Prenatal Downregulation of CB₁Cannabinoid Receptors in the Mouse Prefrontal Cortex Disrupts Cortical Lamination and Induces a Transcriptional Signature Associated with Social Interaction Deficits

Endocannabinoid signaling exerts a neurodevelopmental regulatory role via CB₁ cannabinoid receptors (CB₁Rs), which control pyramidal neuron differentiation, migration, and axonal guidance. Here, we investigated the long-lasting consequences of transient prenatal CB₁R downregulation within the mouse prefrontal cortex by assessing its impact on gene expression, neuronal electrophysiological properties, and animal behavioral traits. Transient loss of CB₁Rs as induced by in-utero small-interference RNA electroporation at Embryonic Day 14.5, when upper-layer neurons are generated, arrested cell migration leading to ectopic neurons that populated deep layers. Whole-cell current-clamp recordings showed that ectopic neurons are less excitable (increased afterhyperpolarization amplitude, decreased sag, lower firing frequency) than deep-layer-native pyramidal neurons. Differentially expressed genes (DEGs), identified by microarray characterization of FACS-sorted electroporated neurons, were significantly enriched in pathways related to cortical development, regulation of cell migration, neurotransmitter secretion, and cytoskeletal organization. Gene set enrichment analysis also supported enrichment in pathways associated with neurodegenerative disorders and synaptic function. The gene expression profile of siCB₁R-derived neurons showed DEGs that had been previously associated with intellectual disability, schizophrenia, and autism. Venn diagrams unveiled one common DEG for neuropsychiatric risk databases and CB₁R expression manipulation, namely, the transcription factor ZBTB20. Prenatal knockdown of CB₁Rs induced long-lasting behavioral alterations in the adult offspring of either sex, with an impairment of social interaction and motor behavior in siCB₁R-derived adult mice. Taken together, these findings highlight the role of CB₁Rs in controlling the development of pyramidal neurons in the prefrontal cortex and support the contribution of altered endocannabinoid signaling to neuropsychiatric vulnerability.