Questions

THE MOLECULAR CIRCUITS OF THE LIVING BRAIN:

 Development, behavior and cognition are at once microscale processes and macroscale interactions between a living organism and its environment. To understand the living brain and its pathologies we must similarly scale from molecular mechanisms to whole brain function, which has proven to be one of the greatest challenges faced in neurobiology. Meeting this challenge demands an approach that can reveal molecular events across the brain, without disrupting the very processes we wish to study. The most powerful technology available for noninvasive whole brain molecular imaging is MRI, with the potential for subsecond and near cellular resolution. It is my overarching goal to realize the potential of MRI to resolve the diverse molecular signaling of development, function, and degeneration across the whole mammalian brain in vivo. To this end, I have established a versatile molecular toolkit for MRI1, including calcium sensors and genetic reporters2–4. Here, I propose hypothesis driven projects that focus the application of my methods on two forms of neurobiological signaling that offer the greatest impact on human health: calcium imaging of neural activity and neuroimmune interactions in regeneration. The work of my lab will establish the field of molecular fMRI, bringing powerful new capabilities to research by bridging functional and mechanistic biology.


PARSING SLEEP CIRCUITS WITH Ca-fMRI:

Figure 1: Multidimensional analysis of MFB stimulation in the dorsal striatum. a) Schematic of stimulus and injection sites. b) Signal change map of MaCaReNa and control particles post stimulus. Nanoparticles create a positive signal change with decreasing [Ca++] c) Deconvolved ROI analysis of stimulus time course (STC) with BM3 sensor. d) STC with MaCaReNa sensor. Positive signal change denotes an initial drop in [Ca++]. e) STC of delta band LFP. f) Single unit recordings of STC divided into “Excited” (n=9/38) and “Inhibited” (n=12/38) (17/38 units were unaffected by stimulus)        

Neurological conditions from autism to Alzheimer’s Disease have characteristic sleep pathologies marked by aberrant “delta waves,” oscillatory brain activity from 0.5-4Hz5. Typically, periods of increased delta are observed in deep sleep and focused attention, suggesting multiple circuits oscillate in this range for network synchronization and gain modulation between cognitive states. Delta oscillations are a reliable biomarker for many neural pathologies, but the causal relationship between delta and cognition is unknown.Few candidate delta oscillating circuits have been identified and the molecular basis of how delta waves initiate, propagate, or terminate remains unclear. Fundamentally, brain-wide, oscillating circuits, are extremely difficult to parse with current methods of functional imaging, making them critically understudied relative to their therapeutic potential.

We have created a breakthrough molecular sensor for functional imaging of calcium with MRI (Ca-fMRI), capable of resolving oscillations in the delta band. In conjunction with electrophysiology, this novel method offers the unprecedented ability to map delta activity and potentially describe long range oscillatory circuits in animal models. The resulting datasets will be compiled into a whole brain atlas of delta oscillating circuits for comparative analysis with genetic models of disease, providing a critical bridge between described molecular pathologies and their less understood, systems level effects. I suspect the molecular dysregulation of delta oscillating circuits is a root cause of the pathologies of sleep and focal attention prominent in neurodevelopmental and degenerative disorders. Determining the mechanisms that shape and regulate these circuits offers new therapeutic targets for diseases of cognition, sleep, and attention.


PROBING THE NEUROIMMUNE REGENERATIVE SIGNALING AXIS WITH MRI REPORTER GENES:

PROBING THE NEUROIMMUNE REGENERATIVE SIGNALING AXIS WITH MRI REPORTER GENES: Stem cells build and replenish the mammalian brain through developmental tissue patterning6 and through adult neuronal regeneration, both basally7, and in response to injury8. Neuroimmune cells, like microglia, have an established neogenic role in healthy brain development, but the same cells are highly susceptible to external factors, leading to pathologies9. Immune challenges affect development10, while regeneration can be inhibited by inflammation, leading to degenerative disorders11. Neuroimmune signaling remains extremely difficult to study in vivo given the sensitivity of microglia to the invasive methods needed to image the brain12. New technologies for noninvasive functional studies would greatly accelerate research on any major neurodegenerative or inflammatory disease.

Figure 2: Ts-Biotag labels Tie2 expressing microglia in an injury model. MRI of an 1d post injury Ts-Biotag mouse shows bright labeling surrounding the site of injury (a) No such labeling is visible in control WT animals (b) IHC shows Ts-Biotag labeling in Iba1+ microglia proximal to the wound site(c) 2d post injury Tie2+(Iba1+) microglia associate with Dcx+(Ang1+) NPCs (d). (Iba1/ Ang1 costaining are not shown and peripheral macrophages were excluded using mCherry+ bone marrow chimeras.)

I have invented several genetic reporters for MRI and generated a transgenic mouse line that labels cell types responding to the neogenic cytokine Ang11,3.  In preliminary studies, I have imaged activated microglia responding to a brain injury model (Fig. 2), an unprecedented functional study of a molecular circuit involved in neuroinflammation and adult neurogenesis. The proposed studies will further define capabilities of genetic imaging with MRI by using this established model to test a hypothesis that adult neogenic neuroimmune interactions recapitulate developmental signaling.The full diversity of cell types and signaling molecules in development and regeneration continues to be described13, however the in vivo signaling dynamics of cytokines are virtually unknown, for lack of in vivo assays of function. Parsing neuroimmune signaling modes offers a new means to determine therapeutic targets for neurodegenerative disorders.

REFERENCES:  *Co-first authorship

1.         Bartelle BB*, Barandov A*, Jasanoff A. Molecular fMRI. J Neurosci. 2016 Apr 13;36(15):4139–4148. PMCID: PMC4829642

2.         Okada S*, Bartelle BB*, Li N, Breton-Provencher V, Lee JJ, Rodriguez E, Melican J, Sur M, Jasanoff A. Calcium-dependent molecular fMRI using a magnetic nanosensor. Nat Nanotechnol. 2018 Jun;13(6):473–477. PMID: 29713073

3.         Bartelle BB, Berríos-Otero CA, Rodriguez JJ, Friedland AE, Aristizábal O, Turnbull DH. Novel genetic approach for in vivo vascular imaging in mice. Circ Res. 2012 Mar 30;110(7):938–947. PMID: 22374133

4.         Bartelle BB, Szulc KU, Suero-Abreu GA, Rodriguez JJ, Turnbull DH. Divalent metal transporter, DMT1: a novel MRI reporter protein. Magn Reson Med. 2013 Sep;70(3):842–850. PMCID: PMC3587026

5.         Knyazev GG. EEG delta oscillations as a correlate of basic homeostatic and motivational processes. Neurosci Biobehav Rev. 2012 Jan;36(1):677–695. PMID: 22020231

6.         Florio M, Huttner WB. Neural progenitors, neurogenesis and the evolution of the neocortex. Development. 2014 Jun;141(11):2182–2194. PMID: 24866113

7.         Alvarez-Buylla A, Kohwi M, Nguyen TM, Merkle FT. The Heterogeneity of Adult Neural Stem Cells and the Emerging Complexity of Their Niche. Cold Spring Harb Symp Quant Biol. 2008 Jan 1;73:357–365. PMID: 19022766

8.         Ohira K. Injury-induced neurogenesis in the mammalian forebrain. Cell Mol Life Sci. 2011 May;68(10):1645–1656. PMID: 21042833

9.         Bilimoria PM, Stevens B. Microglia function during brain development: New insights from animal models. Brain Research. 2015 Aug 18;1617:7–17.

10.       Choi GB, Yim YS, Wong H, Kim S, Kim H, Kim SV, Hoeffer CA, Littman DR, Huh JR. The maternal interleukin-17a pathway in mice promotes autism-like phenotypes in offspring. Science. 2016 Feb 26;351(6276):933–939. PMCID: PMC4782964

11.       Ransohoff RM. How neuroinflammation contributes to neurodegeneration. Science. 2016 Aug 19;353(6301):777–783. PMID: 27540165

12.       Parkhurst CN, Gan W-B. Microglia dynamics and function in the CNS. Current Opinion in Neurobiology. 2010 Oct;20(5):595–600.

13.       Mathys H, Adaikkan C, Gao F, Young JZ, Manet E, Hemberg M, De Jager PL, Ransohoff RM, Regev A, Tsai L-H. Temporal Tracking of Microglia Activation in Neurodegeneration at Single-Cell Resolution. Cell Rep. 2017 Oct 10;21(2):366–380. PMCID: PMC5642107

14.       Czeh M, Gressens P, Kaindl AM. The yin and yang of microglia. Dev Neurosci. 2011;33(3–4):199–209.

15.       Koh S-H, Lo EH. The Role of the PI3K Pathway in the Regeneration of the Damaged Brain by Neural Stem Cells after Cerebral Infarction. Journal of Clinical Neurology. 2015 Oct 1;11(4):297–304.

16.       Heras-Sandoval D, Pérez-Rojas JM, Hernández-Damián J, Pedraza-Chaverri J. The role of PI3K/AKT/mTOR pathway in the modulation of autophagy and the clearance of protein aggregates in neurodegeneration. Cell Signal. 2014 Dec;26(12):2694–2701. PMID: 25173700

17.       Christensen RA, Fujikawa K, Madore R, Oettgen P, Varticovski L. NERF2, a member of the Ets family of transcription factors, is increased in response to hypoxia and angiopoietin-1: A potential mechanism for Tie2 regulation during hypoxia. J Cell Biochem. 2002;85(3):505–515.

18.       Liu D, Martin V, Fueyo J, Lee O-H, Xu J, Cortes-Santiago N, Alonso MM, Aldape K, Colman H, Gomez-Manzano C. Tie2/TEK modulates the interaction of glioma and brain tumor stem cells with endothelial cells and promotes an invasive phenotype. Oncotarget. 2010 Dec;1(8):700–709. PMID: 21321379

19.       Lim WA. Designing customized cell signalling circuits. Nat Rev Mol Cell Biol. 2010 Jun;11(6):393–403. PMID: 20485291