Author:

Keywords:

micronails, neurite outgrowth

Abstract:

Multi electrode arrays have been used since the seventies for the study of electrophysiological properties of neuronal cells and their connectivity in a neuronal network. This technology avoids invasiveness encountered by patch clamp methods, and allows for high throughput measurements. However, the signal-to-noise ratio (SNR) and the lack of control on the neuronal connectivity are thus far still inadequate. Imec works towards a subcellular-sized, three-dimensional micronail electrode array platform to achieve a better contact between the neuronal membrane and the electrodes in order to increase the SNR and to be able to interface on the subcellular scale. Underneath the surface topology of the electrodes an active chip is present to measure and process neuronal signals. With these new technologies it should become possible to study synaptic strengths and neuronal connectivity on the single-cell level, a much-desired feature in present neuronal research. One of the challenges in the development of the chips is to design the micronails in such a way that neurite outgrowth and connectivity can be accurately controlled. Therefore, we performed a morphological analysis of E17 mouse hippocampal neurons on 3 µm-high micronail arrays. Observations indicate that neuronal morphology alters when growing on nail substrates: intracellular cytoskeleton proteins such as actin filaments and microtubules densify around the nail shaft, membrane staining shows this close contact as well. Initial speed of neurite outgrowth (4-30h after plating) was investigated on micronail arrays with different parameters (diameter and spacing of the nails). On the surfaces that appear to provide maximal guidance, total neurite outgrowth is 1.5 times faster in the first 30h compared to a flat surface. Also, the angles of neurite growth patterns relative to the substrate were quantitatively examined. Because of the hexagonal shape of the nails, neurite outgrowth is maximally confined at an angle of 60°, whereas neurites on flat surfaces show random spreading. These results show that our surfaces have a clear impact on the direction of neuronal growth and behavior. In the future, these engineered surfaces could be used for axon guidance and regeneration studies, and implemented in a platform for the study of neuronal connectivity.