Aberrant axo-axonic synaptic reorganization in the phosphorylated L1-CAM/calcium channel subunit alpha2 delta-1-containing central terminals of injured c-fibers in the spinal cord of a neuropathic pain model

Abstract

In the dorsal horn of the spinal cord, peripheral nerve injury induces structural and neurochemical alterations through which aberrant synaptic signals contribute to the formation of neuropathic pain. However, the role of injured primary afferent terminals in such plastic changes remain unclear. In this study, we investigated the effect of nerve injury on the morphology of cell adhesion molecule L1-CAM (total L1-CAM [tL1-CAM])-positive primary afferent terminals and on the synaptic contact pattern in the dorsal horn. In the confocal images, the tL1-CAM-positive terminals showed morphological changes leading to the formation of hypertrophic varicosities in the c-fiber terminal. These hypertrophic varicosities in the dorsal horn were co-labeled with phosphorylated (Ser1181) L1-CAM (pL1-CAM) and shown to store neurotransmitter peptides, but not when co-labeled with the pre-synaptic marker, synaptophysin. Quantitative analyses based on three-dimensional reconstructed confocal images revealed that peripheral nerve injury reduced dendritic synaptic contacts but promoted aberrant axo-axonic contacts on the tL1-CAM-positive hypertrophic varicosities. These tL1-CAM-positive varicosities co-expressed the injury-induced alpha2delta-1 (α2δ-1) subunit of the calcium channel in the dorsal horn. Administration of the anti-allodynic drug, pregabalin, inhibited accumulation of α2δ-1 and pL1-CAM associated with a reduction in hypertrophic changes of tL1-CAM-positive varicosities, and normalized injury-induced alterations in synaptic contacts in the dorsal horn. Our findings highlight the formation of aberrant spinal circuits that mediate the convergence of local neuronal signals onto injured c-fibers, suggesting that these hypertrophic varicosities may be important contributors to the pathological mechanisms underlying neuropathic pain.

Significance Statement We describe, for the first time, morphological changes in L1-CAM-positive injured c-fiber terminals that lead to the formation of hypertrophic varicosities, in which we found phosphorylation of L1-CAM and increased expression of the calcium channel subunit α2δ-1. Moreover, we found alterations in synaptic contacts and discovered that peripheral nerve injury increased axo-axonic contacts onto L1-CAM-positive varicosities while decreasing axo-dendritic contacts. These plastic changes indicate the convergence of neuronal signals onto the pain pathway and may represent a pathological mechanism underlying neuropathic pain. Administration of the anti-allodynic drug pregabalin reversed the injury-induced synaptic alternations. These data highlight the unique role of injured c-fibers in peripheral nerve injury and their role as a possible therapeutic target for neuropathic pain.