P2Y12 regulates microglia activation and excitatory synaptic transmission in spinal lamina II neurons during neuropathic pain in rodents

Abstract

Peripheral nerve injury causes neuropathic pain and microglia activation. P2Y12 receptors on microglia are thought to be a key player in the surveillance of the local environment, but whether or how these receptors are engaged in the cross-talk between microglia and neurons of the dorsal horn remain ambiguous. Using a rodent model of nerve injury-induced pain, we investigated the roles of P2Y12 in microglia activation, excitatory synaptic transmission, and nociceptive allodynia. We found that spinal nerve ligation (SNL) significantly increased the level of P2Y12 receptors specifically in the microglia of the ipsilateral dorsal horn. Injections of P2Y12 antagonists (MRS2395 or clopidogrel) attenuated microglia activation and increased the paw withdrawal latency in response to thermal stimuli on the ipsilateral side without affecting the basal threshold on the contralateral side. These effects on pain behaviors were replicated in P2Y12 knockout mice. Patch-clamp recordings further revealed that partial sciatic nerve ligation (PSNL)-induced excessive miniature excitatory postsynaptic currents (mEPSCs) were significantly attenuated in P2Y12 knockout mice. Moreover, we found that SNL activates the GTP-RhoA/ROCK2 signaling pathway and elevates the level of phosphorylated p38 mitogen-activated protein kinase (MAPK), which was inhibited by the P2Y12 antagonist. The phosphorylation of p38 MAPK was inhibited by a ROCK inhibitor, but not vice versa, suggesting that p38 MAPK is downstream of ROCK activation. Our findings suggest that nerve injury engages the P2Y12 receptor-dependent GTP-RhoA/ROCK2 signaling pathway to upregulate excitatory synaptic transmission in the dorsal horn. This cross-talk ultimately participates in the manifestation of nociceptive allodynia, implicating P2Y12 receptor as a potential target for alleviating neuropathic pain.

Inhibition of Microglial Activation in the Amygdala Reverses Stress-Induced Abdominal Pain in the Male Rat

“…Our findings showed that chronic stress increased phospho-p38 MAPK expression in Iba-1–positive cells, which is consistent with previous reports in the spinal cord after injury or chronic stress. 18 , 47 Our double immunofluorescence labelling showed that phospho-p38 also is expressed in neurons but not in astrocytes. Although we are not able to identify which cell type initially activated p38 MAPK in our model, one possibility is that after chronic stress, neuronal phospho-p38 expression is triggered by activated microglia.…”

Temporal Changes of Spinal Transcriptomic Profiles in Mice With Spinal Nerve Ligation

“…In addition, the BP analysis showed that the hub genes involved in synapse organization, excitatory postsynaptic potential, modulation of chemical synaptic transmission, and synaptic transmission. Nerve injury upregulated excitatory synaptic transmission in the dorsal horn, and then ultimately leaded to NP (Yu et al, 2019). The positive allosteric modulator (adenosine A1 receptor) selectively suppressed primary afferent synaptic transmission and thereby relieved syndromes in a NP model (Imlach et al, 2015).…”

Ca2+-dependent recruitment of voltage-gated sodium channels underlies bilirubin-induced overexcitation and neurotoxicity

“…Samples containing target protein that were resolved on SDS–PAGE were transferred onto polyvinylidene difluoride membranes (Millipore, Billerica, MA, USA) as previously described 36 . The membranes were blocked with 5% nonfat milk in Tris-buffered saline supplemented with Tween (TBST) for 2 h at room temperature.…”