Abstracts

Rationale: Idiopathic generalized epilepsy (IGE) influences almost 3 million peoples in US and has been indicated to have genetic causes in IGE patients, with elusive seizure onset mechanisms. Moreover, IGE animal model studies have identified wide-field epileptic networks. Our previous works indicate that in somatosensory(S1) cortex neurons slow-wave oscillations(SWOs) can drive up unbalanced excitability. However, it remains unknown whether unbalanced SWO-homeostatic potentiation of synaptic excitability holds true to neurons within epileptic networks. Thus we hypothesized that SWO (0.5 Hz) can amplify and exacerbate the imbalance between synaptic excitation and inhibition in S1 cortex neurons within epileptic networks. Methods: Using wt littermates and het GABAR gamma 2 subunit(Gabrg2) Q390X mutation KI mice(crossed with cFos-GFP mice from Jackson laboratory 014135), cFos-GFP neuron networks were examined in paraformaldehyde-perfused brain sections(70µm). Same cortical and subcortical landmarks in wt and het KI mice were used to select same brain sections and count cFos-GFP neurons with ImageJ program. Using ex vivo brain slices and whole-cell recordings, spontaneous(s) excitatory (EPSC) and inhibitory (IPSC) synaptic currents were recorded in cFos-GFP layer V/VI pyramidal neurons in S1 and other cortices. sEPSCs were recorded at Cl- reversal potentials (-55.8mV, K-gluconate based intracellular solution), while sIPSCs recorded at -60 mV (KCl/K-gluconate based solution) with 20 µM NBQX in ACSF. SWOs(5~10 min)(current-clamp mode) were induced by injecting 0.5 Hz cosine oscillating/depolarizing currents into neurons(around -75 mV) with 4-5 spikes riding at oscillation peaks. Results: Compared with only a few cFos-GFP positive neurons from wt littermates, more cFos-GFP positive neurons were clearly identified from het KI mice within S1 cortex, anterior cingulate cortex(ACC), motor cortex, hippocampus CA1 and hypothalamic preoptic area(VLPO/MnPO) and medial amygdala(AmygMeA), as indicated by cFos-GFP neuron counting in ((expressed as #per section per side), n=4 mice each) S1 cortex wt 24+-15 vs het 560+-51, t-test p=0.001; ACC wt 14+-21 vs het 287+-61, p=0.005; VLPO wt 14+-15 vs het 165+-25, p=0.002; AmygMeA wt 56+-21 vs het 870+-103, p=0.001. Moreover, in S1 cortex cFos-GFP neurons from wt littermates, SWOs potentiated both sEPSCs (from -15.46+-1.20 to -28.30+-2.65 pA, n=4, pair t-test p=0.005) and sIPSCs (from -25.38+-2.12 to -39.12+-3.12 pA, n=4; paired t-test p=0.011), while in S1 cortex cFos-GFP neurons from het Q390X KI mice, SWOs potentiated only sEPSCs (from -21.45+-1.43 to -32.33+-3.16 pA, n=4, paired t-test p= 0.02), not sIPSCs(from -13.61+-1.22 to -13.31+-1.62 pA, n=5, paired t-test p=0.886), suggesting that the balance between synaptic sEPSCs and sIPSCs in cFos-GFP cortical neurons was disrupted within epileptic networks from het KI mice. Conclusions: cFos-GFP neuron counting with our IGE mice carrying Gabrg2 Q390X mutation indicated wide-field epileptic networks distributing within cortex and subcortical structures. Moreover, in cFos-GFP cortical neurons from wt littermates, SWOs generated balanced sEPSC and sIPSCs, while in cFos-GFP cortical neurons from het KI mice, SWOs could drive up run-away sEPSCs (unbalanced by sIPSCs). This suggests that SWOs will likely amplify excitability of epileptic network cortical neurons and initiate seizures in a synchronizing way in-vivo in this IGE model during sleep-wake transition or quiet-wake period. Funding: NINDS R21NS096483, R01NS107424 (Zhou) and Vanderbilt Medical Center Dept. Neurology development fund