Abstracts

Rationale:
Dravet syndrome (DS) is a rare, catastrophic form of genetic epilepsy diagnosed within the first year of life. DS is caused by a genetic mutation in the SCN1A gene, which encodes for the voltage gated sodium channel, Nav1.1. Recent evidence has suggested that this mutation primarily affects inhibitory interneurons, leading to a loss of function in these cells. As a result, imbalance between inhibition and excitation occurs, resulting in a seizure. Several studies have shown a transient development in the loss of excitability in interneurons and increased hyperexcitability in hippocampal pyramidal neurons. To determine if CA1 pyramidal neurons exhibit hyperexcitability and if this is dependent on age in a mouse model of DS, whole cell patch clamp technique was used to record electroresponsive membrane properties.
Method:
Experimental animals were generated by breeding a floxed stop male Scn1aA1783V (B6(Cg)-Scn1aatm1.1Dsf/J, Jax#026133) with a Sox2-cre (B6.Cg-Edil3Tg(Sox2-cre)1Amc/J) female mouse to produce both female and male heterozygous (Scn1aA1783/WT) and wildtype offspring. Two age groups were used: P18-25 and P35-60.  Whole cell patch clamping was conducted to determine the electroresponsive membrane properties of CA1 neurons in the hippocampus. 350 µm coronal slices containing the hippocampus were prepared in a sucrose-based aCSF. Patch electrodes (~3MΩ) were filled with potassium gluconate-based internal solution. Cells were blindly patched in the CA1 region of hippocampus. To record electroresponsive membrane properties, a graded series of hyperpolarizing and depolarizing current pulses (50 pA increments, 2s in duration) were applied in current clamp mode. Parameters compared between Scn1aA1783V/WT and wild type mice included: resting membrane potential, membrane resistance, time constant, and action potential properties.
Results:
No differences were present between the resting membrane potential, membrane resistance, and action potential properties (action potential threshold, max depolarization, max depolarization and repolarization slopes) between Scn1aA1783V/WT and wild type mice, independent of age (P18-25 vs. P35-60). There was a trend suggesting the time constant may differ between P18-25 and P35-60 Scn1aA1783V/WT mice, suggesting that CA1 neurons recorded in slices from younger mice respond more rapidly to a stimulus. Additionally, P18-25 Scn1aA1783V/WT mice showed increased action potential generation across the current injection range when action potentials were evoked as compared to P35-60 Scn1aA1783V/WT mice. No differences in any parameter was present between P18-25 and P35-60 wild type mice.
Conclusion:
Overall, Scn1aA1783V/WT and wild type mice have similar CA1 neuron electroresponsive membrane properties, suggesting a lack of hyperexcitability. However, in previous experiments, we have shown that Scn1aA1783V/WT mice have spontaneous seizure through adulthood, suggesting hyperexcitability may exist in a different subset of cells. Future experiments include investigating EPSCs and IPSCs of CA1 neurons and electroresponsive membrane properties and hyperexcitability of dentate granule cells.
Funding:
:HHSN271201600048C