Carbon Nanotube Sites for Neural-Network Patterning and Recording Tamir Gabay 1, Itshak Kalifa 1, Lisa Ezra 1, Eshel Ben-Jacob 2, Yael Hanein 1 1 School of Electrical Engineering, department of physical electronics, 2 School of Physics and Astronomy Tel-Aviv University, Tel-Aviv 69978, Israel Extra-cellular recordings with multi electrode array (MEAs) systems have been used for the last several decades to study the formation and behavior of in- vitro neuronal networks. It is widely accepted that improved MEAs, with high resolution and better control over cell density and patterning, are expected to be useful to expand our understanding of high brain functions and to facilitate novel neuro-chip sensors. This work presents a new MEA configuration, which enables the formation of electrically viable engineered neuronal networks with high-resolution extracellular recording. The networks are engineered according to lithographically defined carbon nanotube (CNT) templates. These CNT templates strongly anchor cells and enable the formation of stable networks. Carbon nanotube coated surfaces are biocompatible, provide an excellent surface for cell growth and offer high specific capacitance crucial in electrochemical applications. According to the new MEA scheme molybdenum lines electrically connect the CNT templates and external instrumentation. Inverted microscope images of interconnected network formed with CNT islands. (Left) after 96 hours, (Center) after 128 hours and (Right) after 150 hours after plating. Cell density was 1x10 -3 cells/µm 2. Arrows indicate well defined connections between neuronal clusters. (Scale bar is 150µm) Inverted microscope images of interconnected networks formed with CNT islands at a cell density of 4.5×10-3 cells/µm2 at (Left) 96 hours, (Center) 128 hours and (Right) 150 hours after plating the cells on the substrate. At this high cell density mechanical tension overpowers cell adhesion to the CNT islands. Scale bar is 150 µm. Inverted microscope images of interconnected neuronal systems formed with CNT islands. (Left) One hour after cell deposition cells are randomly distributed (scale bar is 150 µm). (Center) After four days neurons form clusters at the CNT sites, which form connections between these islands (scale bar is 150 µm). Some islands are not yet connected at this stage and several short and unconnected dendrites and axons are apparent. (Right) Neuron cluster at a CNT island (scale bar is 100 µm). Cell density was 4.5×10 -3 µm -2. Assembly Mechanisms Abstract Neural network collapse into isolated clusters due to tension. (Left) The first step of this process is based mainly of self-wiring of the neurons with only limited neuronal migration. (Right) In a later stage the network segregates into separate clusters. Neural network formation from nucleation centers. (Left) In the first step neuronal migration from low affinity surface results in cell aggregation at high affinity regions (large, dark disks). (Right) In a second step high density neuronal islands form well defined interconnection following the shortest link between islands. Tel Aviv Nano-Technology Workshop, Maagan, April 2005 Micro and Nanofabrication Products Top – Optical microscope image of regular array of molybdenum pads (Dia. 150µm) coated by carbon nanotube. Uncoated molybdenum traces are shown also. Middle – HRSEM (El-Mul Corp.) image of one of the molybdenum pads surfaces, and Bottom – HRSEM (El-Mul Corp.) image of a typical edge of such a molybdenum pad. CVD system Cyclic Voltammetry Cyclic voltammetry experiment results of two 100 µm diameter molybdenum electrodes. Dashed line – electrode coated by carbon nanotube layer; continues line – bare