Evidence that intrathecal morphine-3-glucuronide may cause pain enhancement via toll-like receptor 4/MD-2 and interleukin-1β

Abstract

Morphine-3-glucoronide (M3G) is a major morphine metabolite detected in cerebrospinal fluid of humans receiving systemic morphine. M3G has little-to-no affinity for opioid receptors and induces pain by unknown mechanisms. The pain-enhancing effects of M3G have been proposed to significantly and progressively oppose morphine analgesia as metabolism ensues. We have recently documented that morphine activates toll-like receptor-4 (TLR4), beyond its classical actions on μ-opioid receptors. This suggests that M3G may similarly activate TLR4. This activation could provide a novel mechanism for M3G-mediated pain enhancement, as (a) TLR4 is predominantly expressed by microglia in spinal cord and (b) TLR4 activation releases pain-enhancing substances, including interleukin-1 (IL1). We present in vitro evidence that M3G activates TLR4, an effect blocked by TLR4 inhibitors, and that M3G activates microglia to produce IL1. In vivo, intrathecal M3G (0.75 μg) induced potent allodynia and hyperalgesia, blocked or reversed by interleukin-1 receptor antagonist, minocycline (microglial inhibitor), and (+)-and (−)-naloxone. This latter study extends our prior demonstrations that TLR4 signaling is inhibited by naloxone nonstereoselectively. These results with (+)-and (−)-naloxone also demonstrate that the effects cannot be accounted for by actions at classical, stereoselective opioid receptors. Hyperalgesia (allodynia was not tested) and in vitro M3G-induced TLR4 signaling were both blocked by 17-DMAG, an inhibitor of heat shock protein 90 (HSP90) that can contribute to TLR4 signaling. Providing further evidence of proinflammatory activation, M3G upregulated TLR4 and CD11b (microglial/macrophage activation marker) mRNAs in dorsal spinal cord as well as IL1 protein in the lumbosacral cerebrospinal fluid. Finally, in silico and in vivo data support that the glucuronic acid moiety is capable of inducing TLR4/MD-2 activation and enhanced pain. These data provide the first evidence for a TLR4 and IL1 mediated component to M3G-induced effects, likely of at least microglial origin.

Glial and Neuroimmune Mechanisms as Critical Modulators of Drug Use and Abuse

“…For instance, the morphine metabolite morphine-3-glucuronide exhibits strong affinity for the TLR4 receptor, while morphine-6-glucuronide exhibits no TLR4 activation (Lewis et al, 2010;Wang et al, 2012a). Alternative synthetic opioids would produce different metabolites, which could be an important factor when considering similarities and differences in behavioral outcomes to these opioid drugs.…”

Exploring the Neuroimmunopharmacology of Opioids: An Integrative Review of Mechanisms of Central Immune Signaling and Their Implications for Opioid Analgesia

“…The converse, however, is also true; for example, the modification of the functional group at position 3, often by conjugation, results in a profound loss of opioid receptor activity (e.g., morphine-3-glucuronide) (Chen et al, 1991). However, although this loss of opioid function may signify the end of this molecule's direct opioid receptor activity, it is apparent that metabolites such as morphine-3-glucuronide are capable of having long-term indirect consequences for opioid analgesia and nociceptive hypersensitivity that involve aspects of central immune signaling (Komatsu et al, 2009;Hutchinson et al, 2010c;Lewis et al, 2010).…”

“…Such opioid-induced adaptations, in both microglial and astrocytic reactive phenotypes, in these CNS locations indicate sensitivity of pain systems to central immune signaling alterations in opioid responses. It is noteworthy that nonclassic opioids, such as (ϩ)-morphine and morphine-3-glucuronide, are also capable of inducing similar microglial reactivity, as demonstrated by up-regulation of CD11b (Hutchinson et al, 2010a;Lewis et al, 2010) or other microglial activation markers such as TLR4 (Hutchinson et al, 2010a;Lewis et al, 2010).…”

“…Finally, morphine elevation of TNF-␣ (Johnston et al, 2004) is dependent on TLR2 , ceramide synthase ), p38, and CGRP ) and is sensitive to amitriptyline (Tai et al, 2006;Tai et al, 2009), naltrexone (Bokhari et al, 2009), naloxone, and ␤-funaltrexamine (Sawaya et al, 2009). Other nonclassic opioid ligands such as (ϩ)-morphine, (ϩ)-methadone and morphine-3-glucuronide are also capable of inducing upregulation of IL-1␤, IL-6, and TNF-␣ (Hutchinson et al, 2010a;Lewis et al, 2010), suggesting a possible role for nonclassic opioid pathways in inducing opioid proinflammatory responses.…”

Intrathecal Studies on Animal Pain Models

“…Recently, many studies have revealed that intrathecal morphine could induce glial activation and neuro-inflammation in the spinal cord (Muscoli et al, 2010;Zhang et al, 2011). Several therapeutic targets have been found, including cytokine receptors, kappa-opioid receptors, N-methyl-D-aspartate receptors, and Toll-like receptors (Hameed et al, 2010;Lewis et al, 2010). Recently, tumor necrosis factor (TNF)-antagonist etanercept was found to reverse morphine-induced tolerance and block morphine-induced neuroinflammation in the microglia (Shen et al, 2011).…”