PRRT2 controls neuronal excitability by negatively modulating Na+ channel 1.2/1.6 activity

See Lerche (doi:10.1093/brain/awy073) for a scientific commentary on this article. PRRT2 mutations cause heterogeneous paroxysmal neurological disorders. Using iPSC-derived neurons from patients homozygous for a nonsense PRRT2 mutation and cortical neurons from PRRT2-knockout mice, Fruscione et al. show that PRRT2 is a negative modulator of voltage-dependent NaV1.2/1.6 channels. Increased neuronal excitability may contribute to the paroxysmal nature of PRRT2-linked diseases. Abstract See Lerche (doi:10.1093/brain/awy073) for a scientific commentary on this article. Proline-rich transmembrane protein 2 (PRRT2) is the causative gene for a heterogeneous group of familial paroxysmal neurological disorders that include seizures with onset in the first year of life (benign familial infantile seizures), paroxysmal kinesigenic dyskinesia or a combination of both. Most of the PRRT2 mutations are loss-of-function leading to haploinsufficiency and 80% of the patients carry the same frameshift mutation (c.649dupC; p.Arg217Profs*8), which leads to a premature stop codon. To model the disease and dissect the physiological role of PRRT2, we studied the phenotype of neurons differentiated from induced pluripotent stem cells from previously described heterozygous and homozygous siblings carrying the c.649dupC mutation. Single-cell patch-clamp experiments on induced pluripotent stem cell-derived neurons from homozygous patients showed increased Na+ currents that were fully rescued by expression of wild-type PRRT2. Closely similar electrophysiological features were observed in primary neurons obtained from the recently characterized PRRT2 knockout mouse. This phenotype was associated with an increased length of the axon initial segment and with markedly augmented spontaneous and evoked firing and bursting activities evaluated, at the network level, by multi-electrode array electrophysiology. Using HEK-293 cells stably expressing Nav channel subtypes, we demonstrated that the expression of PRRT2 decreases the membrane exposure and Na+ current of Nav1.2/Nav1.6, but not Nav1.1, channels. Moreover, PRRT2 directly interacted with Nav1.2/Nav1.6 channels and induced a negative shift in the voltage-dependence of inactivation and a slow-down in the recovery from inactivation. In addition, by co-immunoprecipitation assays, we showed that the PRRT2-Nav interaction also occurs in brain tissue. The study demonstrates that the lack of PRRT2 leads to a hyperactivity of voltage-dependent Na