Generation of doxycycline-inducible human induced pluripotent stem cells for production of patient-derived neurons to investigate of the role of ankyrin-G in GABAergic deficits related to bipolar disorder

Abstract

The neuronal role of the scaffolding protein ankyrin-G, and specifically the 480 kDa isoform product of the ANK3 gene, has been established in mouse models as the primary organizer of the axon-initial segment throughout development. A more recently discovered role for ankyrin-G shows that it is necessary for stabilizing GABAA receptors on the axonal and somatodendritic domains of pyramidal neurons through its interaction with the GABAA receptor-associated protein. A specific variant, ANK3 p.W1989R, abolishes this interaction and leads to dysfunction in inhibitory signaling and loss of GABAergic synapses in mouse models. Interestingly, patients with bipolar disorder (BD) present with deficits in inhibitory signaling, and a cohort of these patients has been identified who express the ANK3 p.W1989R variant. However, findings related to this variant in mouse models cannot be applied to human bipolar disorder patients, as the complex genetic background of BD is not conserved in mice. Instead, human induced pluripotent stem cells (hiPSCs), derived from fibroblast cells of a bipolar disorder patient expressing the ANK3 p.W1989R variant, can be used to generate neurons in vitro that retain the patient genetic background. Here I demonstrate the generation of hiPSCs of BD patient and control genetic backgrounds, that are capable of doxycycline-inducible differentiation to both excitatory and inhibitory neuron populations. These will be used in future experiments to assess the necessity and sufficiency of the ANK3 p.W1989R variant to confer deficits in GABAergic signaling and synapse formation in a human-derived BD model, providing insight into a potential molecular mechanism of bipolar disorder.