Effects of organosilane monolayer films on the geometrical guidance of CNS neurons

This work reports on the alignment of central nerve processes using surfaces of modified organosilane monolayer films. Two types of organosilane monolayer films, formed of decyldimethylsiloxane (DDMS) or trimethylsiloxane (TMS), were covalently formed on glass substrates. The films were pattern modified with a synthetic peptide (P1543) derived from mouse laminin, an extracellular matrix (ECM) protein, via a combination of ultraviolet lithography and chemical modification technique. The modification procedure generated a pattern of 10-mu m-wide peptide stripes that were surrounded by either DDMS films or TMS films. A significant difference between the two patterned substrates (DDMS/P1543 and TMS/P1543) was in hydrophobic properties of the surrounding surfaces: whereas the DDMS surrounding surfaces were more hydrophobic compared with the peptide surfaces, the TMS surrounding surfaces showed an equivalent hydrophobic property to that of the peptide surfaces. Effects of the surrounding surfaces on the alignment of central nerve processes were investigated by growing neurons dissociated from embryonic rat hippocampi on the patterned DDMS/P1543 and TMS/P1543 substrates at a low density (40 cells/mm(2)) in a chemically defined culture medium. The time-lapse video microscopy revealed that although a similar bipolar morphology was developed by the hippocampal neurons grown on both patterned substrates, the growth behaviors of the nerve tips were highly affected by physicochemical characteristics of the surrounding surfaces. The growing tips advanced straightforward along the peptide stripes when the surrounding surfaces were formed of DDMS films, whereas those often explored the surrounding surfaces formed of TMS films. Our work directly shows that although the attachment of neurons on patterned substrates is affected by hydrophobic characteristics of surrounding surfaces, the hydrophobicity is not a necessary factor for the neurite guidance. We presume that localized chemical cues, such as laminin synthetic peptide, are rather crucial constituents that affect the directional outgrowth of central nervous system neurons.