Background & Aims
Chronic low back pain (CLBP) is a major unmet clinical need with significant socioeconomic impact, and a large contributor to disability[1,2]. The endocannabinoid system (ECS) is involved in the modulation of nociception, and research has revealed significant alterations in the endocannabinoid system at a genetic level in patients with CLBP[3,4]. Intervertebral disc (IVD) degeneration is a common cause of low back pain and radiating leg pain[5]. The prefrontal cortex (PFC) is a crucial region for top-down modulation of nociceptive processing[6,7]. Here, we investigate the cellular and synaptic properties of layer 5 (L5) pyramidal neurons of the PFC in a novel rodent model of IVD injury (IVDI) in the rat tail, which is associated with a pain phenotype[8]. This work will help elucidate the role of the ECS in this region and is key to understanding the complex interplay of neocortical pain processes in developing CLBP.
Methods
20 male 12-week-old Sprague-Dawley rats underwent IVDI at the base-of-the-tail (1mm diameter punch of the nucleus pulposus of the IVD at Co4-Co5 and Co5-Co6), or sham surgery (n=10/group). Mechanical (electronic von Frey-eVF) and heat (Hargreaves-HG) hypersensitivity were assessed at the base-of-the-tail 48/72hrs, respectively, and weekly thereafter, until post-surgery day (PSD)20. Following euthanasia on PSD21, levels of endocannabinoids/N-acylethanolamines (ECBs/NAEs) were analysed in spinal cord tissue and plasma via LC-MS/MS. The acute ex-vivo brain slice preparation was used to investigate cellular and synaptic properties of layer 5 (L5) pyramidal neurons PFC neurons via patch-clamp electrophysiology.
Time-course data were analysed by two-way RM-ANOVA, followed by post-hoc Tukey’s test. Groups were tested by Student’s unpaired or paired t-tests. Association of ECB alterations, electrophysiological data and pain behaviour were analysed by Pearson correlations.
Results
IVDI rats displayed hypersensitivity to radiant heat and mechanical stimuli, compared to sham rats, from PSD3 and PSD7, respectively, which was sustained until euthanasia on PSD21. Levels of plasma 2-AG were lower in IVDI rats compared to shams on PSD21. Plasma 2-AG levels correlated positively with PSD19/20 eVF/HG withdrawal thresholds, while HG withdrawal thresholds from PSD3 onwards (AUC) correlated negatively with NAE levels in the lumbar spinal cord. L5 pyramids were more excitable in the IVDI model due to an increased input resistance compared to sham. Depolarisation-induced suppression of excitation (DSE), a phenomenon dependent on presynaptic activation of the cannabinoid receptor-1, was present in both conditions, but significantly enhanced in IVDI animals in terms of amplitude and duration. DSE amplitude correlated positively with plasma 2-AG levels after 10s depolarisation, while eVF and HG withdrawal thresholds correlated positively after 5s and 10s depolarisations.
Conclusions
Circulating and spinal ECB/NAE levels were associated with alterations in nociceptive behaviour in this model of low back pain. Endocannabinoid-mediated DSE in L5 pyramidal neurons was potentiated in this pain model. Furthermore, an association of DSE amplitude with circulating 2-AG levels and with nociceptive behaviour was determined. Therefore, we have established IVDI-associated alterations in the ECS, and demonstrated the role of the ECS in synaptic activity in the PFC, a region implicated in the emotional processing of pain. The thorough investigation of the synaptic circuitry of nociception holds the potential to uncover novel strategies for significantly improved therapeutic management of pain.
Source of Financial Support: Funding provided by the Irish Research Council Postgraduate Scholarships (GOIPG/2020/1496).
References
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Presenting Author
Mary Hopkins
Poster Authors
Topics
- Models: Transition to Chronic Pain