Early-life seizures alter synaptic calcium-permeable AMPA receptor function and plasticity

Abstract

Calcium (Ca2+)-mediated1 signaling pathways are critical to synaptic plasticity. In adults, the NMDA glutamate receptor (NMDAR) represents a major route for activity-dependent synaptic Ca2+ entry. However, during neonatal development, when synaptic plasticity is high, many AMPA glutamate receptors (AMPARs) are also permeable to Ca2+ (CP-AMPAR) due to low GluA2 subunit expression, providing an additional route for activity- and glutamate-dependent Ca2+ influx and subsequent signaling. Therefore, altered hippocampal Ca2+ signaling may represent an age-specific pathogenic mechanism. We thus aimed to assess Ca2+ responses 48 hours after hypoxia-induced neonatal seizures (HS) in postnatal day (P)10 rats, a post-seizure time point at which we previously reported LTP attenuation. We found that Ca2+ responses were higher in brain slices from post-HS rats than in controls and this increase was CP-AMPAR-dependent. To determine whether synaptic CP-AMPAR expression was also altered post-HS, we assessed the expression of GluA2 at hippocampal synapses and the expression of long-term depression (LTD), which has been linked to the presence of synaptic GluA2. Here we report a decrease 48 hours after HS in synaptic GluA2 expression at synapses and LTD in hippocampal CA1. Given the potentially critical role of AMPAR trafficking in disease progression, we aimed to establish whether post-seizure in vivo AMPAR antagonist treatment prevented the enhanced Ca2+ responses, changes in GluA2 synaptic expression, and diminished LTD. We found that NBQX treatment prevents all three of these post-seizure consequences, further supporting a critical role for AMPARs as an age-specific therapeutic target.

AMPA Receptor Dysregulation and Therapeutic Interventions in a Mouse Model of CDKL5 Deficiency Disorder

“…Sections incubated without primary and only with secondary antibody were imaged to verify antibody specificity. We have previously confirmed the surface specificity of the GluA2 antibody used here in permeabilized and nonpermeabilized sections incubated with an antibody to MAP2 in addition to GluA2 (Lippman-Bell et al, 2016). Blinded sections were placed on an LSM710 confocal microscope (Carl Zeiss) with an oil-immersion 63 ϫ 1.7 NA objective.…”

“…First, a mask was created from the MAP2 staining to define an ROI so that only puncta on the surface of dendrites would be incorporated in our analysis. To avoid a thresholding artifact, colocalization was first measured at the maximum threshold incorporating all GluA2 puncta; then the GluA2 channel was thresholded at multiple intervals of the max threshold (95%, 90%, 85%, 80%, 75%, and 70%), and colocalization was determined again at each threshold (Nie et al, 2010;Zhou et al, 2011;Lippman-Bell et al, 2016). After thresholding each channel, the overlap of GluA2 with synapsin puncta was quantified using the plugin coloc2 to determine the Manders coefficient (Manders et al, 1993).…”

“…Receptor subunits are dynamically expressed, altering receptor composition and biophysical properties to regulate the excitatory-inhibitory balance throughout the lifetime (Luján et al, 2005;Rakhade and Jensen, 2009). Dysregulated glutamate receptor subunit expression is often associated with epilepsy, autism, and ID (Loddenkemper et al, 2014;Mignogna et al, 2015;Lippman-Bell et al, 2016). In particular, even modest changes in the tightly regulated GluA2 AMPAR subunit have a profound effect on excitability and signal transduction due to the distinct properties of GluA2 (Lippman-Bell et al, 2013;Uzunova et al, 2014;Stephenson et al, 2017).…”

A physiologically-validated rat model of term birth asphyxia with seizure generation after, not during, brain hypoxia

“…These seizures are likely to be mechanistically heterogeneous between and even within individuals, as is evident from EEG recordings and from the highly variable (and usually inadequate) therapeutic responses to standard anticonvulsant agents Connell et al, 1989;Lloyd et al, 2017;Shellhaas and Clancy, 2007;van Rooij et al, 2013). In fact, while much work on rodent models has focused on neuronal molecules and signaling mechanism that are affected as a consequence of seizures (Lippman-Bell et al, 2016;Rakhade et al, 2011;Sun et al, 2013;Wang et al, 2011;Zhou et al, 2015), next to nothing is known about the proximate causes in vivo which are responsible for rendering the neonate brain prone to seizures after BA. Given the steep dependence of neuronal excitability on brain pH, it is reasonable to assume that the large pH changes which take place during and following birth asphyxia in human neonates (Uria-Avellanal and Robertson, 2014) are likely to contribute to the multiple mechanisms that lead to the generation of post-asphyxia seizures.…”

Ocular hypertension drives remodeling of AMPA receptors in select populations of retinal ganglion cells

“…Remodeling of AMPARs is a well established consequence of a wide spectrum of neuronal dysfunction, including drug dependence (Wolf, 2016), seizures (Rakhade et al, 2008; Lippman-Bell et al, 2016), and ischemia (Noh et al, 2005; Dias et al, 2013; Hwang et al, 2013). While the evidence for increased expression of CP-AMPAR is often clear, the underlying mechanism, either a decrease in ADAR2 levels and consequently of GluA2 editing, or a downregulation of GluA2 transcription, is not.…”