We conducted a morphological assay on human neuronal cells in our platform using the IN CeII Analyzer. Previous studies have found that neuroligin-3 (NL3), a protein encoded by the NLGN3 gene in humans, plays an important role in facilitating synapse formation and that overexpression of NL3 can increase synapse number in mouse neurons.31 Due to the fact that mutations in human neuroligin genes are implicated in neurodevelopmental disorders such as autism, we sought to use the role of NL3 on synapse formation as a paradigm to demonstrate the utility of the 96 well plate microdevice in performing a synaptogenesis/morphological assay. We used lentiviruses to overexpress NL3-WT in the excitatory neurons while control iPS cell lines were infected to produce excitatory neurons. Devices were cultured for at least 5 weeks prior to staining for MAP2 and synapsin. We used Intellicount to identify synapsin-positive puncta co-localized within a defined region of MAP2 signal (1.5 μm away from dendrite), used as the metric for synapse number. The quantification reproduced a trend consistent with previous literature, showing an increase in puncta density with NL3
