Controlling Neuronal Growth on Au Surfaces by Directed Assembly of Proteins
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Controlling Neuronal Growth on Au Surfaces by Directed Assembly of Proteins Cristian Staii1, Chris Viesselman2, Jason Ballweg2, Steven Hart4, Justin C. Williams3, Erik W. Dent2, Susan N. Coppersmith4 and Mark A. Eriksson4 1 Physics and Astronomy, Tufts University, 4 Colby Street, Medford, MA, 02155, U.S.A. 2 Anatomy, University of Wisconsin-Madison, Madison, WI, 53706, U.S.A. 3 Biomedical Engineering, University of Wisconsin-Madison, Madison, WI, 53706, U.S.A. 4 Physics, University of Wisconsin-Madison, Madison, WI, 53706, U.S.A.
ABSTRACT Studying how individual neuronal cells grow and interact with each other is of fundamental importance for understanding the functions of the nervous system. However, the mechanism of axonal navigation to their target region and their specific interactions with guidance factors such as membrane-bound proteins, chemical and temperature gradients, mechanical guidance cues, etc. are not well understood. Here we describe a new approach for controlling the adhesion, growth and interconnectivity of cortical neurons on Au surfaces. Specifically, we use Atomic Force Microscopy (AFM) nanolithography to immobilize growth-factor proteins at well-defined locations on Au surfaces. These surface-immobilized proteins act as a) adhesion proteins for neuronal cells (i.e. well-defined locations where the cells “stick” to the surface), and b) promoters/inhibitors for the growth of neurites. Our results show that protein patterns can be used to confine neuronal cells and to control their growth and interconnectivity on Au surfaces. We also show that AFM nanolithography presents unique advantages for this type of work, such as high degree of control over location and shape of the protein patterns, and application of proteins in aqueous solutions (protein buffers), such that the proteins are very likely to retain their folding conformation/bioactivity. INTRODUCTION The basic working unit of the brain is the neuron, a specialized cell designed to transmit information to other neurons, muscle, or gland cells. It consists of a cell body plus long threadlike axons that transmit electrical impulses, and shorter, thicker dendrites, which receive messages from other cells. A daunting task in neuroscience is to figure out how as many as 100 billion neurons are produced, grow, and organize themselves into the truly wonderful information-processing machine which is the human brain. Traditional biological techniques have provided valuable insights into neuronal growth, development, and operation. For example, it is now known that specialized proteins (extracellular matrix proteins and guidance factors) provide cues that influence transmission of information among neurons [1]. The precise spatial arrangement of these proteins plays a crucial role in guiding axons/dendrites to their targets. However, the local environment faced by a growing neuron in vivo is so rich and complex that the collective influence of many guidance cues is extremely hard to decipher [2-3]. Because of these difficulties, an alt
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