Abstracts

Rationale: Mutations in SCN1B, encoding the voltage-gated sodium channel (VGSC) β1/β1B subunits, result in pediatric epilepsies including Dravet syndrome (DS), a severe epileptic encephalopathy. While the majority of DS patients have de novo mutations in SCN1A, patients with two inherited loss-of-function SCN1B alleles also have DS. Brain VGSCs are heterotrimers, composed of one pore-forming α subunit and two modulatory β subunits. While β1 and β1B have been characterized as cell adhesion molecules in vivo, their roles in sodium current (INa) modulation and neuronal excitability in vivo are not well understood.Methods: Here, we investigated the effects of Scn1b deletion on specific pyramidal neuron populations in mouse visual and motor cortex. We used whole cell and nucleated patch clamp to record action potentials (APs), excitatory postsynaptic currents, and somatic INa density in pyramidal neurons in acute brain slices of postnatal (P)14-21 Scn1b null and wildtype (WT) mice. We also performed 3H-saxitoxin binding to assess total levels of tetrodotoxin-sensitive VGSC expression in null and WT cortical membrane preparations.Results: The excitability of null pyramidal neurons of visual cortex layers 2/3, 5, and motor cortex layer 5 were not different from WT. In contrast, layer 6 neurons of visual cortex exhibited a decreased threshold for AP firing, a 5 mV depolarized resting membrane potential, increased AP frequency, decreased magnitude of after-hyperpolarization, and increased frequency of spontaneous excitatory post-synaptic currents compared to WT. To investigate the effect of Scn1b deletion on INa density, we recorded INa in nucleated patches from visual cortex layer 6 and motor cortex layer 5. INa density was decreased to ~60% of WT in visual cortex layer 6 but increased to ~50% above WT in motor cortex layer 5. There was a ~5 mV hyperpolarizing shift in the voltage dependence of VGSC availability in nulls compared to WT with no effect on the voltage dependence of conductance in both layer 5 and 6 of null cortex. The τ of VGSC recovery from inactivation was prolonged in both populations of null pyramidal neurons. 3H-STX binding showed that total levels of VGSCs were reduced in null cortex.Conclusions: These data show layer dependent heterogeneity in the effects of Scn1b deletion on cortical pyramidal neuron excitability and suggest that layer 6 is an epileptogenic region in spite of INa density reduction. We propose that potassium channel, rather than sodium channel, modulation may underlie Scn1b pyramidal neuron hyperexcitability and future work will test this hypothesis. The observation that the effects of SCN1B on excitability are cell type and brain region specific may help to identify which microcircuit components underlie the mechanism of SCN1B-linked DS. Supported by NIH grants R01-NS-076752 and U01-NS-090364 to LLI, the University of Michigan Systems and Integrative Biology Training Grant T32 GM008322-26 (funding for JMH), and a Pre-doctoral Fellowship from the Michigan Brain Initiative to JMH.