Oxygen and Glucose Deprivation in an Organotypic Hippocampal Slice Model of the Developing Rat Brain: The Effects on N-Methyl-d-Aspartate Subunit Composition

Abstract

Background Oganotypic hippocampal slices (OHS) are commonly used to screen for neuroprotective effects of pharmacologic agents relevant to pediatric brain injury. The importance of donor rat pup age and N-methyl-d-aspartate (NMDA) receptor subunit composition have not been addressed. In this study we evaluated the age-dependent effect of oxygen-glucose deprivation (OGD) in the developing rat brain and determined whether OGD modulates the NMDA receptor subunit composition. Methods OHS were prepared from rat pups on postnatal days (PND) 4, 7, 14 and 21 and cultured 7 days in vitro. The slices were exposed to OGD for durations of 5– 60 min. After 24 and 72hours, OHS survival and NMDA subunit composition were assessed. Results Cell death was evident in OHS prepared from PND 14 and 21 rat pups (p<0.001) with OGD durations of 5 and 10 min, respectively. In OHS prepared from PND7 rat pups, neurodegeneration was not evident until 20 min OGD (p<0.001). Exposure to OGD in OHS prepared from PND4 and PND7 rat pups was associated with a transition in the NMDA receptor subunit composition from NR2B predominant to NR2A predominant subunit composition. Conclusions This in vitro neonatal rat pup investigation using OHS supports both an age and an NMDA receptor subunit composition-dependent relationship between OGD and neuronal cell death.

Validation of organotypical hippocampal slice cultures as an ex vivo model of brain ischemia: different roles of NMDA receptors in cell death signalling after exposure to NMDA or oxygen and glucose deprivation

“…Thus, slice cultures represent an advantage over acute slices, as the slice cultures will have recovered from the stress and damage introduced by slice preparation at the time they are used in experimental settings. Our data are in line with results from a study on organotypical hippocampal slices cultivated on membranes (Stoppini et al1991), demonstrating a postnatal age-related change in NR2A and NR2B subunits composition that parallels what is seen in vivo in rodents (Wise-Faberowski et al, 2009). In our study, we additionally investigated the total level of NMDAR indicated by NR1 expression.…”

A lasting effect of postnatal sevoflurane anesthesia on the composition of NMDA receptor subunits in rat prefrontal cortex

“…Comparison between groups receiving different treatments was carried out by determining the protein ratio of the protein of interest with tubulin using the immunoreactive area by densitometry. Data are expressed as a ratio of the NMDA subunits to tubulin (Wise‐Faberowski et al, 2009).…”

Glutamate receptor and transporter modifications in rat organotypic hippocampal slice cultures exposed to oxygen–glucose deprivation: The contribution of cyclooxygenase-2

“…These reports contrast with our data showing that meloxicam provides a noticeable neuroprotection against OGD in the hippocampal organotypic slice cultures together with increases in the expression of GluN2B. We think that these unexpected increases in GluN2B as a consequence of the meloxicam treatment could be dependent on the stage when sections were obtained: In this regard, levels of GluN2B in hippocampal organotypic slice culture from 4-and 7-day-old rats present noticeably high levels of GluN2B, compared with the strongly reduced levels for those of 14-and 21-day-old rats (Wise-Faberowski et al, 2009). A serial study of the response of GluN2B is being performed in our laboratories to check this hypothesis.…”