Bridget M. Lumb

A Rapamycin-Sensitive Signaling Pathway Is Essential for the Full Expression of Persistent Pain States

Translational control through the mammalian target of rapamycin (mTOR) is critical for synaptic plasticity, cell growth, and axon guidance. Recently, it was also shown that mTOR signaling was essential for the maintenance of the sensitivity of subsets of adult sensory neurons. Here, we show that persistent pain states, but not acute pain behavior, are substantially alleviated by centrally administered rapamycin, an inhibitor of the mTOR pathway. We demonstrate that rapamycin modulates nociception by acting on subsets of primary afferents and superficial dorsal horn neurons to reduce both primary afferent sensitivity and central plasticity. We found that the active form of mTOR is present in a subpopulation of myelinated dorsal root axons, but rarely in unmyelinated C-fibers, and heavily expressed in the dorsal horn by lamina I/III projection neurons that are known to mediate the induction and maintenance of pain states. Intrathecal injections of rapamycin inhibited the activation of downstream targets of mTOR in dorsal horn and dorsal roots and reduced the thermal sensitivity of A-fibers. Moreover, in vitro studies showed that rapamycin increased the electrical activation threshold of Aδ-fibers in dorsal roots. Together, our results imply that central rapamycin reduces neuropathic pain by acting both on an mTOR-positive subset of A-nociceptors and lamina I projection neurons and suggest a new pharmacological route for therapeutic intervention in persistent pain states.

Local Translation in Primary Afferent Fibers Regulates Nociception

Recent studies have demonstrated the importance of local protein synthesis for neuronal plasticity. In particular, local mRNA translation through the mammalian target of rapamycin (mTOR) has been shown to play a key role in regulating dendrite excitability and modulating long-term synaptic plasticity associated with learning and memory. There is also increased evidence to suggest that intact adult mammalian axons have a functional requirement for local protein synthesis in vivo. Here we show that the translational machinery is present in some myelinated sensory fibers and that active mTOR-dependent pathways participate in maintaining the sensitivity of a subpopulation of fast-conducting nociceptors in vivo. Phosphorylated mTOR together with other downstream components of the translational machinery were localized to a subset of myelinated sensory fibers in rat cutaneous tissue. We then showed with electromyographic studies that the mTOR inhibitor rapamycin reduced the sensitivity of a population of myelinated nociceptors known to be important for the increased mechanical sensitivity that follows injury. Behavioural studies confirmed that local treatment with rapamycin significantly attenuated persistent pain that follows tissue injury, but not acute pain. Specifically, we found that rapamycin blunted the heightened response to mechanical stimulation that develops around a site of injury and reduced the long-term mechanical hypersensitivity that follows partial peripheral nerve damage - a widely used model of chronic pain. Our results show that the sensitivity of a subset of sensory fibers is maintained by ongoing mTOR-mediated local protein synthesis and uncover a novel target for the control of long-term pain states.

The representation of prolonged and intense, noxious somatic and visceral stimuli in the ventrolateral orbital cortex of the cat

&NA; The responses of single neurones in the ventrolateral orbital (VLO) cortex to noxious pinch, heating of the skin, twisting of the joints and distension of the gall bladder were studied in cats anaesthetized with halothane. Of 60 neurones studied, 44 responded to prolonged ( > 10 sec) stimuli that were well within the noxious range. Neurones were relatively unresponsive to innocuous stimuli or to the transient application of noxious stimuli. Many single neurones responded to a variety of modalities of noxious stimuli (e.g., skin heating and gall bladder distension). Many neurones studied showed a fluctuating level (5–15 Hz) of ongoing spontaneous activity. Neurones responded with either an increased frequency of spikes (excitation) or an inhibition of spontaneous discharge, irrespective of the source of noxious stimulation. Noxious stimuli delivered simultaneously to two different tissues (e.g., skin and visceral) sometimes produced excitation of the neurone under study, to levels above that produced by the application a noxious stimulus to only one of the tissues. Receptive fields were often large involving both contralateral and ipsilateral areas of the body, as well as both fore and hind limbs. No evidence of somatotopic organization was obtained. The responses of some neurones outlasted the application of the stimuli by many minutes. It is concluded that single neurones in the ventrolateral orbital cortex respond to the prolonged application of intensely noxious stimuli to a variety of body tissues, in a manner that is in keeping with the involvement of this cortical area in both the physiological, autonomic and experiential components of the affective‐motivational aspect of pain. Furthermore, from the consequences of lesion studies in man and animals, it is proposed that the activation of cells in the orbital cortex by a variety of noxious stimuli reflects its more general role in the development and maintenance of behaviour in response to negative reinforcement of both social and physical origins.

Inescapable and Escapable Pain is Represented in Distinct Hypothalamic‐Midbrain Circuits: Specific Roles for aδ‐ and C‐Nociceptors

The affective responses to pain arising from deep somatic and visceral tissues differ markedly from those evoked by brief cutaneous insults. Deep pain evokes passive emotional coping that includes quiescence and vasodepression. In contrast, cutaneous pain evokes an active emotional coping: the fight or flight response. There is now considerable evidence to support the notion that nociceptive inputs arising from different peripheral domains drive the different functional columns of the periaqueductal grey (PAG) that co‐ordinate either active or passive coping strategies. Nociceptive inputs from deep structures drive neurones in the ventrolateral columns that co‐ordinate passive emotional coping whereas brief cutaneous insults activate the dorsolateral/lateral columns that co‐ordinate active coping strategies. An emerging concept, as presented in the preceding article by Keay & Bandler, is that it is the behavioural significance of the nociceptive input, rather than its organ of origin per se, that determines the characteristics of the affective response. These authors provide evidence that brief, escapable stimuli activate neurones in the dorsolateral/lateral columns of the PAG and that inescapable, persistent pain, irrespective of its organ of origin, activates the ventrolateral column. This review will present recent evidence that differential representation of escapable and inescapable pain in the PAG extends to distinct representations of ‘first’ and ‘second’ pain, as indicated by the columnar distribution of neurones activated by inputs from Aδ‐ and C‐nociceptors. Furthermore, the functional organisation of projections from circumscribed regions of the hypothalamus to the different columns of the PAG indicates that the behavioural significance of the pain signal is represented in brain regions other than the PAG.

Systemic inhibition of the mammalian target of rapamycin (mTOR) pathway reduces neuropathic pain in mice

Summary Systemic inhibition of mTORC1 with rapalog CCI‐779 or a ATP‐competitive inhibitor Torin1 alleviates the mechanical hypersensitivity associated with inflammation and neuropathy without damaging sensory neurons. ABSTRACT The management of neuropathic pain is unsatisfactory, and new treatments are required. Because the sensitivity of a subset of fast‐conducting primary afferent nociceptors is thought to be regulated by the mammalian target of rapamycin complex 1 (mTORC1) signaling pathway, selectively targeting mTORC1 represents a new strategy for the control of chronic pain. Here we show that activated mTOR was expressed largely in myelinated sensory fibers in mouse and that inhibiting the mTORC1 pathway systemically alleviated mechanical hypersensitivity in mouse models of inflammatory and neuropathic pain. Specifically, systemic administration of mTORC1 inhibitor temsirolimus (CCI‐779), both acutely (25 mg/kg i.p.) and chronically (4 daily 25 mg/kg i.p.), inhibited the mTORC1 pathway in sensory axons and the spinal dorsal horn and reduced mechanical and cold hypersensitivity induced by nerve injury. Moreover, systemic treatment with CCI‐779 also reduced mechanical but not heat hypersensitivity in an inflammatory pain state. This treatment did not influence nociceptive thresholds in naive or sham‐treated control animals. Also, there was no evidence for neuronal toxicity after repeated systemic treatment with CCI‐779. Additionally, we show that acute and chronic i.p. administration of Torin1 (20 mg/kg), a novel ATP‐competitive inhibitor targeting both mTORC1 and mTORC2 pathways, reduced the response to mechanical and cold stimuli in neuropathic mice. Our findings emphasize the importance of the mTORC1 pathway as a regulator of nociceptor sensitivity and therefore as a potential target for therapeutic intervention, particularly in chronic pain.

Optoactivation of Locus Ceruleus Neurons Evokes Bidirectional Changes in Thermal Nociception in Rats

Pontospinal noradrenergic neurons are thought to form part of a descending endogenous analgesic system that exerts inhibitory influences on spinal nociception. Using optogenetic targeting, we tested the hypothesis that excitation of the locus ceruleus (LC) is antinociceptive. We transduced rat LC neurons by direct injection of a lentiviral vector expressing channelrhodopsin2 under the control of the PRS promoter. Subsequent optoactivation of the LC evoked repeatable, robust, antinociceptive (+4.7°C ± 1.0, p < 0.0001) or pronociceptive (−4.4°C ± 0.7, p < 0.0001) changes in hindpaw thermal withdrawal thresholds. Post hoc anatomical characterization of the distribution of transduced somata referenced against the position of the optical fiber and subsequent further functional analysis showed that antinociceptive actions were evoked from a distinct, ventral subpopulation of LC neurons. Therefore, the LC is capable of exerting potent, discrete, bidirectional influences on thermal nociception that are produced by specific subpopulations of noradrenergic neurons. This reflects an underlying functional heterogeneity of the influence of the LC on the processing of nociceptive information.

A reliable method for the preferential activation of C- or A-fibre heat nociceptors

There is strong evidence that A- and C-fibre nociceptors evoke significantly different sensory experiences, are differentially sensitive to pharmacological intervention, and play different roles in pain pathology. It is therefore of considerable interest to be able to selectively activate one fibre type or the other in studies of nociceptive processing. Here, we report significant modifications to a non-invasive technique, first described by Yeomans et al. [Pain 59 (1994) 85; Pain 68 (1996) 141; Pain 68 (1996) 133], which uses different rates of skin heating to preferentially activate A- or C-nociceptors. A copper disk (diameter: 4mm) was used to transfer heat evenly across the dorsal surface of the rat hindpaw. Initial experiments established the relationship between the temperature at the skin surface and the sub-epidermal temperature. Subsequently, the vanilloid capsaicin, which sensitises unmyelinated C-mechanoheat nociceptors, was shown to decrease the thresholds of reflex responses evoked by slow rates of heating. In contrast thresholds of responses to fast rates of skin heating were unchanged, indicating that nociceptors activated by this stimulus were capsaicin-insensitive A-fibre heat nociceptors.

Descending control of spinal nociception from the periaqueductal grey distinguishes between neurons with and without C-fibre inputs

&NA; Information about noxious events in the periphery is conveyed to the spinal cord in A‐ and C‐fibre nociceptive afferents, which have largely distinct electrical and chemical properties and which convey different qualities of the pain signal. Descending control that originates in the different functional columns of the midbrain periaqueductal grey (PAG) has important roles in the modulation of spinal nociception in different behavioural and emotional states and, it is now believed, in animal models of chronic pain. However, few studies of descending control have considered differential modulation of A‐ versus C‐nociceptor‐evoked responses. Here, we report that descending inhibitory control from the rostrocaudal extent of the dorsolateral/lateral and ventrolateral columns of the PAG preferentially targets Class 2 deep dorsal horn neurons with C‐fibre inputs. Pinch‐evoked responses of these neurons were depressed significantly by −37 ± 4.2% (P < 0.0001). In contrast, the pinch‐evoked responses of Class 2 neurons without C‐fibre inputs (presumably A‐fibre mediated) were enhanced significantly by +34 ± 11.8% (P < 0.01). Further experiments indicated these facilitatory effects were at least partly due to a reduction in C‐fibre‐mediated segmental inhibition. We suggest this differential control of spinal nociception would be appropriate in many of the varied situations in which the PAG is believed to become active, whether short term (e.g. fight or flight) or long term (e.g. chronic pain). Additionally, the pro‐nociceptive effects observed in a subset of spinal neurons may be related to the descending facilitation that has been reported in animal models of chronic pain.

Cyclooxygenase-1-Derived Prostaglandins in the Periaqueductal Gray Differentially Control C- versus A-Fiber-Evoked Spinal Nociception

Nonsteroidal anti-inflammatory drugs (NSAIDs) exert analgesic effects by inhibiting peripheral cyclooxygenases (COXs). It is now clear that these drugs also have central actions that include the modulation of descending control of spinal nociception from the midbrain periaqueductal gray (PAG). Descending control is a powerful determinant of the pain experience and is thus a potential target for analgesic drugs, including COX inhibitors. Noxious information from the periphery is conveyed to the spinal cord in A- and C-fiber nociceptors, which convey different qualities of the pain signal and have different roles in chronic pain. This in vivo study used different rates of skin heating to preferentially activate A- or C-heat nociceptors to further investigate the actions of COX inhibitors and prostaglandins in the PAG on spinal nociceptive processing. The results significantly advance our understanding of the central mechanisms underlying the actions of NSAIDs and prostaglandins by demonstrating that (1) in the PAG, it is COX-1 and not COX-2 that is responsible for acute antinociceptive effects of NSAIDs in vivo; (2) these effects are only evoked from the opioid-sensitive ventrolateral PAG; and (3) prostaglandins in the PAG exert tonic facilitatory control that targets C- rather than A-fiber-mediated spinal nociception. This selectivity of control is of particular significance given the distinct roles of A- and C-nociceptors in acute and chronic pain. Thus, effects of centrally acting prostaglandins are pivotal, we suggest, to both the understanding of nociceptive processing and the development of new analgesic drugs.

Endogenous analgesic action of the pontospinal noradrenergic system spatially restricts and temporally delays the progression of neuropathic pain following tibial nerve injury

Summary A role for the pontospinal noradrenergic system to dynamically restrict the spatiotemporal expression of the neuropathic pain phenotype in a nerve injury model. ABSTRACT Pontospinal noradrenergic neurons form part of an endogenous analgesic system that suppresses acute pain, but there is conflicting evidence about its role in neuropathic pain. We investigated the chronology of descending noradrenergic control during the development of a neuropathic pain phenotype in rats following tibial nerve transection (TNT). A lumbar intrathecal cannula was implanted at the time of nerve injury allowing administration of selective &agr;‐adrenoceptor (&agr;‐AR) antagonists to sequentially assay their effects upon the expression of allodynia and hyperalgesia. Following TNT animals progressively developed mechanical and cold allodynia (by day 10) and subsequently heat hypersensitivity (day 17). Blockade of &agr;2‐AR with intrathecal yohimbine (30 &mgr;g) revealed earlier ipsilateral sensitization of all modalities while prazosin (30 &mgr;g, &agr;1‐AR) was without effect. Established allodynia (by day 21) was partly reversed by the re‐uptake inhibitor reboxetine (5 &mgr;g, i.t.) but yohimbine no longer had any sensitising effect. This loss of effect coincided with a reduction in the descending noradrenergic innervation of the ipsilateral lumbar dorsal horn. Yohimbine reversibly unmasked contralateral hindlimb allodynia and hyperalgesia of all modalities and increased dorsal horn c‐fos expression to an innocuous brush stimulus. Contralateral thermal hyperalgesia was also reversibly uncovered by yohimbine administration in a contact heat ramp paradigm in anaesthetised TNT rats. Following TNT there is an engagement of inhibitory &agr;2‐AR‐mediated noradrenergic tone which completely masks contralateral and transiently suppresses the development of ipsilateral sensitization. This endogenous analgesic system plays a key role in shaping the spatial and temporal expression of the neuropathic pain phenotype after nerve injury.