Arati Sridharan

Long-term changes in the material properties of brain tissue at the implant–tissue interface

OBJECTIVE Brain tissue undergoes dramatic molecular and cellular remodeling at the implant-tissue interface that evolves over a period of weeks after implantation. The biomechanical impact of such remodeling on the interface remains unknown. In this study, we aim to assess the changes in the mechanical properties of the brain-electrode interface after chronic implantation of a microelectrode. APPROACH Microelectrodes were implanted in the rodent cortex at a depth of 1 mm for different durations-1 day (n = 4), 10-14 days (n = 4), 4 weeks (n = 4) and 6-8 weeks (n = 7). After the initial duration of implantation, the microelectrodes were moved an additional 1 mm downward at a constant speed of 10 µm s(-1). Forces experienced by the microelectrode were measured during movement and after termination of movement. The biomechanical properties of the interfacial brain tissue were assessed from measured force-displacement curves using two separate models-a two-parameter Mooney-Rivlin hyperelastic model and a viscoelastic model with a second-order Prony series. MAIN RESULTS Estimated shear moduli using a second-order viscoelastic model increased from 0.5-2.6 kPa (day 1 of implantation) to 25.7-59.3 kPa (after 4 weeks of implantation) and subsequently decreased to 0.8-7.9 kPa after 6-8 weeks of implantation in 6 of the 7 animals. The estimated elastic modulus increased from 4.1-7.8 kPa on the day of implantation to 24-44.9 kPa after 4 weeks. The elastic modulus was estimated to be 6.8-33.3 kPa in 6 of the 7 animals after 6-8 weeks of implantation. The above estimates suggest that the brain tissue surrounding the microelectrode evolves from a stiff matrix with maximal shear and elastic modulus after 4 weeks of implantation into a composite of two different layers with different mechanical properties-a stiff compact inner layer surrounded by softer brain tissue that is biomechanically similar to brain tissue-during the first week of implantation. Tissue micromotion-induced stresses on the microelectrode constituted 12-55% of the steady-state stresses on the microelectrode on the day of implantation (n = 4), 2-21% of the steady-state stresses after 4 weeks of implantation (n = 4), and 4-10% of the steady-state stresses after 6-8 weeks of implantation (n = 7). SIGNIFICANCE Understanding biomechanical behavior at the brain-microelectrode interface is necessary for the long-term success of implantable neuroprosthetics and microelectrode arrays. Such quantitative physical characterization of the dynamic changes in the electrode-tissue interface will (a) drive the design and development of more mechanically optimal, chronic brain implants, and (b) lead to new insights into key cellular and molecular events such as neuronal adhesion, migration and function in the immediate vicinity of the brain implant.

Long-Term Neural Recordings Using MEMS Based Movable Microelectrodes in the Brain

One of the critical requirements of the emerging class of neural prosthetic devices is to maintain good quality neural recordings over long time periods. We report here a novel MEMS (Micro Electro Mechanical Systems) based technology that can move microelectrodes in the event of deterioration in neural signal to sample a new set of neurons. Microscale electro-thermal actuators are used to controllably move microelectrodes post-implantation in steps of approximately 9 μm. In this study, a total of 12 movable microelectrode chips were individually implanted in adult rats. Two of the twelve movable microelectrode chips were not moved over a period of 3 weeks and were treated as control experiments. During the first 3 weeks of implantation, moving the microelectrodes led to an improvement in the average signal to noise ratio (SNR) from 14.61 ± 5.21 dB before movement to 18.13 ± 4.99 dB after movement across all microelectrodes and all days. However, the average root-mean-square values of noise amplitudes were similar at 2.98 ± 1.22 μV and 3.01 ± 1.16 μV before and after microelectrode movement. Beyond 3 weeks, the primary observed failure mode was biological rejection of the PMMA (dental cement) based skull mount resulting in the device loosening and eventually falling from the skull. Additionally, the average SNR for functioning devices beyond 3 weeks was 11.88 ± 2.02 dB before microelectrode movement and was significantly different (p < 0.01) from the average SNR of 13.34 ± 0.919 dB after movement. The results of this study demonstrate that MEMS based technologies can move microelectrodes in rodent brains in long-term experiments resulting in improvements in signal quality. Further improvements in packaging and surgical techniques will potentially enable movable microelectrodes to record cortical neuronal activity in chronic experiments.

Adaptive Movable Neural Interfaces for Monitoring Single Neurons in the Brain

Implantable microelectrodes that are currently used to monitor neuronal activity in the brain in vivo have serious limitations both in acute and chronic experiments. Movable microelectrodes that adapt their position in the brain to maximize the quality of neuronal recording have been suggested and tried as a potential solution to overcome the challenges with the current fixed implantable microelectrodes. While the results so far suggest that movable microelectrodes improve the quality and stability of neuronal recordings from the brain in vivo, the bulky nature of the technologies involved in making these movable microelectrodes limits the throughput (number of neurons that can be recorded from at any given time) of these implantable devices. Emerging technologies involving the use of microscale motors and electrodes promise to overcome this limitation. This review summarizes some of the most recent efforts in developing movable neural interfaces using microscale technologies that adapt their position in response to changes in the quality of the neuronal recordings. Key gaps in our understanding of the brain–electrode interface are highlighted. Emerging discoveries in these areas will lead to success in the development of a reliable and stable interface with single neurons that will impact basic neurophysiological studies and emerging cortical prosthetic technologies.

Immobilization of functional light antenna structures derived from the filamentous green bacterium Chloroflexus aurantiacus.

The integration of highly efficient, natural photosynthetic light antenna structures into engineered systems while their biophotonic capabilities are maintained has been an elusive goal in the design of biohybrid photonic devices. In this study, we report a novel technique to covalently immobilize nanoscaled bacterial light antenna structures known as chlorosomes from Chloroflexus aurantiacus on both conductive and nonconductive glass while their energy transducing functionality was maintained. Chlorosomes without their reaction centers (RCs) were covalently immobilized on 3-aminoproyltriethoxysilane (APTES) treated surfaces using a glutaraldehyde linker. AFM techniques verified that the chlorosomes maintained their native ellipsoidal ultrastructure upon immobilization. Results from absorbance and fluorescence spectral analysis (where the Stokes shift to 808/810 nm was observed upon 470 nm blue light excitation) in conjunction with confocal microscopy confirm that the functional integrity of immobilized chlorosomes was also preserved. In addition, experiments with electrochemical impedance spectroscopy (EIS) suggested that the presence of chlorosomes in the electrical double layer of the electrode enhanced the electron transfer capacity of the electrochemical cell. Further, chronoamperometric studies suggested that the reduced form of the Bchl- c pigments found within the chlorosome modulate the conduction properties of the electrochemical cell, where the oxidized form of Bchl- c pigments impeded any current transduction at a bias of 0.4 V within the electrochemical cell. The results therefore demonstrate that the intact chlorosomes can be successfully immobilized while their biophotonic transduction capabilities are preserved through the immobilization process. These findings indicate that it is feasible to design biophotonic devices incorporating fully functional light antenna structures, which may offer significant performance enhancements to current silicon-based photonic devices for diverse technological applications ranging from CCD devices used in retinal implants to terrestrial and space fuel cell applications.

Packaging and Non-Hermetic Encapsulation Technology for Flip Chip on Implantable MEMS Devices

We report here a successful demonstration of a flip-chip packaging approach for a microelectromechanical systems (MEMS) device with in-plane movable microelectrodes implanted in a rodent brain. The flip-chip processes were carried out using a custom-made apparatus that was capable of the following: creating Ag epoxy microbumps for first-level interconnect; aligning the die and the glass substrate; and creating nonhermetic encapsulation (NHE). The completed flip-chip package had an assembled weight of only 0.5 g significantly less than the previously designed wire-bonded package of 4.5 g. The resistance of the Ag bumps was found to be negligible. The MEMS microelectrodes were successfully tested for its mechanical movement with microactuators generating forces of 450 with a displacement resolution of 8.8 . An NHE on the front edge of the package was created by patterns of hydrophobic silicone microstructures to prevent contamination from cerebrospinal fluid while simultaneously allowing the microelectrodes to move in and out of the package boundary. The breakdown pressure of the NHE was found to be 80 cm of water, which is significantly (4.5-11 times) larger than normal human intracranial pressures. Bench top tests and in vivo tests of the MEMS flip-chip packages for up to 75 days showed reliable NHE for potential long-term implantation.

Voltage Preconditioning Allows Modulated Gene Expression in Neurons Using PEI-complexed siRNA

We present here a high efficiency, high viability siRNA-delivery method using a voltage-controlled chemical transfection strategy to achieve modulated delivery of polyethylenimine (PEI) complexed with siRNA in an in vitro culture of neuro2A cells and neurons. Low voltage pulses were applied to adherent cells before the administration of PEI-siRNA complexes. Live assays of neuro2a cells transfected with fluorescently tagged siRNA showed an increase in transfection efficiency from 62 ± 14% to 98 ± 3.8% (after −1 V). In primary hippocampal neurons, transfection efficiencies were increased from 30 ± 18% to 76 ± 18% (after −1 V). Negligible or low-level transfection was observed after preconditioning at higher voltages, suggesting an inverse relationship with applied voltage. Experiments with propidium iodide ruled out the role of electroporation in the transfection of siRNAs suggesting an alternate electro-endocytotic mechanism. In addition, image analysis of preconditioned and transfected cells demonstrates siRNA uptake and loading that is tuned to preconditioning voltage levels. There is approximately a fourfold increase in siRNA loading after preconditioning at −1 V compared with the same at ±2–3 V. Modulated gene expression is demonstrated in a functional knockdown of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) in neuro2A cells using siRNA. Cell density and dendritic morphological changes are also demonstrated in modulated knockdown of brain derived neurotrophic factor (BDNF) in primary hippocampal neurons. The method reported here has potential applications in the development of high-throughput screening systems for large libraries of siRNA molecules involving difficult-to-transfect cells like neurons.

Optoelectronic Energy Transfer at Novel Biohybrid Interfaces Using Light Harvesting Complexes from Chloroflexus aurantiacus

In nature, nanoscale supramolecular light harvesting complexes initiate the photosynthetic energy collection process at high quantum efficiencies. In this study, the distinctive antenna structure from Chloroflexus aurantiacusthe chlorosomeis assessed for potential exploitation in novel biohybrid optoelectronic devices. Electrochemical characterization of bacterial fragments containing intact chlorosomes with the photosynthetic apparatus show an increase in the charge storage density near the working electrode upon light stimulation and suggest that chlorosomes contribute approximately one-third of the overall photocurrent. Further, isolated chlorosomes (without additional photosynthetic components, e.g., reaction centers, biochemical mediators) produce a photocurrent (approximately 8-10 nA) under light saturation conditions. Correlative experiments indicate that the main chlorosome pigment, bacteriochlorophyll-c, contributes to the photocurrent via an oxidative mechanism. The results reported herein are the first to demonstrate that isolated chlorosomes (lipid-enclosed sacs of pigments) directly transduce light energy in an electrochemical manner, laying an alternative, biomimetic approach for designing photosensitized interfaces in biofuel cells and biomedical devices, such as bioenhanced retinal prosthetics.

Work in progress - using case studies to increase the retention of female doctoral students in STEM Fields

This paper discusses part of the NSF funded project, CareerWISE: Internet-delivered resilience training to increase the persistence of Women Ph.D. Students in STEM Fields. Research shows that rates of doctoral student attrition (i.e., beginning and then not completing a doctoral degree program) are substantially higher for women than for men, including in the physical sciences and engineering. The CareerWISE project addresses this problem through the development of a Web-based resource consisting of training materials that will build the resilience skills, problem solving skills, and persistence of women in science and engineering PhD programs. CareerWISE is currently in the second of three years, and this paper focuses primarily on the portion of the project involving written case studies. The differences between the types of case studies traditionally used in science and engineering courses and the CareerWISE case studies are discussed, along with the plans for evaluating and integrating the cases with the other pieces of the CareerWISE project.

Biohybrid photoelectrochemical nanoengineered interfaces

Incorporation of biophotonic components in artificial devices is an emerging trend in exploring biomimetic approaches for green technologies. In this study, highly efficient, nanoscaled light antenna structures from green photosynthetic bacteria, known as chlorosomes, comprised of bacteriochlorophyll-c pigment arrays that are stable in aqueous environments are studied in an electrochemical environment for their photoelectrogenic capacity. Biohybrid electrochemical cells containing chlorosomes coupled to the native bacterial photosynthetic apparatus have a higher dark charge storage density (at least 10-fold) than electrochemical cells with decoupled chlorosomes. Nevertheless, upon light stimulation, the charge storage density, also known as charge injection capacity, for both electrochemical systems increased the charge stored near the electrode. Decoupled chlorosome-based systems showed a light-intensity dose-dependent response reaching a maximum change of ˜300 nC/cm 2 at near sunlight intensities (˜80-100mW/cm 2 ). Chronoamperometric studies under light stimulation conditions confirmed the photo-induced effect. Current studies are focused on optimization of the electrode/chlorosome interfacial properties across various heterogeneous interfaces. Successful implementation of harvesting photo-energy using the chlorosome or its derivatives may lead to substantial innovations in current biophotonic technologies, such as biofuel cells and retinal prosthetics.