Abstract
Neuronal cultures are widely used in neuroscience research. However, the randomness of circuits in conventional cultures prevents accurate in vitro modeling of cortical development and of the pathogenesis of neurological and psychiatric disorders. A basic feature of cortical circuits that is not captured in standard cultures of dissociated cortical cells is directional connectivity. In this work, a polydimethylsiloxane (PDMS)-based device that achieves directional connectivity between micro 3D cultures is demonstrated. The device consists of through-holes for micro three-dimensional (μ3D) clusters of cortical cells connected by microtrenches for axon and dendrite guidance. The design of the trenches relies in part on the concept of axonal edge guidance, as well as on the novel concept of specific dendrite targeting. This replicates dominant excitatory connectivity in the cortex, enables the guidance of the axon after it forms a synapse in passing (an “en passant” synapse), and ensures that directional selectivity is preserved over the lifetime of the culture. The directionality of connections was verified morphologically and functionally. Connections were dependent on glutamatergic synapses. The design of this device has the potential to serve as a building block for the reconstruction of more complex cortical circuits in vitro.
Highlights
Neuronal cultures are a widely used tool in basic and translational neuroscience research
Device design and optimization The device (Fig. 2) contained two through-holes serving as the target compartment (TC) and source compartment (SC) to hold μ3D neuronal cultures
Staining results showed that both dendrites and axons from Target compartment (TC) turned toward SC when the joint of trenches was too close to TC (Supplemental Fig. S1b)
Summary
Neuronal cultures are a widely used tool in basic and translational neuroscience research. Their advantages include ease of experimental access and compatibility with high-throughput screening technologies. In vitro neuronal cell culture models have been used for studies into neurite outgrowth[1], synaptic strength and scaling[2], cell interactions[3,4,5], and drug screening[6,7], among many others. The randomness of cultured circuits prevents accurate in vitro modeling of the brain’s development and pathogenesis of neurological and psychiatric disorders. The ability to create precise neural circuits in a high-throughput compatible and accessible platform could have a transformative effect on investigations into the mechanisms of brain development and on drug discovery
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