Anti-inflammatory treatment with a soluble epoxide hydrolase inhibitor attenuates seizures and epilepsy-associated depression in the LiCl-pilocarpine post-status epilepticus rat model

Highlights

• TPPU alleviates seizures and depression in the LiCl-pilocarpine post- SE rat model.

• TPPU attenuates inflammations in the brain of LiCl-pilocarpine post-SE rat model.

• TPPU might be a new treatment for epilepsy and comorbidities, especially depression.

Abstract

Purpose

This study aimed to investigate whether 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), a soluble epoxide hydrolase inhibitor with anti-inflammatory effects, could alleviate spontaneous recurrent seizures (SRS) and epilepsy-associated depressive behaviours in the lithium chloride (LiCl)-pilocarpine-induced post-status epilepticus (SE) rat model.

Methods

The rats were intraperitoneally (IP) injected with LiCl (127 mg/kg) and pilocarpine (40 mg/kg) to induce SE. A video surveillance system was used to monitor SRS in the post-SE model for 6 weeks (from the onset of the 2nd week to the end of the 7th week after SE induction). TPPU (0.1 mg/kg/d) was intragastrically given for 4 weeks from the 21st day after SE induction in the SRS + 0.1 TPPU group. The SRS + PEG 400 group was given the vehicle (40% polyethylene glycol 400) instead, and the control group was given LiCl and PEG 400 but not pilocarpine. The sucrose preference test (SPT) and forced swim test (FST) were conducted to evaluate the depression-like behaviours of rats. Immunofluorescent staining, enzyme-linked immunosorbent assay, and western blot analysis were performed to measure astrocytic and microglial gliosis, neuronal loss, and levels of soluble epoxide hydrolase (sEH), cytokines [tumour necrosis factor alpha (TNF-α), interleukin (IL)-1β, and IL-6], and cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB).

Results

The frequency of SRS was significantly decreased at 6 weeks and 7 weeks after SE induction in the 0.1TPP U group compared with the SRS + PEG 400 group. The immobility time (IMT) evaluated by FST was significantly decreased, whereas the climbing time (CMT) was increased, and the sucrose preference rate (SPR) evaluated by SPT was in an increasing trend. The levels of sEH, TNF-α, IL-1β, and IL-6 in the hippocampus (Hip) and prefrontal cortex (PFC) were all significantly increased in the SRS + PEG 400 group compared with the control group; neuronal loss, astrogliosis, and microglial activation were also observed. The astrocytic and microglial activation and levels of the pro-inflammatory cytokines in the Hip and PFC were significantly attenuated in the TPPU group compared with the SRS + PEG 400 group; moreover, neuronal loss and the decreased CREB expression were significantly alleviated as well.

Conclusion

TPPU treatment after SE attenuates SRS and epilepsy-associated depressive behaviours in the LiCl-pilocarpine induced post-SE rat model, and it also exerts anti-inflammatory effects in the brain. Our findings suggest a new therapeutic approach for epilepsy and its comorbidities, especially depression.

Introduction

Epilepsy is a chronic brain disease, which not only has the clinical feature of recurrent seizures but also has cognitive and psychological comorbidities, especially when patients have active refractory seizures (Josephson et al., 2017). There is a strong bidirectional relationship between epilepsy and depression (Keezer et al., 2016). Certain common neurobiological mechanisms are found to contribute to the comorbidity of epilepsy and depression (Kanner, 2017). Selective serotonin reuptake inhibitors (SSRIs) are the antidepressants most recommended to alleviate depressive symptoms in patients with epilepsy, but they have a high risk of exacerbating seizures, especially when using in overdose (Maguire et al., 2014, Mula, 2017). There currently is no effective therapeutic method for simultaneously treating the epilepsy and depression.

Emerging evidence indicates that neuroinflammation plays an important role in epileptogenesis in both humans with epilepsy and animal models of epilepsy (Vezzani et al., 2013, Butler et al., 2016). Over-production of inflammatory factors such as interleukin (IL)-1β, IL-6, tumour necrosis factor alpha (TNF-α), and prostaglandin E2 (PGE2) contributes to the progression of seizures (Vezzani et al., 2008, Wang et al., 2018). Activation of inflammatory mediators such as cyclooxygenase (COX)-2 was also found to induce neuronal damage and facilitate seizures (Kulkarni and Dhir, 2009, Rojas et al., 2019). Glial cell activation may potentiate seizures by increasing pro-inflammatory cytokines and inducing the dysfunction of neuron-glial communication (Devinsky et al., 2013, Alyu and Dikmen, 2017). Moreover, Mazarati et al. found that the hippocampal IL-1β was a contributing factor for depressive behaviours in a pilocarpine-induced status epilepticus (SE) model, and blockade of hippocampal IL-1 receptor (IL-1R) exerted an anti-depressant effect in the post-SE model (Mazarati et al., 2010), indicating that an inflammatory mechanism may be closely involved in epilepsy-associated depression as well. Treatments targeting neuroinflammation might present a novel therapeutic strategy for patients with epilepsy and neurobehavioural comorbidities (Paudel et al., 2018).

In recent years, the arachidonic acid (AA) metabolic pathway and its roles in inflammation have been widely studied. AA is an abundant unsaturated fatty acid stored in membrane phospholipids. Free AA is metabolised into active intermediate substrates by COX, lipoxygenase (LOX), and cytochrome P450 (CYP450) epoxygenases, of which CYP450 epoxygenases metabolise AA into different types of epoxyeicosatrienoid acids (EETs). EETs have a variety of beneficial functions including anti-inflammatory effects, vasodilation, and even exerting neuroprotective effects (Iliff et al., 2010). However, EETs are easily hydrolysed by soluble epoxide hydrolase (sEH) to form the corresponding diols with reduced biological activity (Wang et al., 2018). CYP450 epoxygenases and sEH are widely expressed in neurons, astrocytes, and microvascular endothelial cells in the cortex and hippocampus (Bianco et al., 2009). Studies showed that sEH gene knockout or inhibiting the activity of sEH could enhance the beneficial effects of EETs, and that inhibitors of sEH (sEHI) have potent anti-inflammatory effects (Harris and Hammock, 2013).

It has been demonstrated that the level of sEH was significantly elevated in the temporal cortex and hippocampal complexes of patients with temporal lobe epilepsy (TLE) (Ahmedov et al., 2017). In a mouse model of acute tetramethylenedisulfotetramine intoxication, post-exposure administration of sEHI in combination with diazepam effectively prevented progression to tonic seizures and animal death probably mediated by the potent anti-inflammatory effects of sEHI (Vito et al., 2014). Another study found that the expression of sEH protein was higher in the brain of chronically stressed mice and post-mortem brain samples of patients with psychiatric diseases than their controls, and it showed that pre-treatment with sEHI prevented depression-like behaviours in an inflammation-induced depression model, indicating that sEH also plays a key role in the pathophysiology of depression (Ren et al., 2016). In this study, we aimed to investigate the effects of treatment with 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU) 21 days after SE, a type of sEHI that can cross the blood–brain barrier (Liu et al., 2013), on seizures and the epilepsy-associated depressive behaviours in the lithium chloride (LiCl)-pilocarpine-induced post-SE rat model. Moreover, markers for inflammation and cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB) in the neuronal survival pathway were measured to explore the underlying mechanism of TPPU.