Cortical EEG and in vivo calcium imaging to investigate DEE9-relevant phenotypes of a novel Pcdh19 cKO mouse: focus on parvalbumin-positive interneurons

Developmental and epileptic encephalopathy 9 (DEE9) is a neurodevelopmental disorder caused by mutations in the PCDH19 gene, which encodes protocadherin-19 (PCDH19), a synaptic adhesion molecule broadly expressed in the central nervous system. DEE9 is characterized by epilepsy, intellectual disability, autism spectrum disorder (ASD), and other neuropsychiatric comorbidities, often accompanied by pharmacoresistance. The cellular and network mechanisms underlying DEE9 remain poorly understood, limiting the development of targeted therapeutic strategies. Recent findings highlighted functional alterations and imbalanced excitatory and inhibitory (E/I) synaptic transmission in DEE9 mouse model. Given the particular involvement of GABAergic inhibitory circuitry, in this study, I investigated the role of PCDH19 in parvalbumin-positive inhibitory interneurons (PVIs) using a novel PV-Pcdh19 conditional knockout (PV-Pcdh19cKO) mouse model. In the first part of my project I performed in vivo electroencephalographic (EEG) recordings and a comprehensive battery of behavioural tests. EEG recordings revealed male-specific and age-dependent alterations in cortical oscillatory activity and disrupted E/I balance. Both sexes showed increased susceptibility to chemically induced seizures, suggesting a fragile network state. Behavioural analyses demonstrated ASD-like phenotypes in males and females, including impaired social preference, reduced social motivation, and repetitive behaviours, without deficits in general cognition, spatial memory, or learning. In the second part, I focused on the medial prefrontal cortex (mPFC), a region enriched in PVIs and implicated in both ASD and epilepsy. At the microcircuit level, structural and functional analyses revealed an increased number of excitatory synapses with altered ultrastructure, alongside age-dependent changes in the intrinsic excitability of both PVIs and pyramidal neurons (PNs). To investigate whether mPFC PVIs and PNs contribute to social cognition deficits, in vivo single-cell calcium imaging and found that PV-Pcdh19cKO mice displayed aberrant, cell type-specific activity patterns in response to social interaction, reflecting impaired temporal coordination of PVI output. Together, these findings support a model in which PCDH19 loss in PVIs triggers a developmental-dependent E/I imbalance, linked to altered cortical coordination and social behaviour deficits, providing the first evidence for the critical contribution of PVIs to DEE9 pathophysiology.

Developmental and epileptic encephalopathy 9 (DEE9) is a neurodevelopmental disorder caused by mutations in the PCDH19 gene, which encodes protocadherin-19 (PCDH19), a synaptic adhesion molecule broadly expressed in the central nervous system. DEE9 is characterized by epilepsy, intellectual disability, autism spectrum disorder (ASD), and other neuropsychiatric comorbidities, often accompanied by pharmacoresistance. The cellular and network mechanisms underlying DEE9 remain poorly understood, limiting the development of targeted therapeutic strategies. Recent findings highlighted functional alterations and imbalanced excitatory and inhibitory (E/I) synaptic transmission in DEE9 mouse model. Given the particular involvement of GABAergic inhibitory circuitry, in this study, I investigated the role of PCDH19 in parvalbumin-positive inhibitory interneurons (PVIs) using a novel PV-Pcdh19 conditional knockout (PV-Pcdh19cKO) mouse model. In the first part of my project I performed in vivo electroencephalographic (EEG) recordings and a comprehensive battery of behavioural tests. EEG recordings revealed male-specific and age-dependent alterations in cortical oscillatory activity and disrupted E/I balance. Both sexes showed increased susceptibility to chemically induced seizures, suggesting a fragile network state. Behavioural analyses demonstrated ASD-like phenotypes in males and females, including impaired social preference, reduced social motivation, and repetitive behaviours, without deficits in general cognition, spatial memory, or learning. In the second part, I focused on the medial prefrontal cortex (mPFC), a region enriched in PVIs and implicated in both ASD and epilepsy. At the microcircuit level, structural and functional analyses revealed an increased number of excitatory synapses with altered ultrastructure, alongside age-dependent changes in the intrinsic excitability of both PVIs and pyramidal neurons (PNs). To investigate whether mPFC PVIs and PNs contribute to social cognition deficits, in vivo single-cell calcium imaging and found that PV-Pcdh19cKO mice displayed aberrant, cell type-specific activity patterns in response to social interaction, reflecting impaired temporal coordination of PVI output. Together, these findings support a model in which PCDH19 loss in PVIs triggers a developmental-dependent E/I imbalance, linked to altered cortical coordination and social behaviour deficits, providing the first evidence for the critical contribution of PVIs to DEE9 pathophysiology.