Involvement of PKA-dependent upregulation of nNOS-CGRP in adrenomedullin-initiated mechanistic pathway underlying CFA-induced response in rats

We have previously shown that intrathecal administration of the adrenomedullin (AM) receptor antagonist AM22-52 produces a long-lasting anti-hyperalgesia effect. This study examined the hypothesis that AM recruits other pronociceptive mediators in complete Freund's adjuvant (CFA)-induced inflammation. Injection of CFA in the hindpaw of rat produced an increase in the expression of nNOS in dorsal root ganglion (DRG) and the spinal dorsal horn. An intrathecal administration of AM22-52, but not the CGRP antagonist BIBN4096BS, abolished the CFA-induced increase of nNOS. Moreover, AM-induced increase of CGRP was inhibited by the nNOS inhibitors L-NAME and 7-nitroindazole in cultured ganglion explants. Addition of AM to ganglion cultures induced an increase in nNOS protein, which was attenuated by the PKA inhibitor H-89. Treatment with AM also concentration-dependently increased cAMP content and pPKA protein level, but not its non-phosphorylated form, in cultured ganglia. In addition, nNOS was shown to be co-localized with the AM receptor components calcitonin receptor-like receptor and receptor activity-modifying protein 2- and 3 in DRG neurons. The present study suggests that the enhanced activity of nitric oxide (NO) mediates the biological action of AM at the spinal level and that AM recruits NO-CGRP via cAMP/PKA signaling in a mechanistic pathway underlying CFA-induced hyperalgesia.

► Intrathecal adrenomedullin/AM receptor antagonist AM22-52 abolished CFA-induced nNOS. ► AM-induced increase of CGRP was inhibited by the nNOS inhibitors. ► AM induced an increase in nNOS protein in ganglion cultures. ► AM induced increase in nNOS was attenuated by PKA inhibitor H-89. ► AM increased cAMP and pPKA, but not its non-phosphorylated form, in cultured ganglia.