Blood-brain barrier (BBB) dysfunction plays an important role in cellular damage in neurological diseases and brain injuries. This project employs an innovative in vivo imaging technology that explores how BBB injury causes negative tissue response to neural probes. This in turn directs future probe designs.

Neural activity is not just impacted by BBB leakage and inflammation, but also disruption of metabolic supply. We demonstrated that tissue deoxygenation occurs around chronically implanted microelectrodes and that neurons around microelectrodes can be silenced1. We showed with in vivo widefield imaging that basal GCaMP signal was present around the electrode (Fig. 1a, blue Region-of-Interest) and was roughly the same as distant control regions (green) (i.e., no injury). However, decreases in the GCaMP coefficient of variance (COV), which represents loss or silencing of ongoing neuronal activity, were visible on the day of implantation compared to areas further away from the implanted area (green). Interestingly, over days, this silencing of neuronal activity coincided with a loss of Blood Oxygenation Level Dependent - Optical Intrinsic Signal (BOLD-OIS or OIS-BOLD) (Fig. 1c). In this subject, the decrease in blood oxygenation dissipated by day 4 and began returning to normal levels by day 7 (Fig. 1c). This decrease and subsequent improvement over the next several days may suggest healing. In vivo two-photon microscopy confirmed in another GCaMP mice four weeks post-implant that only neurons > 80 µm were activated by visual stimulation (LED flashes to contralateral eye) (Fig. 2 Green). However, we also show the presence of silenced neurons (Fig. 2 Red) near the electrode. These neurons were only activated with strong, direct electrical stimulation. It is possible that these neurons are silenced to decrease metabolic consumption while angiogenesis and wound healing restores the metabolic supply chain.

Figure 1: Loss of neuronal activity is associated to loss of tissue oxygenation without the loss of neurons. (a) No large changes in overall neuronal fluorescence or superficial injury is observed. Temporal COV for neuronal GCaMP activity (b) and OIS-BOLD (c) signals show that loss of neural activity (b: dark region) coincides with loss of tissue oxygenation (c: dark region). The changes in OIS-BOLD suggest there was a transient decrease in blood oxygen supply post-implantation. (Michelson et al JNE 2018)

Current and future research areas. The immediate translatable results are that Neuralink is developing an image guiding system for their ‘sewing machine inserter’ to implant individual ultrasmall “Neuralace” polymer probes while avoiding blood vessels. However, more broadly, we begin to uncover growing evidence that neural activity dysfunction around implants are, at least in part, due to metabolic stress1-3. There is a growing body of literature on the role of small vessel (capillary) degeneration, especially pericytes, in neurodegenerative disease outcomes4-32. We have shown in multiple publications that red blood cell flow rates in multiple capillaries around implanted arrays can greatly decrease or become completely occluded over the course of hours after implantation1,33,34. Moreover, we discuss13 that pericyte density is stable around microelectrode over the first 7 days post-implant and only significantly decreases at 28 days35. However, if the flow rate decrease occurs on the order of hours, why does the neural activity decrease on the order of days? In addition, current clinical invasive BCI participants are not being screened for familial history of Alzheimer’s Disease. With our Alzheimer’s Disease/Alzheimer’s Disease Related Dementia (AD/ADRD) supplement to this R01, we are conducting pilot studies to examine how microelectrode implantation affects neurodegeneration in genetically ‘vulnerable’ populations.

Figure 2: Neuronal Silencing around Microelectrode arrays 4 wks post implant. V1 Neurons > 80 μm from the electrode can be evoked by drifting grating visual stimulation in the contralateral eye (Green). In contrast, there are many neurons near the microelectrode that are silenced (Red).

1 Michelson, N. J. et al. Multi-scale, multi-modal analysis uncovers complex relationship at the brain tissue-implant neural interface: New Emphasis on the Biological Interface. Journal of Neural Engineering 15 (2018).

2 Wellman, S. M. et al. Cuprizone-induced oligodendrocyte loss and demyelination impairs recording performance of chronically implanted neural interfaces. Biomaterials, 119842 (2020).

3 Golabchi, A. et al. Melatonin improves quality and longevity of chronic neural recording. Biomaterials 180, 225-239, doi: (2018).

4 Staals, J., Makin, S. D., Doubal, F. N., Dennis, M. S. & Wardlaw, J. M. Stroke subtype, vascular risk factors, and total MRI brain small-vessel disease burden. Neurology 83, 1228-1234 (2014).

5 Hammer, D. X. et al. Longitudinal vascular dynamics following cranial window and electrode implantation measured with speckle variance optical coherence angiography. Biomed Opt Express 5, 2823-2836, doi:10.1364/BOE.5.002823 (2014).

6 Klohs, J., Rudin, M., Shimshek, D. R. & Beckmann, N. Imaging of cerebrovascular pathology in animal models of Alzheimer's disease. Front Aging Neurosci 6, 32, doi:10.3389/fnagi.2014.00032 (2014).

7 Marchesi, V. T. Alzheimer's dementia begins as a disease of small blood vessels, damaged by oxidative-induced inflammation and dysregulated amyloid metabolism: implications for early detection and therapy. Faseb J 25, 5-13, doi:10.1096/fj.11-0102ufm (2011).

8 Hall, C. N. et al. Capillary pericytes regulate cerebral blood flow in health and disease. Nature 508, 55 (2014).

9 Wardlaw, J. Blood-brain barrier and cerebral small vessel disease. Journal of the neurological sciences 299, 66-71 (2010).

10 Jokinen, H. et al. Brain atrophy accelerates cognitive decline in cerebral small vessel disease: the LADIS study. Neurology 78, 1785-1792 (2012).

11 Bennett, C. et al. Neuroinflammation, oxidative stress, and blood-brain barrier (BBB) disruption in acute Utah electrode array implants and the effect of deferoxamine as an iron chelator on acute foreign body response. Biomaterials 188, 144-159, doi: (2019).

12 Cheng, J. et al. Targeting pericytes for therapeutic approaches to neurological disorders. Acta Neuropathol 136, 507-523, doi:10.1007/s00401-018-1893-0 (2018).

13 Wellman, S. M. & Kozai, T. D. Y. Understanding the Inflammatory Tissue Reaction to Brain Implants To Improve Neurochemical Sensing Performance. ACS Chemical Neuroscience, doi:10.1021/acschemneuro.7b00403 (2017).

14 Rustenhoven, J., Jansson, D., Smyth, L. C. & Dragunow, M. Brain pericytes as mediators of neuroinflammation. Trends in pharmacological sciences 38, 291-304 (2017).

15 Attwell, D., Mishra, A., Hall, C. N., O'Farrell, F. M. & Dalkara, T. What is a pericyte? J Cereb Blood Flow Metab 36, 451-455 (2016).

16 Tsai, H. H. et al. Oligodendrocyte precursors migrate along vasculature in the developing nervous system. Science 351, 379-384, doi:10.1126/science.aad3839 (2016).

17 Sakuma, R. et al. Brain pericytes serve as microglia-generating multipotent vascular stem cells following ischemic stroke. Journal of neuroinflammation 13, 57 (2016).

18 Sweeney, M. D., Ayyadurai, S. & Zlokovic, B. V. Pericytes of the neurovascular unit: key functions and signaling pathways. Nature neuroscience 19, 771 (2016).

19 Maki, T. et al. Potential interactions between pericytes and oligodendrocyte precursor cells in perivascular regions of cerebral white matter. Neurosci Lett 597, 164-169, doi:10.1016/j.neulet.2015.04.047 (2015).

20 Choe, Y., Huynh, T. & Pleasure, S. J. Migration of oligodendrocyte progenitor cells is controlled by transforming growth factor beta family proteins during corticogenesis. J Neurosci 34, 14973-14983, doi:10.1523/JNEUROSCI.1156-14.2014 (2014).

21 Özen, I. et al. Brain pericytes acquire a microglial phenotype after stroke. Acta Neuropathol 128, 381-396 (2014).

22 Muoio, V., Persson, P. B. & Sendeski, M. M. The neurovascular unit – concept review. Acta Physiologica 210, 790-798, doi:10.1111/apha.12250 (2014).

23 Farahani, R. M., Sarrafpour, B., Simonian, M., Li, Q. & Hunter, N. Directed gliaassisted angiogenesis in a mature neurosensory structure: Pericytes mediate an adaptive response in human dental pulp that maintains bloodbarrier function. Journal of Comparative Neurology 520, 3803-3826 (2012).

24 Winkler, E. A., Bell, R. D. & Zlokovic, B. V. Central nervous system pericytes in health and disease. Nature neuroscience 14, 1398-1405, doi:10.1038/nn.2946 (2011).

25 Zlokovic, B. V. Neurovascular pathways to neurodegeneration in Alzheimer's disease and other disorders. Nat Rev Neurosci 12, 723-738, doi:10.1038/nrn3114 (2011).

26 Takata, F. et al. Brain pericytes among cells constituting the blood-brain barrier are highly sensitive to tumor necrosis factor-alpha, releasing matrix metalloproteinase-9 and migrating in vitro. Journal of neuroinflammation 8, 106, doi:10.1186/1742-2094-8-106 (2011).

27 Armulik, A. et al. Pericytes regulate the blood–brain barrier. Nature 468, 557 (2010).

28 Daneman, R., Zhou, L., Kebede, A. A. & Barres, B. A. Pericytes are required for blood-brain barrier integrity during embryogenesis. Nature 468, 562-566, doi:10.1038/nature09513 (2010).

29 Bell, R. D. et al. Pericytes control key neurovascular functions and neuronal phenotype in the adult brain and during brain aging. Neuron 68, 409-427, doi:10.1016/j.neuron.2010.09.043 (2010).

30 Diaz-Flores, L. et al. Pericytes. Morphofunction, interactions and pathology in a quiescent and activated mesenchymal cell niche. Histol Histopathol 24, 909-969 (2009).

31 Balabanov, R. et al. Interferon-gamma-oligodendrocyte interactions in the regulation of experimental autoimmune encephalomyelitis. The Journal of neuroscience : the official journal of the Society for Neuroscience 27, 2013-2024 (2007).

32 Peppiatt, C. M., Howarth, C., Mobbs, P. & Attwell, D. Bidirectional control of CNS capillary diameter by pericytes. Nature 443, 700-704, doi:10.1038/nature05193 (2006).

33 Kozai, T. D., Jaquins-Gerstl, A. S., Vazquez, A. L., Michael, A. C. & Cui, X. T. Brain tissue responses to neural implants impact signal sensitivity and intervention strategies. ACS Chem Neurosci 6, 48-67, doi:10.1021/cn500256e (2015).

34 Kozai, T. D. Y., Vazquez, A. L., Weaver, C. L., Kim, S. G. & Cui, X. T. In vivo two photon microscopy reveals immediate microglial reaction to implantation of microelectrode through extension of processes. J Neural Eng 9 (2012).

35 Wellman, S. M., Li, L., Yaxiaer, Y., McNamara, I. N. & Kozai, T. D. Revealing spatial and temporal patterns of cell death, glial proliferation, and blood-brain barrier dysfunction around implanted intracortical neural interfaces. Frontiers in Neuroscience 13, 493 (2019).