Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Full-bandwidth electrophysiology of seizures and epileptiform activity enabled by flexible graphene microtransistor depth neural probes

Abstract

Mapping the entire frequency bandwidth of brain electrophysiological signals is of paramount importance for understanding physiological and pathological states. The ability to record simultaneously DC-shifts, infraslow oscillations (<0.1 Hz), typical local field potentials (0.1–80 Hz) and higher frequencies (80–600 Hz) using the same recording site would particularly benefit preclinical epilepsy research and could provide clinical biomarkers for improved seizure onset zone delineation. However, commonly used metal microelectrode technology suffers from instabilities that hamper the high fidelity of DC-coupled recordings, which are needed to access signals of very low frequency. In this study we used flexible graphene depth neural probes (gDNPs), consisting of a linear array of graphene microtransistors, to concurrently record DC-shifts and high-frequency neuronal activity in awake rodents. We show here that gDNPs can reliably record and map with high spatial resolution seizures, pre-ictal DC-shifts and seizure-associated spreading depolarizations together with higher frequencies through the cortical laminae to the hippocampus in a mouse model of chemically induced seizures. Moreover, we demonstrate the functionality of chronically implanted devices over 10 weeks by recording with high fidelity spontaneous spike-wave discharges and associated infraslow oscillations in a rat model of absence epilepsy. Altogether, our work highlights the suitability of this technology for in vivo electrophysiology research, and in particular epilepsy research, by allowing stable and chronic DC-coupled recordings.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Buy article

Get time limited or full article access on ReadCube.

$32.00

All prices are NET prices.

Fig. 1: Flexible gDNP technology and characterization.
Fig. 2: Validation of in vivo full-bandwidth recording capabilities of gDNPs.
Fig. 3: Electrophysiological recording of characteristic epilepsy biomarkers.
Fig. 4: Chronic assessment of the stability, functionality and biocompatibility of gDNPs.

Data availability

All relevant data obtained to evaluate the main findings of the paper are openly available in Zenodo at https://doi.org/10.5281/zenodo.5655535. All other raw data are available from the corresponding author upon reasonable request.

Code availability

Python v.3.7 packages (Matplotlib v.3.2.0 and Numpy v.1.17.4) and the following Python library were used for electric characterization of the gSGFET arrays: https://github.com/aguimera/PyGFET. A custom Simulink model was used for graphene microtransistor electrophysiological data acquisition; contact g.tec medical engineering for code access. Electrophysiological data were analysed using Python v.3.7 packages (Matplotlib v.3.2.0, Numpy v.1.17.4, Pandas v.0.25.3, seaborn v.0.9.0, Neo v.0.8.0 and Elephant) and the custom library PhyREC (https://github.com/aguimera/PhyREC/tree/PhyREC4). Custom scripts can be found at Zenodo (https://doi.org/10.5281/zenodo.5655535). Immunohistochemical data analysis was performed using Python v.3.7 script (https://github.com/kebarr/biocompatibility_study).

References

  1. Perucca, P., Dubeau, F. & Gotman, J. Intracranial electroencephalographic seizure-onset patterns: effect of underlying pathology. Brain 137, 183–196 (2014).

    Google Scholar 

  2. Modur, P. N. High frequency oscillations and infraslow activity in epilepsy. Ann. Indian Acad. Neurol. 17, S99–S106 (2014).

    Google Scholar 

  3. Revankar, G. S. et al. in Seizures in Critical Care: A Guide to Diagnosis and Therapeutics (eds Varelas, P. N. and Claassen, J.) 77–90 (Humana Press, 2017); https://doi.org/10.1007/978-3-319-49557-6_5

  4. Dreier, J. P. The role of spreading depression, spreading depolarization and spreading ischemia in neurological disease. Nat. Med. 17, 439–447 (2011).

    CAS  Google Scholar 

  5. Staba, R. J., Stead, M. & Worrell, G. A. Electrophysiological biomarkers of epilepsy. Neurotherapeutics 11, 334–346 (2014).

    CAS  Google Scholar 

  6. Dell, K. L., Cook, M. J. & Maturana, M. I. Deep brain stimulation for epilepsy: biomarkers for optimization. Curr. Treat. Options Neurol. 21, 47 (2019).

    Google Scholar 

  7. Kuhlmann, L., Lehnertz, K., Richardson, M. P., Schelter, B. & Zaveri, H. P. Seizure prediction—ready for a new era. Nat. Rev. Neurol. 14, 618–630 (2018).

    Google Scholar 

  8. Chari, A., Thornton, R. C., Tisdall, M. M. & Scott, R. C. Microelectrode recordings in human epilepsy: a case for clinical translation. Brain Commun. 2, fcaa082 (2020).

    Google Scholar 

  9. Lee, S. et al. DC shifts, high frequency oscillations, ripples and fast ripples in relation to the seizure onset zone. Seizure 77, 52–58 (2020).

    Google Scholar 

  10. Li, C. et al. Evaluation of microelectrode materials for direct-current electrocorticography. J. Neural Eng. 13, 16008 (2015).

    Google Scholar 

  11. Hartings, J. A. How slow can you go? Nat. Mater. 18, 194–196 (2019).

    CAS  Google Scholar 

  12. Major, S., Gajovic-Eichelmann, N., Woitzik, J. & Dreier, J. P. Oxygen-induced and pH-induced direct current artifacts on invasive platinum/iridium electrodes for electrocorticography. Neurocrit. Care 35, 146–159 (2021).

    CAS  Google Scholar 

  13. Khodagholy, D. et al. In vivo recordings of brain activity using organic transistors. Nat. Commun. 4, 1575 (2013).

    Google Scholar 

  14. Kostarelos, K., Vincent, M., Hebert, C. & Garrido, J. A. Graphene in the design and engineering of next-generation neural interfaces. Adv. Mater. 29, 1700909 (2017).

    Google Scholar 

  15. Blaschke, B. M. et al. Mapping brain activity with flexible graphene micro-transistors. 2D Mater. 4, 025040 (2017).

    Google Scholar 

  16. Hébert, C. et al. Flexible graphene solution-gated field-effect transistors: efficient transducers for micro-electrocorticography. Adv. Funct. Mater. 28, 1703976 (2017).

  17. Masvidal-Codina, E. et al. High-resolution mapping of infraslow cortical brain activity enabled by graphene microtransistors. Nat. Mater. 18, 280–288 (2019).

    CAS  Google Scholar 

  18. Weltman, A., Yoo, J. & Meng, E. Flexible, penetrating brain probes enabled by advances in polymer microfabrication. Micromachines 7, 180 (2016).

    Google Scholar 

  19. Tien, L. W. et al. Silk as a multifunctional biomaterial substrate for reduced glial scarring around brain-penetrating electrodes. Adv. Funct. Mater. 23, 3185–3193 (2013).

    CAS  Google Scholar 

  20. Coenen, A. M. L. & Van Luijtelaar, E. L. J. M. Genetic animal models for absence epilepsy: a review of the WAG/Rij strain of rats. Behav. Genet. 33, 635–655 (2003).

    CAS  Google Scholar 

  21. Terlau, J. et al. Spike-wave discharges in absence epilepsy: segregation of electrographic components reveals distinct pathways of seizure activity. J. Physiol. https://doi.org/10.1113/JP279483 (2020).

  22. Zijlmans, M. et al. High-frequency oscillations as a new biomarker in epilepsy. Ann. Neurol. 71, 169–178 (2012).

    Google Scholar 

  23. Jacobs, J. et al. High-frequency oscillations (HFOs) in clinical epilepsy. Prog. Neurobiol. 98, 302–315 (2012).

    CAS  Google Scholar 

  24. Ikeda, A. et al. Focal ictal direct current shifts in human epilepsy as studied by subdural and scalp recording. Brain 122, 827–838 (1999).

    Google Scholar 

  25. Wu, S. et al. Role of ictal baseline shifts and ictal high-frequency oscillations in stereo-electroencephalography analysis of mesial temporal lobe seizures. Epilepsia 55, 690–698 (2014).

    Google Scholar 

  26. Vanhatalo, S. et al. Very slow EEG responses lateralize temporal lobe seizures: an evaluation of non-invasive DC-EEG. Neurology 60, 1098–1104 (2003).

    CAS  Google Scholar 

  27. Duan, X. et al. Intracellular recordings of action potentials by an extracellular nanoscale field-effect transistor. Nat. Nanotechnol. 7, 174–179 (2012).

    CAS  Google Scholar 

  28. Hess, L. H. Graphene transistors for biosensing and bioelectronics. Proc. IEEE 101, 1780–1792 (2013).

    CAS  Google Scholar 

  29. Hess, L. H. et al. High-transconductance graphene solution-gated field effect transistors. Appl. Phys. Lett. 99, 033503 (2011).

    Google Scholar 

  30. Garcia-Cortadella, R. et al. Distortion-free sensing of neural activity using graphene transistors. Small 16, 1906640 (2020).

    CAS  Google Scholar 

  31. Lecomte, A. et al. Silk and PEG as means to stiffen a parylene probe for insertion in the brain: toward a double time-scale tool for local drug delivery. J. Micromech. Microeng. 25, 125003 (2015).

    Google Scholar 

  32. Fueta, Y. & Avoli, M. Effects of antiepileptic drugs on 4-aminopyridine-induced epileptiform activity in young and adult rat hippocampus. Epilepsy Res. 12, 207–215 (1992).

    CAS  Google Scholar 

  33. Padmanabhan, K. & Urban, N. N. Disrupting information coding via block of 4-AP-sensitive potassium channels. J. Neurophysiol. 112, 1054–1066 (2014).

    CAS  Google Scholar 

  34. Zakharov, A., Chernova, K., Burkhanova, G., Holmes, G. L. & Khazipov, R. Segregation of seizures and spreading depolarization across cortical layers. Epilepsia 60, 2386–2397 (2019).

    CAS  Google Scholar 

  35. Hartings, J. A. et al. Direct current electrocorticography for clinical neuromonitoring of spreading depolarizations. J. Cereb. Blood Flow Metab. 37, 1857–1870 (2017).

    Google Scholar 

  36. Harriott, A. M., Takizawa, T., Chung, D. Y. & Chen, S. P. Spreading depression as a preclinical model of migraine. J. Headache Pain 20, 45 (2019).

    Google Scholar 

  37. Buzsáki, G. & Lopes da Silva, F. L High frequency oscillations in the intact brain. Prog. Neurobiol. 98, 241–249 (2012).

    Google Scholar 

  38. Ikeda, A. et al. Active direct current (DC) shifts an “Red slow”: two new concepts for seizure mechanisms and identification of the epileptogenic zone. Neurosci. Res. 156, 95–101 (2020).

    Google Scholar 

  39. Kamarajan, C., Pandey, A. K., Chorlian, D. B. & Porjesz, B. The use of current source density as electrophysiological correlates in neuropsychiatric disorders: a review of human studies. Int. J. Psychophysiol. 97, 310–322 (2015).

    Google Scholar 

  40. Flynn, S. P., Barrier, S., Scott, R. C., Lenck-Santini, P. P. & Holmes, G. L. Status epilepticus induced spontaneous dentate gyrus spikes: in vivo current source density analysis. PLoS ONE 10, e0132630 (2015).

    Google Scholar 

  41. Coenen, A. M. L. & Van Luijtelaar, E. L. J. M. The WAG/Rij rat model for absence epilepsy: age and sex factors. Epilepsy Res. 1, 297–301 (1987).

    CAS  Google Scholar 

  42. Orlowska-Feuer, P. et al. Infra-slow modulation of fast beta/gamma oscillations in the mouse visual system. J. Physiol. 599, 1631–1650 (2021).

    CAS  Google Scholar 

  43. Garcia-Cortadella, R. et al. Graphene active sensor arrays for long-term and wireless mapping of wide frequency band epicortical brain activity. Nat. Commun. 12, 211 (2021).

    CAS  Google Scholar 

  44. Bahari, F. et al. Seizure-associated spreading depression is a major feature of ictal events in two animal models of chronic epilepsy. Preprint at bioRxiv https://doi.org/10.1101/455519 (2020).

  45. Dreier, J. P. et al. Spreading convulsions, spreading depolarization and epileptogenesis in human cerebral cortex. Brain 135, 259–275 (2012).

    Google Scholar 

  46. Dreier, J. P. et al. Recording, analysis, and interpretation of spreading depolarizations in neurointensive care: review and recommendations of the COSBID research group. J. Cereb. Blood Flow Metab. 37, 1595–1625 (2017).

    Google Scholar 

  47. De Tisi, J. et al. The long-term outcome of adult epilepsy surgery, patterns of seizure remission, and relapse: a cohort study. Lancet 378, 1388–1395 (2011).

    Google Scholar 

  48. Kanazawa, K. et al. Intracranially recorded ictal direct current shifts may precede high frequency oscillations in human epilepsy. Clin. Neurophysiol. 126, 47–59 (2015).

    Google Scholar 

  49. Lauritzen, M. et al. Clinical relevance of cortical spreading depression in neurological disorders: migraine, malignant stroke, subarachnoid and intracranial hemorrhage, and traumatic brain injury. J. Cereb. Blood Flow Metab. 31, 17–35 (2011).

    Google Scholar 

  50. Schaefer, N. et al. Improved metal-graphene contacts for low-noise, high-density microtransistor arrays for neural sensing. Carbon 161, 647–655 (2020).

    CAS  Google Scholar 

  51. Jin, H. J. et al. Water-stable silk films with reduced β-sheet content. Adv. Funct. Mater. 15, 1241–1247 (2005).

    CAS  Google Scholar 

  52. Rockwood, D. N. et al. Materials fabrication from Bombyx mori silk fibroin. Nat. Protoc. 6, 1612–1631 (2011).

    CAS  Google Scholar 

  53. Vepari, C. & Kaplan, D. L. Silk as a biomaterial. Prog. Polym. Sci. 32, 991–1007 (2007).

    CAS  Google Scholar 

  54. Cao, Y. & Wang, B. Biodegradation of silk biomaterials. Int. J. Mol. Sci. 10, 1514–1524 (2009).

    CAS  Google Scholar 

  55. Gobin, A. S., Froude, V. E. & Mathur, A. B. Structural and mechanical characteristics of silk fibroin and chitosan blend scaffolds for tissue regeneration. J. Biomed. Mater. Res. A 74, 465–473 (2005).

    Google Scholar 

  56. Russo, E. et al. Upholding WAG/Rij rats as a model of absence epileptogenesis: hidden mechanisms and a new theory on seizure development. Neurosci. Biobehav. Rev. 71, 388–408 (2016).

    CAS  Google Scholar 

  57. van Luijtelaar, G. & van Oijen, G. Establishing drug effects on electrocorticographic activity in a genetic absence epilepsy model: advances and pitfalls. Front. Pharmacol. 11, 395 (2020).

    CAS  Google Scholar 

Download references

Acknowledgements

This work has received funding from the European Union’s Horizon 2020 research and innovation programme under Grant Agreement No. 881603 (GrapheneCore3). ICN2 is supported by the Severo Ochoa Centres of Excellence programme, funded by the Spanish Research Agency (AEI, grant no. SEV-2017–0706), and by the CERCA Programme/Generalitat de Catalunya. A.B.C. is supported by the International PhD Programme La Caixa-Severo Ochoa (Programa Internacional de Becas ‘la Caixa’-Severo Ochoa). This work has made use of the Spanish ICTS Network MICRONANOFABS, partially supported by MICINN and the ICTS ‘NANBIOSIS’, more specifically by the Micro-NanoTechnology Unit of the CIBER in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN) at the IMB-CNM. We also acknowledge funding from the Generalitat de Catalunya (2017 SGR 1426), and the 2DTecBio project (FIS2017-85787-R) funded by the Ministerio de Ciencia, Innovación y Universidades of Spain, the Agencia Estatal de Investigación (AEI) and the Fondo Europeo de Desarrollo Regional (FEDER/UE). Part of this work was co-funded by the European Regional Development Funds (ERDF) allocated to the Programa operatiu FEDER de Catalunya 2014–2020, with the support of the Secretaria d’Universitats i Recerca of the Departament d’Empresa i Coneixement of the Generalitat de Catalunya for emerging technology clusters devoted to the valorization and transfer of research results (GraphCAT 001-P-001702). A.B.C. acknowledges that this work has been carried out within the framework of a PhD in Electrical and Telecommunication Engineering at the Universitat Autònoma de Barcelona. R.C.W. is funded by a Senior Research Fellowship awarded by the Worshipful Company of Pewterers. D.R. is a Biotechnology and Biological Sciences Research Council (BBSRC) LIDo sponsored PhD student. We thank M. Walker and L. Lemieux (UCL Queen Square Institute of Neurology) for their comments on the manuscript.

Author information

Authors and Affiliations

Authors

Contributions

A.B.C. carried out most of the fabrication and characterization of the gDNPs, contributed to the design and performance of the in vivo experiments, analysed the data and wrote the manuscript. E.M.-C. contributed to the design and planning of the in vivo experiments, to the data analysis and particularly to the DC-shift and SD analysis of the in vivo data. R.C.W. and T.M.S. performed the in vivo experiments. D.R. contributed to the in vivo experiments and DC-coupled recordings with the glass micropipette. N.S., E.R.-L., X.I. and J.M.D.l.C. contributed to the fabrication and characterization of the gDNPs. E.D.C., J.B. and C.H. contributed to the growth, transfer and characterization of the CVD graphene used in the gDNPs. E.P.-A., A.H. and E.R.-L. contributed to the optimization of the SF stiffening protocol of the gDNPs. J.M.-A. contributed to the fabrication of the custom electronic instrumentation and development of a Python-based user interface. D.V. contributed to the Python scripts and technical discussions. J.R.S. reviewed the manuscript. J.F. and J.S. contributed to the mechanical assessment of the SF and the SF back-coated gDNPs. M.D. performed all surgeries for the biocompatibility study. A.D. and K.B. contributed to the capture of histological images and image processing and analysis. S.S. and K.B. contributed to the preparation and review of the manuscript. A.G.-B. contributed to the design and fabrication of the custom electronic instrumentation, the development of a custom gSGFET Python library and analysis of the data. R.V., K.K., R.C.W., A.G.-B. and J.A.G. participated in the design of all experiments and thoroughly reviewed the manuscript. All authors read and reviewed the manuscript.

Corresponding authors

Correspondence to Rob C. Wykes, Anton Guimerà-Brunet or Jose A. Garrido.

Ethics declarations

Competing interests

C.G. is the owner of g.tec medical engineering and Guger Technologies. J.A.G, K.K and A.G.-B declare financial interest in INBRAIN Neuroelectronics. All other authors have no competing interests.

Additional information

Peer review information Nature Nanotechnology thanks Matthew Nelson, Bozhi Tian and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary Methods, Figs. 1–23 and Table 1.

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Bonaccini Calia, A., Masvidal-Codina, E., Smith, T.M. et al. Full-bandwidth electrophysiology of seizures and epileptiform activity enabled by flexible graphene microtransistor depth neural probes. Nat. Nanotechnol. 17, 301–309 (2022). https://doi.org/10.1038/s41565-021-01041-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41565-021-01041-9

This article is cited by

Search

Quick links

Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research