PKC-Dependent Signaling Pathways within PAG and Thalamus Contribute to the Nitric Oxide-Induced Nociceptive Behavior

Nitric oxide (NO) is an important molecule involved in nociceptive processing in the central nervous system. The release of NO within the spinal cord has long been implicated in the mechanisms underlying exaggerated pain sensitivity, and administration of NO donors can induce hyperalgesia. To elucidate the supraspinal mechanism responsible for NO-induced nociceptive hypersensitivity, we investigated the modulation of protein kinase C (PKC) and downstream effectors following treatment with the NO donors nitroglycerin and sodium nitroprusside. Both compounds induced a prolonged cold allodynia and heat hyperalgesia, increased levels of c-Fos and IL-1β, and activated NF-κB within periaqueductal grey matter and thalamus. Simultaneously, an increased expression and phosphorylation of PKC γ and ε were detected. To clarify the cellular mechanism involved in the NO-induced hypernociception, we examined the expression of transcription factors that act as PKC downstream effectors. A dramatic hyperphosphorylation of CREB and STAT1 was observed. The i.c.v. administration of the PKC blocker calphostin C prevented the NO-induced hypernociception, the hyperphosphorylation of CREB and STAT1, and partially reduced NF-κB activation. Conversely, the increase of IL-1β was unmodified by calphostin C. These results suggest the relevance of cerebral PKC-mediated CREB and STAT1 activation in the NO donor-induced nociceptive behavior. 1. Introduction Nitric oxide (NO) is a molecule importantly involved in pain processing within central nervous system. The release of NO within the spinal cord has long been implicated in the mechanisms underlying exaggerated pain sensitivity [1]. A reduced nociceptive behavior was observed in inflammatory and neuropathic pain after intrathecal administration of NO synthase (NOS) inhibitors [2, 3] and after genetic deletion of the NOS isoforms [4–6]. In addition, NO donors induced thermal [7] and mechanical [8] hyperalgesia. Studies on the possible mechanism of action of NO and subsequent steps in the NO-induced cascade show that NO signals by various mechanisms including cGMP synthesis, nitrosylation of ion channels, ADP-ribosylation, and the interaction with molecular oxygen and superoxide radicals to produce reactive nitrogen species that can modify a number of macromolecules [9]. A major signaling mechanism of NO in spinal nociceptive processing initiates with activation of NO sensitive guanylyl cyclase and subsequent cGMP production [10, 11]. It has been speculated that activation of cGMP-dependent protein kinase I (cGKI), in turn, is the