Involvement of adrenomedullin in spinal glial activation following chronic administration of morphine in rats.

Adrenomedullin (AM) belongs to the calcitonin gene-related peptide (CGRP) family. Our previous studies show that chronic exposure to morphine increases spinal AM bioactivity, contributing to the development and maintenance of morphine tolerance. This study investigated the possible involvement of AM in morphine-evoked gliosis. Real-time polymerase chain reaction was performed to determine interleukin-1β (IL-1β), IL-6 and tumour necrosis factor-α (TNF-α) mRNAs in the spinal dorsal horn and cultured sensory ganglion explants. Immunohistochemistry was performed to identify spinal microglia and astrocytes. Repetitive intrathecal (i.t.) injection of morphine (20 μg) increased the expression of IL-1β, IL-6 and TNF-α mRNAs in the spinal dorsal horn. The co-administration of the selective AM receptor antagonist AM22-52 (36 μg) markedly attenuated chronic morphine-evoked increase in IL-1β and IL-6, but not TNF-α, mRNA levels. Exposure of cultured dorsal root ganglion (DRG) explants to morphine (3.3 μmol/L) for 6 days up-regulated IL-1β and IL-6 mRNA expressions. The depletion of AM gene using small interfering RNA (siRNA) approach abolished morphine-evoked increase in IL-1β and IL-6 syntheses in the cultured DRG. The blockade of AM receptors by i.t. AM22-52 also inhibited chronic morphine-evoked cell hypertrophy of microglia and astrocytes as well as an increase in OX-42 and GFAP (glial fibrillary acidic protein) immunoreactivities. Furthermore, the 6-day treatment with AM (10 μg, i.t.) induced morphological changes of microglia and astrocytes as well as an increase in IL-1β, IL-6 and TNF-α mRNA levels in the spinal dorsal horn. The present study supports the idea that up-regulation of the pronociceptive mediator AM can recruit spinal glial cells, resulting in an increase in cytokines during chronic use of morphine.