Astrocytes and neurons collaboratively shape neural networks, influencing metabolism, synapse formation, and network dynamics. Dysfunctions in either cell type are implicated in diverse brain disorders. In order to gain insight into the transcriptional profile of hiPSC-derived astrocytes and neurons, we performed scRNA analysis and compared astrocytes and neurons before and after co-culture.Sequencing with Illumina Novaseq6000, preprocessing with 10x Genomics CellRanger. Custom R/Seurat pipeline for data analysis.NGN2 neurons were identified as excitatory. Within the astrocyte population we observed subtypes, finding radial glia in mono-culture and additionally identifying astrocytes in co-culture. Astrocyte and neuron-specific gene panels consistently showed increased expression in co-culture compared to mono-culture, implying increased specification of both cell-types in co-culture. For astrocytes this was strengthened by the pseudotime trajectory moving from dividing cells to radial glia to astrocytes, indicating cell type development upon co-culture. Gene set enrichment analysis on these astrocytes from co-culture versus monoculture showed enrichment of functional GOBP related to cell differentiation, immune system and neurogenesis.Through co-culturing, the cellular environment contributes to development of both cell types. Astrocytes showed signs of increased specification, as demonstrated by transcriptomic development of these cells from their pseudotime trajectory, peaking in the co-culture cells. Neurons showed less changes in transcriptomic profile, which could be explained by the forced nature of differentiation through NGN2-overexpression. These data were further validated by functional analyses, showing increased synapse formation and synaptic activity of neuronal networks in co-culture with human astrocytes compared to neuronal mono-cultures and co-cultures with mouse astrocytes.