Customizable, wireless and implantable neural probe design and fabrication via 3D printing

Kyle E. Parker, Juhyun Lee, Jenny R. Kim, Chinatsu Kawakami, Choong Yeon Kim, Raza Qazi, Kyung-In Jang, Jae-Woong Jeong, Jordan G. McCall

Published: 2022-10-20 DOI: 10.1038/s41596-022-00758-8

Extended

Extended Data Fig. 1 Technical drawing with dimensions for 3D-printed stereotaxic adapter for cannula holder.

a , b , Top ( a ) and side ( b ) view of 3D-printed stereotaxic adapter.

Extended Data Fig. 2 Bulk preparation of Metabond quick set cement kit.

a , Selected components of the Metabond kit used to secure the implant in its position. b , Add one scoop of Metabond radiopaque L-powder to the chilled ceramic dish. c , Add four drops of Quick Base liquid. d , Add one drop of Catalyst liquid. e , Mix cement preparation with rounded spatula. f , Use spatula to apply it directly to the skull surface.

Extended Data Fig. 3 Technical drawing with dimensions for a 5-mm-long unilateral 3D-POP and a probe holder.

a , Top view (left) and cross-section view (right) of 3D-POP. b , Top view (left) and cross-section view (right) of probe holder.

Extended Data Fig. 4 Technical drawing with dimensions for a supporter and a stencil mask used for PDMS screen printing.

a , Top view (left) and cross-section view (right) of the supporter. b , Top view of stencil mask.

Extended Data Fig. 5 Circuit diagram for the BLE control circuit.

The circuit includes a BLE SoC, a voltage regulator, indicator LEDs (red, green) and connectors for the integration of a lithium polymer (LiPo) battery in a 3D-POP. The voltage regulator converts fluctuating input voltage from the LiPo battery into constant output voltage (3 V) and supplies it to the BLE SoC. The BLE SoC enables wireless control by smartphone and regulates the photostimulation condition of 3D-POPs. Indicator LEDs help to recognize the communication status of the wireless system.

Extended Data Fig. 6 Preparation of the behavioral assay for optogenetic stimulation.

Preparation for wireless operation. Integrate a 3D-POP and the BLE wireless control module. Check LED operation using the smartphone app. b , c , Preparation for a wired operation: solder wires to a male pin connector ( b ), and connect a signal generator to a 3D-POP through the wire and apply electrical pulse signals for testing ( c ).

Supplementary information

Supplementary Data 1

STL file for the stereotaxic adapter

Supplementary Data 2

STL files for the 1D probe

Supplementary Data 3

STL files for PDMS screen printing

Supplementary Data 4

Gerber and CAD files for the wireless control circuit

Supplementary Data 5

Program code for the BLE SoC

Supplementary Data 6

Source codes for the smartphone application

Supplementary Data 7

STL files for the 3 × 3 probe

References

This Protocol Extension describes the low-cost production of rapidly customizable optical neural probes for invivo optogenetics. We detail the use of a 3D printer to fabricate minimally invasive microscale inorganic light-emitting-diode-based neural prob
Introduction
Materials
Procedure 1
- Fabrication of 3D-POPs
Procedure 2
Procedure 3
Troubleshooting
Timing
Anticipated results
- References
- References
References
ACKNOWLEDGMENTS
ACKNOWLEDGMENTS

Deisseroth, K. Optogenetics: 10 years of microbial opsins in neuroscience. Nat. Neurosci. 18, 1213–1225 (2015).
Adamantidis, A. et al. Optogenetics: 10 years after ChR2 in neurons—views from the community. Nat. Neurosci. 18, 1202–1212 (2015).
Janak, P. H. & Tye, K. M. From circuits to behaviour in the amygdala. Nature 517, 284–292 (2015).
Lee, J. et al. Rapidly customizable, scalable 3D-printed wireless optogenetic probes for versatile applications in neuroscience. Adv. Funct. Mater. 30, 2004285 (2020).
McCall, J. G. et al. Fabrication and application of flexible, multimodal light-emitting devices for wireless optogenetics. Nat. Protoc. 8, 2413–2428 (2013).
McCall, J. G. et al. Preparation and implementation of optofluidic neural probes for in vivo wireless pharmacology and optogenetics. Nat. Protoc. 12, 219–237 (2017).
Kim, T. et al. Injectable, cellular-scale optoelectronics with applications for wireless optogenetics. Science 340, 211–216 (2013).
Jeong, J.-W. et al. Wireless optofluidic systems for programmable in vivo pharmacology and optogenetics. Cell 162, 662–674 (2015).
Byun, S.-H. et al. Mechanically transformative electronics, sensors, and implantable devices. Sci. Adv. 5, eaay0418 (2019).
Poher, V. et al. Micro-LED arrays: a tool for two-dimensional neuron stimulation. J. Phys. Appl. Phys. 41, 094014 (2008).
Grossman, N. et al. Multi-site optical excitation using ChR2 and micro-LED array. J. Neural Eng. 7, 016004 (2010).
Gerhardt, K. P. et al. An open-hardware platform for optogenetics and photobiology. Sci. Rep. 6, 35363 (2016).
Aharoni, D., Khakh, B. S., Silva, A. J. & Golshani, P. All the light that we can see: a new era in miniaturized microscopy. Nat. Methods 16, 11–13 (2019).
Siegle, J. H. et al. Open Ephys: an open-source, plugin-based platform for multichannel electrophysiology. J. Neural Eng. 14, 045003 (2017).
Yang, Y. et al. Wireless multilateral devices for optogenetic studies of individual and social behaviors. Nat. Neurosci. 24, 1035–1045 (2021).
Shin, G. et al. Flexible near-field wireless optoelectronics as subdermal implants for broad applications in optogenetics. Neuron 93, 509–521.e3 (2017).
Park, S. I. et al. Soft, stretchable, fully implantable miniaturized optoelectronic systems for wireless optogenetics. Nat. Biotechnol. 33, 1280–1286 (2015).
Park, S. I. et al. Ultraminiaturized photovoltaic and radio frequency powered optoelectronic systems for wireless optogenetics. J. Neural Eng. 12, 056002 (2015).
Noh, K. N. et al. Miniaturized, battery-free optofluidic systems with potential for wireless pharmacology and optogenetics. Small 14, 1702479 (2018).
Kim, C. Y. et al. Soft subdermal implant capable of wireless battery charging and programmable controls for applications in optogenetics. Nat. Commun. 12, 535 (2021).
Qazi, R. et al. Wireless optofluidic brain probes for chronic neuropharmacology and photostimulation. Nat. Biomed. Eng. 3, 655–669 (2019).
Qazi, R., Kim, C. Y., Byun, S.-H. & Jeong, J.-W. Microscale inorganic LED based wireless neural systems for chronic in vivo optogenetics. Front. Neurosci. 12, 764 (2018).
Sparta, D. R. et al. Construction of implantable optical fibers for long-term optogenetic manipulation of neural circuits. Nat. Protoc. 7, 12–23 (2011).
Dagnew, R. et al. CerebraLux: a low-cost, open-source, wireless probe for optogenetic stimulation. Neurophotonics 4, 045001 (2017).
Tye, K. M. et al. Amygdala circuitry mediating reversible and bidirectional control of anxiety. Nature 471, 358–362 (2011).
Pisanello, F. et al. Dynamic illumination of spatially restricted or large brain volumes via a single tapered optical fiber. Nat. Neurosci. 20, 1180–1188 (2017).
Pisanello, M. et al. Tailoring light delivery for optogenetics by modal demultiplexing in tapered optical fibers. Sci. Rep. 8, 4467 (2018).
Wu, F. et al. Monolithically integrated μLEDs on silicon neural probes for high-resolution optogenetic studies in behaving animals. Neuron 88, 1136–1148 (2015).
Burton, A. et al. Wireless, battery-free subdermally implantable photometry systems for chronic recording of neural dynamics. Proc. Natl Acad. Sci. USA 117, 2835–2845 (2020).
Implantable Biomedical Microsystems - 1st Edition. http://www.elsevier.com/books/implantable-biomedical-microsystems/bhunia/978-0-323-26208-8
Lu, L. et al. Wireless optoelectronic photometers for monitoring neuronal dynamics in the deep brain. Proc. Natl Acad. Sci. USA 115, E1374–E1383 (2018).
Yin, M. et al. Wireless neurosensor for full-spectrum electrophysiology recordings during free behavior. Neuron 84, 1170–1182 (2014).
Burton, A. et al. Wireless, battery-free, and fully implantable electrical neurostimulation in freely moving rodents. Microsyst. Nanoeng. 7, 1–12 (2021).
Lee, W. et al. Microfabrication and in vivo performance of a microdialysis probe with embedded membrane. Anal. Chem. 88, 1230–1237 (2016).
Mickle, A. D. et al. A wireless closed loop system for optogenetic peripheral neuromodulation. Nature 565, 361–365 (2019).
Zhang, Y. et al. Battery-free, fully implantable optofluidic cuff system for wireless optogenetic and pharmacological neuromodulation of peripheral nerves. Sci. Adv. 5, eaaw5296 (2019).
Scharf, R. et al. Depth-specific optogenetic control in vivo with a scalable, high-density μLED neural probe. Sci. Rep. 6, 28381 (2016).
Shim, E., Chen, Y., Masmanidis, S. & Li, M. Multisite silicon neural probes with integrated silicon nitride waveguides and gratings for optogenetic applications. Sci. Rep. 6, 22693 (2016).
Zorzos, A. N., Scholvin, J., Boyden, E. S. & Fonstad, C. G. Three-dimensional multiwaveguide probe array for light delivery to distributed brain circuits. Opt. Lett. 37, 4841–4843 (2012).
Delcasso, S., Denagamage, S., Britton, Z. & Graybiel, A. M. HOPE: hybrid-drive combining optogenetics, pharmacology and electrophysiology. Front. Neural Circuits 12, (2018).
Kim, C. K. et al. Simultaneous fast measurement of circuit dynamics at multiple sites across the mammalian brain. Nat. Methods 13, 325–328 (2016).
Kim, K. et al. HectoSTAR microLED optoelectrodes for large-scale, high-precision in invo opto-electrophysiology. Preprint at http://doi.org/10.1101/2020.10.09.334227 (2020).
Wentz, C. T. et al. A wirelessly powered and controlled device for optical neural control of freely-behaving animals. J. Neural Eng. 8, 046021 (2011).
Ameli, R., Mirbozorgi, A., Neron, J.-L., LeChasseur, Y. & Gosselin, B. A wireless and batteryless neural headstage with optical stimulation and electrophysiological recording. In 2013 35th Annual International Conference of the IEEE Engineering in Medicine and Biology Society (EMBC) 5662–5665 (2013).
Hashimoto, M., Hata, A., Miyata, T. & Hirase, H. Programmable wireless light-emitting diode stimulator for chronic stimulation of optogenetic molecules in freely moving mice. Neurophotonics 1, 011002–011002 (2014).
Montgomery, K. L. et al. Wirelessly powered, fully internal optogenetics for brain, spinal and peripheral circuits in mice. Nat. Methods 12, 969–974 (2015).
de Groot, A. et al. NINscope, a versatile miniscope for multi-region circuit investigations. eLife 9, e49987 (2020).
Barbera, G., Liang, B., Zhang, L., Li, Y. & Lin, D.-T. A wireless miniScope for deep brain imaging in freely moving mice. J. Neurosci. Methods 323, 56–60 (2019).
Jacob, A. D. et al. A compact head-mounted endoscope for in vivo calcium imaging in freely behaving mice. Curr. Protoc. Neurosci. 84, e51 (2018).
Qazi, R. et al. Scalable and modular wireless-network infrastructure for large-scale behavioural neuroscience. Nat. Biomed. Eng. http://doi.org/10.1038/s41551-021-00814-w (2021).
Barbera, G. et al. An open source motorized swivel for in vivo neural and behavioral recordings. MethodsX 7, 101167 (2020).
Andrews, C. D. & Hutson, P. H. A reliable multichannel commutator for making electrical contact with conscious, freely moving rats. J. Neurosci. Methods 5, 73–76 (1982).
Fee, M. S. & Leonardo, A. Miniature motorized microdrive and commutator system for chronic neural recording in small animals. J. Neurosci. Methods 112, 83–94 (2001).
Roh, M., McHugh, T. J. & Lee, K. A video based feedback system for control of an active commutator during behavioral physiology. Mol. Brain 8, 61 (2015).
Shansky, R. M. & Murphy, A. Z. Considering sex as a biological variable will require a global shift in science culture. Nat. Neurosci. http://doi.org/10.1038/s41593-021-00806-8 (2021).
Miller, L. R. et al. Considering sex as a biological variable in preclinical research. FASEB J. 31, 29–34 (2017).
Butler, T. R., Karkhanis, A. N., Jones, S. R. & Weiner, J. L. Adolescent social isolation as a model of heightened vulnerability to comorbid alcoholism and anxiety disorders. Alcohol. Clin. Exp. Res. 40, 1202–1214 (2016).
Ma, X. et al. Social isolation-induced aggression potentiates anxiety and depressive-like behavior in male mice subjected to unpredictable chronic mild stress. PloS ONE 6, e20955 (2011).
Oehler, J., Jähkel, M. & Schmidt, J. Neuronal transmitter sensitivity after social isolation in rats. Physiol. Behav. 41, 187–191 (1987).
Tuboly, G., Benedek, G. & Horvath, G. Selective disturbance of pain sensitivity after social isolation. Physiol. Behav. 96, 18–22 (2009).
Tomova, L. et al. Acute social isolation evokes midbrain craving responses similar to hunger. Nat. Neurosci. 23, 1597–1605 (2020).
Matthews, G. A. et al. Dorsal raphe dopamine neurons represent the experience of social isolation. Cell 164, 617–631 (2016).
Nonogaki, K., Nozue, K. & Oka, Y. Social isolation affects the development of obesity and type 2 diabetes in mice. Endocrinology 148, 4658–4666 (2007).
Yizhar, O., Fenno, L. E., Davidson, T. J., Mogri, M. & Deisseroth, K. Optogenetics in neural systems. Neuron 71, 9–34 (2011).
Marcotte, M. et al. Handling techniques to reduce stress in mice. JoVE J. Vis. Exp. http://doi.org/10.3791/62593 (2021).
Gouveia, K. & Hurst, J. L. Improving the practicality of using non-aversive handling methods to reduce background stress and anxiety in laboratory mice. Sci. Rep. 9, 20305 (2019).
Sensini, F. et al. The impact of handling technique and handling frequency on laboratory mouse welfare is sex-specific. Sci. Rep. 10, 17281 (2020).