Activation of adult rat CNS endothelial cells by opioid-induced toll-like receptor 4 (TLR4) signaling induces proinflammatory, biochemical, morphological, and behavioral sequelae

Neuroscience. 2014 Nov 7:280:299-317. doi: 10.1016/j.neuroscience.2014.09.020. Epub 2014 Sep 18.

Abstract

CNS immune signaling contributes to deleterious opioid effects including hyperalgesia, tolerance, reward, and dependence/withdrawal. Such effects are mediated by opioid signaling at toll-like receptor 4 (TLR4), presumptively of glial origin. Whether CNS endothelial cells express TLR4 is controversial. If so, they would be well positioned for activation by blood-borne opioids, contributing to opioid-induced pro-inflammatory responses. These studies examined adult primary rat CNS endothelial cell responses to (-)-morphine or its mu opioid receptor (MOR)-inactive metabolite morphine-3-glucuronide (M3G), both known TLR4 agonists. We demonstrate that adult rat CNS endothelial cells express functional TLR4. M3G activated nuclear factor kappaB (NF-κB), increased tumor necrosis factor-α (TNFα) and cyclooxygenase-2 (COX2) mRNAs, and released prostaglandin E2 (PGE2) from these cells. (-)-Morphine-induced upregulation of TNFα mRNA and PGE2 release were unmasked by pre-treatment with nalmefene, a MOR antagonist without TLR4 activity (unlike CTAP, shown to have both MOR- and TLR4-activity), suggestive of an interplay between MOR and TLR4 co-activation by (-)-morphine. In support, MOR-dependent Protein Kinase A (PKA) opposed TLR4 signaling, as PKA inhibition (H-89) also unmasked (-)-morphine-induced TNFα and COX2 mRNA upregulation. Intrathecal injection of CNS endothelial cells, stimulated in vitro with M3G, produced TLR4-dependent tactile allodynia. Further, cortical suffusion with M3G in vivo induced TLR4-dependent vasodilation. Finally, endothelial cell TLR4 activation by lipopolysaccharide and/or M3G was blocked by the glial inhibitors AV1013 and propentofylline, demonstrating endothelial cells as a new target of such drugs. These data indicate that (-)-morphine and M3G can activate CNS endothelial cells via TLR4, inducing proinflammatory, biochemical, morphological, and behavioral sequelae. CNS endothelial cells may have previously unanticipated roles in opioid-induced effects, in phenomena blocked by presumptive glial inhibitors, as well as TLR4-mediated phenomena more broadly.

Keywords: CTAP; M3G; TLR4; endothelial cell; morphine; nalmefene.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Central Nervous System / drug effects*
  • Central Nervous System / physiology
  • Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Cyclooxygenase 2 / metabolism
  • Dinoprostone / metabolism
  • Endothelial Cells / drug effects*
  • Endothelial Cells / physiology
  • Hyperalgesia / drug therapy
  • Hyperalgesia / physiopathology
  • Male
  • Morphine / pharmacology*
  • Morphine Derivatives / pharmacology*
  • NF-kappa B / metabolism
  • Narcotics / pharmacology*
  • Neuroglia / drug effects
  • Neuroglia / physiology
  • Neuroimmunomodulation / drug effects
  • Neuroimmunomodulation / physiology
  • RNA, Messenger / metabolism
  • Rats, Sprague-Dawley
  • Receptors, Opioid, mu / antagonists & inhibitors
  • Receptors, Opioid, mu / metabolism
  • Toll-Like Receptor 4 / agonists
  • Toll-Like Receptor 4 / metabolism*
  • Tumor Necrosis Factor-alpha / metabolism
  • Vasodilation / drug effects
  • Vasodilation / physiology

Substances

  • Morphine Derivatives
  • NF-kappa B
  • Narcotics
  • RNA, Messenger
  • Receptors, Opioid, mu
  • Tlr4 protein, rat
  • Toll-Like Receptor 4
  • Tumor Necrosis Factor-alpha
  • Morphine
  • Cyclooxygenase 2
  • Ptgs2 protein, rat
  • Cyclic AMP-Dependent Protein Kinases
  • Dinoprostone
  • morphine-3-glucuronide