Efficacy of dual enkephalinase inhibition in a preclinical migraine model is mediated by activation of peripheral delta opioid receptors.

OBJECTIVE: The aim of this study was to evaluate whether elevating levels of enkephalin by inhibiting their degradation can attenuate stress-induced migraine-like behaviors in mice. BACKGROUND: Previous studies in animals have suggested the delta opioid receptor (DOR) as a novel migraine target. The primary endogenous ligands for DOR are enkephalins and their levels can be increased by pharmacological inhibition of enkephalinases; however, it is not clear whether enkephalinase inhibition can be efficacious in preclinical migraine models through activation of DOR or whether other opioid receptors might be involved. Further, it is not clear whether opioid receptors in the central nervous system are necessary for these effects. METHODS: This study used a model of repetitive restraint stress in mice that induces periorbital hypersensitivity and priming to the nitric oxide donor sodium nitroprusside (SNP; 0.1 mg/kg). Von Frey filaments were used to measure periorbital mechanical thresholds and grimace scores were evaluated by observing mouse facial features. Animals were treated with the dual enkephalinase inhibitor (DENKI) PL37. RESULTS: On day two post-stress, PL37 given to mice via either intravenous injection (10 mg/kg) or oral gavage (20 mg/kg) significantly attenuated stress-induced periorbital hypersensitivity and facial grimace responses. Additionally, both intravenous (10 mg/kg) and oral gavage (20 mg/kg) of PL37 prior to SNP (0.1 mg/kg) administration on day 14 post-stress significantly reduced SNP-induced facial hypersensitivity. Injection of the DOR antagonist naltrindole (0.1 mg/kg) but not the mu-opioid receptor antagonist CTAP (1 mg/kg) prior to PL37 treatment blocked the effects. Finally, pretreatment of mice with the peripherally restricted opioid receptor antagonist naloxone methiodide (5 mg/kg) blocked the effects of PL37. CONCLUSIONS: These data demonstrate that inhibiting enkephalinases, and thus protecting enkephalins from degradation, attenuates stress-induced migraine-like behavior via activation of peripheral DOR. Peripheral targeting of endogenous opioid signaling may be an effective therapeutic strategy for migraine.

Soluble Epoxide Hydrolase Inhibition Attenuates Excitotoxicity Involving 14,15-Epoxyeicosatrienoic Acid-Mediated Astrocytic Survival and Plasticity to Preserve Glutamate Homeostasis.

Astrocytes play pivotal roles in regulating glutamate homeostasis at tripartite synapses. Inhibition of soluble epoxide hydrolase (sEHi) provides neuroprotection by blocking the degradation of 14,15-epoxyeicosatrienoic acid (14,15-EET), a lipid mediator whose synthesis can be activated downstream from group 1 metabotropic glutamate receptor (mGluR) signaling in astrocytes. However, it is unclear how sEHi regulates glutamate excitotoxicity. Here, we used three primary rat cortical culture systems, neuron-enriched (NE), astrocyte-enriched glia-neuron mix (GN), and purified astrocytes, to delineate the underlying mechanism by which sEHi and 14,15-EET attenuate excitotoxicity. We found that sEH inhibitor 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA) and 14,15-EET both attenuated N-methyl-D-aspartate (NMDA)-induced neurite damage and cell death in GN, not NE, cortical cultures. The anti-excitotoxic effects of 14,15-EET and AUDA were both blocked by the group 1 mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP), as were their protective effects against NMDA-disrupted perineuronal astrocyte processes expressing glutamate transporter-1 (GLT-1) and subsequent glutamate uptake. Knockdown of sEH expression also attenuated NMDA neurotoxicity in mGluR5- and GLT-1-dependent manners. The 14,15-EET/AUDA-preserved astroglial integrity was confirmed in glutamate-stimulated primary astrocytes along with the reduction of the c-Jun N-terminal kinase 1 phosphorylation, in which the 14,15-EET effect is mGluR5-dependent. In vivo studies validated that sEHi and genetic deletion of sEH (Ephx2-KO) ameliorated excitotoxic kainic acid-induced seizure, memory impairment, and neuronal loss while preserving GLT-1-expressing perineuronal astrocytes in hippocampal CA3 subregions. These results suggest that 14,15-EET mediates mGluR5-dependent anti-excitotoxicity by protecting astrocytes to maintain glutamate homeostasis, which may account for the beneficial effect of sEH inhibition in excitotoxic brain injury and diseases.

Correction to: Soluble Epoxide Hydrolase Inhibition Attenuates Excitotoxicity Involving 14,15-Epoxyeicosatrienoic Acid-Mediated Astrocytic Survival and Plasticity to Preserve Glutamate Homeostasis.

The original version of this article unfortunately contained a mistake. The authors observed inadvertent error in Fig. 7d, in which the image of the GFAP/DAPI in the WT saline treated mice was rotated left 90-degree by mistake. The corrected representative image is given below.