Correction NEUROSCIENCE Correction for “Regulation of myelin structure and conduc- tion velocity by perinodal astrocytes,” by Dipankar J. Dutta, Dong Ho Woo, Philip R. Lee, Sinisa Pajevic, Olena Bukalo, William C. Huffman, Hiroaki Wake, Peter J. Basser, Shahriar SheikhBahaei, Vanja Lazarevic, Jeffrey C. Smith, and R. Douglas Fields, which was first published October 29, 2018; 10.1073/ pnas.1811013115 (Proc. Natl. Acad. Sci. U.S.A. 115,11832–11837). The authors note that the following statement should be added to the Acknowledgments: “We acknowledge Dr. Hae Ung Lee for preliminary experiments that informed the ultimate experimental approach.” Published under the PNAS license. Published online June 10, 2019. www.pnas.org/cgi/doi/10.1073/pnas.1908361116 12574 | PNAS | June 18, 2019 | vol. 116 | no. 25 www.pnas.org Downloaded by guest on October 2, 2021 Regulation of myelin structure and conduction velocity by perinodal astrocytes Dipankar J. Duttaa,b, Dong Ho Wooa, Philip R. Leea, Sinisa Pajevicc, Olena Bukaloa, William C. Huffmana, Hiroaki Wakea, Peter J. Basserd, Shahriar SheikhBahaeie, Vanja Lazarevicf, Jeffrey C. Smithe, and R. Douglas Fieldsa,1 aSection on Nervous System Development and Plasticity, The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892; bThe Henry M. Jackson Foundation for the Advancement of Military Medicine, Inc., Bethesda, MD 20817; cMathematical and Statistical Computing Laboratory, Office of Intramural Research, Center for Information Technology, National Institutes of Health, Bethesda, MD 20892; dSection on Quantitative Imaging and Tissue Sciences, The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892; eCellular and Systems Neurobiology Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892; and fExperimental Immunology Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892 Edited by Terrence J. Sejnowski, Salk Institute for Biological Studies, La Jolla, CA, and approved September 25, 2018 (received for review June 26, 2018) The speed of impulse transmission is critical for optimal neural glial contact can be modified by vesicular release of substances from circuit function, but it is unclear how the appropriate conduction astrocytes and that this could enable astrocytes to influence nodal velocity is established in individual axons. The velocity of impulse structure, myelin sheath thickness, and impulse conduction speed. transmission is influenced by the thickness of the myelin sheath and the morphology of electrogenic nodes of Ranvier along axons. Results Here we show that myelin thickness and nodal gap length are Astrocyte Exocytosis Inhibits Thrombin-Mediated Cleavage of NF155. reversibly altered by astrocytes, glial cells that contact nodes of Our bioinformatic analysis revealed a potential proteolytic cleavage Ranvier. Thrombin-dependent proteolysis of a cell adhesion molecule site in the extracellular domain of mouse NF155 after arginine, amino that attaches myelin to the axon (neurofascin 155) is inhibited by acids 924–926 (glycine–arginine–glycine), that is specific for the en- vesicular release of thrombin protease inhibitors from perinodal zyme thrombin (SI Appendix,Fig.S1A) (7). Molecular modeling astrocytes. Transgenic mice expressing a dominant-negative fragment showed that the cleavage site is on the surface of the protein (Fig. 1C of VAMP2 in astrocytes, to reduce exocytosis by 50%, exhibited and Movie S1), accessible to thrombin (SI Appendix,Fig.S1B), and NEUROSCIENCE detachment of adjacent paranodal loops of myelin from the axon, thrombin treatment of subcortical white matter in vitro was found to increased nodal gap length, and thinning of the myelin sheath in the cleave NF155 at this site (SI Appendix,Fig.S1C). Cleavage here optic nerve. These morphological changes alter the passive cable would sever the Ig 5–6 domains that interact with Contactin1 (Fig. properties of axons to reduce conduction velocity and spike-time 1C), thus breaking the attachment of paranodal loops to the axon (8). arrival in the CNS in parallel with a decrease in visual acuity. All effects Under normal conditions thrombin is secreted by neurons and were reversed by the thrombin inhibitor Fondaparinux. Similar results enters the CNS from the vascular system (9). In the CNS, as- were obtained by viral transfection of tetanus toxin into astrocytes of rat corpus callosum. Previously, it was unknown how the myelin trocytes are the primary source of the thrombin inhibitor protease sheath could be thinned and the functions of perinodal astrocytes nexin1 (PN1) (also called glial-derived nexin-1 and SERPINE2) were not well understood. These findings describe a form of nervous (10), suggesting that perinodal astrocytes could modulate myelin system plasticity in which myelin structure and conduction velocity are adjusted by astrocytes. The thrombin-dependent cleavage of neuro- Significance fascin 155 may also have relevance to myelin disruption and repair. Proper communication between brain regions, via white mat- myelin plasticity | node of Ranvier | thrombin | neurofascin 155 | ter tracts, allows us to carry out complex cognitive and motor spike-time–dependent plasticity tasks. Impulses traveling must arrive at relay points almost si- multaneously for such communication to be effective. Myelin enables saltatory conduction of impulses and hence is a candi- onduction velocity (CV) is increased in axons with thicker date for modulation of impulse conduction velocity. But myelin Cmyelin sheaths, and the length of the nodal gap and distri- + structure, once formed, has been considered static. In this study, bution of Na channels in the node of Ranvier have a strong we show that mature myelin structure is dynamic. The mature influence on the action potential firing threshold, frequency of myelin sheath thickness and nodal gap length can be reversibly firing, and CV (1). More than 95% of nodes of Ranvier in the modulated to optimize the speed of axonal impulses. This CNS are contacted by astrocytes (2), but the function of peri- modulation of myelin is regulated by exocytosis of thrombin nodal astrocytes remains a long-standing question (3). The pos- protease inhibitors from astrocytes at the node of Ranvier. sibility that astrocytes could participate in maintaining or remodeling myelin structure was explored using transgenic mice Author contributions: R.D.F. designed research; D.J.D., D.H.W., P.R.L., S.P., O.B., W.C.H., and by viral gene transfection in rats. P.J.B., S.S., V.L., and R.D.F. performed research; J.C.S. contributed new reagents/analytic The myelin sheath attaches to the axon by forming a spiral tools; D.J.D., D.H.W., P.R.L., S.P., O.B., W.C.H., H.W., P.J.B., S.S., V.L., and R.D.F. analyzed junction in the paranodal region flanking the node of Ranvier data; and D.J.D. and R.D.F. wrote the paper. (Fig. 1A), which has the appearance of a series of loops when Conflict of interest statement: The United States Government applied for an international “ sectioned longitudinally for electron microscopy (EM). Paranodal patent (US Patent PCT/US2016/027776) titled, Methods of treating or preventing demy- + elination using thrombin inhibitors and methods of detecting demyelination using Neu- loops promote action potential propagation by concentrating Na ” + rofascin 155, on April 15, 2016, as the sole beneficiary. R.D.F., D.J.D., and D.H.W. are channels in the node, preventing juxtaparanodal K channels from listed as inventors in the patent application. diffusing into the node (4), and narrowing the nodal gap length to This article is a PNAS Direct Submission. increase membrane resistivity and reduce capacitance (5). Paranodal This open access article is distributed under Creative Commons Attribution-NonCommercial- loops are attached to the axon via septate-like junctions (Fig. 1B) NoDerivatives License 4.0 (CC BY-NC-ND). composed of a complex of three intercellular proteins, with neuro- 1To whom correspondence should be addressed. Email: [email protected]. fascin155 (NF155) on the paranodal loop interacting with the This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. Contactin1-associated protein1 (Caspr1)/Contactin1 complex on the 1073/pnas.1811013115/-/DCSupplemental. axon (Fig. 1C) (6). We tested the hypothesis that this point of axo- www.pnas.org/cgi/doi/10.1073/pnas.1811013115 PNAS Latest Articles | 1of6 By hypothesis, the 50% reduction in exocytosis of PN1 and Oligodendrocyte Neuron D A 0.2 other serine protease inhibitors from astrocytes should increase * NF155 cleavage at AA924 in white matter of mice expressing 0.1 p=0.01 (gfap)dnVAMP2, and this was confirmed by immunoblot and g total protein) µ amino acid sequencing by mass spectroscopy (Fig. 1 E and F). No n=8 n=4 n=8 n = cell-culture 0.0 replicate change was found in the other neurofascin family member at the Myelin sheath Wild-type - + PN1 (pg/ node, neurofascin 186 (NF186), which lacks a thrombin cleavage Astrocyte (gfap)dnVAMP2 E site. Many substances can be released by exocytosis from astrocytes, (gfap)dnVAMP2 F 60 - + p=0.007 but cleavage of NF155 in animals expressing (gfap)dnVAMP2 was NF155 * Myelin sheath NF125 40 prevented by daily s.c. injections of the highly specific thrombin Axon Astrocyte NF 30 inhibitor Fondaparinux, indicating that thrombin proteolysis is re- Node Paranode NF186 G H Axon 20 sponsible for the increased cleavage of NF155 (Fig. 1 and ). Enolase Fondaparinux treatment in the absence of (gfap)dnVAMP2 gene N=5 N=8 N = mice EGFP % NF155 (A.U.) cleavage 0 + expression had no significant effect (SI Appendix,Fig.S5). B (gfap)dnVAMP2) (gfap)dnVAMP2- G+ PBS + Fond. H 60 * Astrocyte Exocytosis Regulates Detachment of Paranodal Loops of NF155 Myelin at the Node of Ranvier. Paranodal NF125 Cleavage of NF155 by thrombin * 40 loops NF 30 p=0.044 should disrupt the axoglial junction holding paranodal loops to NF186 20 the axon, causing them to detach.