Inflammatory processes in peripheral tissues lead to central sensitization in the spinal cord, which contributes to hyperalgesia and allodynia typically associated with inflammatory pain. Although evidence suggests that plastic changes in the spinal dorsal horn account for central sensitization, the relative contribution of pre- and postsynaptic mechanisms and of peripheral and supraspinal factors are not entirely clear. The superficial dorsal horn of the spinal cord, particularly substantia gelatinosa (SG), is a major projection site of small-diameter afferent nerve fibers that predominantly transmit nociceptive signals [1
]. SG neurons also receive descending inputs from the brainstem [1
]. Therefore, in addition to intraspinal neuroplastic changes, peripheral as well as supraspinal factors may contribute to central sensitization.
Pain-related neuroplastic changes in central nervous system (CNS) structures can be shown definitively by the electrophysiological analysis of synaptic transmission and neuronal excitability in spinal cord or brain slice preparations obtained from animals in which an experimental pain state has been induced [4
]. The slice preparation allows the analysis of pain-related plasticity because it is disconnected from the site of peripheral injury (inflammation) and from other CNS areas, be it supraspinal sites (spinal cord slice) or spinal cord (brain slices). Therefore, changes measured in the slice preparation are maintained independently of continuous inputs to the area of interest. Accordingly, changes of synaptic circuitry in SG neurons were shown in slices from animals with complete Freund's adjuvant induced hindpaw inflammation [4
] and synaptic plasticity was demonstrated in amygdala neurons from animals with knee joint arthritis [7
The kaolin and carrageenan (K/C) induced knee joint arthritis is a well established model of inflammatory pain. Electrophysiological, pharmacological, neurochemical and behavioral studies have used this model to analyze pain mechanisms at different levels of the nervous system and showed the sensitization of primary afferent nerve fibers, spinal dorsal horn neurons and neurons in the central nucleus of the amygdala (CeA) [12
]. Using slice preparations, synaptic plasticity was demonstrated in the CeA, but not yet in the spinal cord, in the K/C arthritis pain model.
The purpose of this study was to compare synaptic transmission and neuronal excitability in SG neurons in spinal cord slices from normal and from arthritic animals using patch-clamp recordings. Another goal was to determine the role of calcitonin gene-related peptide (CGRP) in pain-related spinal plasticity since CGRP has emerged as an important molecule at different levels of the pain neuraxis in the arthritis pain model.
CGRP is a 37 amino acid peptide that activates adenylyl cyclase and protein kinase A through G-protein-coupled receptors, including the CGRP1 receptor for which selective antagonists are available [18
]. CGRP is involved in peripheral and spinal pain mechanisms [22
]. We showed recently that CGRP also plays an important role in the transmission of nociceptive information to the amygdala through the spino-parabrachio-amygdaloid pathway [10
The source of CGRP in the spinal cord dorsal horn is primary afferents. CGRP coexists with substance P in small-diameter afferent fibers, and CGRP containing terminals and CGRP receptors are found in the dorsal horn, including SG [30
]. CGRP is released in the spinal dorsal horn by noxious stimulation and peripheral inflammation such as the K/C arthritis [26
]. Peripheral inflammation also leads to changes in CGRP binding sites in the dorsal horn [32
Spinal application of CGRP facilitates nociceptive behavior [24
] and sensitizes the responses of dorsal horn neurons to innocuous and noxious peripheral stimulation [28
] and to intraspinally administered excitatory amino acids [23
] and substance P [39
]. In a slice preparation, CGRP produced a slow depolarization and enhanced excitability of dorsal horn neurons; the effect on evoked synaptic transmission was not studied [40
]. Conversely, block of spinal CGRP receptors with an antagonist (CGRP8-37) or antiserum induced antinociception in animal models of inflammatory [25
] or central neuropathic pain [45
]. CGRP8-37 also inhibited the responses of spinal dorsal horn neurons to transdermial electrical stimulation of the hindpaw [46
] and to noxious mechanical stimulation of the knee joint [29
]. CGRP8-37 prevented or reversed central sensitization of dorsal horn neurons in the arthritis and capsaicin pain models [28
]. Arthritic CGRP knockout mice showed reduced nociceptive behavioral responses [47
Although it is widely accepted that CGRP plays an important role in the modulation of spinal nociceptive processing, the cellular mechanisms and pre- or post-synaptic sites of action through which CGRP contributes to central sensitization remain to be determined. The present study addressed the role of CGRP in synaptic plasticity in the superficial dorsal horn in vitro in a model of arthritic pain induced in vivo. Our data show for the first time synaptic plasticity and increased excitability of SG neurons in the K/C arthritis pain model. A CGRP receptor antagonist inhibits synaptic plasticity whereas CGRP itself facilitates synaptic transmission through a postsynaptic mechanism that involves direct membrane effects on SG neurons.