Orchestrating cells on a chip: Employing surface acoustic waves towards the formation of neural networks

For the investigation of cell-cell interaction in general and for neural communication and future applications of neural networks, a controllable and well-defined network structure is crucial. We here propose the implementation of an acoustically driven system for tunable and deliberate stimulation and manipulation of cell growth on a chip. This piezoelectric chip allows us to generate a checkerboard-like standing surface acoustic wave pattern coupled to a fluid layer in a microfluidic chamber on top. Such a dynamically induced patterning lattice is shown to allow for the active positioning of the neurons and subsequent guided neurite outgrowth, thus finally overcoming the limitations of static approaches. After thorough characterization of the resulting tunable potential landscape, we successfully demonstrate cell adhesion and even growth of the such positioned cells within the well-defined pressure nodes. We demonstrate neuron growth at predetermined positions and observe a subsequent neurite outgrowth, even being correlated with the artificial potential landscape. For the very delicate and sensitive primary neural cells, this is a change of paradigm! Our experimental findings give us confidence that our hybrid lab-on-a-chip system in the near future will allow researchers to study cell-cell interaction of primary neurons. If scaled to a true network level, it will enable us to control and study how neural networks connect, interact, and communicate.

Figure 1

The experimental setup, dynamically variable in space in time to allow single-particle trapping to align neurons separately. (a), (b) Technical drawing of the SAW chip with an aligned microchannel. (c) AFM micrograph of the SSAW force field. (d–f) SAW-mediated alignment of $d=2\mu \mathrm{m}$ diameter polystyrene beads ($P_{\mathrm{IN}}=12\mathrm{dBm}$). Starting from a homogenous suspension at $t=0\mathrm{s}$ (d), after $t=21\mathrm{s}$, the bead accumulation reached a total maximum patterning efficiency (e). (f) Time to accumulate the beads as a function of the applied SAW power level $P_{\mathrm{IN}}$. (g) Particle accumulation using a “chirped” IDT ($d=300\mathrm{nm}$, $f_{\mathrm{SAW}}=147.9\mathrm{MHz}$). (h) Intensity along the red line in D. (i) Nodal distance as a function of the $f_{\mathrm{SAW}}$. (j)–(l) Single-particle trapping of $d=7.38\mu \mathrm{m}$ $\mathrm{Si}{\mathrm{O}}_2$ beads with a FFT inset. For $\frac{{\lambda }_{\mathrm{SAW}}}{D}\ge 2.4$ the trapping for single particles is achieved.

Figure 2

Neuronal cell growth inside the bioreactor to validate that SAW irradiation is not harmful. The numbers denote the same cells at different times. The arrows point at newly grown neurites, indicating healthy cells. (a) No SAW, (b) $P_{\mathrm{IN}}=6\mathrm{dBm}$, (c), $P_{\mathrm{IN}}=12\mathrm{dBm}$.

Figure 3

Adhesion of living cells at nodal positions of the SSAW. (a) SaOs-2 cells in a SSAW field. (b) Primary neurons being laterally aligned by SSAW using a SAW chip with ${\lambda }_{\mathrm{SAW}}=25\mu \mathrm{m}$ at a power level of $P_{\mathrm{IN}}=21\mathrm{dBm}$. (c) SaOs-2 cell adhesion after successful SSAW-mediated alignment. The insets show a selection of cells moving and spreading as a function of time.

Figure 4

(a) Neurons after the SSAW at $P_{\mathrm{IN}}=18\mathrm{dBm}$ was activated for $t=11\mathrm{h}$. The adhered neurons even started growing extensions. (b) Relative frequency $P$ of the outgrowth direction.