Figure 1. Cytoarchitectures of neurons in vivo compared with dish or microfluidic-device (µFD) cultures.

(a) The cytoarchitecture of brain tissue is remarkably complex, as seen in cell packing and interwoven processes of the cerebral cortex in situ imaged by array tomography. The three-dimensional space of a mouse cortical neuron (green) is surrounded by other cells (marked by DAPI-labeled nuclei, blue) and myriad presynaptic boutons (synapsin-labeled, red). Figure modified and used with permission.^[2] Scale bar = 10 µm. (b) Electron micrograph of layer IV of the rat somatosensory cortex reveals densely packed structures with essentially no free space around the cell body (nucleus, N), myelinated axons (ma), axon bundles (ab), dendritic spines (sp), and dendrites (d); mitochrondria also are shown (m). Scale bar = 5 µm. Original image contributed by Graham Knott. (c) At low densities in a conventional dish culture, neurons develop complex cytoarchitectures and connections despite open space between neurites and cell bodies. Pre-synaptic terminals (synapsin, green) stud the dendritic arbor (MAP 2, blue) of a primary hippocampal neuron (33 days in vitro); rhodamine-phalloidin labels filamentous actin (red), scale bar = 20 µm. Modified and used with permission.^[80] (d) A µFD fabricated to organize cellular connections and control the local fluidic micro-environment. Depicted is a two-channel µFD (left) with tapering interconnects (enlarged at right), which facilitate directional orientation of neurons for analyzing interactions. Neurons can be cultured in each channel; interactions can be restricted to and guided by the interconnecting tunnels (i.e., interconnects). Cortical neurons were seeded in one channel (left, blue) and striatal neurons in another channel (right, blue). Interactions between channels are restricted to the interconnects (orange, enlarged on right). Tapered interconnects funnel cortical axons toward the channel on the right, while restricting the reverse migration of afferent axons back through the interconnects. Reproduced and modified by permission of The Royal Society of Chemistry (RSC).^[52] (e) Complex and elaborate architectures of neurons can be guided and organized using a µFD. A single striatal neuron (yellow, right channel) is innervated by organized axon bundles from cortical neurons (green) cultured in the adjacent channel and directed via the interconnecting tunnels. Striatal neurons immunolabeled for MAP2 (red), cortical neurons labeled for α-tubulin (green). Reproduced and modified by permission of the RSC.^[52]