Sustained synchronized neuronal network activity in a human astrocyte co-culture system

Abstract

Impaired neuronal network function is a hallmark of neurodevelopmental and neurodegenerative disorders such as autism, schizophrenia, and Alzheimer’s disease and is typically studied using genetically modified cellular and animal models. Weak predictive capacity and poor translational value of these models urge for better human derived in vitro models. The implementation of human induced pluripotent stem cells (hiPSCs) allows studying pathologies in differentiated disease-relevant and patient-derived neuronal cells. However, the differentiation process and growth conditions of hiPSC-derived neurons are non-trivial. In order to study neuronal network formation and (mal)function in a fully humanized system, we have established an in vitro co-culture model of hiPSC-derived cortical neurons and human primary astrocytes that recapitulates neuronal network synchronization and connectivity within three to four weeks after final plating. Live cell calcium imaging, electrophysiology and high content image analyses revealed an increased maturation of network functionality and synchronicity over time for co-cultures compared to neuronal monocultures. The cells express GABAergic and glutamatergic markers and respond to inhibitors of both neurotransmitter pathways in a functional assay. The combination of this co-culture model with quantitative imaging of network morphofunction is amenable to high throughput screening for lead discovery and drug optimization for neurological diseases.

Astrocyte lineage cells are essential for functional neuronal differentiation and synapse maturation in human iPSC‐derived neural networks

“…Addition of astrocyte‐like cells to hiPSC‐derived neural cultures has been used previously to overcome limited differentiation and slow functional maturation of hiPSC‐derived neural cultures (Dodla et al, ; Kuijlaars et al, ; Muratore, Srikanth, Callahan, & Young‐Pearse, ; Odawara et al, ; Shi et al, ; Tang et al, ). For example, artificial coculture with mouse astrocytes led to strongly increased spontaneous synaptic activity within an 8 weeks cultivation period (Tang et al, ).…”

Role of human induced pluripotent stem cell-derived spinal cord astrocytes in the functional maturation of motor neurons in a multielectrode array system

“…To establish a hiPSC platform of spinal cord astrocyte/MN co-culture, we first optimized the sequence of co-culture. As observed by others 35 Important to utilizing this human spinal cord-specific co-culture system was ensuring that hiPSC-derived neuron identity did not change with time in culture, nor did the density of either astrocytes or neurons, as this could influence the interpretation of MEA recordings. Our data suggest that the majority of our cells are spinal cord motor neurons (positive for ChAT and, to a lesser extent, for ISL1/2), expressing appropriate neurotransmitter receptors, with AMPA receptors being more represented than GABA and glycine receptors, as would be expected in the spinal cord.…”

Advances and Current Challenges Associated with the Use of Human Induced Pluripotent Stem Cells in Modeling Neurodegenerative Disease

“…Specifically, spontaneous firing rates in coculture on day 35 were twice the level reported in neuron-only controls, and mixed populations could be maintained for more than 120 days. More recently, coculture of hiPSC-derived cortical neurons with human astrocytes showed more frequent and synchronized spike trains as well as more dynamic changes in overall spike patterns compared with neuron-only controls [Kuijlaars et al, 2016;Kayama et al, 2018]. The occurrence of temporally coordinated spiking patterns may represent physiological signs of organized circuits within the cultured hiPSC-derived neuronal population, indicating the value of astrocytes on neuronal organization DOI: 10.1159/000493018 and function in vitro.…”