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BackgroundAlzheimer’s disease (AD) is a progressive neurodegenerative disorder characterized by an abnormal accumulation of amyloid-β (Aβ) plaques, neuroinflammation, and impaired neurogenesis. Urolithin A (UA), a gut-microbial metabolite of ellagic acid, has been reported to exert anti-inflammatory effects in the brain. However, it is unknown whether UA exerts its properties of anti-inflammation and neuronal protection in the APPswe/PS1ΔE9 (APP/PS1) mouse model of AD.MethodsMorris water maze was used to detect the cognitive function. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was performed to detect neuronal apoptosis. Immunohistochemistry analyzed the response of glia, Aβ deposition, and neurogenesis. The expression of inflammatory mediators were measured by enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR). The modulating effects of UA on cell signaling pathways were assayed by Western blotting.ResultsWe demonstrated that UA ameliorated cognitive impairment, prevented neuronal apoptosis, and enhanced neurogenesis in APP/PS1 mice. Furthermore, UA attenuated Aβ deposition and peri-plaque microgliosis and astrocytosis in the cortex and hippocampus. We also found that UA affected critical cell signaling pathways, specifically by enhancing cerebral AMPK activation, decreasing the activation of P65NF-κB and P38MAPK, and suppressing Bace1 and APP degradation.ConclusionsOur results indicated that UA imparted cognitive protection by protecting neurons from death and triggering neurogenesis via anti-inflammatory signaling in APP/PS1 mice, suggesting that UA might be a promising therapeutic drug to treat AD.

Background and Purpose Considerable effort has recently been directed at developing multifunctional opioid drugs to minimize the unwanted side effects of opioid analgesics. We have developed a novel multifunctional opioid agonist, DN‐9. Here, we studied the analgesic profiles and related side effects of peripheral DN‐9 in various pain models. Experimental Approach Antinociceptive effects of DN‐9 were assessed in nociceptive, inflammatory, and neuropathic pain. Whole‐cell patch‐clamp and calcium imaging assays were used to evaluate the inhibitory effects of DN‐9 to calcium current and high‐K+‐induced intracellular calcium ([Ca2+]i) on dorsal root ganglion (DRG) neurons respectively. Side effects of DN‐9 were evaluated in antinociceptive tolerance, abuse, gastrointestinal transit, and rotarod tests. Key Results DN‐9, given subcutaneously, dose‐dependently produced antinociception via peripheral opioid receptors in different pain models without sex difference. In addition, DN‐9 exhibited more potent ability than morphine to inhibit calcium current and high‐K+‐induced [Ca2+]i in DRG neurons. Repeated treatment with DN‐9 produced equivalent antinociception for 8 days in multiple pain models, and DN‐9 also maintained potent analgesia in morphine‐tolerant mice. Furthermore, chronic DN‐9 administration had no apparent effect on the microglial activation of spinal cord. After subcutaneous injection, DN‐9 exhibited less abuse potential than morphine, as was gastroparesis and effects on motor coordination. Conclusions and Implications DN‐9 produces potent analgesia with minimal side effects, which strengthen the candidacy of peripherally acting opioids with multifunctional agonistic properties to enter human studies to alleviate the current highly problematic misuse of classic opioids on a large scale.

In a previously described chimeric peptide, we reported that the multifunctional opioid/neuropeptide FF (NPFF) receptor agonist 0 (BN-9) produced antinociception for 1.5 h after supraspinal administration. Herein, four cyclic disulfide analogs containing l- and/or d-type cysteine at positions 2 and 5 were synthesized. The cyclized analogs and their linear counterparts behaved as multifunctional agonists at both opioid and NPFF receptors in vitro and produced potent analgesia without tolerance development. In comparison to 0, cyclized peptide 6 exhibited sevenfold more potent μ-opioid receptor agonistic activity in vitro. Interestingly, the cyclized analog 6 possessed an improved stability in the brain and an increased blood–brain barrier permeability compared to the parent peptide 0 and produced more potent analgesia after supraspinal or subcutaneous administration with improved duration of action of 4 h. In addition, antinociceptive tolerance of analog 6 was greatly reduced after subcutaneous injection compared to fentanyl, as was the rewarding effect, withdrawal reaction, and gastrointestinal inhibition.

The opioid and neuropeptide FF pharmacophore-containing chimeric peptide 0 (BN-9) was recently developed and produced potent nontolerance forming analgesia. In this study, 11 analogues of 0 were designed and synthesized. An in vitro cAMP assay demonstrated that these analogues behaved as multifunctional agonists at both opioid and NPFF receptors. In mouse tail-flick test, most of the analogues produced potent nontolerance forming antinociception. Notably, 11 (DN-9) was 33-fold more potent than 0 at analgesic effects, which was mediated by μ- and κ-opioid receptors. In addition, 11 also produced powerful analgesic effects in the formalin pain and CFA-induced chronic inflammatory pain models. Strikingly, following its repeated administration for 6 days, 11 did not produce antinociceptive tolerance in the tail-flick test and CFA-induced pain model. The present work indicates that it is reasonable to design multifunctional peptide ligands for opioid and NPFF receptors in a single molecule producing effective nontolerance forming antinociception.