Multielectrode array analysis of human iPSC-motor neuron maturation following coculture with iPSC-spinal cord astrocytes (P4.4-031)

Objective: Develop a fully humanized co-culture platform of induced pluripotent stem cell-derived spinal cord astrocytes (hiPSC-A) and motor neurons (hiPSC-MN) for multi-electrode array (MEA) recording. Background: MEA has been previously utilized in electrophysiological recordings of rodent MNs and hiPSC-cortical neurons. However, there is no literature on hiPSC-A influences on hiPSC-MN electrophysiological properties. Design/Methods: Through the critical steps of caudalization and ventralization, we generated spinal cord hiPSC-A and -MNs. We compared hiPSC-MN monocultures to hiPSC-MN and hiPSC-A co-cultures and investigated astrocyte variables that could influence MN maturation. MEA recordings were performed weekly over a four-week period, to obtain electrophysiological parameters including spike and burst rate and the percentage of spiking and bursting electrodes. Immunocytochemistry was used to analyze astrocyte and neuronal populations. We tested the effect of neurotransmitter agonists/antagonists, including kainate, CNQX and bicuculline. Results: The culture of hiPSC-MN alone resulted in large aggregates of cells with delayed electrophysiological maturation. The addition of hiPSC-A resulted in reduced neuronal aggregation accompanied by the accelerated electrophysiological maturation of hiPSC-MN across all time points examined. These electrophysiological measures following hiPSC-A/hiPSC-MN co-culture were mirrored morphologically by increased diameter of neuronal processes, mean soma size, and complexity of neuronal connections. We also found that hiPSC-MN and hiPSC-neurons expressed appropriate neurotransmitter receptors which were modulated by known agonist/antagonists. Using several lines of hiPSC-A, we also found that the electrophysiological properties are consistent across lines. When compared with rodent astrocytes, hiPSC-A also appear to induce a more rapid maturation of hiPSC-MN both morphologically and electrophysiologically. Conclusions: We demonstrate that this humanized co-culture method allows for the long-term, non-invasive, serial recording of spinal cord neuronal populations, and for the electrophysiological analysis of the effects of pharmacologic manipulation of these cultures. This platform will be of use for studying astrocyte/neuron interactions in the context of motor neuron disease. Disclosure: Dr. Taga has nothing to disclose. Dr. Dastgheyb has nothing to disclose. Dr. Joseph has nothing to disclose. Dr. Habela has nothing to disclose. Dr. Richard has nothing to disclose. Dr. Haughey has nothing to disclose. Dr. Maragakis has nothing to disclose.