Search

Xylella fastidiosa is a xylem-limited nonflagellated bacterium that causes economically important diseases of plants by developing biofilms that block xylem sap flow. How the bacterium is translocated downward in the host plant's vascular system against the direction of the transpiration stream has long been a puzzling phenomenon. Using microfabricated chambers designed to mimic some of the features of xylem vessels, we discovered that X. fastidiosa migrates via type IV-pilus-mediated twitching motility at speeds up to 5 [micro]m [min-.sup.1] against a rapidly flowing medium (20,000 [micro]m [min-.sup.1]). Electron microscopy revealed that there are two length classes of pili, long type IV pili (1.0 to 5.8 [micro]m) and short type I pili (0.4 to 1.0 [micro]m). We further demonstrated that two knockout mutants (pilB and pilQ mutants) that are deficient in type IV pili do not twitch and are inhibited from colonizing upstream vascular regions in planta. In addition, mutants with insertions in pilB or pilQ (possessing type I pili only) express enhanced biofilm formation, whereas a mutant with an insertion in fimA (possessing only type IV pili) is biofilm deficient.

Model silicon intracortical probes with microfluidic channels were fabricated and tested to examine the feasibility of using diffusion-mediated delivery to deliver therapeutic agents into the volume of tissue exhibiting reactive responses to implanted devices. Three-dimensional probe structures with microfluidic channels were fabricated using surface micromachining and deep reactive ion etching (DRIE) techniques. In vitro functional tests of devices were performed using fluorescence microscopy to record the transient release of Texas Red labeled transferrin (TR-transferrin) and dextran (TR-dextran) from the microchannels into 1% w/v agarose gel. In vivo performance was characterized by inserting devices loaded with TR-transferrin into the premotor cortex of adult male rats. Brain sections were imaged using confocal microscopy. Diffusion of TR-transferrin into the extracellular space and uptake by cells up to 400 [micro]m from the implantation site was observed in brain slices taken 1 h postinsertion. The reactive tissue volume, as indicated by the presence of phosphorylated mitogen-activated protein kinases (MAPKs), was characterized using immunohistochemistry and confocal microscopy. The reactive tissue volume extended 600, 800, and 400 [micro]m radially from the implantation site at 1 h, 24 h, and 6 weeks following insertion, respectively. These results indicate that diffusion-mediated delivery can be part of an effective intervention strategy for the treatment of reactive tissue responses around chronically implanted intracortiral probes. Index Terms--Biocompatibility, drug delivery, microfluidics, neural prostheses, reactive response.

Neuronal cell networks have been reconstructed on planar microelectrode arrays (MEAs) from dissociated hippocampal pyramidal neurons. Microcontact printing ([micro]CP) and a photoresist-liftoff method were used to selectively localize poly-L-lysine (PLL) on the surface of MEAs. Haptotaxis led to the organization of the neurons into networks localized adjacent to microelectrodes. Various grids of PLL with 2-25-[micro]m-wide lines spaced by 50-200 [micro]m with 15-25-[micro]m nodes at intersection points were used to guide cell body attachment and neurite outgrowth. Bursting activity with spike amplitude attenuation was observed and multichannel recordings detected instances of coincident firing activity. Finally, we present here an extracellular recording from a ~2 [micro]m bundle of guided neurites. Index Terms--Hippocampal pyramidal neuron, microcontact printing, microelectrode array, neuron networks.