Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 99, Issue 20, Pages 12542-12547Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.192457199
Keywords
microfluidics; neuronal polarity; hippocampal
Categories
Funding
- NIGMS NIH HHS [R37 GM030367, R01 GM030367, GM 30367] Funding Source: Medline
- NINDS NIH HHS [NS039059, R01 NS039059] Funding Source: Medline
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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 del[laminin] > about 0.06 mug (ml.mum)(-1) (defined by dividing the change of concentration of laminin in solution over the distance of the gradient) orient axon specification, whereas those with del[laminin] < about 0.06 mug (ml.mum)(-1) have no effect.
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