2009 WILEY-VCH Verlag Gm Carbon nanotubes (CNTs) have been the subject of extensive research ever since their discovery because of their unique electrical, mechanical, physiochemical, and 1D structural properties. Novel applications based on CNTs have been explored in a variety of fields, such as nanoelectronic devices, electrochemical detection, and electronic biomolecular sensors. More recently, CNTs were employed to interact with living systems at subcellular, cellular, and tissue levels, due to their molecular size, biocompatibility, and stability in aqueous environments. Amongmany interesting examples, individual CNTs dispersed in aqueous solution have been utilized as nanovectors to deliver macromolecules into live cells without apparently undermining cell viability or eliciting immunoreactions, and as blocking agents to modulate and probe ion channels on cell membranes. Alternatively, CNT solid substrates have been used to interface with live cells for various promising applications. It has been shown that CNTs support the attachment and growth of various cell types, including osteoblasts, cardiac muscle cells, and neurons. Therefore, CNTs, which are mechanically robust and are able to form nanofibrous networks similar to the extracellular matrix (ECM), have been considered to be appealing candidates for the development of 2D and 3D scaffolds for tissue regeneration. In particular, conductive CNT networks have been used to integrate with neurons to promote and guide neuron growth and to stimulate and sense their electrical activities, demonstrating the potential of CNTs in neural prosthesis. Despite the fact that the recent convergence of nanotechnology and cellular biology is creating enormous new opportunities, careful investigation into how nanostructures such as CNTs might interfere with specific cellular functions is still lacking. In the same way that the nanostructured ECM shapes cell structures and functions in diverse and intricate ways, the nanotopographic nanotube substrates may also exert profound effects on cell adhesion, proliferation, morphology, cytoskeleton organization, and intracellular signaling pathways. For instance, it has been shown that the metabolic activities of human osteoblastic cells grown on CNT substrates are enhanced in a manner dependent on the nanotube geometry. In this study, the effects of a nanotopographic CNT substrate on the motion of secretory vesicles in neuronendocrine PC12 cells (which secrete catecholamines via exocytosis of large dense core vesicles) are examined. Exocytosis is a highly dynamic and regulated process, in which secretory vesicles are, upon being triggered, directed to the outer cell membrane, to subsequently discharge their contents (such as hormones in endocrine cells and neurotransmitters in neurons) to the extracellular environment. Prior to the final Ca2þ-dependent vesicle fusion, secretory vesicles have to be transported to and trapped in the subplasmalemmal membrane region by the cortical actin meshwork and associated proteins, where they explore and interact with various secretory proteins, and are eventually released at exocytotic sites upon triggering. How vesicles move in the near-membrane region is directly related to the exocytosis dynamics. Fluorescent-tagged subplasmalemmal vesicles can be individually visualized and tracked in real time using a novel and powerful optical technique: total-internalreflection fluorescence microscopy (TIRFM). By using selective evanescent illumination of the thin section ( 200 nm) just above the interface between the glass coverslip and the adhered cell, TIRFM is instrumental in gaining insight into dynamic events occurring at, or close to, the plasma membrane of living cells, with outstanding optical contrast and resolution. Using TIRFM, it was found that the lateral motion of the subplasmalemmal secretory vesicles in PC12 cells was significantly impeded when a CNT network was used as a cell-growth and -adhesion substrate; this is likely to be caused by membrane deformation on the nanoscale, induced by the underlying nanorough CNT mat. Single-walled carbon nanotubes (SWCNTs) 1 nm in diameter were transfer-printed onto clean glass coverslips. As revealed by atomic force microscopy (AFM, Fig. 1A), SWCNT bundles with diameters on the order of 10 nm formed a thin-film network with mesh size much less than 1mm. It is worth noting that such a CNT net has a surface roughness comparable to the nanoscale of native ECM constituents such as collagen fibers, and is at least two orders of magnitude thinner than a typical cell. CNT coverslips were washed, sterilized, and then coated with poly(L-lysine) to promote cell adhesion. As verified by AFM imaging, the poly(L-lysine) coating did not cause any appreciable alteration to the surface roughness. PC12 cells cultured on the CNT coverslips (Fig. 1B, bottom) were indistinguishable from those cultured on bare glass coverslips coated with poly(L-lysine)