Spatiotemporal progression of ubiquitin-proteasome system inhibition after status epilepticus suggests protective adaptation against hippocampal injury

Abstract

BackgroundThe ubiquitin-proteasome-system (UPS) is the major intracellular pathway leading to the degradation of unwanted and/or misfolded soluble proteins. This includes proteins regulating cellular survival, synaptic plasticity and neurotransmitter signaling; processes controlling excitability thresholds that are altered by epileptogenic insults. Dysfunction of the UPS has been reported to occur in a brain region- and cell-specific manner and contribute to disease progression in acute and chronic brain diseases. Prolonged seizures, status epilepticus, may alter UPS function but there has been no systematic attempt to map when and where this occurs in vivo or to determine the consequences of proteasome inhibition on seizure-induced brain injury.MethodTo determine whether seizures lead to an impairment of the UPS, we used a mouse model of status epilepticus whereby seizures are triggered by an intra-amygdala injection of kainic acid. Status epilepticus in this model causes cell death in selected brain areas, in particular the ipsilateral CA3 subfield of the hippocampus, and the development of epilepsy after a short latent period. To monitor seizure-induced dysfunction of the UPS we used a UPS inhibition reporter mouse expressing the ubiquitin fusion degradation substrate ubiquitinG76V-green fluorescent protein. Treatment with the specific proteasome inhibitor epoxomicin was used to establish the impact of proteasome inhibition on seizure-induced pathology.Results and conclusionsOur studies show that status epilepticus induced by intra-amygdala kainic acid causes select spatio-temporal UPS inhibition which is most evident in damage-resistant regions of the hippocampus, including CA1 pyramidal and dentate granule neurons then appears later in astrocytes. In support of this exerting a beneficial effect, injection of mice with the proteasome inhibitor epoxomicin protected the normally vulnerable hippocampal CA3 subfield from seizure-induced neuronal death in the model.These studies reveal brain region- and cell-specific UPS impairment occurs after seizures and suggest UPS inhibition can protect against seizure-induced brain damage. Identifying networks or pathways regulated through the proteasome after seizures may yield novel target genes for the treatment of seizure-induced cell death and possibly epilepsy.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-017-0163-2) contains supplementary material, which is available to authorized users.

Bi-directional genetic modulation of GSK-3β exacerbates hippocampal neuropathology in experimental status epilepticus

“…To explore the response of GSK-3β to prolonged seizures, we used a well-characterized model of intra-amygdala KA-induced SE in mice 59 . As previously reported, SE resulted in hippocampal damage that was mainly localized to the ipsilateral CA3 subfield, although scattered cell death was present in the CA1 and the hilus regions (Fig.…”

Expression and function of the metabotropic purinergic P2Y receptor family in experimental seizure models and patients with drug‐refractory epilepsy

“…We have shown that experimental status epilepticus leads to an inhibition of the ubiquitin‐proteasome system (UPS), thereby potentially increasing P2Y 1 levels. UPS function is also altered during chronic epilepsy in mice . Seizure‐induced changes in receptor stability or internalization may also account for the increase in P2Y 1 levels .…”

Context-Specific Switch from Anti- to Pro-epileptogenic Function of the P2Y1 Receptor in Experimental Epilepsy

“…RNA extraction and qPCR. RNA extraction was performed using the Trizol method, as described previously (Engel et al, 2017b). Quantity and quality of RNA were measured using a Nanodrop Spectrophotometer (Thermo Fisher Scientific).…”

“…Western blot analysis. Western blot analysis was performed as described previously (Engel et al, 2017b). Lysis buffer containing phosphatase and protease inhibitors was used to homogenize hippocampal tissue and extract protein, which was quantified using a Tecan plate reader at 560 nm; 30 g of protein samples was loaded onto an acrylamide gel and separated by SDS-PAGE.…”