Abstracts

Rationale:
In human epileptic patients, seizure activity incidence prefers non-rapid eye movement (NREM) sleep period to rapid-eye movement (REM) sleep period, suggesting the possible link between NREM sleep and seizure incidence. Our previous works in one mouse model of idiopathic generalized epilepsy (IGE) have indicated that slow-wave oscillations (SWOs, 0.5 Hz) can generate un-balanced synaptic excitation in neurons (AES 2019, 3.026). Meanwhile, it is known that female human subjects seem to have more SWO EEG activity with large amplitudes during NREM sleep than male human subjects. Thus, using an IGE mouse model with GABAR gamma 2 subunit (Gabrg2) Q390X mutation, we hypothesized that epileptic spike-wave discharge (SWD) incidence in female mice would be more frequent than that in male mice.
Method:
With all procedures in accordance with guidelines set by the Institutional Animal Care and Use Committee of Vanderbilt University Medical Center, both wild-type (wt) and heterozygous (het) Gabrg2+/Q390X KI (adult male and female, postnatal 90-120 days) mouse underwent EEG mouse surgery (anesthesia 1–3% isoflurane(vol/vol)). One screw EEG electrode was planted on each side of somatosensory cortex brain skulls (anterior-posterior between -1.82 and -0.46 mm reference to bregma, midline-lateral -4.0 or +4.0 mm) and third ground electrode over cerebellum surface (#8201 Pinnacle Technology, Lawrence KS). One pair of stainless-steel wires were planted in neck trapezius muscles. After surgery, mice were returned to animal facility for recovery for at least 1 week, with normal day-light and dark circadian cycle. EEG (0.1~100 Hz, epidural) and electromyogram (EMG) activity (10~400 Hz)  were recorded by using two Multiclamp 700B amplifiers and one DigiData 1400 digitizer (at 20 KHz) (Molecular Device Inc.) (current-clamp mode, two channels for EEG recordings, one channel for EMG recordings). Mouse epileptic behaviors were video-recorded with simultaneous EEG activity to classify seizure stages based on Racine scales. Bilateral synchronous SWDs (typical absence epilepsy SWD, 6-12 Hz) and slow SWD (atypical absence epilepsy SSWD, 3-6 Hz) were defined as trains ( >1 s) of rhythmic biphasic spikes, with a voltage amplitude at least two-fold higher than baseline amplitudes. Data were expressed as mean ± SEM (standard error of mean).
Results:
Our results did not show much SWD incidence in both wt male and wt female mice. However, female het mice exhibited significantly more epileptic SWD incidence and longer SWD duration, compared with female wt mice (SWD#/hour (hr): wt 2.57 ± 1.25, n=7 mice vs het 73.16 ± 14.73, n = 6 mice, t-test p = 0.0001) (total SWD duration/hr: wt 6.05 ± 3.43 second (s), n = 7 vs het 420.39 ± 122.14 s, n = 6, t-test p = 0.0014) (averaged single SWD duration: wt 1.37 ± 0.40 s, n = 7 vs het 4.85 ± 1.04 s, n=6, t-test p=0.0029). Furthermore, compared with male het mice, female het mice exhibited significantly more epileptic SWD incidence and longer SWD duration (SWD#/hr: male het 36.90 ± 7.05, n = 7 mice vs female het 73.16 ± 14.73, n = 6, t-test p = 0.039) (total SWD duration/ hr: male het 115.91 ± 27.67 s, n = 7 vs het 420.39 ± 122.14 s, n = 6, t-test p = 0.023) (averaged SWD duration: male het 2.59 ± 0.27 s, n = 7 vs het 4.85 ± 1.04 s, n = 6, t-test p = 0.046).
Conclusion:
Consistent with our previous findings on SWO-induced seizure onset in het Gabrg2 Q390X mice, Gabrg2 Q390X mutation in female mice could cause seizure incidence. Moreover, the gender difference in potential sleep SWS period between male het and female het mice further contributes to more epileptic SWD incidence and longer SWD duration in female het mice than male het mice. This indicates that sleep activity due to gender factor likely plays a role in seizure incidence difference.
Funding:
:NIH R21 NS096483 and R01 NS107424-01