Gradients of Substrate-Bound Laminin Orient Axonal Specification of Neurons

Author(s): Dertinger, Stephan K. W., Xingyu Jiang, Zhiying Li, Venkatesh N. Murthy, George M. Whitesides

Journal: Proc. Natl. Ac. Sci. (2002) 99(20): 12542-12547.

Abstract:
Little is known about the influence of substrate-bound gradients on neuronal development, since it has been difficult to fabricate gradients over the distances typically required for biological studies (a few hundred micrometers). This article demonstrates a generally applicable technique for the fabrication of substrate bound gradients of proteins with complex shapes, using laminar flows in microchannels. Gradients that range from pure laminin to pure BSA were formed in solution by using a network of microchannels, and these proteins were allowed to adsorb onto a homogeneous layer of poly-L-lysine. Rat hippocampal neurons were cultivated on these substrate-bound gradients. Analysis of optical images of these neurons showed that axon specification is oriented in the direction of increasing surface density of laminin. Linear gradients in laminin adsorbed from a gradient in solution having a slope of ∇[laminin] > about 0.06 µg (ml-µm)^-1 (defined by dividing the change of concentration of laminin in solution over the distance of the gradient) orient axon specification, whereas those with ∇[laminin] < about 0.06 µg (ml-µm)^-1 have no effect.