Nanoenabled Direct Contact Interfacing of Syringe-Injectable Mesh Electronics.

Polymer-based electronics with low bending stiffnesses and high flexibility, including recently reported macroporous syringe-injectable mesh electronics, have shown substantial promise for chronic studies of neural circuitry in the brains of live animals. A central challenge for exploiting these highly flexible materials for in vivo studies has centered on the development of efficient input/output (I/O) connections to an external interface with high yield, low bonding resistance, and long-term stability. Here we report a new paradigm applied to the challenging case of injectable mesh electronics that exploits the high flexibility of nanoscale thickness two-sided metal I/O pads that can deform and contact standard interface cables in high yield with long-term electrical stability. First, we describe the design and facile fabrication of two-sided metal I/O pads that allow for contact without regard to probe orientation. Second, systematic studies of the contact resistance as a function of I/O pad design and mechanical properties demonstrate the key role of the I/O pad bending stiffness in achieving low-resistance stable contacts. Additionally, computational studies provide design rules for achieving high-yield multiplexed contact interfacing in the case of angular misalignment such that adjacent channels are not shorted. Third, the in vitro measurement of 32-channel mesh electronics probes bonded to interface cables using the direct contact method shows a reproducibly high yield of electrical connectivity. Finally, in vivo experiments with 32-channel mesh electronics probes implanted in live mice demonstrate the chronic stability of the direct contact interface, enabling consistent tracking of single-unit neural activity over at least 2 months without a loss of channel recording. The direct contact interfacing methodology paves the way for scalable long-term connections of multiplexed mesh electronics neural probes for neural recording and modulation and moreover could be used to facilitate a scalable interconnection of other flexible electronics in biological studies and therapeutic applications.

Syringe-injectable Mesh Electronics for Stable Chronic Rodent Electrophysiology.

Implantable brain electrophysiology probes are valuable tools in neuroscience due to their ability to record neural activity with high spatiotemporal resolution from shallow and deep brain regions. Their use has been hindered, however, by mechanical and structural mismatches between the probes and brain tissue that commonly lead to micromotion and gliosis with resulting signal instability in chronic recording experiments. In contrast, following the implantation of ultraflexible mesh electronics via syringe injection, the mesh probes form a seamless, gliosis-free interface with the surrounding brain tissue that enables stable tracking of individual neurons on at least a year timescale. This protocol details the key steps in a typical mouse neural recording experiment using syringe-injectable mesh electronics, including the fabrication of mesh electronics in a standard photolithography-based process possible at many universities, loading mesh electronics into standard capillary needles, stereotaxic injection in vivo, connection of the mesh input/output to standard instrumentation interfaces, restrained or freely moving recording sessions, and histological sectioning of brain tissue containing mesh electronics. Representative neural recordings and histology data are presented. Investigators familiar with this protocol will have the knowledge necessary to incorporate mesh electronics into their own experiments and take advantage of the unique opportunities afforded by long-term stable neural interfacing, such as studies of aging processes, brain development, and the pathogenesis of brain disease.

Syringe-Injectable Electronics with a Plug-and-Play Input/Output Interface.

Syringe-injectable mesh electronics represent a new paradigm for brain science and neural prosthetics by virtue of the stable seamless integration of the electronics with neural tissues, a consequence of the macroporous mesh electronics structure with all size features similar to or less than individual neurons and tissue-like flexibility. These same properties, however, make input/output (I/O) connection to measurement electronics challenging, and work to-date has required methods that could be difficult to implement by the life sciences community. Here we present a new syringe-injectable mesh electronics design with plug-and-play I/O interfacing that is rapid, scalable, and user-friendly to nonexperts. The basic design tapers the ultraflexible mesh electronics to a narrow stem that routes all of the device/electrode interconnects to I/O pads that are inserted into a standard zero insertion force (ZIF) connector. Studies show that the entire plug-and-play mesh electronics can be delivered through capillary needles with precise targeting using microliter-scale injection volumes similar to the standard mesh electronics design. Electrical characterization of mesh electronics containing platinum (Pt) electrodes and silicon (Si) nanowire field-effect transistors (NW-FETs) demonstrates the ability to interface arbitrary devices with a contact resistance of only 3 Ω. Finally, in vivo injection into mice required only minutes for I/O connection and yielded expected local field potential (LFP) recordings from a compact head-stage compatible with chronic studies. Our results substantially lower barriers for use by new investigators and open the door for increasingly sophisticated and multifunctional mesh electronics designs for both basic and translational studies.

Syringe-injectable mesh electronics integrate seamlessly with minimal chronic immune response in the brain.

Implantation of electrical probes into the brain has been central to both neuroscience research and biomedical applications, although conventional probes induce gliosis in surrounding tissue. We recently reported ultraflexible open mesh electronics implanted into rodent brains by syringe injection that exhibit promising chronic tissue response and recording stability. Here we report time-dependent histology studies of the mesh electronics/brain-tissue interface obtained from sections perpendicular and parallel to probe long axis, as well as studies of conventional flexible thin-film probes. Confocal fluorescence microscopy images of the perpendicular and parallel brain slices containing mesh electronics showed that the distribution of astrocytes, microglia, and neurons became uniform from 2-12 wk, whereas flexible thin-film probes yield a marked accumulation of astrocytes and microglia and decrease of neurons for the same period. Quantitative analyses of 4- and 12-wk data showed that the signals for neurons, axons, astrocytes, and microglia are nearly the same from the mesh electronics surface to the baseline far from the probes, in contrast to flexible polymer probes, which show decreases in neuron and increases in astrocyte and microglia signals. Notably, images of sagittal brain slices containing nearly the entire mesh electronics probe showed that the tissue interface was uniform and neurons and neurofilaments penetrated through the mesh by 3 mo postimplantation. The minimal immune response and seamless interface with brain tissue postimplantation achieved by ultraflexible open mesh electronics probes provide substantial advantages and could enable a wide range of opportunities for in vivo chronic recording and modulation of brain activity in the future.

Stable long-term chronic brain mapping at the single-neuron level.

Stable in vivo mapping and modulation of the same neurons and brain circuits over extended periods is critical to both neuroscience and medicine. Current electrical implants offer single-neuron spatiotemporal resolution but are limited by such factors as relative shear motion and chronic immune responses during long-term recording. To overcome these limitations, we developed a chronic in vivo recording and stimulation platform based on flexible mesh electronics, and we demonstrated stable multiplexed local field potentials and single-unit recordings in mouse brains for at least 8 months without probe repositioning. Properties of acquired signals suggest robust tracking of the same neurons over this period. This recording and stimulation platform allowed us to evoke stable single-neuron responses to chronic electrical stimulation and to carry out longitudinal studies of brain aging in freely behaving mice. Such advantages could open up future studies in mapping and modulating changes associated with learning, aging and neurodegenerative diseases.

Syringe Injectable Electronics: Precise Targeted Delivery with Quantitative Input/Output Connectivity.

Syringe-injectable mesh electronics with tissue-like mechanical properties and open macroporous structures is an emerging powerful paradigm for mapping and modulating brain activity. Indeed, the ultraflexible macroporous structure has exhibited unprecedented minimal/noninvasiveness and the promotion of attractive interactions with neurons in chronic studies. These same structural features also pose new challenges and opportunities for precise targeted delivery in specific brain regions and quantitative input/output (I/O) connectivity needed for reliable electrical measurements. Here, we describe new results that address in a flexible manner both of these points. First, we have developed a controlled injection approach that maintains the extended mesh structure during the "blind" injection process, while also achieving targeted delivery with ca. 20 µm spatial precision. Optical and microcomputed tomography results from injections into tissue-like hydrogel, ex vivo brain tissue, and in vivo brains validate our basic approach and demonstrate its generality. Second, we present a general strategy to achieve up to 100% multichannel I/O connectivity using an automated conductive ink printing methodology to connect the mesh electronics and a flexible flat cable, which serves as the standard "plug-in" interface to measurement electronics. Studies of resistance versus printed line width were used to identify optimal conditions, and moreover, frequency-dependent noise measurements show that the flexible printing process yields values comparable to commercial flip-chip bonding technology. Our results address two key challenges faced by syringe-injectable electronics and thereby pave the way for facile in vivo applications of injectable mesh electronics as a general and powerful tool for long-term mapping and modulation of brain activity in fundamental neuroscience through therapeutic biomedical studies.