Activation of dorsal horn cannabinoid CB2 receptor suppresses the expression of P2Y12 and P2Y13 receptors in neuropathic pain rats

Abstract

BackgroundMore evidence suggests that dorsal spinal cord microglia is an important site contributing to CB2 receptor-mediated analgesia. The upregulation of P2Y12 and P2Y13 purinoceptors in spinal dorsal horn microglia is involved in the development of pain behavior caused by peripheral nerve injury. However, it is not known whether the expression of P2Y12 and P2Y13 receptors at spinal dorsal horn will be influenced after CB2 receptor activation in neuropathic pain rats.MethodsChronic constriction injury (CCI) and intrathecal ADPbetaS injection were performed in rats to induce neuropathic pain. The paw withdrawal latency (PWL) was used to evaluate thermal hyperalgesia in neuropathic rats. The expression of P2Y12 and P2Y13 receptors, p-p38MAPK, and NF-kappaBp65 was detected with RT-PCR and western blotting analysis.ResultsTreatment with AM1241 produces a pronounced inhibition of CCI-induced thermal hyperalgesia and significantly inhibited the increased expression of P2Y12 and P2Y13 receptors at the mRNA and protein levels, which open up the possibility that P2Y12 and P2Y13 receptor expression are downregulated by CB2 receptor agonist AM1241 in CCI rats. Western blot analysis demonstrated that AM1241 reduced the elevated expression of p-p38MAPK and NF-κBp65 in the dorsal spinal cord induced by CCI. After administration with either SB203580 (p38MAPK inhibitor) or PDTC (NF-kappaB inhibitor), the levels of P2Y13 receptor expression in the dorsal spinal cord were lower than those in the CCI group. However, in CCI rats, the increased expression of P2Y12 receptor was prevented by intrathecal administration of PDTC but not by SB203580. In addition, minocycline significantly decreased the increased expression of P2Y12 and P2Y13 receptors. The similar results can be observed in ADPbetaS-treated rats. Intrathecal injection of ADPbataS causes thermal hyperalgesia and increased expression of P2Y12 and P2Y13 receptors in the dorsal spinal cord of naive rats. Moreover, intrathecal injection of AM1241 alleviates pain response and reduces the elevated expression of P2Y12 and P2Y13 receptors, p-p38MAPK, and NF-κBp65 in the dorsal spinal cord of ADPbetaS-treated rats. Intrathecal injection of SB203580 significantly inhibited the ADPbetaS-induced P2Y13 receptor expression, without affecting P2Y12 receptor expression. However, treatment with either SB203580 or PDTC effectively inhibited P2Y13 receptor expression compared to ADPbetaS-treated rats.ConclusionsIn CCI- and ADPbetaS-treated rats, AM1241 pretreatment could efficiently activate CB2 receptor, while inhibiting p38MAPK and NF-kappaB activation in the dorsal spinal cord. CB2 receptor stimulation decreased P2Y13 receptor expression via p38MAPK/NF-kappaB signaling. On the other hand, CB2 receptor activation decreased P2Y12 receptor expression via p38MAPK-independent NF-kappaB signaling pathway.

P2Y12 deficiency in mouse impairs noradrenergic system in brain, and alters anxiety‐like neurobehavior and memory

“…Therefore, it is possible that shortened travel distance in central area of P2Y 12 KO mice might be caused by reduced overall locomotor activities (see Figure S1, Supporting Information). Moreover, as overwhelming evidences indicate an important role for P2Y 12 in neuropathic pain, we chose more than one mild test, respectively, to measure the performance of the mice, but we could not completely exclude the possibility that the sensory impairments could affect behavior of KO animals in this study.…”

P2Y13 receptors regulate microglial morphology, surveillance, and resting levels of interleukin 1β release

“…Haynes et al () and Stefani et al () found that, despite confirming P2Y 13 mRNA expression in microglia (Zhang et al, ), its expression could not be detected at the protein level. The inability to antibody‐label P2Y 13 may reflect a high rate of constitutive proteasomal degradation (Pons et al, ), or may alternatively suggest that P2Y 13 protein is only expressed at high levels in microglial culture (Quintas, Vale, Goncalves, & Queiroz, ) or upon microglial activation in situ (Niu et al, ).…”

“…However, P2Y 13 protein expression may be low (Haynes et al, 2006), possibly due to constitutive ubiquitination and proteasomal degradation (Pons et al, 2014). P2Y 13 mRNA is upregulated in pathological conditions such as demyelination (Bedard, Tremblay, Chernomoretz, & Vallieres, 2007) and neuropathic pain (Kobayashi, Yamanaka, Yanamoto, Okubo, & Noguchi, 2012;Niu et al, 2017), suggesting a role of this receptor in neuroinflammation. However, its function in microglia, especially under physiological conditions, is still unclear, although it may evoke a rise of [Ca 2+ ] i in response to ADP (Zeng et al, 2014) and be involved in the release of proinflammatory cytokines (Liu et al, 2017).…”

Spontaneous, anecdotal, retrospective, open-label study on the efficacy, safety and tolerability of cannabis galenical preparation (Bedrocan)

“…Cannabis contains more than 60 endogenous and exogenous compounds defined as cannabinoids (CBs) that act primarily through specific cannabinoid receptors: CB 1 and CB 2 receptors. The CB 1 are distributed in the central nervous system and involved in learning, memory and cognitive processes, while the CB 2 receptors are located in the peripheral nervous system, including brainstem, cerebellum, microglia and the immune system . In the past decade, preclinical studies and case reports have shown the therapeutic potential of two main CBs, delta‐9‐tetrahydrocannabinol acid (Δ 9 ‐THCA) and cannabidiol (CBD) against a range of pathologies .…”

“…The CB 1 are distributed in the central nervous system and involved in learning, memory and cognitive processes, while the CB 2 receptors are located in the peripheral nervous system, including brainstem, cerebellum, microglia and the immune system. [8][9][10][11][12][13][14][15] In the past decade, preclinical studies and case reports have shown the therapeutic potential of two main CBs, delta-9tetrahydrocannabinol acid (D 9 -THCA) and cannabidiol (CBD) against a range of pathologies. [16,17] D 9 -THCA is the main constituent in raw cannabis, it converts to D 9 -THC when heated over a certain temperature, binding at both CB receptors.…”