Abstract
In the stargazer mouse model of absence epilepsy, altered corticothalamic excitation of reticular thalamic nucleus (RTN) neurons has been suggested to contribute to abnormal synchronicity in the corticothalamic-thalamocortical circuit, leading to spike-wave discharges, the hallmark of absence seizures. AMPA receptor expression and function are decreased in stargazer RTN, due to a mutation of AMPAR auxiliary subunit stargazin. It is unresolved and debated, however, if decreased excitation of RTN is compatible with epileptogenesis. We tested the hypothesis that relative NMDAR expression may be increased in RTN and/or thalamic synapses in stargazers using Western blot on dissected thalamic nuclei and biochemically isolated synapses, as well as immunogold cytochemistry in RTN. Expression of main NMDAR subunits was variable in stargazer RTN and relay thalamus; however, mean expression values were not statistically significantly different compared to controls. Furthermore, no systematic changes in synaptic NMDAR levels could be detected in stargazer thalamus. In contrast, AMPAR subunits were markedly decreased in both nucleus-specific and synaptic preparations. Thus, defective AMPAR trafficking in stargazer thalamus does not appear to lead to a ubiquitous compensatory increase in total and synaptic NMDAR expression, suggesting that elevated NMDAR function is not mediated by changes in protein expression in stargazer mice.
Highlights
Epilepsy is one of the most prevalent chronic neurological disorders, affecting approximately 65 million people worldwide[1], one third of whom are unresponsive to currently available anti-epileptic medication[2]
In view of its pivotal role and specific impairment in stargazer mice, NMDAR total protein expression was further explored at CT-reticular thalamic nucleus (RTN) synapses using post-embedding immunogold cytochemistry for comparative analysis of NR1 immunogold particles associated with single postsynaptic densities (PSD) of presumptive CT synapses in control and stargazer RTN single sections
VGlut[1] was downregulated in the epileptic ventral posterior (VP), which corresponds with the decreased frequency of miniature excitatory postsynaptic currents in adult stargazer VP shown before[26]
Summary
Epilepsy is one of the most prevalent chronic neurological disorders, affecting approximately 65 million people worldwide[1], one third of whom are unresponsive to currently available anti-epileptic medication[2]. The cortical input to RTN appears to be of critical importance; in the Gria4−/− mouse model[16], for instance, a decrease in AMPA receptor function at corticothalamic synapses on RTN neurons leads to SWDs. The loss of Gria4-containing AMPARs at corticothalamic (CT)-RTN synapses results in decreased feed-forward inhibition of relay thalamus, but reveals a new pathway for network hyperexcitability[17], as the weakened inhibition of the relay nuclei via the CT-RTN-VP route in effect leads to increased direct excitation of VP neurons via corticothalamic projections. We used Western blot for comparative analysis of NMDAR subunits in dissected RTN and VP nuclei to determine if there are region specific changes in stargazers; Western blot was used to examine AMPA- and NMDAR expression in biochemically isolated, pooled thalamic synapses in stargazers. Protein expression of AMPAR subunit GluA4 was analysed in parallel to demonstrate region specificity and reproducibility of previous findings in stargazers
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