Abstracts

Rationale:
Dravet syndrome (DS) is a severe form of epilepsy with a high incidence of sudden unexpected death in epilepsy (SUDEP). Respiratory failure is a leading cause of SUDEP despite it is unclear how DS-associated genetic variants or seizure activity disrupts respiratory control. Most DS cases are caused by mutations in the Scn1a gene which encodes Nav1.1 channels that preferentially regulate inhibitory neurons early in development. Previously we showed that expression of a Scn1a loss of function variant (A1783V) in all inhibitory neurons resulted in seizures, breathing problems and premature death. Despite this severe phenotype, it is unclear how loss of Scn1a function disrupts brainstem respiratory centers. To address this, we conditionally expressed Scn1a^A1783V in subcortical brainstem respiratory centers by crossing floxed stop Scn1a^A1783V mice with GlyT2-cre mice to generate GlyT2::Scn1a^A1783V/+ and control (floxed only or cre only) animals.  

Methods:
The novel open field assay was used to assess locomotor behavior and anxiety, EEG activity was measured by radio telemetry to characterize spontaneous and heat induced seizures, and whole-body plethysmography was used to access baseline breathing and the ventilatory response to 3, 5 and 7% CO₂ (balance O₂). All experiments were performed in mice two months of age, no sex differences were detected in behaviors of interest so genders were pooled for analysis. 

Results:

GlyT2::Scn1a^A1783V/+ mice were obtained at the expected frequency and exhibited grossly normal motor and anxiety behavior.  These animals also did not exhibit spontaneous seizures; however, 11 of 17 (~70%) GlyT2::Scn1a^A1783V/+ mice exhibited febrile seizures (at 42 ± .3 ^oC) shortly thereafter followed by death.  Conversely, control mice do not show seizure activity over this same temperature range (n=12).  GlyT2::Scn1a^A1783V/+ mice also showed reduced respiratory frequency (n=10) and increased post-sigh apnea (frequency and duration; n=7) under room air conditions and they showed a blunted ventilatory response to CO₂ compared to control mice.

Conclusions:
Together, these results show that loss of Scn1a function in glycinergic neurons disrupts breathing and increases febrile seizure propensity. Also, since febrile seizures are caused by respiratory alkalosis, these results suggest baseline breathing problems may serve as an early biomarker of febrile seizure propensity and risk of mortality in DS. Significance Statement: These results show for the first time that disruption of glycinergic signaling contributes to clinically important aspects of DS. 

Funding:

F31 NS120467; R01HL137094; R01HL104101