Fig. 1

Experimental plan. (1) Human iPSCs were induced to generate neural stem cells (NSCs) and patterned to develop medial ganglionic eminence cells (MGE) and GABAergic interneurons (IN). These cells were molecularly characterized for stages of development. (2) EVs from NSCs, MGE cells and interneurons were isolated, characterized and loaded with GABA using different methods. The amount of EV encapsulated GABA from each source and method was quantified using ELISA. 3 and 4. Rats with epilepsy (REs) were generated using the Kainic acid model in F344 rats (n = 32), and seizures were scored continuously from 3–6 months with video recording. At 4 months, REs were intranasally treated with interneuron-derived EV-encapsulated GABA (IN-EV-GABA) daily for 7 days. At 5 months, the same rats were treated with MGE-derived EV encapsulated GABA (MGE-EV-GABA) daily for 7 days, and at 6 months, the same rats were treated with NSC-derived EV-encapsulated GABA (NSC-EV-GABA) daily for 7 days. In another set of experiments, MGE-EVs, IN-EVs and GABA were intranasally administered daily for a week to REs separately at 4 months, and seizures were recorded. 5. Another batch of REs was treated with MGE-EV-GABA for 7 days and tested for detailed behavioral analysis before and during the treatment. GABA was conjugated with o-phthaldialdehyde (OPA), and EV-encapsulated GABA-OPA was intranasally administered and traced in the brain. Following brain isolation and fixation, immunostaining for presynaptic marker synaptophysin was performed, and 0.7 μm confocal Z-sections were evaluated at 630 magnifications