Abstract
The cardiac autonomic nervous system plays a critical role in heart function, and its dysregulation is associated with various cardiac disorders, including arrhythmias and heart failure. However, the study of neuro-cardiac interactions in humans is limited by the lack of proper physiologically relevant experimental models [1].
Induced pluripotent stem cell (iPSC) -derived sympathetic neurons (iPSC-SNs) and iPSCs- derived cardiomyocytes (iPSCs-CMs) were first fully characterized in monoculture and then used to establish the neurocardiac co-culture model. The two cell populations were initially cultured in separate chambers of a two-chamber insert. Upon its removal, iPSC-SNs extended axonal projections toward iPSC-CMs, forming strong direct neuro-cardiac connections. The co-culture was analyzed using a multi electrode array (MEA) system, revealing that after seven days, the beat rate of iPSC-CMs remained stable, whereas iPSC-SNs exhibited a significant increase in firing activity (P value: 0.010). To assess the functional connection between hiPSC-SNs and hiPSC-CMs, nicotine was used to stimulate iPSC-SNs, resulting in a significant increase in the beat rate of iPSC-CMs (P value: 0.016), while no effect was observed in iPSC-CMs maintained in monoculture. The nicotine-induced increase in iPSC-CM beat rate was abolished by β-blocker propranolol (P value:0.048) but remained unaffected by α-bungarotoxin, which blocks nicotinic receptors in iPSC-SNs. Additionally, the β-adrenergic agonist isoproterenol was used to enhance the beat rate of iPSC-CMs, an effect that was significantly reduced by propranolol administration (P value: 0.002). After seven days of co-culture, nicotine-treated cells exhibited a marked reduction in FFN270-stained vesicles, a fluorescent tracer for norepinephrine, indicating effective neurotransmitter release and functional synaptic exocytosis (P value: 0.004).
This human iPSCs-derived model represents a robust approach to studying disorders characterized by impaired cardiac innervation, paving the way for new insights into cardiac autonomic dysfunction and personalized medicine. Furthermore it is an innovative tool for drug screening, allowing the evaluation of pharmacological compounds targeting autonomic heart regulation. Finally, this system can be scaled to 3D co-culture models, further enhancing its physiological relevance for disease modeling and therapeutic testing.