Highly precise optogenetic and electrophysiological studies provide an approach to correlate the neural activity of specific cells to observed behaviors, which is important for the explorations of neural circuits in the brain and the pathologies of the nervous system. Embedded optical fibers and electrode probes are the most common invasive devices in the studies of these areas. Previous studies have shown that foreign body reactions due to chronic tissue damage lead to gradual encapsulation of the implanted probes, resulting in dysfunction of the devices. Over the past two decades, researchers have experimented with advanced materials to fabricate invasive electrodes and optical fiber devices that are mechanically compatible with soft brain tissue, thereby reducing the relative micromotion and damage between the device interface and brain tissue. Extensive efforts have been made in this area; however, challenges have yet to be overcome for practical applications. First, the assembly of the optical fiber and electrode(s) generally results in a significant increase in the volume of the implant, which produces additional tissue damage. Second, an alternative strategy is accommodating recording and waveguide channels in fibers through a thermally drawn process. However, this method has difficulty breaking the limit of the number of recorded sites (usually less than 30). Third, existing optical waveguide materials, such as polycarbonate and quartz, have a Young's modulus far greater than that of brain tissue. Probes made of softer biocompatible materials, on the other hand, are mechanically incapable of embedding themselves into brain tissue. Moreover, the lack of rigidity poses challenges in the implantation into living brains. A rigid shuttle device is usually required for the implantation of a flexible probe. However, the insertion and retraction of the shuttle device inevitably cause extra damage to the brain tissue. Natural silk has remarkable application prospects in biomedical and bio-optical devices due to its excellent physical and biomedical properties, such as transparency, mechanical strength and crucial bio-compatibility. Here, we present an optoelectronic probe, Silk-Optrode, with natural silk as an optical waveguide material, which can simultaneously achieve high-precision intracranial light stimulation and electrophysiological recording. Through the attachment of the flexible electrode array to the surface of the silk optical fiber, 128 recording channels can be integrated on a single probe. Meanwhile, the diameter of the probe does not notably increase. Compared with polylactic acid (PLA) and other polymer materials as previously reported for making implantable flexible optical fibers, silk has the advantage of being adjustable under different hydration conditions. The Silk-Optrode can actively transform from a hard brittle state to a flexible and high elongation state after implantation (bending stiffness decreased by ~11 times), thus simultaneously achieving the simplest implant operation and causing less chronic tissue damage. Low optical transmission loss makes silk-based optical fibers suitable for optogenetic studies as waveguides. In these studies, Silk-Optrode achieved the effective regulation of neural activity and behavior while recording more than 50 high-quality isolated unit activities within one experiment. We envision that this technology will provide new opportunities for the combination of multifunctional biomaterial devices with neurological disease research.

The introduction section extensively reviews the existing literature on optogenetic and electrophysiological neural probes. It highlights the challenges associated with current technologies, such as the difficulty in combining optical fibers and electrodes without increasing implant volume, limitations in the number of recording sites achievable with thermally drawn fibers, and the mismatch in Young's modulus between existing waveguide materials and brain tissue. The review emphasizes the need for biocompatible, flexible probes that can be implanted easily while minimizing tissue damage. The authors specifically mention previous work using advanced materials for electrodes and optical fibers, but point out that significant challenges remain. The literature review then introduces natural silk as a promising alternative due to its transparency, biocompatibility, and adjustable mechanical properties.

The methodology section details the fabrication and characterization of the Silk-Optrode probe. First, the fabrication of uniform, high-transparency silk optical fibers is described, emphasizing the challenges of eliminating air bubbles and the process used to achieve this. The mechanical properties of the silk fibers in both hydrated and dehydrated states are characterized through tensile-fracture experiments, showing a significant decrease in elastic modulus and bending stiffness upon hydration. The authors then describe the fabrication of the flexible electrode array using microfabrication techniques, highlighting its ultra-flexibility and low bending stiffness. The integration of the electrode array onto the silk optical fiber is explained, including the use of a biological binder (PEG) to secure the array to the fiber's surface. The electrodeposition of PEDOT onto the recording sites is also detailed, along with its effect on reducing interfacial impedance. Finally, the methodology section describes the in vivo experimental setup, including virus injection into the mouse brain, implantation of the Silk-Optrode probe, and the recording and analysis of neuronal activity. Finite element analysis (FEA) is used to simulate probe-tissue interactions and compare the mechanical properties of silk, polycarbonate, silica, and steel probes during brain micromotion.

The key findings demonstrate the successful fabrication and in vivo application of the Silk-Optrode probe. The silk optical fibers exhibited low optical transmission loss (3.0 dB/cm), suitable for optogenetic stimulation. The hydration of the silk fiber enabled a self-adaptive change in mechanical properties, transitioning from a rigid state suitable for implantation to a flexible state that conforms to the surrounding brain tissue. FEA simulations confirmed the superior mechanical compatibility of the silk-based probe compared to probes made of other materials (steel, silica, PC), exhibiting significantly reduced strain and stress within the brain tissue during micromotion. The 128-channel flexible electrode array demonstrated high fabrication yield (85%) and successfully recorded high-quality neuronal signals in awake, freely moving mice. In one experiment, 57 well-isolated units were recorded, yielding 0.52 units per channel, a significant improvement over previous technologies. Long-term implantation studies (two months) showed reduced immunoreactive glial responses and tissue lesions compared to conventional probes.

The findings address the research question by demonstrating a novel, biocompatible, and highly functional neural probe. The self-adaptive nature of the silk optical fiber allows for accurate implantation with minimal tissue damage, while maintaining excellent optical and electrical performance. The high channel count and successful recording of numerous well-isolated units highlight the probe's potential for advanced neurophysiological studies. The results demonstrate significant advancements in the field of neural interfaces, providing a superior alternative to existing technologies. The superior biocompatibility, as indicated by reduced glial response and tissue damage, suggests that the Silk-Optrode could potentially enhance the longevity and reliability of chronic neural recordings. The successful integration of optogenetic stimulation and multichannel electrophysiological recording opens up new avenues for investigating brain function and treating neurological disorders.

This study successfully demonstrated a novel silk-based self-adaptive flexible opto-electro neural probe (Silk-Optrode) capable of high-fidelity simultaneous optogenetic stimulation and multichannel neural recording. The unique properties of silk, combined with advanced microfabrication techniques, enabled the creation of a probe with superior biocompatibility, flexibility, and high channel count. The results suggest that the Silk-Optrode offers significant advantages over existing neural probes and holds substantial promise for advancing the understanding of brain function and treating neurological diseases. Future research could focus on further miniaturization of the probe, increasing the number of recording channels, and exploring the application of this technology in different brain regions and animal models.

While the Silk-Optrode shows significant promise, there are limitations to consider. The study primarily focused on the medial prefrontal cortex (mPFC) in mice, and the generalizability to other brain regions and species needs further investigation. Long-term studies beyond two months are needed to fully assess the long-term biocompatibility and stability of the implant. The current fabrication process might require optimization for large-scale production. Furthermore, while the PEG binder was effective, exploring alternative bio-compatible adhesives could further improve integration and long-term stability.