A grid micropattern of neuronal cells was formed on a free-standing collagen film (35 μm thickness) by directing migration and extension of neurons along a Matrigel pattern previously prepared on the film by the microcontact printing method. The neurons migrated to reach the nodes on the grid pattern and extended neurites to bridge cell bodies at the nodes. The resulting neuronal micropattern on the collagen film containing culture medium can be handled and deformed with tweezers with maintenance of physiological activity of the neurons, as examined by response of cytosolic Ca2+ concentration to a dose of bradykinin. This portability is the unique advantage of the present system that will open novel possibility of cellular engineering including the on-demand combination with analytical devices. The repetitive lamination of the film on a microelectrode chip was demonstrated for local electrical stimulation of a specific part of the grid micropattern of neurons, showing Ca2+ wave propagation along the neurites. The molecular permeability is the further advantage of the free-standing hydrogel substrate, which allows external supply of nutrients and dosing with chemical stimulants through the film even under rolled and laminated conditions.
Keywords: Portable neuronal micropattern; hydrogel film; neuronal tissue engineering.