Ryan Patel

α2δ-1 Gene Deletion Affects Somatosensory Neuron Function and Delays Mechanical Hypersensitivity in Response to Peripheral Nerve Damage

The α2δ-1 subunit of voltage-gated calcium channels is upregulated after sensory nerve injury and is also the therapeutic target of gabapentinoid drugs. It is therefore likely to play a key role in the development of neuropathic pain. In this study, we have examined mice in which α2δ-1 gene expression is disrupted, to determine whether α2δ-1 is involved in various modalities of nociception, and for the development of behavioral hypersensitivity after partial sciatic nerve ligation (PSNL). We find that naive α2δ-1−/− mice show a marked behavioral deficit in mechanical and cold sensitivity, but no change in thermal nociception threshold. The lower mechanical sensitivity is mirrored by a reduced in vivo electrophysiological response of dorsal horn wide dynamic range neurons. The CaV2.2 level is reduced in brain and spinal cord synaptosomes from α2δ-1−/− mice, and α2δ-1−/− DRG neurons exhibit lower calcium channel current density. Furthermore, a significantly smaller number of DRG neurons respond to the TRPM8 agonist menthol. After PSNL, α2δ-1−/− mice show delayed mechanical hypersensitivity, which only develops at 11 d after surgery, whereas in wild-type littermates it is maximal at the earliest time point measured (3 d). There is no compensatory upregulation of α2δ-2 or α2δ-3 after PSNL in α2δ-1−/− mice, and other transcripts, including neuropeptide Y and activating transcription factor-3, are upregulated normally. Furthermore, the ability of pregabalin to alleviate mechanical hypersensitivity is lost in PSNL α2δ-1−/− mice. Thus, α2δ-1 is essential for rapid development of mechanical hypersensitivity in a nerve injury model of neuropathic pain.

Mechanisms of the gabapentinoids and α2δ‐1 calcium channel subunit in neuropathic pain

The gabapentinoid drugs gabapentin and pregabalin are key front‐line therapies for various neuropathies of peripheral and central origin. Originally designed as analogs of GABA, the gabapentinoids bind to the α2δ‐1 and α2δ‐2 auxiliary subunits of calcium channels, though only the former has been implicated in the development of neuropathy in animal models. Transgenic approaches also identify α2δ‐1 as key in mediating the analgesic effects of gabapentinoids, however the precise molecular mechanisms remain unclear. Here we review the current understanding of the pathophysiological role of the α2δ‐1 subunit, the mechanisms of analgesic action of gabapentinoid drugs and implications for efficacy in the clinic. Despite widespread use, the number needed to treat for gabapentin and pregabalin averages from 3 to 8 across neuropathies. The failure to treat large numbers of patients adequately necessitates a novel approach to treatment selection. Stratifying patients by sensory profiles may imply common underlying mechanisms, and a greater understanding of these mechanisms could lead to more direct targeting of gabapentinoids.

Diffuse noxious inhibitory controls and nerve injury: restoring an imbalance between descending monoamine inhibitions and facilitations.

Abstract Diffuse noxious inhibitory controls (DNICs) utilize descending inhibitory controls through poorly understood brain stem pathways. The human counterpart, conditioned pain modulation, is reduced in patients with neuropathy aligned with animal data showing a loss of descending inhibitory noradrenaline controls together with a gain of 5-HT3 receptor-mediated facilitations after neuropathy. We investigated the pharmacological basis of DNIC and whether it can be restored after neuropathy. Deep dorsal horn neurons were activated by von Frey filaments applied to the hind paw, and DNIC was induced by a pinch applied to the ear in isoflurane-anaesthetized animals. Spinal nerve ligation was the model of neuropathy. Diffuse noxious inhibitory control was present in control rats but abolished after neuropathy. &agr;2 adrenoceptor mechanisms underlie DNIC because the antagonists, yohimbine and atipamezole, markedly attenuated this descending inhibition. We restored DNIC in spinal nerve ligated animals by blocking 5-HT3 descending facilitations with the antagonist ondansetron or by enhancing norepinephrine modulation through the use of reboxetine (a norepinephrine reuptake inhibitor, NRI) or tapentadol (&mgr;-opioid receptor agonist and NRI). Additionally, ondansetron enhanced DNIC in normal animals. Diffuse noxious inhibitory controls are reduced after peripheral nerve injury illustrating the central impact of neuropathy, leading to an imbalance in descending excitations and inhibitions. Underlying noradrenergic mechanisms explain the relationship between conditioned pain modulation and the use of tapentadol and duloxetine (a serotonin, NRI) in patients. We suggest that pharmacological strategies through manipulation of the monoamine system could be used to enhance DNIC in patients by blocking descending facilitations with ondansetron or enhancing norepinephrine inhibitions, so possibly reducing chronic pain.

Novel TRPM8 Antagonist Attenuates Cold Hypersensitivity after Peripheral Nerve Injury in Rats

Abnormal cold sensitivity is a common feature of a range of neuropathies. In the murine somatosensory system, multiple aspects of cold sensitivity are dependent on TRPM8, both short term and in response to peripheral nerve injury. The specialized nature of cold-sensitive afferents and the restricted expression of TRPM8 render it an attractive target for the treatment of cold hypersensitivity. This current study examines the effect of a novel TRPM8 antagonist (M8-An) in naive and spinal nerve-ligated rats through behavioral and in vivo electrophysiological approaches. In vitro, M8-An inhibited icilin-evoked Ca2+ currents in HEK293 cells stably expressing human TRPM8 with an IC50 of 10.9 nM. In vivo, systemic M8-An transiently decreased core body temperature. Deep dorsal horn recordings were made in vivo from neurons innervating the hind paw. M8-An inhibited neuronal responses to innocuous and noxious cooling of the receptive field in spinal nerve-ligated rats but not in naive rats. No effect on neuronal responses to mechanical and heat stimulation was observed. In addition, M8-An also attenuated behavioral responses to cold but not mechanical stimulation after nerve ligation without affecting the uninjured contralateral response. The data presented here support a contribution of TRPM8 to the pathophysiology of cold hypersensitivity in this model and highlight the potential of the pharmacological block of TRPM8 in alleviating the associated symptoms.

Anti-hyperalgesic effects of a novel TRPM8 agonist in neuropathic rats: A comparison with topical menthol

&NA; An in vivo electrophysiological characterisation of the pro‐ and anti‐nociceptive effects of TRPM8 agonists reveals menthol and a novel TRPM8 agonist have selective inhibitory effects on cold sensitivity in neuropathic rats. &NA; Menthol has historically been used topically to alleviate various pain conditions. At low concentrations, this non‐selective TRPM8 agonist elicits a cooling sensation, however higher concentrations result in cold hyperalgesia in normal subjects and paradoxically analgesia in neuropathic patients. Through behavioural and electrophysiological means, we examined whether this back‐translated into a pre‐clinical rodent model. Menthol was applied topically to the hind paws of naive and spinal nerve‐ligated (SNL) rats. In behavioural assays, menthol did not affect withdrawal thresholds to mechanical stimulation and 10% and 40% menthol rarely sensitised withdrawals to innocuous cooling in naïve rats. However, in SNL rats, 10% and 40% menthol alleviated cold hypersensitivity. This was partly corroborated by in vivo electrophysiological recordings of dorsal horn lamina V/VI neurones. As several studies have implicated TRPM8 in analgesia, we examined whether a novel systemically available TRPM8 agonist, M8‐Ag, had more potent anti‐hyperalgesic effects than menthol in neuropathic rats. In vitro, M8‐Ag activates TRPM8, expressed in HEK293 cells, with an EC50 of 44.97 nM. In vivo, M8‐Ag inhibited neuronal responses to innocuous and noxious cooling in SNL rats with no effect in sham‐operated rats. This effect was modality selective; M8‐Ag did not alter neuronal responses to mechanical, heat or brush stimulation. In addition, M8‐Ag attenuated behavioural hypersensitivity to innocuous cooling but not mechanical stimulation. These data suggest that menthol induced hyperalgesia is not consistently replicable in the rat and that the analgesic properties are revealed by injury. Systemic TRPM8 agonists might be beneficial in neuropathy without affecting normal cold sensitivity.

Neuronal hyperexcitability in the ventral posterior thalamus of neuropathic rats: modality selective effects of pregabalin

Studies on brain mechanisms of neuropathic pain are lacking. This study characterizes the properties of rat ventral posterior thalamic wide dynamic range (WDR) and nociceptive-specific (NS) neurons, the latter of which are uncharacterized in a neuropathic state. We provide evidence of phenotypic changes in neuronal sensitivity that may underlie cold and brush hypersensitivity, and that WDR neurons, and not NS neurons, encode hypersensitivity to low-intensity stimuli. Pregabalin reversed neuronal hyperexcitability in spinal nerve-ligated rats in a modality-selective manner.

Calcium channel modulation as a target in chronic pain control

Neuropathic pain remains poorly treated for large numbers of patients, and little progress has been made in developing novel classes of analgesics. To redress this issue, ziconotide (Prialt™) was developed and approved as a first‐in‐class synthetic version of ω‐conotoxin MVIIA, a peptide blocker of Cav2.2 channels. Unfortunately, the impracticalities of intrathecal delivery, low therapeutic index and severe neurological side effects associated with ziconotide have restricted its use to exceptional circumstances. Ziconotide exhibits no state or use‐dependent block of Cav2.2 channels; activation state‐dependent blockers were hypothesized to circumvent the side effects of state‐independent blockers by selectively targeting high‐frequency firing of nociceptive neurones in chronic pain states, thus alleviating aberrant pain but not affecting normal sensory transduction. Unfortunately, numerous drugs, including state‐dependent calcium channel blockers, have displayed efficacy in preclinical models but have subsequently been disappointing in clinical trials. In recent years, it has become more widely acknowledged that trans‐aetiological sensory profiles exist amongst chronic pain patients and may indicate similar underlying mechanisms and drug sensitivities. Heterogeneity amongst patients, a reliance on stimulus‐evoked endpoints in preclinical studies and a failure to utilize translatable endpoints, all are likely to have contributed to negative clinical trial results. We provide an overview of how electrophysiological and operant‐based assays provide insight into sensory and affective aspects of pain in animal models and how these may relate to chronic pain patients in order to improve the bench‐to‐bedside translation of calcium channel modulators.

An investigation into the inhibitory function of serotonin in diffuse noxious inhibitory controls in the neuropathic rat

Following neuropathy α2‐adrenoceptor‐mediated diffuse noxious inhibitory controls (DNIC), whereby a noxious conditioning stimulus inhibits the activity of spinal wide dynamic range (WDR) neurons, are abolished, and spinal 5‐HT7 receptor densities are increased. Here, we manipulate spinal 5‐HT content in spinal nerve ligated (SNL) animals and investigate which 5‐HT receptor mediated actions predominate.

Spinal neuronal correlates of tapentadol analgesia in cancer pain: A back-translational approach

Pain is a common and highly debilitating complication for cancer patients significantly compromising their quality of life. Cancer‐induced bone pain involves a complex interplay of multiple mechanisms including both inflammatory and neuropathic processes and also some unique changes. Strong opioids are a mainstay of treatments but side effects are problematic and can compromise optimal pain control. Tapentadol is a novel dual‐action drug, both stimulating inhibitory μ‐opioid receptors (MOR) and mediating noradrenaline reuptake inhibition (NRI) leading to activation of the inhibitory α‐2 adrenoceptor. It has been demonstrated to treat effectively both acute and chronic pain. We here demonstrate the efficacy in a model of cancer‐induced bone pain.

Genes, molecules and patients--emerging topics to guide clinical pain research.

This review selectively explores some areas of pain research that, until recently, have been poorly understood. We have chosen four topics that relate to clinical pain and we discuss the underlying mechanisms and related pathophysiologies contributing to these pain states. A key issue in pain medicine involves crucial events and mediators that contribute to normal and abnormal pain signaling, but remain unseen without genetic, biomarker or imaging analysis. Here we consider how the altered genetic make-up of familial pains reveals the human importance of channels discovered by preclinical research, followed by the contribution of receptors as stimulus transducers in cold sensing and cold pain. Finally we review recent data on the neuro-immune interactions in chronic pain and the potential targets for treatment in cancer-induced bone pain.