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ABSTRACT: The beam profile of an electron beam (EB) can be focused onto less than a nanometer spot and scanned over a wide field with extremely high speed sweeping. Thus, EB is employed for nano scale lithography in applied physics research studies and in fabrication of semiconductors. We applied a scanning EB as a control system for a living cell membrane which is representative of large scale complex systems containing nanometer size components. First, we designed the opposed co-axial dual optics containing an inverted electron beam lithography (I-EBL) system and a fluorescent optical microscope. This system could provide in situ nano processing for a culturing living cell on a 100-nm-thick SiN nanomembrane, which was placed between the I-EBL and the fluorescent optical microscope. Then we demonstrated the EB-induced chemical direct nano processing for a culturing cell with hundreds of nanometer resolution and visualized real-time images of the scanning spot of the EB-induced luminescent emission and chemical processing using a high sensitive camera mounted on the optical microscope. We concluded that our closed-loop in situ nano processing would be able to provide a nanometer resolution display of virtual molecule environments to study functional changes of bio-molecule systems.
Biochemical and Biophysical Research Communications 02/2013; · 2.48 Impact Factor
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ABSTRACT: A pair of exoskeletal limbs for a rat has been developed based on a vehicular Brain-Machine Interface "Rat-Car". The "RatCar" is a whole-body motor prosthesis system for a rat developed by the authors, estimating locomotion velocity according to neural signals pattern to move the rat body by the vehicle instead of its original limbs. In this paper, exoskeletal limbs have displaced the wheels for more natural modality of body control. The system was tested by applying peripheral nerve signals from a behaving rat.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 08/2012; 2012:1707-10.
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ABSTRACT: We present a simple method to regulate the direction of axon development in cultured neurons using microfabrication and microfluidics techniques. We fabricate a PDMS-based device and place it onto a chemically micropatterned glass substrate. We confirm that cultured neurons extend neurites along the medium flow direction and the micropatterned regions.
Journal of Bioscience and Bioengineering 04/2012; 114(1):92-5. · 1.79 Impact Factor
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ABSTRACT: We improved adhesiveness of a neuron-like cell, PC12, on a Parylene-C surface by O(2) plasma treatment which changes the surface from hydrophobic to hydrophilic. Neural cell adhesiveness on the plasma-treated Parylene-C was more than twenty times better compared to non-treated Parylene-C and it was close to that on a conventional polystyrene tissue-culture dish.
Journal of Bioscience and Bioengineering 12/2011; 113(3):395-8. · 1.79 Impact Factor
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11/2011; , ISBN: 978-953-307-607-2
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ABSTRACT: We investigated neuronal cell differentiation, particularly neurite outgrowth, on the surface of diX H and diX AM using an in vitro examination of a neuron-like rat pheochromocytoma cell line, PC12. diX H and diX AM are in the parylene family of diX C (or Parylene-C), which is widely used as a novel coating material to insulate neural electrodes, and they have been recently commercialized; diX H and diX AM offer different features of biocompatibility. Previously, we found that these new parylene materials have high cell adhesiveness to neuronal cells whereas the adhesiveness of diX C is extremely low. However, their cell differentiation remains unknown although neuronal cell differentiation plays a crucial role in their development and regeneration. This study showed that almost all PC12 cells adhering to the surface of diX AM and diX H were differentiated, but the neurite outgrowth was significantly larger on diX H than that on diX AM and a conventional polystyrene culture dish. The result suggests that diX H may be advantageous as a biocompatible coating material for a scaffold, which can be used on virtually any substrate to get various configurations in neural devices.
Biotechnology Progress 10/2011; 28(2):587-90. · 2.34 Impact Factor
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ABSTRACT: Precise control of differentiation processes of pluripotent stem cells is a key component for the further development of regenerative medicine. For this purpose, combining a cell-aggregate-size treatment for regulating intercellular signal transmissions and an electrical stimulation technique for inducing cellular responses is a promising approach. In the present study, we developed microfabricated electrode substrates that allow simultaneous stimulation of embryoid bodies (EBs) of P19 cells. Mouse embryonal carcinoma P19 cells can be induced to differentiate into three germ layers and serve as a promising stem cell model. Microcavity-array patterns were fabricated onto indium-tin-oxide (ITO) substrates using a standard photo-lithography technique, and uniform-sized EBs of P19 cells were inserted into each microcavity. Electrical stimulation was applied to the EBs through substrate electrodes and stimulus-induced intracellular calcium transients were monitored. We confirmed that the developed electrode device could simultaneously stimulate smaller (200μm diameter) and larger (500μm diameter) EBs inserted in the microcavities and induce specific spatio-temporal patterns of intracellular calcium transients in the EBs with fine reproducibility. We concluded that the developed microcavity array with embedded electrodes could simultaneously and effectively stimulate uniform-sized EBs inserted in it. Therefore, it is a promising experimental tool for precisely controlling cell differentiation processes.
Journal of Bioscience and Bioengineering 09/2011; 112(6):624-9. · 1.79 Impact Factor
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ABSTRACT: Regeneration of damaged central nervous systems (CNS) is an important topic in neuroscience and neuroengineering. Grafting new neurons derived from pluripotent stem cells into damaged regions can be done to restore functions after injury. Little is known, however, about network-wide interactions between stem-cell-derived neurons and CNS neurons. In this study, we developed a co-culture method of stem cell-derived neuronal networks and CNS networks and observed spontaneous activity in the co-culture samples. By using a microfabricated poly(dimethylsiloxane) device having two culture compartments and 20 connecting microconduits, we are able to compartmentalize P19-derived neurons and mouse cortical neurons and connect them via the microconduits. Furthermore, we combined the co-culture device and a microelectrode array (MEA)-based recording system and recorded spontaneous activity in the co-cultured networks. We found that periodic synchronized bursting spreading over both neuronal networks occurred during the second week in vitro and that P19-derived neurons in the co-cultured networks had different developmental processes compared with those grown in monoculture. These findings suggest that functional interactions form between P19-dervived neurons and mouse cortical neurons and that the co-culture method is useful for exploring the network-wide integrations between stem cell-derived neurons and CNS neurons.
Bio Systems 08/2011; 107(1):1-8. · 1.27 Impact Factor
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ABSTRACT: The cultured neuronal monolayer has been a promising model system for studying the neuronal dynamics, from single cell to network-wide level. Randomness in the reconstituted network structure has, however, hindered regulated signal transmissions from one neuron to another or from one neuronal population to another. Applying microfabrication-based cell patterning techniques is a promising approach to handling these problems. In the present study, we attempt to regulate the direction of axon development and the pathway of signal transmissions in cultured neuronal networks using micro-fabrication and - fluidic techniques. We created a PDMS-based culture device, which consisted of arrays of U-shaped cell trapping microwells, and placed it onto a chemically micropatterned glass substrate. After 6 days in vitro, we confirmed that cortical neurons extended neurites along the medium flow direction and the micropatterned regions.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 08/2011; 2011:3613-6.
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ABSTRACT: Heat-shock protein90 (HSP90) plays an essential role in maintaining stability and activity of its clients. HSP90 is involved in cell differentiation and survival in a variety of cell types. To elucidate the possible role of HSP90 in myogenic differentiation and cell survival, we examined the time course of changes in the expression of myogenic regulatory factors, intracellular signaling molecules, and anti-/pro-apoptotic factors when C2C12 cells were cultured in differentiation condition in the presence of a HSP90-specific inhibitor, geldanamycin. Furthermore, we examined the effects of geldanamycin on muscle regeneration in vivo. Our results showed that geldanamycin inhibited myogenic differentiation with decreased expression of MyoD, myogenin and reduced phosphorylation levels of Akt1. Geldanamycin had little effect on the phosphorylation levels of p38MAPK and ERK1/2 but reduced the phosphorylation levels of JNK. Along with myogenic differentiation, geldanamycin increased apoptotic nuclei with decreased expression of Bcl-2. The skeletal muscles forced to regenerate in the presence of geldanamycin were of poor repair with small regenerating myofibers and increased connective tissues. Together, our findings suggest that HSP90 may modulate myogenic differentiation and may be involved in cell survival.
Molecular and Cellular Biochemistry 07/2011; 358(1-2):265-80. · 2.06 Impact Factor
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ABSTRACT: The abundance, morphology, and functional properties of mitochondria become altered in response to denervation. To gain insight into the regulation of this process, mitochondrial enzyme activities and gene expression involved in mitochondrial biogenesis and dynamics in mouse gastrocnemius muscle was investigated. Sciatic nerve transactions were performed on mice, and then gastrocnemius muscles were isolated at days 5 and 30 after surgery. Muscle weight was decreased significantly by 15% and 62% at days 5 and 30 after surgery, respectively. The activity of citrate synthase, a marker of oxidative enzyme, was reduced significantly by 31% and 53% at days 5 and 30, respectively. Enzyme histochemical analysis revealed that subsarcolemmal mitochondria were largely lost than intermyofibrillar mitochondria at day 5, and this trend was further progressed at day 30 after surgery. Expression levels of peroxisome proliferator-activated receptor, γ coactivator 1 (PGC-1)α, estrogen-related receptor α (ERRα), and mitofusin 2 were down-regulated throughout the experimental period, whereas those of PGC-1β, PRC, nuclear respiratory factor (NRF)-1, NRF-2, TFAM, and Lon protease were down-regulated at day 30 after surgery. These results suggest that PGC-1α, ERRα, and mitofusin 2 may be important factors in the process of denervation-induced mitochondrial adaptation. In addition, other PGC-1 family of transcriptional coactivators and DNA binding transcription factors may also contribute to mitochondrial adaptation after early response to denervation.
Journal of physiology and biochemistry 03/2011; 67(3):359-70. · 1.71 Impact Factor
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ABSTRACT: In this study, we suggest a new guideline for regeneration microelectrode to be implanted between the severed stumps of peripheral nerves, the microelectrode designed particularly for connecting the signal line of an artificial hand directly to the nerve system. The nerve regeneration microelectrode is an interface device expected to realize a BMI (brain-machine interface).
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 01/2010; 2010:1531-4.
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ABSTRACT: An online neuro-robotic platform in the form of a small vehicle, the "RatCar" has been developed. First, a rat had neural electrodes implanted in the motor cortices to record extracellular potentials. Then, our system combined the rat and its vehicle body by hanging the rat under its floor. In this paper, an experimental platform is proposed to observe and analyze motor commands by correlating neural signals with locomotion states. It was designed to operate in 2 modes; (a) adaptively correlating neural signals and locomotion states to determine motor commands, and (b) applying the estimated motor commands to control the vehicle according to the intention of the rat. As a result, time-varying correlation between neural and locomotion activities has been adaptively visualized in real time to analyze motor commands in various body conditions. In addition, a control of the vehicle has been improved.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 01/2010; 2010:4168-71.
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ABSTRACT: Electromyographic (EMG) signals have been used to control active prosthetic arms for amputees. One of the obstacles in making such prosthetic arms is the timed estimation of posture, because EMG signals and muscle movements are not necessarily synchronized. We estimated the finger motions for trumpet players by using both surface EMG (sEMG) and the timing information using body motion. The algorithms consisted of Principal Component Analysis (PCA), and Support Vector Machine (SVM). The results showed that applying the timing information using body motion increases how precisely the motion of the fingers is estimated.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 01/2010; 2010:5847-50.
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ABSTRACT: A typical prosthetic limb has sensory shortcomings, one of which is lack of kinesthesia. Conventional methods to evoke kinesthesia, which would be required for a precise control of prosthetic limbs, include tendon vibration and skin stretch, but these produce only the sensation of small movements. In this study, tendon vibration is extended to evoke sensations of a more rapid movement. A stimulation method in which vibration is applied to two tendons is proposed, in contrast to most studies in which a single tendon was vibrated. Experimental results indicated that vibration applied to both ends of a muscle produces sensations of more rapid movement than vibration to just one. However, no significant difference in sensation was found between vibrating tendons of synergistic muscles and a single tendon.
Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference 01/2010; 2010:5851-3.
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ABSTRACT: The present study introduces an approach to detect and classify extracellulary recorded action potentials of neurons, usually termed spike sorting. Our approach is based on template matching, which is an optimal filter under Gaussian noise; however, this approach is usually expensive in computational time.To speed up the filter, it is important to curtail the matching process when the distance between template and waveform exceeds some threshold. We approach this aspect of the problem using the frame of similarity detection algorithms (SSDA) and Davies-Bouldin validation indices (DBVI). Windowing pair of the filter was selected in DBVI-based order and a signal which has a rapidly increasing error was discarded to reduce the computational time. DBVI is a function of the ratio of the sum of within-cluster scatter to between-cluster separation; thus, using this order we can expect to separate a signal and a noise in fewer window point calculation than full point matching. This matching process performed well, with a shortter computational time and fewer incorrect classifications than other ordering methods such as time-based or amplitude-based order. © 2009 Wiley Periodicals, Inc. Electron Comm Jpn, 92(11): 57–63, 2009; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecj.10066
Electronics and Communications in Japan 10/2009; 92(11):57 - 63. · 0.08 Impact Factor
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ABSTRACT: DiX C (or Parylene-C) has been widely used as a coating material to insulate neural electrodes in recent decades. However, its uses are limited due to its extremely low adhesiveness with neuronal cells. Other functional materials in the diX family, such as diX A, diX AM, and diX H, have been commercialized recently and would offer different features in biocompatibility from diX C. However, their cell adhesiveness remains unknown. In this work, we used an in vitro approach to investigate how the surface of each material in the diX family affects the degree of neuronal cell adhesiveness compared with a conventional culture dish of polystyrene (PS). The neuronal cell adhesiveness on diX AM and diX H was almost equivalent to that for the PS dish, whereas neuronal cells did not settle on the surface of diX C and diX A. Our results suggest that diX AM and diX H could provide another practical feature as a coating material for a scaffold in a substrate with any configuration in neural devices.
Neuroscience Letters 09/2009; 464(1):26-8. · 2.11 Impact Factor
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ABSTRACT: Neural interface devices that will allow signals from the human nervous system to control external equipment are extremely important for the next generation of prosthetic systems. A novel multichannel regeneration-type nerve electrode designed to record from and stimulate peripheral nerves has been developed to allow the control of artificial hands and to generate artificial sensations. In this study a novel flexible regeneration microelectrode based on the nerve regeneration principle was designed and fabricated using MEMS technologies. The electrode, which was fabricated on a 25-µm-thick parylene C substrate, has multiple fluidic channels. Each fluidic channel was 100µm wide×30µm high×1500µm long and featured multiple electrodes inside them as recording and stimulating sites. They also served as guidance channels for the regenerating axons. © 2009 Wiley Periodicals, Inc. Electron Comm Jpn, 92(4): 29–34, 2009; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/ecj.10059
Electronics and Communications in Japan 04/2009; 92(4):29 - 34. · 0.08 Impact Factor
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ABSTRACT: The undulation pump (formerly called the pre-cessional displacement pump) is a continuous flow displacement-type blood pump that is being developed as an implantable total artificial heart. A new undulation pump was developed for chronic use and was examined with animal experiment and flow visualization studies. In the animal experiment using a left ventricular bypass in goats, severe hemolysis occurred. After driving for 12 h, thrombus formation inside the pump was found. In the flow visualization studies, the flow pattern showed that the flow inside the pump was a very complicated turbulent flow. Improvement of hemolysis and thrombus formation is important to realize implantable total artificial hearts using undulation pumps.
Artificial Organs 11/2008; 19(7):691 - 693. · 2.00 Impact Factor
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ABSTRACT: An online brain-machine interface (BMI) in the form of a small vehicle, the “RatCar,” has been developed. A rat had neural electrodes implanted in its primary motor cortex and basal ganglia regions to continuously record neural signals. Then, a linear state space model represents a correlation between the recorded neural signals and locomotion states (i.e., moving velocity and azimuthal variances) of the rat. The model parameters were set so as to minimize estimation errors, and the locomotion states were estimated from neural firing rates using a Kalman filter algorithm. The results showed a small oscillation to achieve smooth control of the vehicle in spite of fluctuating firing rates with noises applied to the model. Major variation of the model variables converged in a first 30 seconds of the experiments and lasted for the entire one hour session.
Engineering in Medicine and Biology Society, 2008. EMBS 2008. 30th Annual International Conference of the IEEE; 09/2008
