociceptors in vivo, and causes increases in intracellular cellular ,[Ca+2]in , and enhances nociceptor excitability, by favoring the opening of a tetrodotoxin-resistant voltage-gated Na+ channel found exclusively in nociceptors, as well as causing a reduction of the currents carried by delayed-rectifier type K+ channels. All these effects are mediated by the ETA sub-type of receptors for ET-1. In addition, acute ET-1 injection under the skin enhances the amount of both glutamate and CGRP in the epidermis, accounting for another pathway of rapid nociceptor sensitization, and also enhances the expression of the transducing receptor TRPV-1, a molecule that senses noxious heat and the elevated [H+] that accompanies inflammation. All these actions are conducted through ETA receptors.
The ETB receptor sub-type, in contrast, makes two different contributions to pain; ETB receptors on neurons are pro-algesic and sensitize nociceptors to mechanical stimuli, thereby elevating pain behavior, whereas ETB receptors on keratinocytes, in the skin's epidermis, are anti-hyperalgesic, countering the effects of pain stimuli and pain sensitizers. This anti-algesic action occurs by the release of the opioid peptide beta-endorphin from the keratinocytes that synthesize and store it. The eventual likelihood that a nociceptor will generate afferent impulses to signal pain is critically dependent on the dynamic changes wrought by pro-algesic neuronal ETA and ETB receptors and the anti-algesic ETB receptors on keratinocytes, a dynamic which differs among different tissues, e.g. between hairy skin and glabrous, hairless skin, and also which changes after injury and inflammation.
Current work is focused on the role of the ET receptors in skin, and thus in the role of skin as a sensory transducing organ rather than just a tissue thought provides structural support and protection. Our lab has collaborated with the Skin Disease Research Center laboratory of Dr. James Rheinwald to grow basal keratinocytes and to differentiate them and study the effects of ET-1. Such in vitro cultures allow a degree of autonomy and manipulation that is difficult to achieve in vivo, and provides the opportunity to study basic cellular processes in keratinocytes that contribute to peripheral pain processing, by examining gene expression (RT-PCR), cellular physiology (Ca-imaging and whole cell voltage-clamp) and protein expression (Western blot and immuno-cytochemistry). Findings in this system indicate that ETA receptors are expressed primarily by basal, proliferating keratinocytes and ETB receptors by differentiated, suprabasal keratinocytes. ET-1 stimulates these cells to effect an increase in intracellular calcium, and also to stimulate the enzyme adenylyl cyclase and thereby elevate the signaling molecule cyclic AMP. Western blot analysis using antibodies that selectively bind to different parts of the ETB receptor indicate that much of this receptor is cleaved and then both remains on the surface, where it can still bind ET-1, and resides somewhere inside the cell, where it may or may not have some residual function.
Nerve Growth Factor and pain.
In collaboration with the laboratory of Professor Grant Nicol at the University of Indiana Medical center, we have been exploring the pathways that couple the injection of nerve growth factor (NGF) in the skin to the increased sensitivity to tactile and thermal stimulation at the injection site. In support of Dr. Nicol’s observations on isolated sensory neurons in vitro, we have found that much of this pain sensitization from NGF in vivo occurs via the low affinity p75ntr (neurotrophin) receptor, a finding that disputes the commonly held assumption that the high affinity trkA (tyrosine kinase) receptor is responsible for the pain from peripheral NGF. The activation of the p75ntr pathway involves the critical participation of an unusual enzyme called PKMzeta, an enzyme that is controlled by its biosynthesis and degradation rather than by a regulatory sub-unit, as are most other typical PK (protein kinase) enzymes. PKMzeta in the brain (hippocampus) has been shown to be important for memory and so it may have an analogous function in the development of long-lasting hypersensitivity in pain pathways, but through its actions in the periphery, skin, and in the spinal cord.
Mechanisms and Models of Chronic Post-operative Pain
Chronic post-operative pain is an important clinical consequence following many procedures, including thoracotomies, mastectomies, gynecological surgery, obstetrical surgery and most forms of amputation. Our laboratory has been studying this phenbomenon by developing new animal models that mimic surgical procedures. Using the hairy skin of the rat we have found that skin incisions through the back, dorso-lumbar region, cause both primary (near the wound) and secondary (far from the wound) mechanical hypersensitivity, but that these elevated “pain” responses can be prevented by application of local anesthetics that are released slowly from synthetic “microspheres”. Incisions through the medial thigh, followed by an hour-long retraction of the skin and muscle that entraps the saphenous nerve (skin-muscle incision and retraction, SMIR), leads to a profound and long-lasting (4-5 week) tactile hypersensitivity of the ipsilateral plantar paw surface, a striking example of secondary hyperalgesia since this region is not directly innervated by the saphenous nerve. Microscopic examination documents the absence of nerve damage at the retraction site. But investigations of spinal cord segments that process signals from incoming nerves show that a signaling molecule called p38 MAPKinase is phosphorylated, and thus activated, with a time-course that parallels the development of pain after the SMIR procedure. Inhibitors of the p38 activated pathway injected into the spinal fluid next to these segments are able to prevent the pain from developing if given at the time of surgery, but once the pain has developed, a week or two after surgery, these same agents are largely ineffective.
Current research on chronic pain in our laboratory uses a rat model of thoracotomy with rib retraction. Such a procedure results in at least several months of tactile sensitivity, that is spread over much of the back. After surgery the pre-operative “pain” responses, local twitches and quick single withdrawal “reflexes”, are replaced by more complex behavior that indicates a type of processing in the brain of true, perceived pain. We have converted these complex responses to a quantifiable Qualitative Pain Index, shown that it is responsive to the selective analgesic morphine, and that it is diminished in parallel to the reduction in mechano-sensitive area and threshold reduction in rats that have received spinal Resolvins at the time of surgery.
Gary Stirchartz, Ph.D. is Co-Director of the Pain Research Center and is a Professor of Anaesthesia at Harvard Medical School.
- Strichartz GR. “Novel ideas of local anaesthetic actions on various ion channels to ameliorate post-operative pain.” Br J Anaesth 2008; doi: 10.1093/bja/aen101
- Khodorova A, Strichartz G. “Contralateral Paw Sensitization Following Injection of Endothelin-1: Effects of local anesthetics differentiate peripheral and central processes.” Neuroscience 2010;165:553-60. DOI: 10.1016/j.neuroscience.2009.10.049
- Khodorova A, Strichartz G. “Remarkably Long-Lasting Tachyphylaxis of Pain Responses to ET-1: Evidence Against Central Nervous System Involvement.” Can J Physiol Pharmacol 2010 ;88:668-75.
- Gerner P, Wang CF, Lee BS, Suzuki S, Degirolami U, Gandhi A, Knaack D, Strichartz G. “The relationship between functional sciatic nerve block duration and the rate of release of lidocaine from a controlled-release matrix.” Anesth Analg 2010;111:221-9.
- Wang, C-F, Pancaro C, Gerner P, Strichartz, G. “Prolonged suppression of post-incisional pain by a slow-release formulation of lidocaine.” Anesthesiology, 2011; 114:135-49.
- Huang L, Wang C-F, Serhan CN, Strichartz G. “Enduring prevention and transient reduction of post-operative pain by intrathecal Resolvin D1.” Pain 2011;152 : 557–565
- Colvin A, Wang C-F, Soens MA, Mitani AA, Strichartz G, Gerner P. “Prolonged cutaneous analgesia from transdermal penetration of amitriptyline and capsaicin.” Reg Anesth Pain Med 2011; 36: 236-40.
- Montmayeur JP, Barr TP, Kam SA, Packer SJ, Strichartz GR. “Elevation of intracellular calcium in clonal neuronal and embryonic kidney cells involves endogenous endothelin-A receptors linked to phospholipase C through Gαq/11.” Pharmacol Res 2011; 64:258-67.
- Huang L, Gao Y-J, Wang J, Strichartz G. “Shifts in cell-type expression accompany a diminishing role of spinal p38-MAPKinase activation over time during prolonged postoperative pain.” Anesthesiology 2011; 115:1281-90.
- Ohri R, Blaskovich P, Wang J, Pham L, Nichols G, Hildebrand W, Costa D , Scarborough N , Herman C, Strichartz G. “Prolonged nerve block by microencapsulated bupivacaine prevents acute post-operative pain in rats.” Reg Anesth Pain Med 2012; 37:607-615.
- Barreveld AM, Witte J, Chahal H, Durieux ME, Strichartz G. “Preventive analgesia by local anesthetics: The reduction of post-operative pain by peripheral nerve blocks and intravenous drugs.” Anesth Analg (in press)
- Leeson S, Strichartz GR. “Kinetics of Uptake and Washout of Lidocaine in Rat Sciatic Nerve in vitro.” (in press)