Mechanisms of antiepileptic drug action underlying neural tube defects
Revealing the regulatory mechanisms that coordinate neural plate folding and neural tube closure could improve preventative measures for neural tube defects (NTDs). Among the most prevalent and serious birth defects, NTDs occur in 1 in 1000 births in the United States. Multiple causes, both genetic and environmental, have been recognized, including the use of antiepileptic drugs (AEDs) during pregnancy. There is a 10- to 20-fold increase in NTD incidence associated with the use of AEDs during pregnancy, but the teratogenic mechanisms are unknown. The prevalent paradigm that neural activity is not evident at early stages of development suggests that AED teratogenic effects are due to off-target mechanisms. However, recent evidence of embryonic excitability during neural tube formation has challenged this theory and supports the essential need to investigate the presence and regulation of neural activity during nervous system development. Voltage-gated sodium channels (Nav) are one of the most common and efficacious cellular target of AEDs in the mature nervous system yet remain relatively unexplored during neural tube formation. My findings demonstrate that Nav is expressed by Xenopus laevis neural plate cells throughout neural plate folding and neural tube closure and its insertion in neural plate cell membranes increases with the progression of neural tube formation, suggesting that Nav may play an important role in this process. Indeed, I demonstrate that both Nav blockers and Nav-AEDs elicit neural tube defects when exposed to neurulating Xenopus laevis embryos. In addition, Nav blockers and Nav-AEDs comparably reduce neural plate cell calcium activity and reverse the developmental progression of neural plate cell calcium dynamics during neurulation suggesting that AEDs may inhibit calcium transients by impairing Nav function. This work establishes FluoVoltTM membrane potential dye as a useful probe for studying neural plate cell resting membrane potential in Xenopus laevis embryos and demonstrates that the resting membrane potential of neural plate cells is dependent on K+, Na+, and Cl- gradients. Moreover, I find that Nav blockers and Nav-AEDs hyperpolarize neural plate cells. Here I propose a model of mechanisms of Nav-AED induced NTDs. Resting membrane potential of neural plate cells allows for spontaneous activation of Nav which in turn leads to further depolarization of neural plate cells and activation of NMDAR and calcium dynamics. This calcium activity is important for regulating neural plate cell cycle and neural tube formation. Nav-AEDs interfere with Na+ currents and calcium dynamics leading to NTDs. This work significantly contributes to the mechanistic understanding of neural activity regulating neural tube formation to advance the discovery of safer therapeutic options for pregnant women with epilepsy.