Patients with mutations in the PRRT2 gene (proline-rich transmembrane protein 2) are characterized by a large spectrum of neurological disorders, including mainly benign epilepsy and paroxysmal kinesigenic dyskinesia. PRRT2 is a neuronal protein implicated in the modulation of the voltage-gated sodium channel, in particular NaV 1.2 and 1.6. Indeed, a dysregulation of this gene could lead to hyperexcitability due to the loss of the Na+ channel modulation. To better understand the mechanism, we generated glutamatergic induced neurons (glutamatergic iNeurons) from PRRT2 induced pluripotent stem cells (IPSC) obtained from patients’ fibroblasts by the inducible overexpression of NGN2. Firstly, we generated IPSC from fibroblast from two homozygous and one heterozygous male PRRT2 patients. Then, we optimized NGN2 overexpression protocol by adding FACS sorting and pre-differentiation steps to generate a homogeneous population of glutamatergic neurons that were plated together with Rat astrocytes in multielectrode array (MEA) for functional analysis or in Petri dishes for patch clamp and immunostaining. Preliminary results showed a higher level of firing activity in the homozygous patients compared to the heterozygous ones and the controls both in MEA and in patch clamp experiments starting from 21 days of differentiation. Our model showed promising results to generate active glutamatergic iNeurons that can be used as platform for drug screening leading to new insights into new potential treatments for PRRT2 related disorders.