Behavioural and neuronal correlates of central pain processing in a rat model of osteoarthritis

Osteoarthritis (OA) of the knee join is a chronic condition characterized by the loss of articular cartilage around the joint leading to changes in joint capsule. Clinically, OA is manifested by joint stiffness, swelling, bone tenderness, discomfort upon movement and joint pain. The latter is the main reason why patients seek medical treatment. OA pain is often classified as nociceptive due to tissue damage leading to inflammation. However, a subgroup of OA patients exhibits pain with neuropathic pain-like features. Thus, it is important to understand the differences in pain processing between these two groups of patients, as it will have implications on treatment. 2mg of monosodium iodoacetate (MIA) was intra-articularly injected into the left knee of male Sprague-Dawley rats in order to study behavioural and electrophysiological changes that appear during the development of OA pain. The MIA model provides two distinct stages of OA pain. An early acute inflammatory stage (2-4 days after MIA injection) and a late stage that presents neuropathic pain-like features (14-21 days post MIA injection). Cartilage damage scores differ between the early and late stage MIA animals with the latter exhibiting a more severe maximal OA score. Behaviourally, paw withdrawal thresholds decrease in the acute inflammatory stage and returned almost back to baseline in the late stage. Weight bearing deficits are present in both stages suggesting that both groups exhibit on going pain. CaV2.2 is present in the pre-synaptic terminals of primary afferent fibers in the spinal dorsal horn and mediates neurotransmitter release. ω-conotoxin GVIA is a small peptide that acts as a state independent blocker of CaV2.2, while TROX-1 is a state dependent blocker of the channel. In-vivo electrophysiological recordings of WDR neurones revealed that ω–conotoxin was able to significantly inhibit neuronal evoked responses to electrical, dynamic brush, mechanical and thermal stimuli in the late stage MIA animals but had little or no effects in the early stage MIA animals. State dependent blocker TROX-1, had no effects in neuronal responses of WDR neurones, in any group. Additionally, in-vivo electrophysiological revealed an increased descending serotonergic drive in a subgroup of late stage MIA animals. Additionally, Descending noxious inhibitory controls (DNIC) disappeared in this stage of the model as observed after neuropathy. Suggesting that in the late stage descending modulation is altered

Neuropathy following spinal nerve injury shares features with the irritable nociceptor phenotype: A back-translational study of oxcarbazepine

BACKGROUND: The term 'irritable nociceptor' was coined to describe neuropathic patients characterised by evoked hypersensitivity and preservation of primary afferent fibres. Oxcarbazepine is largely ineffectual in an overall patient population but has clear efficacy in a sub-group with the irritable nociceptor profile. We examine whether neuropathy in rats induced by spinal nerve injury shares overlapping pharmacological sensitives with the irritable nociceptor phenotype using drugs that target sodium channels. METHODS: In vivo electrophysiology was performed in anaesthetised spinal nerve ligated (SNL) and sham-operated rats to record from wide dynamic range (WDR) neurones in the ventral posterolateral thalamus (VPL) and dorsal horn. RESULTS: In neuropathic rats, spontaneous activity in the VPL was substantially attenuated by spinal lidocaine, an effect that was absent in sham rats. The former measure was in part dependent on ongoing peripheral activity as intraplantar lidocaine also reduced aberrant spontaneous thalamic firing. Systemic oxcarbazepine had no effect on wind-up of dorsal horn neurones in sham and SNL rats. However, in SNL rats, oxcarbazepine markedly inhibited punctate mechanical, dynamic brush and cold-evoked neuronal responses in the VPL and dorsal horn, with minimal effects on heat-evoked responses. In addition, oxcarbazepine inhibited spontaneous activity in the VPL. Intraplantar injection of the active metabolite licarbazepine replicated the effects of systemic oxcarbazepine supporting a peripheral locus of action. CONCLUSIONS: We provide evidence that ongoing activity in primary afferent fibres drives spontaneous thalamic firing after spinal nerve injury, and that oxcarbazepine through a peripheral mechanism exhibits modality-selective inhibitory effects on sensory neuronal processing. This article is protected by copyright. All rights reserved

α‑Actinin Promotes Surface Localization and Current Density of the Ca2+ Channel CaV1.2 by Binding to the IQ Region of the α1 Subunit

The voltage-gated L-type Ca2+ channel CaV1.2 is crucial for initiating heartbeat and control of a number of neuronal functions such as neuronal excitability and long-term potentiation. Mutations of CaV1.2 subunits result in serious health problems, including arrhythmia, autism spectrum disorders, immunodeficiency, and hypoglycemia. Thus, precise control of CaV1.2 surface expression and localization is essential. We previously reported that α-actinin associates and colocalizes with neuronal CaV1.2 channels and that shRNA-mediated depletion of α-actinin significantly reduces localization of endogenous CaV1.2 in dendritic spines in hippocampal neurons. Here we investigated the hypothesis that direct binding of α-actinin to CaV1.2 supports its surface expression. Using two-hybrid screens and pull-down assays, we identified three point mutations (K1647A, Y1649A, and I1654A) in the central, pore-forming α11.2 subunit of CaV1.2 that individually impaired α-actinin binding. Surface biotinylation and flow cytometry assays revealed that CaV1.2 channels composed of the corresponding α-actinin-binding-deficient mutants result in a 35-40% reduction in surface expression compared to that of wild-type channels. Moreover, the mutant CaV1.2 channels expressed in HEK293 cells exhibit a 60-75% decrease in current density. The larger decrease in current density as compared to surface expression imparted by these α11.2 subunit mutations hints at the possibility that α-actinin not only stabilizes surface localization of CaV1.2 but also augments its ion conducting activity

α‑Actinin Promotes Surface Localization and Current Density of the Ca²⁺ Channel Ca_V1.2 by Binding to the IQ Region of the α1 Subunit

The voltage-gated L-type Ca²⁺ channel Ca_V1.2 is crucial for initiating heartbeat and control of a number of neuronal functions such as neuronal excitability and long-term potentiation. Mutations of Ca_V1.2 subunits result in serious health problems, including arrhythmia, autism spectrum disorders, immunodeficiency, and hypoglycemia. Thus, precise control of Ca_V1.2 surface expression and localization is essential. We previously reported that α-actinin associates and colocalizes with neuronal Ca_V1.2 channels and that shRNA-mediated depletion of α-actinin significantly reduces localization of endogenous Ca_V1.2 in dendritic spines in hippocampal neurons. Here we investigated the hypothesis that direct binding of α-actinin to Ca_V1.2 supports its surface expression. Using two-hybrid screens and pull-down assays, we identified three point mutations (K1647A, Y1649A, and I1654A) in the central, pore-forming α₁1.2 subunit of Ca_V1.2 that individually impaired α-actinin binding. Surface biotinylation and flow cytometry assays revealed that Ca_V1.2 channels composed of the corresponding α-actinin-binding-deficient mutants result in a 35–40% reduction in surface expression compared to that of wild-type channels. Moreover, the mutant Ca_V1.2 channels expressed in HEK293 cells exhibit a 60–75% decrease in current density. The larger decrease in current density as compared to surface expression imparted by these α₁1.2 subunit mutations hints at the possibility that α-actinin not only stabilizes surface localization of Ca_V1.2 but also augments its ion conducting activity