Intradermal injection of capsaicin induces acute substance P release from rat spinal cord dorsal horn.

Increased release of substance P (SP) from the dorsal horn following noxious stimuli, such as spinal administration of capsaicin, has been demonstrated in previous studies. However, changes in the release of SP in response to intradermal injection of capsaicin still remain unknown. This study was designed to demonstrate in vivo spinal SP release following intradermal injection of capsaicin (3%, 50 microl), using polyimide tubing with a single hole introduced into the rat dorsal horn. The changes in the content of SP in the rat dorsal horn tissues before and after capsaicin (3%, 50 microl) injection were also investigated. The SP concentration in the samples was analyzed using an enzyme-linked immunosorbent assay (ELISA). We found that intradermal injection of capsaicin induced a quick SP release within the dorsal horn. The peak of the release appeared around 10 min after the injection. In contrast, intradermal injection of capsaicin had no significant effect on the SP content in the dorsal horn. This study has provided direct evidence of the effect of intradermal injection of capsaicin on SP release within the dorsal horn, with the major source being from the central terminals of primary afferents.

Activation of protein kinase B/Akt signaling pathway contributes to mechanical hypersensitivity induced by capsaicin.

We investigated the involvement of the protein kinase B/Akt (PKB/Akt) signaling pathway in the mechanical hypersensitivity induced in rats by capsaicin. Intradermal injection of capsaicin results in activation of PKB/Akt in the lumbar spinal cord, most prominently in the dorsal horn, starting by 5 min after capsaicin injection and lasting at least 1h. The activated PKB/Akt in the spinal cord is in neurons, since phospho-PKB/Akt (p-PKB/Akt) colocalizes with the neuronal marker, neuronal-specific nuclear protein (NeuN). The mechanical hypersensitivity is shown by the enhanced paw withdrawal frequency to applications of von Frey filaments with different bending forces (30, 100, 200 mN) on the rat paw. Pre-treatment with several different PKB/Akt inhibitors, including SH-6, Akt inhibitor IV, and Akt inhibitor V, blocked the mechanical hypersensitivity induced by intradermal injection of capsaicin, a measure of spinal cord central sensitization. Two structurally unrelated phosphoinositide 3-Kinase (PI3K, upstream of PKB/Akt) inhibitors, Wortmannin and LY294002, also prevented the mechanical hypersensitivity induced by intradermal injection of capsaicin. Furthermore, post-treatment with the PI3K inhibitor, Wortmannin, or PKB/Akt inhibitors, such as NL-71-101, SH-6, Akt inhibitor IV, and inhibitor V significantly reduced the established mechanical hypersensitivity induced by capsaicin. The PKB/Akt signaling pathway in the spinal cord is therefore involved in pain hypersensitivity.

Calcitonin gene-related peptide receptor activation produces PKA- and PKC-dependent mechanical hyperalgesia and central sensitization.

Calcitonin gene-related peptide (CGRP), acting through CGRP receptors, produces behavioral signs of mechanical hyperalgesia in rats and sensitization of wide dynamic range (WDR) neurons in the spinal cord dorsal horn. Although involvement of CGRP receptors in central sensitization has been confirmed, the second-messenger systems activated by CGRP receptor stimulation and involved in pain transmission are not clear. This study tested whether the hyperalgesia and sensitizing effects of CGRP receptor activation on WDR neurons are mediated by protein kinase A or C (PKA or PKC) signaling. Intrathecal injection of CGRP in rats produced mechanical hyperalgesia, as shown by paw withdrawal threshold tests. CGRP-induced hyperalgesia was attenuated significantly by the CGRP1 receptor antagonist, CGRP8-37. The effect was also attenuated significantly by a PKA inhibitor (H89) or a PKC inhibitor (chelerythrine chloride). Electrophysiological experiments demonstrated that superfusion of the spinal cord with CGRP-induced sensitization of spinal dorsal horn neurons. The CGRP effect could be blocked by CGRP8-37. Either a PKA or PKC inhibitor (H89 or chelerythrine) also attenuated this effect of CGRP. These results are consistent with the hypothesis that CGRP produces hyperalgesia by a direct action on CGRP1 receptors in the spinal cord dorsal horn and suggest that the effects of CGRP are mediated by both PKA and PKC second-messenger pathways.