Lipid Emulsion Improves Functional Recovery in an Animal Model of Stroke.

Stroke is a life-threatening condition that leads to the death of many people around the world. Reperfusion injury after ischemic stroke is a recurrent problem associated with various surgical procedures that involve the removal of blockages in the brain arteries. Lipid emulsion was recently shown to attenuate ischemic reperfusion injury in the heart and to protect the brain from excitotoxicity. However, investigations on the protective mechanisms of lipid emulsion against ischemia in the brain are still lacking. This study aimed to determine the neuroprotective effects of lipid emulsion in an in vivo rat model of ischemic reperfusion injury through middle cerebral artery occlusion (MCAO). Under sodium pentobarbital anesthesia, rats were subjected to MCAO surgery and were administered with lipid emulsion through intra-arterial injection during reperfusion. The experimental animals were assessed for neurological deficit wherein the brains were extracted at 24 h after reperfusion for triphenyltetrazolium chloride staining, immunoblotting and qPCR. Neuroprotection was found to be dosage-dependent and the rats treated with 20% lipid emulsion had significantly decreased infarction volumes and lower Bederson scores. Phosphorylation of Akt and glycogen synthase kinase 3-ß (GSK3-ß) were increased in the 20% lipid-emulsion treated group. The Wnt-associated signals showed a marked increase with a concomitant decrease in signals of inflammatory markers in the group treated with 20% lipid emulsion. The protective effects of lipid emulsion and survival-related expression of genes such as Akt, GSK-3ß, Wnt1 and ß-catenin were reversed by the intra-peritoneal administration of XAV939 through the inhibition of the Wnt/ß-catenin signaling pathway. These results suggest that lipid emulsion has neuroprotective effects against ischemic reperfusion injury in the brain through the modulation of the Wnt signaling pathway and may provide potential insights for the development of therapeutic targets.

Neuroprotection from Excitotoxic Injury by Local Administration of Lipid Emulsion into the Brain of Rats.

Lipid emulsion was recently shown to attenuate cell death caused by excitotoxic conditions in the heart. There are key similarities between neurons and cardiomyocytes, such as excitability and conductibility, which yield vulnerability to excitotoxic conditions. However, systematic investigations on the protective effects of lipid emulsion in the central nervous system are still lacking. This study aimed to determine the neuroprotective effects of lipid emulsion in an in vivo rat model of kainic acid-induced excitotoxicity through intrahippocampal microinjections. Kainic acid and/or lipid emulsion-injected rats were subjected to the passive avoidance test and elevated plus maze for behavioral assessment. Rats were sacrificed at 24 h and 72 h after kainic acid injections for molecular study, including immunoblotting and qPCR. Brains were also cryosectioned for morphological analysis through cresyl violet staining and Fluorojade-C staining. Anxiety and memory functions were significantly preserved in 1% lipid emulsion-treated rats. Lipid emulsion was dose-dependent on the protein expression of ß-catenin and the phosphorylation of GSK3-ß and Akt. Wnt1 mRNA expression was elevated in lipid emulsion-treated rats compared to the vehicle. Neurodegeneration was significantly reduced mainly in the CA1 region with increased cell survival. Our results suggest that lipid emulsion has neuroprotective effects against excitotoxic conditions in the brain and may provide new insight into its potential therapeutic utility.

Analgesic effects of FAAH inhibitor in the insular cortex of nerve-injured rats.

The insular cortex is an important region of brain involved in the processing of pain and emotion. Recent studies indicate that lesions in the insular cortex induce pain asymbolia and reverse neuropathic pain. Endogenous cannabinoids (endocannabinoids), which have been shown to attenuate pain, are simultaneously degraded by fatty acid amide hydrolase (FAAH) that halts the mechanisms of action. Selective inhibitor URB597 suppresses FAAH activity by conserving endocannabinoids, which reduces pain. The present study examined the analgesic effects of URB597 treatment in the insular cortex of an animal model of neuropathic pain. Under pentobarbital anesthesia, male Sprague-Dawley rats were subjected to nerve injury and cannula implantation. On postoperative day 14, rodents received microinjection of URB597 into the insular cortex. In order to verify the analgesic mechanisms of URB597, cannabinoid 1 receptor (CB1R) antagonist AM251, peroxisome proliferator-activated receptor alpha (PPAR alpha) antagonist GW6471, and transient receptor potential vanilloid 1 (TRPV1) antagonist Iodoresiniferatoxin (I-RTX) were microinjected 15 min prior to URB597 injection. Changes in mechanical allodynia were measured using the von-Frey test. Expressions of CB1R, N-acyl phosphatidylethanolamine phospholipase D (NAPE-PLD), and TRPV1 significantly increased in the neuropathic pain group compared to the sham-operated control group. Mechanical threshold and expression of NAPE-PLD significantly increased in groups treated with 2 nM and 4 nM URB597 compared with the vehicle-injected group. Blockages of CB1R and PPAR alpha diminished the analgesic effects of URB597. Inhibition of TRPV1 did not effectively reduce the effects of URB597 but attenuated expression of NAPE-PLD compared with the URB597-injected group. In addition, optical imaging demonstrated that neuronal activity of the insular cortex was reduced following URB597 treatment. Our results suggest that microinjection of FAAH inhibitor into the insular cortex causes analgesic effects by decreasing neural excitability and increasing signals related to the endogenous cannabinoid pathway in the insular cortex.

Plasticity-Related PKMζ Signaling in the Insular Cortex Is Involved in the Modulation of Neuropathic Pain after Nerve Injury.

The insular cortex (IC) is associated with important functions linked with pain and emotions. According to recent reports, neural plasticity in the brain including the IC can be induced by nerve injury and may contribute to chronic pain. Continuous active kinase, protein kinase Mζ (PKMζ), has been known to maintain the long-term potentiation. This study was conducted to determine the role of PKMζ in the IC, which may be involved in the modulation of neuropathic pain. Mechanical allodynia test and immunohistochemistry (IHC) of zif268, an activity-dependent transcription factor required for neuronal plasticity, were performed after nerve injury. After ζ-pseudosubstrate inhibitory peptide (ZIP, a selective inhibitor of PKMζ) injection, mechanical allodynia test and immunoblotting of PKMζ, phospho-PKMζ (p-PKMζ), and GluR1 and GluR2 were observed. IHC demonstrated that zif268 expression significantly increased in the IC after nerve injury. Mechanical allodynia was significantly decreased by ZIP microinjection into the IC. The analgesic effect lasted for 12 hours. Moreover, the levels of GluR1, GluR2, and p-PKMζ were decreased after ZIP microinjection. These results suggest that peripheral nerve injury induces neural plasticity related to PKMζ and that ZIP has potential applications for relieving chronic pain.