Progress in brain research, volume 214

Koleske Department of Molecular Biophysics and Biochemistry; Department of Neurobiology; Interdepartmental Neuroscience Program, and Program in Cellular Neuroscience, Neurodegeneration,

New Haven, Connecticut

USA

Donald G SteinAsa G Candler Professor

Department of Emergency Medicine

Emory UniversityAtlanta, GeorgiaUSA

Dick F SwaabProfessor of Neurobiology

Medical Faculty, University of Amsterdam;Leader Research team Neuropsychiatric DisordersNetherlands Institute for Neuroscience

AmsterdamThe Netherlands

Howard L Fields

Professor of NeurologyEndowed Chair in Pharmacology of AddictionDirector, Wheeler Center for the Neurobiology of Addiction

University of CaliforniaSan Francisco, California

USA

First edition 2014

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Notices

Knowledge and best practice in this field are constantly changing As new research andexperience broaden our understanding, changes in research methods, professional practices, ormedical treatment may become necessary

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Hayder Amin

Istituto Italiano di Tecnologia, NetS3Laboratory, Neuroscience and Brain

Technologies Dpt., Genova, Italy

Pavle Andjus

Center for Laser Microscopy, Institute for Physiology and Biochemistry, Faculty of

Biology, University of Belgrade, Belgrade, Serbia

Eleonora Aronica

Department of (Neuro)Pathology, Academic Medical Center and Swammerdam

Institute for Life Sciences, Center for Neuroscience, University of Amsterdam,

Amsterdam, and SEIN—Stichting Epilepsie Instellingen Nederland, Heemstede,

The Netherlands

Ke´vin Baranger

Aix Marseille Universite´, CNRS, UMR 7259, NICN, 13344, and Neurology and

Neuropsychology Department, AP-HM, Marseille, France

Martin Bastmeyer

Institute of Zoologie, Karlsruhe, and Institute of Functional Interfaces (IFG),

Karlsruhe Institute of Technology (KIT), Eggenstein-Leopoldshafen, Germany

Luca Berdondini

Istituto Italiano di Tecnologia, NetS3Laboratory, Neuroscience and Brain

Technologies Dpt., Genova, Italy

Vladimir Berezin

Laboratory of Neural Plasticity, Department of Neuroscience and Pharmacology,

University of Copenhagen, Symbion, Fruebjergvej 3, Box 39, Copenhagen Ø,

Program in Developmental Biology, and Department of Neuroscience, Baylor

College of Medicine, One Baylor Plaza, Houston, TX, USA

Lorenzo A Cingolani

Department of Neuroscience and Brain Technologies, Istituto Italiano di

Tecnologia, Genoa, Italy

v

Natasˇa Jovanov Milosˇevic´

Croatian Institute for Brain Research, and Department of Medical Biology,University of Zagreb School of Medicine, Zagreb, Croatia

Milosˇ Judasˇ

Croatian Institute for Brain Research, University of Zagreb School of Medicine,Zagreb, Croatia

Leszek Kaczmarek

Department of Molecular and Cellular Neurobiology, Nencki Institute, Warsaw,

Poland

Meghan E Kerrisk

Department of Molecular Biophysics and Biochemistry, Yale University,

New Haven, CT, USA

Michel Khrestchatisky

Aix Marseille Universite´, CNRS, UMR 7259, NICN, 13344, Marseille, France

Anthony J Koleske

Department of Molecular Biophysics and Biochemistry; Department of

Neurobiology; Interdepartmental Neuroscience Program, and Program in Cellular

Neuroscience, Neurodegeneration, and Repair, Yale University, New Haven, CT,

USA

Svetlana Korotchenko

Laboratory for Brain Extracellular Matrix Research, University of Nizhny

Novgorod, Nizhny Novgorod, Russia; Department of Neuroscience and Brain

Technologies; Istituto Italiano di Tecnologia, Genova, Italy

Ivica Kostovic

Croatian Institute for Brain Research, University of Zagreb School of Medicine,

Zagreb, Croatia

Jessica C.F Kwok

John van Geest Centre for Brain Repair, University of Cambridge, Forvie Site,

Robinson Way, Cambridge, UK

Tomasz Lebitko

Department of Molecular and Cellular Neurobiology, Nencki Institute, Warsaw,

Poland

Stephen L Leib

Neuroinfection Laboratory, Institute for Infectious Diseases, University of Bern,

Bern, and Biology Division, Spiez Laboratory, Swiss Federal Office for Civil

Protection, Spiez, Switzerland

Department of Molecular & Cellular Neurobiology, Center for Neurogenomics &

Cognitive Research, Neuroscience Campus Amsterdam, VU University

Amsterdam, HV Amsterdam, The Netherlands

University of Leipzig, EU-ESF Transnational Junior Research Group

“MESCAMP”, Paul Flechsig Institute for Brain Research, Leipzig, GermanyMariusz Mucha

University of Exeter, Exeter, UK

University of Exeter, Exeter, UK

Center for Behavioral Brain Sciences (CBBS), and Department of

Neurochemistry and Molecular Biology, Leibniz Institute for Neurobiology,

Magdeburg, Germany

Oleg Senkov

Molecular Neuroplasticity Group, German Center for Neurodegenerative

Diseases (DZNE), Magdeburg, Germany

Alessandro Simi

Istituto Italiano di Tecnologia, NetS3Laboratory, Neuroscience and Brain

Technologies Dpt., Genova, Italy

August B Smit

Department of Molecular & Cellular Neurobiology, Center for Neurogenomics &

Cognitive Research, Neuroscience Campus Amsterdam, VU University

Amsterdam, HV Amsterdam, The Netherlands

Eduardo Soriano

Department of Cell Biology, University of Barcelona; Centro de Investigacio´n en

Red sobre Enfermedades Neurodegenerativas (CIBERNED), ISCIII, Madrid, and

Vall d’Hebron Institut de Recerca (VHIR), Barcelona, Spain

Sabine Spijker

Department of Molecular & Cellular Neurobiology, Center for Neurogenomics &

Cognitive Research, Neuroscience Campus Amsterdam, VU University

Amsterdam, HV Amsterdam, The Netherlands

Vera Stamenkovic

Center for Laser Microscopy, Institute for Physiology and Biochemistry, Faculty of

Biology, University of Belgrade, Belgrade, Serbia

Michal Stawarski

Laboratory of Cell Biophysics, Department of Molecular and Cellular

Neurobiology, Nencki Institute of Experimental Biology, Warsaw, Poland

Teresa Tarrago

Iproteos S.L., and Institute for Research in Biomedicine (IRB Barcelona),

Barcelona, Spain

Ayse Tekinay

UNAM-National Nanotechnology Research Center, Institute of Materials Science

and Nanotechnology, Bilkent University, Ankara, Turkey

Ursula Theocharidis

Department of Cell Morphology and Molecular Neurobiology, Ruhr-University

Bochum, Bochum, Germany

Department of Molecular & Cellular Neurobiology, Center for Neurogenomics &Cognitive Research, Neuroscience Campus Amsterdam, VU University

Amsterdam, HV Amsterdam, The Netherlands

Jennifer Vandooren

Department of Microbiology and Immunology, Laboratory of Immunobiology,Rega Institute for Medical Research, University of Leuven, Leuven, BelgiumLydia Vargova

Charles University, 2nd Faculty of Medicine, and Institute of ExperimentalMedicine AS CR, v.v.i., Department of Neuroscience, Prague, Czech RepublicNaiara Vazquez

Department of Translational Neuroscience, University of Antwerp, Wilrijk,Belgium

Bernhard Wehrle-Haller

Department of Cell Physiology and Metabolism, Centre Me´dical Universitaire,University of Geneva, Geneva, Switzerland

Hans Werner Mu¨ller

Molecular Neurobiology Laboratory, Department of Neurology, University Medical Center Du¨sseldorf, Du¨sseldorf, Germany

Heinrich-Heine-Grzegorz M Wilczynski

The Nencki Institute of Experimental Biology, Polish Academy of Sciences,Warsaw, Poland

Jakub Wlodarczyk

Laboratory of Cell Biophysics, Department of Molecular and Cellular

Neurobiology, Nencki Institute of Experimental Biology, Warsaw, Poland

Sujeong Yang

John van Geest Centre for Brain Repair, University of Cambridge, Forvie Site,

Robinson Way, Cambridge, UK

The organization of the extracellular matrix (ECM) is a reflection of the role and

function of organs in our bodies The interaction of cells with the ECM determines

their polarity, shape, and form and is providing cues for survival and proliferation

The brain, in comparison with other organs, shows an extremely complex

architec-ture, in which neurons, glial cells, and blood vessels are interacting to create and

maintain a dynamic network, in which beneficial synaptic connections need to be

actively maintained and other remodeled in response to changes in signaling input

Similar to other organ systems, cell–cell interactions based on direct contacts via

cadherins and signaling receptors, as well as cell–matrix interactions with the

ECM scaffold, are controlling the organization of glial cells and neurons as well as

the projections of neurites and location of synapses All these structures are

embedded within an ECM scaffold formed by fiber or network-forming proteins

and membrane-anchored or secreted glycosaminoglycans

Despite recent advances in the ECM field, the importance of neural ECM for

physiological and pathological processes is less widely recognized than that of other

nervous system elements To overcome this, a European consortium “Brain

Extra-cellular Matrix in Health and Disease (ECMNet)” was established in 2010 as a part

of intergovernmental framework for European Cooperation in Science and

Technol-ogy (COST) Now, ECMNet combines more than 200 young and established

re-searchers from 20 European countries (http://www.costbm1001.eu/) Each book

chapter of this volume is prepared involving ECMNet members and other leading

experts from the USA and Japan The chapters cover the broad range of topics,

grouped into four parts, which are devoted to normal physiological functions of

neural ECM, its role in brain diseases, development of methods to image the

ECM, to therapeutically target it, and to generate artificial ECM

FUNCTIONS OF NEURAL ECM

The neural ECM is well recognized to play a key role in neural development and the

first two chapters of the book are devoted to this topic.Theocharidis, Long,

ffrench-Constant, and Faissner (2014)discuss available data on expression of tenascins,

laminins, and proteoglycans in the ECM of the stem cell niche and argue for crucial

importance of ECM for the biology of this cellular compartment Heikkinen,

Pihlajaniemi, Faissner, and Yuzaki (2014) focus on how proteoglycans, tenascin,

and C1q (C1qDC) family proteins regulate synapse formation, maintenance, and

pruning during neural development In the adult central nervous system (CNS),

mul-tiple neural ECM molecules together with astroglial, pre-, and postsynaptic elements

form tetrapartite synapses, and the ECM regulates Hebbian synaptic plasticity

through the modulation of perisomatic GABAergic inhibition, intrinsic neuronal

ex-citability, and intracellular signaling, as presented bySenkov, Andjus, Radenovic,

xiii

Soriano, and Dityatev (2014) This chapter also gives an account on bidirectional ulation of memory acquisition by ECM molecules and highlights that removal of ECMmay promote cognitive flexibility and extinction of fear and drug memories To sta-bilize network dynamics and avoid hypo- and hyperexcitability of neurons, adaptiveHebbian modifications of neurons and synapses must be complemented by homeo-static forms of plasticity.Frischknecht, Chang, Rasband, and Seidenbecher (2014)point to the ECM as a prime candidate to orchestrate and integrate individual cellularstates into the homeostasis of the tissue, which is implemented via synaptic scaling,adjustment in the balance between excitation and inhibition, and axon initial segmentplasticity Many effects of ECM molecules are mediated by their interactions withcognate ECM receptors, first of all, integrins Kerrisk, Cingolani, and Koleske(2014)discuss how activation of ECM receptors modulates downstream signaling cas-cades that control cytoskeletal dynamics and synaptic activity to regulate neuronalstructure and function and thereby impact animal behavior Tsilibary andcolleagues (2014)focus on the role of extracellular proteolysis and put forward a chal-lenging view that the main function of proteolysis is not the degradation of ECM andthe loosening of perisynaptic structures, but rather a release of signaling moleculesfrom the ECM, transsynaptic proteins, and latent forms of growth factors.

mod-NEURAL ECM IN BRAIN DISEASES

As summarized in the first part of this volume, various components of the ECM play

a significant role in maintenance of the environmental milieu for different cell types

in the CNS and in regulation of cellular responses to physiological stimuli ling evidence collected over recent years, however, demonstrate that plasticity in theECM can also be triggered by genetic or acquired pathological stimuli to the brain.Moreover, the ECM is an active player in the CNS repair process by forming a scaf-fold, which orchestrates the cellular plasticity events toward either favorable or un-favorable outcome over the lifespan.Milosˇevic´, Judasˇ, Aronica, and Kostovic (2014)discuss the expression pattern of major components of the fetal ECM in the humanbrain and the role they play during normal laminar and connectivity development aswell as in the neurodevelopmental disorders.Kwok, Yang, and Fawcett (2014)ad-dress current progresses of chondroitin sulfate proteoglycans in regulating plasticity

Compel-in neurodegenerative diseases, braCompel-in tumors, and CNS Compel-injury They also Compel-investigatethe opportunities of manipulating ECM to facilitate postinjury recovery.Vandooren,Damme, and Ghislain Opdenakker (2014)discuss the mechanisms of matrix metal-loproteinase MMP-9 in neuroinflammation, and the use of MMP-9-specific inhibitors

as anti-inflammatory agents Morawski, Filippov, Tzinia, Tsilibary, and Vargova(2014)review the information on age-related changes in the ECM, how they couldcontribute to pathophysiology of neurodegenerative diseases, such as Alzheimer’sdisease, and what could be the therapeutic approaches targeted to the ECM to combat,for example, amyloid clearance Pitka¨nen et al (2014) review the role of uPAR-interactome, MMPs and TIMPs, tenascin-R, and LG1 in different epilepsy syndromes

and how they contribute to epileptogenesis and ictogenesis In addition, the role of the

ECM in epilepsy-related comorbidies and the current status ofin vivo imaging of

ECM-related molecules in patients are discussed.Lubbers, Smit, Spijker, and van

den Oever (2014)review neurodevelopmental and other mechanisms affecting

differ-ent compondiffer-ents of the ECM, which could lead to the expression of neuropsychiatric

disorders, in particular, addiction, schizophrenia, and mood disorders

NEURAL ECM-TARGETING TOOLS AND THERAPEUTICS

There is a growing interest to develop methodology allowing for detailed structural

and functional analysis of ECM, particularlyin vivo, to be able to follow ECM

remo-deling during plasticity and in diseased brains.Zeug et al (2014)provide a detailed

overview of current microscopic methods used for ECM analysis and also describe

general labeling strategies for ECM visualization and imaging of the proteolytic

re-organization of ECM as well as applications of F€orster resonance energy

transfer-based approaches to monitor ECM functions with a high spatiotemporal resolution

Baranger et al (2014)discuss data on the endogenous MMP inhibitors in the CNS

and regulation of MMP-mediated proteolysis in inflammatory, neurodegenerative

and infectious diseases, and synthetic inhibitors of MMPs and the perspective of their

therapeutic use.Berezin, Walmod, Filippov, and Dityatev (2014)provide a

compre-hensive overview of multiple strategies for targeting the ECM molecules and their

metabolizing enzymes and receptors with antibodies, peptides, glycosaminoglycans,

and other natural and synthetic compounds They also discuss application of

devel-oping ECM-targeting drugs in Alzheimer’s disease, epilepsy, schizophrenia,

addic-tion, multiple sclerosis, Parkinson’s disease, and cancer

NEURAL ECM SCAFFOLDS

The unique electrochemical connection at synapses is backed up by multiple

me-chanical connections linking the pre- and postsynaptic membranes to each other

as well as to the surrounding ECM Because of this intimate link between neurites

and their synapses and the unique 3D architecture of the brain, it is so far impossible

to artificially reconstruct the brain Nevertheless, in the last part of this volume, we

would like to address the questions how one could mimic a scaffold that can be used

by neurons and glial cells to create neuronal connections that can be used to

func-tionally replace damaged tissues (Estrada, Tekinay, & Mu¨ller, 2014) To do this,

one does not only need to develop ways of creating surfaces or scaffolds, which

would allow the growth of neurites and glia, but also ways to create electrochemical

connections between the healthy brain tissue and implanted neuronal networks, as

discussed bySimi, Amin, Maccione, Nieus, and Berdondini (2014) An alternative

approach to create new functional brain tissue would be to implant neuronal stem

cells in such a way that glial cells and neurons can rebuild the damaged scaffolds

In order to do this, we require however precise information how a stem cell

compartment is maintained and how differentiating neurons can be instructed to grate, to stop, and to send out axons and dendrites in a stereotype and reproduciblemanner One class of receptors that can read both structural and mechanical infor-mation that are preprogrammed within the ECM are integrins However, to instructneuronal or glial cell behavior via the extracellular scaffold, we need to understandhow integrins are mediating adhesion to ECM and provide specific signaling forneurite extension or maturation of synapses, the aspects discussed by Wehrle-Haller and Bastmeyer (2014).

mi-This volume is the first book focusing on the neural ECM, which is an attempt tosynthesize the views of basic scientists, medical doctors, and engineers how it worksunder normal conditions and in diseased brains and how to repair or reconstitute it

We expect that this volume will become a reference book for PhD students to itate their entry in this complex and dynamic field and will be highly beneficial forestablished neuroscientists to better understand the role of ECM in their “favorite”functions of neural cells, for pharma industry and doctors to include the ECM in theshortlist of therapeutically attractive targets and for tissue engineers to learn how tobetter mimic the complexity of neural ECM and design new functional ECMscaffolds

facil-Alexander DityatevMagdeburg, GermanyBernhard Wehrle-HallerGeneva, SwitzerlandAsla Pitka¨nenKuopio, Finland

Estrada, V., Tekinay, A., Mu¨ller, H.W., 2014 Neural ECM mimetics Prog Brain Res 214,391–413

Frischknecht, R., Chang, K.-J., Rasband, M.N., Seidenbecher, C.I., 2014 Neural ECM ecules in axonal and synaptic homeostatic plasticity Prog Brain Res 214, 81–100.Heikkinen, A., Pihlajaniemi, T., Faissner, A., Yuzaki, M., 2014 Neural ECM and synaptogen-esis Prog Brain Res 214, 29–51

mol-Kerrisk, M.E., Cingolani, L.A., Koleske, A.J., 2014 ECM receptors in neuronal structure, aptic plasticity, and behavior Prog Brain Res 214, 101–131

syn-Kwok, J.C.F., Yang, S., Fawcett, J.W., 2014 Neural ECM in regeneration and rehabilitation.Prog Brain Res 214, 179–192

Lubbers, B.R., Smit, A.B., Spijker, S., van den Oever, M.C., 2014 Neural ECM in addiction,

schizophrenia, and mood disorder Prog Brain Res 214, 263–284

Milosˇevic´, N.J., Judasˇ, M., Aronica, E., Kostovic, I., 2014 Neural ECM in laminar

organiza-tion and connectivity development in healthy and diseased human brain Prog Brain Res

214, 159–178

Morawski, M., Filippov, M., Tzinia, A., Tsilibary, E., Vargova, L., 2014 ECM in brain aging

and dementia Prog Brain Res 214, 207–227

Pitka¨nen, A., Ndode-Ekane, X.E., Łukasiuk, K., Wilczynski, G.M., Dityatev, A., Walker, M.C.,

et al., 2014 Neural ECM and epilepsy Prog Brain Res 214, 229–262

Senkov, O., Andjus, P., Radenovic, L., Soriano, E., Dityatev, A., 2014 Neural ECM molecules

in synaptic plasticity, learning and memory Prog Brain Res 214, 53–80

Simi, A., Amin, H., Maccione, A., Nieus, T., Berdondini, L., 2014 Integration of

micro-structured scaffolds, neurons and multielectrode arrays Prog Brain Res 214, 415–442

Theocharidis, U., Long, K., ffrench-Constant, C., Faissner, A., 2014 Regulation of the neural

stem cell compartment by extracellular matrix constituents Prog Brain Res 214, 3–28

Tsilibary, E., Tzinia, A., Radenovic, L., Stamenkovic, V., Lebitko, T., Mucha, M., et al., 2014

Neural ECM proteases in learning and synaptic plasticity Prog Brain Res 214, 135–157

Vandooren, J., Damme, J.V., Ghislain Opdenakker, G., 2014 On the structure and functions of

gelatinase B/matrix metalloproteinase-9 in neuroinflammation Prog Brain Res 214,

193–206

Wehrle-Haller, B., Bastmeyer, M., 2014 Intracellular signaling and perception of neuronal

scaffold through integrins and their adapter proteins Prog Brain Res 214, 443–460

Zeug, A., Stawarski, M., Bieganska, K., Korotchenko, S., Wlodarczyk, J., Dityatev, A., et al.,

2014 Current microscopic methods for the neural ECM analysis Prog Brain Res 214,

287–312

COST—Matrix for European

research networks

Srec´ko Gajovic´*,1, Roland Pochet{,2

*Croatian Institute for Brain Research, University of Zagreb School of Medicine, Zagreb, Croatia;

COST Domain Committee Biomedicine and Molecular Biosciences, Rapporteur of COST Action

BM1001—ECMNET: Brain Extracellular Matrix in Health and Disease, Brussels, Belgium

{Faculte´ de Me´decine, Universite´ Libre de Bruxelles, Brussels, Belgium

1 Corresponding author: Tel.: +358-1-4566948; Fax: +358-1-4566795,

e-mail address: srecko.gajovic@hiim.hr

Abstract

Today’s researchers are faced with a change from curiosity-driven to mandate-driven research

These two approaches are well combined within scientific networks (Actions) supported by

the European Cooperation in Science and Technology (COST) program The functioning

of COST Actions, although directed only to networking, has a substantial impact on European

science and can be compared to the functioning of the extracellular matrix in the brain, which

although scarce plays a key role in initiation, maintenance, and plasticity of intercellular

in-teractions in the nervous system COST networks enable interdisciplinary approach and

sup-port early-stage researchers, which is a vital asset for the advancement of science

Keywords

curiosity-driven research, mandate-driven research, extracellular matrix, interdisciplinarity,

early-stage researchers

Scientists are frequently considered as a distinct group of people, not sharing the

same characteristics as the general population Although this stereotype is certainly

far from reality, it shapes the public perception Scientists are perceived as people

with broad knowledge dedicated to solving unresolved issues with persistence and

care of the details The origin of the stereotype are famous scientists in history

and different fictional or real-life characters described in the literature, movies,

the-ater plays, or newspaper articles One should note that their success is often

consid-ered to be a result of individual effort and a consequence of some of their personality

2 Past Chair of COST Domain Committee Biomedicine and Molecular Biosciences (2010–2013),

Brussels, Belgium.

xix

features One of these features is curiosity, and curiosity was agreed as a major ing force of science This type of research is referred to as curiosity-driven research

driv-or blue-skies research, and today, scientists will strongly defend their scientific dom and the right to perform research based on their individual curiosity(Linden, 2008)

free-On the other hand, today’s science is a complex network of many interactingelements, which make scientific activities far more demanding than a simple pursuit

of individual curiosities An opposing approach to curiosity-driven research ismandate-driven research The science of today is not only here to provide newknowledge but also to solve the emerging societal issues and contribute to the soci-etal needs (Svalastog, 2014) The turning point in this process was considered to bethe Manhattan project, when governments of several countries engaged numerousscientists as well as other professionals in a joint effort to create the first atomicbombs The project involved more than 130,000 people and the cost of nearly

US$2 billion (in the 1940s; Web page Wikipedia, 2014: http://en.wikipedia.org/wiki/Manhattan_Project, as assessed on June 6, 2014)

Although curiosity is an inevitable element of today’s science, it is clear that rently we have a highly expensive system involving many individuals with differentexpertise If we pursue the extracellular matrix in the brain analogy, in order toachieve the brain complexity, many individual cells should be engaged and their ac-tivities should be supported The brain activities are supported by the activity of thewhole body, which “serves” the brain and which is “governed” by the brain An im-portant part of this support is provided directly in the brain by the extracellular ma-trix As the extracellular matrix in the brain is scarce, at first it was considered as notvery important, but it is more and more recognized as essential component for for-mation, maintenance, and plasticity of synaptic connections and for concerted action

cur-of neuronal, glial, vascular, and immune cells

Although the parallels between biological systems and society are only of trative nature, they are still worth exploring The resources used by the today’s sci-ence are considerable and need to be justified Therefore, the mandate-drivenresearch is frequently imperative for scientists to maintain their activities Theresource allocation by granting agencies is usually organized on the top-down basis,where the need is first identified and then the resources are allocated to the projectbest fitting to the requested mandate In this context, the European Union (EU)defines the needs at the level of the European Parliament and subsequently theEuropean Commission organizes the granting system (previously FrameworkProgrammes, currently Horizon 2020), which addresses the predefined needs withdedicated calls This rationale is clearly supported by the need to justify the resourcescollected from the EU countries and their tax payers Still it is constantly criticized byscientific community as it predicts a novelty before it appears, and leaves manypotentially fruitful research lines unrecognized and consequently unfunded One

illus-of the ways to complement the top-down approach is to apply the bottom-up proach, such as in programs financed through European Commission, the most no-table examples of which are COST (European Cooperation in Science andTechnology) and the European Research Council (ERC)

ap-COST was founded in 1971 long before the establishment of the EU in 1993.

Therefore, COST is an intergovernmental organization, governed by the European

member states, some of which are non-EU countries (currently, COST has 34

mem-ber states), and Israel as a Cooperating State (Web page COST, 2014:http://www

cost.eu/, as assessed on June 6, 2014) The operating costs are currently covered by

grants obtained from the European Commission COST is dedicated to select

net-works using the bottom-up approach The basic form of this network is referred

to as an Action, which is granted for a limited time period to achieve its tasks through

networking Although these tasks could be rather complex, the financing is limited

only to support the networking activities, rather than to provide the resources (e.g.,

equipment and consumables) necessary to achieve this goal The Action financing is

considered as only a top-up to the existing resources enabling research activities, and

the Action should provide a concerted effort using its other resources to achieve more

than an individual group can do by itself

It is obvious that the financial contribution of COST Action is scarce in

compar-ison to the total resources used by the Action members By analogy, it could be

con-sidered negligible in the same way that the scarce extracellular matrix was

considered not important for the brain On the contrary, the cumulative experience

gained throughout the history of COST shows that COST is indeed a very useful

in-strument supporting European Science and Technology Achievements of COST

networks are confirmed not only by the so-called tangible results (e.g., number of

joint scientific publications) but also by many intangible benefits, such as cultivating

a spirit of large-scale interdisciplinary research The opinion of the authors is that

these intangible results are of the outmost importance and have profoundly changed

the European science

As any study of COST would lack a control group, i.e., a group of countries

re-sembling Europe and without COST, it is impossible to verify the contribution of

COST and difficult to demonstrate the benefits of COST to the European science

Here, we can use the example of extracellular matrix in the brain showing its

impor-tance for coordination of interactions between neurons and other cells, aiming to

maintain cellular networks, while supporting their adaptive functional plasticity

In the same way, COST Actions have helped European scientists to get to know each

other, communicate, and coordinate their individual efforts toward a joint

coopera-tion Although Actions are funded for a limited period of time, the strength of these

collaborations is not limited to the Action lifetime, but it lasts long after and

con-tinues in different forms of joint activities, many of them not even related to

re-search, but rather to education, regulatory work, entrepreneurship, and novel

economical activities Very often, the extracellular matrix is formed between cells

of different types/origins and serves to avoid antagonism between them, so to say to

harmonize their relationships One of the well-known examples is the ECM

mole-cule laminin 11, which is a part of extracellular matrix that prevents invasion of

Schwan cell processes into the synaptic cleft between the motor nerve terminal

and the muscle fiber and thus helps to maintain synaptic transmission (Patton,

Chiu, & Sanes, 1998) Similarly, COST Actions help to avoid competition between

groups by stimulating cooperative activities

One of the COST particularities is the support of interdisciplinary research Inthis context, it is noteworthy that neural extracellular matrix molecules are secreted

by neurons and glial cells and form a structure integrating components derived fromthese major cell types Thus, the extracellular matrix is not just an extension of neu-rons or glia into the extracellular space but a unique integrative entity, which canreceive and release specific signals in response to cellular activity Similarly, COSTActions are composed from experts from different fields that bring their expertize toaddress specific scientific and social needs and generate unique knowledge, con-cepts, and products Currently, to make substantial scientific breakthroughs is mainlypossible through interdisciplinary efforts The difficulties to achieve interdisciplin-ary collaboration are considered as a major obstacle to further scientific advance-ment (Bennett & Gadlin, 2012) COST Actions are a unique instrument to solvethis issue The complexity of science clearly dictates the need for specialization,meaning that every individual can gain only a tiny subset of the total knowledgeand skills Combining different expertize in a network is an obvious solution, stillthe interdisciplinary efforts are difficult to achieve One of the major obstacles de-scribed is a communication problem because different disciplines develop differentapproaches and even different vocabularies, hindering the cross talk between disci-plines COST Actions, by gathering experts from different fields, stimulate the crosstalk and offer the dedicated time and recourses to exchange information and opin-ions COST Actions also involve early-stage researchers, and it is of outmost impor-tance to allow young researchers to communicate and understand other disciplines.This can be seen as investment into future that can further promote the advancement

of European science

To maintain COST networks productivity is not an easy task It is achieved byorganizing the scientific committees on the basis of the intergovernmental principle.According to this principle, scientists belonging to the scientific committees couldassist other scientists to develop networks However, the evaluation and monitoringsystem based on standing scientific committees received criticism mainly related tothe standardized procedures and avoidance of conflicts of interest This is anotheraspect of the development of our society, where public institutions simultaneouslypromote and undermine general trust (Robbins, 2012) Frequently, to regain trust,administrative procedures take precedence to free collaboration Paradoxically,the system gets complicated, therefore less transparent (opposite to what wasintended to be achieved), and the public trust further weakens Whether the same willhappen in the future of COST remains to be seen, but it is certain that the currentCOST system based on standing scientific evaluation and monitoring committeeswill expire in September 2014

In conclusion, just as the extracellular matrix is essential to functioning of thebrain, the networking of scientists is essential for further advancement of science.One of the important networking tools is COST, and among the exemplary results

of COST Action networks is the current issue ofProgress in Brain Research Wethink that promoting networking and bottom-up approach represents an optimumbalance between curiosity- and mandate-driven research Networking is a skill that

needs to be developed and that enables the cross talk between the disciplines

There-fore, training of early-stage researchers in networking skills and maintaining

func-tional networks is vital for further advancement of science

REFERENCES

Bennett, L.M., Gadlin, H., 2012 Collaboration and team science: from theory to practice

Journal of Investigative Medicine 60, 768–775

Linden, B., 2008 Basic Blue Skies Research in the UK: Are we losing out? Journal of

Bio-medical Discovery and Collaboration 3, 3

Patton, B.L., Chiu, A.Y., Sanes, J.R., 1998 Synaptic laminin prevents glial entry into the

syn-aptic cleft Nature 393, 698–701

Robbins, B.G., 2012 A blessing and a curse? Political institutions in the growth and decay of

generalized trust: a cross-national panel analysis, 1980–2009 PLoS One 7, e35120

Svalastog, A.L., 2014 The value of bio-objects and policy discourses in Europe Croatian

Medical Journal 55, 167–170

Regulation of the neural

stem cell compartment by

*Department of Cell Morphology and Molecular Neurobiology, Ruhr-University

Bochum, Bochum, Germany

{ MRC Centre for Regenerative Medicine, The University of Edinburgh, Edinburgh, UK

1 Corresponding author: Tel.: +49-234-3223851; Fax: +49-234-3214313,

e-mail address: andreas.faissner@rub.de

Abstract

Neural stem cells (NSCs) derive from the neuroepithelium of the neural tube, develop into

radial glial cells, and recede at later developmental stages In the adult, late descendants of

these embryonic NSCs reside in discretely confined areas of the central nervous system,

the stem cell niches The best accepted canonical niches are the subventricular zone of the

lateral ventricle and the subgranular zone of the dentate gyrus of the hippocampus Stem cell

niches provide a privileged environment to NSCs that supports self-renewal and maintenance

of this cellular compartment While numerous studies have highlighted the importance of

tran-scription factors, morphogens, cytokines, and growth factors as intrinsic and extrinsic factors

of stem cell regulation, less attention has been paid to the molecular micromilieu that

charac-terizes the stem cell niches In this chapter, we summarize increasing evidence that the

extra-cellular matrix (ECM) of the stem cell environment is of crucial importance for the biology of

this cellular compartment A deeper understanding of the molecular composition of the ECM,

the complementary receptors, and the signal transduction pathways engaged may prove highly

relevant for harnessing NSCs in the context of biotechnological applications

Keywords

Asymmetrical division, Extracellular matrix, Glial progenitors, Integrins, Laminin, Neural

stem cell niche, Phosphacan, Proteoglycans, Radial glial cells, Subventricular zone, Tenascin

Progress in Brain Research, Volume 214, ISSN 0079-6123, http://dx.doi.org/10.1016/B978-0-444-63486-3.00001-3

1 NEUROGENESIS UNFOLDS IN DISTINCT STEPS

AND INVOLVES NEURAL STEM CELLS

The neural plate derives from the neuroectoderm and is composed of a layer of roepithelial cells that expand by symmetrical division With increasing growth of theneural tube, the neuroepithelium progressively elongates to give rise to radial glialcells with a radial morphology whose processes span the developing CNS These ra-dial glia give rise to neurons and glial cellsin vivo and thus serve as authentic neuralstem cells (NSCs;Anthony et al., 2004; Hartfuss et al., 2001; Malatesta et al., 2000,2003; Noctor et al., 2001, 2004) They divide by symmetrical divisions at the inner(or ventricular) surface of the developing neural tube—the ventricular zone—at earlystages but later on give birth to neurons by switching to an asymmetrical divisionmode (Kriegstein and Alvarez-Buylla, 2009; Merkle and Alvarez-Buylla, 2006)

neu-In the classical model of radial glial-guided migration, this cell population has alsobeen highlighted as a scaffold for migrating neurons (Rakic, 2007) In mammalianspecies, neurons may be generated via a population of intermediate progenitors (oramplifying precursors) that are generated by the asymmetrical stem cell divisionsand then undergo a restricted number of symmetrical divisions so increasing neuro-genesis and enabling ongoing cortical expansion (Gotz and Huttner, 2005) In spe-cies that are characterized by a substantially enlarged cortical surface such asprimates, additional populations of amplifying precursors are found in a further, dis-tinct neurogenic zone close to the cortical surface, so fueling the further corticalgrowth (Fietz and Huttner, 2011; Hansen et al., 2010)

Following neurogenesis, the stem cells switch to the generation of astrocytes andoligodendrocyte precursors, with the latter migrating to target regions to myelinatethe axonal connections (Nave, 2010) The majority of the radial glia undergo a finalsymmetrical division to generate two differentiated daughters and so vanish from theCNS, with the exception of the Bergmann glia of the cerebellum and the Mu¨ller glia

of the retina Some descendants of the radial glia, however, persist as radial-type trocytes that reside in the subventricular zone (SVZ) of the lateral ventricle and thesubgranular zone (SGZ) of the dentate gyrus of the hippocampus There, the astro-cytes act as adult NSCs and serve neurogenesis in the adult CNS (Kriegstein andAlvarez-Buylla, 2009)

as-2 MOLECULAR DETERMINANTS OF ASYMMETRICAL DIVISION

The transition from symmetrical to asymmetrical division of radial glia accompaniesthe generation of distinct neural cell precursor populations Asymmetrical divisionimplies that the daughter cells of a dividing stem cell adopt different fates that arereflected by distinct patterns of gene expression and consequently differentiationpathways Conceptually the differences of the daughter cells could be caused either

by a differential repartition of intrinsic determinants of the dividing stem cell, or,alternatively, the daughter cells might segregate into different microenvironmentsthat subsequently would engage specific receptors and drive the progeny into distinct

differentiation pathways Evidence for the selective distribution of intrinsic

determi-nants during stem cell division has been obtained in drosophila, for example, the

pro-tein numb (Couturier et al., 2013), male versus female chromosomes, or centrosomes

(Yadlapalli and Yamashita, 2013) In mammalian NSCs, the asymmetrical

distribu-tion of apical membranes of radial glia as a consequence of the inclinadistribu-tion angle of

the cell division plane has been highlighted (Gotz and Huttner, 2005), as well as the

asymmetrical distribution of centrosome-associated primary cilium membrane

(Paridaen et al., 2013)

3 ENVIRONMENTAL ASYMMETRY AND THE STEM CELL NICHE

The alternative interpretation of divisional asymmetry instructed by different

micro-environments that drive the progeny into distinct differentiation pathways

empha-sizes the influence of the cellular microenvironment In the adult nervous system,

NSCs are confined to privileged areas, the so-called stem cell niches Stem cell

niches have been detected in various organs and are characterized by the coexistence

of stem cells, supporting niche cells, the neighborhood of blood vessels, and a

spe-cialized extracellular matrix (ECM;Scadden, 2006) In this microenvironment, stem

cells interact with niche cells and other cellular components via membrane-mediated

cell–cell interactions and respond to morphogens, cytokines, and growth factors; to

autocrine, paracrine, and endocrine signals; and to ECM components

4 THE STEM CELL NICHES OF THE ADULT CNS

In the adult CNS, the two generally accepted canonical regions of neurogenesis are

the SVZ of the lateral ventricle and the SGZ of the hippocampus It is likely that these

neurogenic areas are characterized by specialized environments that sustain NSCs

and, as in niches elsewhere, function as integrative entities for a large number of

physiological stimuli (Scadden, 2006; Zhao et al., 2008) In the CNS, the specialized

niche microenvironment is constituted by astrocytes, endothelia of blood vessels,

leptomeningeal cells, and the cerebrospinal fluid in the case of the SVZ

Morpho-gens, cytokines, ECM constituents, and neurotransmitters are released into the niche

environment (Ihrie and Alvarez-Buylla, 2011; Kazanis and ffrench-Constant, 2011;

Patel et al., 2012) In these niches, a subclass of slowly dividing astrocytes

(descen-dants of the radial glia, the major neural stem/progenitor cell (NSPC) of the

devel-oping nervous system as described earlier) that express the transcription factor Sox2

and the intermediate filament protein GFAP act as stem cells (Ming and Song, 2005)

These astrocytes are also called type B cells and continuously generate

transit-amplifying precursors, type C cells, which rapidly expand the cell pool and develop

further to the type A cells, the neuroblasts These migrate towards the olfactory bulb

through a migration path that is surrounded by (non-stem cell) astrocytes, forming

the rostral migratory stream The morphogen sonic hedgehog (Shh) supports the

proliferation and the maintenance of the neuroblasts When they reach their target,these neuroblasts differentiate into dopaminergic interneurons and contribute to theregeneration of the local olfactory network (Ming and Song, 2011; Zhao et al., 2008).

In the SGZ of the hippocampal dentate gyrus in the adult forebrain (Kempermann

et al., 2004), the granule neurons of the hippocampus are born continuously from theNSPCs It has been proposed that this neurogenesis in the hippocampus plays an im-portant role for memory formation (Garthe and Kempermann, 2013) As in the SVZ,this occurs via the formation of an amplifying precursor population Here, however,the cellular and molecular composition of the niche is poorly established althoughthe observation that increasing blood vessel density in the SGZ increases neurogen-esis (Licht et al., 2011) suggests that blood vessels are an important part of the nichejust as they are in the SVZ

In recent years, increasing evidence has pointed to the versatility and variousfunctions of the ECM in the NSC compartment The ECM consists of glycoproteinsand proteoglycans and assembles to selective and specific macromolecular super-structures in the stem cell microenvironment (Barros et al., 2011; Dityatev et al.,2010; Garwood et al., 2001) A systematic comparison of transcriptomes has under-lined that the ECM microenvironment distinguishes the neurogenic zones of the in-ner and the outer SVZs in the human embryonic brain The data suggested that celladhesion and cell-ECM interactions are relevant for the proliferation and self-renewal

of neural progenitors in the developing human neocortex Important classes of ECMmolecules that have emerged from expression of the stem and amplifying precursor-containing regions include tenascins, collagens, laminins, proteoglycans, and theintegrin receptors (Fietz et al., 2012) In particular, the stimulation of the integrinavb3 promotes the expansion of basal progenitors of the mouse brain by increasingcell cycle reentry of Pax6-negative and Tbr2-positive intermediate progenitors(Stenzel et al., 2014) Next, therefore, we will discuss ECM molecules implicated

by prior work in adult neural niches The primary focus will be the tenascins, afterwhich we will review the work on laminins, proteoglycans, and ECM receptors

5 TENASCIN PROTEINS IN THE NSC NICHE

Tenascin-C proteins were among the first identified ECM proteins of the NSC niche(Gates et al., 1995) The glycoproteins tenascin-C (TN-C), tenascin-R (TN-R),tenascin-X (TN-X), and tenascin-W (TN-W) of the tenascin gene family share aset of structural motifs, namely, a cysteine-rich N-terminus, a sequence of fibronec-tin type III (FNIII) modules, and homologies to fibrinogen-beta at the C-terminus(Tucker et al., 2006) TN-C was of particular interest because it is transientlyexpressed by astrocytes in the developing CNS There, it is distributed in discreteboundary-like patterns, for example, in the barrel field of the somatosensory cortex

of the mouse (Faissner and Steindler, 1995) Numerous studies have highlighted pulsive, inhibitory, or stimulatory effects of TN-C on axon growth and guidance anddemonstrated cell type-dependent differences in TN-C responsiveness (Faissner,

re-1997; Wehrle-Haller and Chiquet, 1993) As a special feature, TN-C monomers

com-prise alternatively spliced FNIII domains between the fifth domain and the sixth

do-main of the constitutive structure, up to six motifs in the mouse and nine motifs in the

human (Fig 1) These domains are spliced independently from one another, which

provides the basis for 2ncombinatorial variants with n individual FNIII domains

Thus, up to 64 combinations are conceivable in the mouse, and 512 variants could

potentially be generated in the human In fact, 24 variants were found on the mRNA

level in an initial screen in the developing mouse cerebellum (Joester and Faissner,

1999, 2001; Theocharidis and Faissner, 2012) There is evidence that the

combina-torial variants are regulated during development (Rigato et al., 2002) and in response

to lesions (Dobbertin et al., 2010; Garwood et al., 2012).TN-C thus belongs in a

cat-egory of genes that can generate a large number of variants by alternative splicing of

structural domains, similar to dsCAMs (Zipursky and Sanes, 2010) or to neurexins

(Craig and Kang, 2007) Unlike the latter two that are expressed in neurons,TN-C so

far is the only glial gene with this remarkable characteristic.Tenascin genes seem to

have emerged in urochordates but not in other invertebrate phyla and hence may be

FIGURE 1

Multimodular tenascin-C structure Tenascin-C appears as oligomeric protein consisting

of six monomers The monomers show a multimodular structure with an N-terminal

cysteine-rich assembly domain and 14,5 EGF-like repeats, followed by a number of fibronectin

type III (FNIII) domains Six of them are constitutively expressed and can be supplemented

with up to six (in the mouse) alternatively spliced FNIII domains This leads to sequences

of different sizes that have been analyzed in detail (Joester and Faissner, 1999; Theocharidis

and Faissner, 2012; von Holst et al., 2007)

specific for chordates (Tucker et al., 2006) Genes with related egf-type repeats havebeen described in drosophila, namely, theTenm genes (Baumgartner et al., 1994).The N-terminus links tenascin monomers to multimers, for example, TN-C assem-bles to hexamers under nonreducing conditions that appear as so-called hexabra-chions upon electron microscopy of rotary-shadowed preparations (Chiquet-Ehrismann and Tucker, 2011).

The adult niche of the SVZ is strongly enriched in TN-C where it is expressed bytype B cells and deposited in an area neighboring the ependymal cell layer (Fig 2) Inthe tenascin-C / knockout, minor structural deficits of the niche could be detected,the number of stem cells and their progeny however was not affected, and the regen-eration of the stem cell compartment upon treatment with cytosine beta-D-arabino-furanoside was not different from the wild type (Kazanis et al., 2007) The astrocytessurrounding the migrating type A neuroblasts are enriched with TN-C that may keepthe neuroblasts on track by its repulsive properties (Faissner and Kruse, 1990;Jankovski and Sotelo, 1996) Additionally, in their destination in the olfactory bulb,the related gene TN-R attracts the neuroblasts out of the stream into the periglomer-ular networks (Saghatelyan et al., 2004)

6 EXPRESSION OF TENASCIN GENES IN RADIAL GLIA

AND ASTROCYTE PROGENITORS

The expression of TN-C in the adult stem cell niche has led to further studies at lier developmental stages In the mouse neural tube, TN-C can be detected at aroundE12–E13 and is clearly expressed in radial glia (Fig 3;Garcion et al., 2001, 2004).Radial glial cells can be identified with selective markers such as vimentin, BLBP,GLAST, and nestin The cells can be cultivated in the model of neurospheres, aggre-gates of cells in suspension culture that comprise NSPCs and various progenitors.These neurospheres express TN-C and 20 isoforms were detected on the messagelevel in cultures derived from embryonic forebrain Among these, the combination

ear-of FNIII domains A1A4BD was novel and has so far only been found in NSPCs.Vector-driven overexpression of the paired-box transcription factor Pax6, a tran-scription factor that is characteristic of radial glia, resulted in enhanced expression

of TN-C variants that contain 4–6 alternatively spliced FNIII domains (von Holst

et al., 2007) In order to study the biological significance of TN-C for NSCs, spheres obtained from wild-type and tenascin-C / knockout mice were compared.These studies showed that the upregulation of the EGF receptor is delayed in the ab-sence of TN-C, which modifies the response of NSPCs to growth factors Overall, thetransition from solely FGF2-responsive towards FGF2- and EGF-responsive NSPCsseems favored by TN-C (Garcion et al., 2004) To understand in more detail potentialunderlying genetic mechanisms, gene trap lines of NSPCs were established and ex-posed to TN-C proteins as stimulus The analysis of responsive genes revealed thatTN-C represses the expression of Sam68, a member of the STAR family of splicingfactors that binds to several mRNA species When overexpressed in NSPCs, Sam68

neuro-drives the expression of high Mr tenascin-C variants, indicative of a reciprocal

reg-ulatory loop (Moritz et al., 2008) These results suggest that TN-C in the SVZ

envi-ronment impinges on the gene expression program of NSPCs (von Holst, 2008) This

conclusion was reinforced when the spinal cord of the tenascin-C / mutant was

studied There, the elimination of TN-C results in enhanced proliferation of

FIGURE 2

Tenascin-C is present in mature stem cell niches Neural stem cells persist in the neurogenic

subventricular zone lining the lateral ventricles in the mature mouse forebrain Type

B cells are slowly dividing stem cells that produce fast-dividing transit-amplifying type C cells

The latter give rise to neuroblasts (A cells) that migrate through the rostral migratory

stream toward the olfactory bulb The extracellular matrix in the stem cell compartment

contains tenascin-C (arrowheads) (Gates et al., 1995; Kazanis et al., 2007) and laminin

(Kazanis and ffrench-Constant, 2011; Kazanis et al., 2010) interacting with the different

cell types in distinct manner Abbreviations: BV, blood vessel; Cor, cortex; LV, lateral ventricle;

SVZ, subventricular zone

FIGURE 3

Tenascin-C is present in the embryonic stem cell niche Radial glial cells are the main stemcell type in the developing forebrain They are positive for the transcription factor Pax6 Theircell bodies are located in the ventricular zone lining the lateral ventricles Radial glial cellsprovide guiding fibers for newborn neurons migrating outward to their destined layers Matureneurons settle in the outer cortical layers and are positive forbIII tubulin The ECM proteintenascin-C is expressed in the ventricular zone where the neural stem cells reside.Abbreviations: CP, cortical plate; LV, lateral ventricle; SVZ, subventricular zone; VZ,ventricular zone

FGF2-responsive progenitors, a delay of EGFR expression, an increase in the

num-ber concomitant with a decrease of migration of FGFR3-positive astrocyte

progen-itors, and a transiently increased number of GFAP-positive astrocytes (Karus et al.,

2011) This modification in the pool of astrocyte progenitors is accompanied by a

shift in the expression territories of several transcription factors that characterize

pro-genitor domains of the spinal cord (Freeman, 2010), for example, a dorsal expansion

of Nkx2.2 and Nkx6.1 A gene array analysis showed that several genes are

upregu-lated in the E15 spinal cord of the TN-C mutant mouse, interestingly the gene

sulfatase-1 This enzyme removes sulfates from complex glycosaminoglycans

(GAGs) and thereby weakens the heparan sulfate-dependant signaling process,

no-tably of FGF2 cytokines and Shh (Lamanna et al., 2008) In conclusion, the

dysre-gulation of sulfatase-1 activity in consequence of TN-C ablation may weaken the

FGF2 signaling in the spinal cord and thereby delay the acquisition of EGF

respon-siveness observed in the spinal cord NSPCs (Karus et al., 2011)

Bergmann glia cells of the cerebellum are another type of radially oriented cells

with glial characteristics They are present in the developing cerebellum with their

cell bodies located in the Purkinje cell layer and radial processes contacting the outer

cerebellar surface During postnatal development, precursor cells in the external

granular layer of the cerebellum proliferate and generate a large number of cells that

in their postmitotic stage leave their original position and start migrating inward

These newborn granular cells get in contact to the radial processes of Bergmann glia

cells and use them as guiding fibers through the tissue towards the inner granular

layer (Xu et al., 2013) There, they start to develop their dendrites and axons and

build connections to their interaction partners

The migration process of granular cells depends on the presence of guiding

mol-ecules in the tissue and on the surface of the interacting cells Molmol-ecules presented by

the glial cells ensure their functional integrity and can be detected by surface

recep-tors in the neuronal membrane TN-C is present in the molecular layer where it is

deposited by Bergmann glia cells (Fig 4) They express TN-C as soon as they appear

from radial glial cells during embryonic development in the ventricular zone of the

cerebellar primordium (Yuasa, 1996) It could be shown that the interaction of TN-C

with the granular cells in the molecular layer can be blocked by pertubating

anti-bodies and that this decreases the neuronal migration along Bergmann glia processes

(Husmann et al., 1992) Antibodies detecting different domains of the TN-C

mole-cule have differential impact on this functional neuron–glia interaction, which

im-plies the heterogeneity of TN-C domain functions

7 EXPRESSION OF TENASCIN GENES IN OLIGODENDROCYTE

PROGENITORS

Tenascins have also been implicated in the regulation of a third stem cell population

in the adult CNS, the adult oligodendrocyte precursors These proliferate throughout

life (Ffrench-Constant and Raff, 1986) and generate new oligodendrocytes in the

FIGURE 4

Bergmann glia cells in the cerebellum express tenascin-C Granular neurons (NeuN-positivenuclei) are born in the periphery and migrate inward along fibers of Bergmann glia cells(vimentin-positive, arrows in insert).In situ hybridizations show the expression of

tenascin-C in the Purkinje cell layer where the cell bodies of Bergmann glia are located Thetenascin-C protein (green) is secreted to the extracellular space and provides the matrix formigrating neurons (positive forbIII tubulin) Abbreviations: EGL, external granular layer; IGL,inner granular layer; ML, molecular layer; PCL, Purkinje cell layer; WM, white matter

normal (Young et al., 2013) and damaged (Zawadzka et al., 2010) CNS Their

emer-gence parallels the late phase of neurogenesis in the murine neural tube and precedes

a later phase of astrocyte progenitor formation (Kriegstein and Alvarez-Buylla,

2009) When this developmental phase was monitored in the tenascin-C / knockout

mutant, it appeared that the proliferation of OPCs was reduced, while the migration

rate into the optic nerve was increased and accompanied by a reduced rate of

apo-ptosis (Garcion et al., 2001) This may be the reason why overall the myelin pattern

in adult tenascin-C / mutants is structurally and functionally normal (Kiernan et al.,

1999) An interesting switch of tenascin gene expression has been described in

OPCs, in that tenascin-C is produced by A2B5-positive OPCs and progressively

downregulated, while TN-R emerges with maturation of oligodendrocytes

(Czopka et al., 2009) The more mature oligodendrocytes produce TN-R that occurs

in two isoforms that differ by one FNIII domain Using oligodendrocyte cultures and

tenascin-C / or tenascin-R / knockout mouse mutants, it could be shown that

TN-C delays while TN-R favors the maturation of OPCs, as revealed by the

expres-sion of the maturation marker myelin basic protein (MBP; Czopka et al., 2009;

Garwood et al., 2004) The inhibitory mechanism engaged by TN-C in OPCs

in-volves the Ig superfamily member contactin (Cntn1) in association with the Src

fam-ily kinase Fyn in lipid rafts and possibly the RNA-binding molecule Sam68

downstream of Fyn Upregulation of Sam68 accelerates while downregulation of

Sam68 retards the maturation of oligodendrocytes (Czopka et al., 2010)

8 REGULATION OF TENASCIN GENES IN NSCs

Tenascin-C is transiently expressed during development and downregulated in most

adult tissues, with the exception of the stem cell niches Furthermore TN-C is

upre-gulated under pathological conditions in wounds, in the mesenchyme of carcinomas,

and in gliomas, in which the level of expression seems to correlate with malignity

and increased metastasis (Chiquet-Ehrismann and Tucker, 2011; Orend and

Chiquet-Ehrismann, 2006) The tight regulation of TN-C begs the question of which

factors are involved on the levels of transcription and receptor-mediated signaling

Several binding sites can be found in the regulatory sequences of theTN-C gene The

binding of the neural transcription factor OTX2 to the humanTN-C promoter has

been described (Briata et al., 1999; Gherzi et al., 1997; Jones et al., 1992), and in

the embryonic stem cell niche, the regulation of theTN-C splice variants by the

homeobox transcription factor Pax6 was shown Large isoforms of the glycoprotein

are favored upon Pax6 overexpression (von Holst et al., 2007) When this important

factor for the development of radial glial cells is missing, the expression of TN-C is

impaired in the developing forebrain (Gotz et al., 1998; Holm et al., 2007; Stoykova

et al., 1997) Some Krox and POU homeodomain transcription factors, the

homeo-box transcription factors evx1 and prx1, and NF-kB and c-Jun directly bind to the

promoter region and regulate the TN-C gene expression (Copertino et al., 1995,

1997; Gherzi et al., 1997; Jones and Jones, 2000; Jones et al., 1990, 1992, 1993,

2001; Mettouchi et al., 1997) Other regulations are predicted by studying possiblebinding sites in the DNA sequence, but a functional significance in NSC develop-ment and maintenance has not yet been clarified.

In the stem cell, niche soluble factors display important extrinsic regulators ofNSC functions Several growth factors and cytokines have been described with animpact on TN-C gene regulation In the adult stem cell niche, the expression ofTN-C is enhanced upon injection of EGF to the lateral ventricle alongside an in-creased proliferation rate of transit-amplifying precursors (Doetsch et al., 2002) As-trocytes can be stimulated and change their expression profile of TN-C and its splicevariants when treated with bFGF or TGF-b1, which resembles the reaction to path-ogenic stimulus but may also be the case for NSCs with astrocytic characteristics(Dobbertin et al., 2010; Meiners et al., 1993)

9 LAMININ PROTEINS IN THE ADULT STEM CELL NICHE

The laminins are the best-described ECM component of the adult stem cell nicheselsewhere in the body They are a family of trimeric proteins, consisting of an a,

b, and g subunit The nomenclature now describes the combination of the known five

a, three b, and three g chains that make up 1 of the 16 isoforms (Aumailley et al.,

2005) In the CNS, NSCs and supporting cells within the niche, such as astrocytes,express laminin (Liesi et al., 1983) The SVZ contains high levels of a wide range oflaminins, both within basement membranes and surrounding the vasculature(Kazanis et al., 2010) Additionally, high levels of laminin expression were found

in fractones, fingerlike projections of ECM that contact the NSCs (Kerever et al.,2007; Mercier et al., 2002) This heterogeneity of laminin expression suggests mul-tiple roles within the NSC niche This has been shown in the embryonic cortex,which has high levels of laminin expression (Drago et al., 1991; Lathia et al.,

2007) Disruption of laminin chains during development results in cortical dysplasia(Halfter et al., 2002; Radner et al., 2013; Tsuda et al., 2010), and the loss of specificlaminin chains a2 and a4 results in radial glial cell detachment, reduced cortical size,and increased apoptosis (Radakovits et al., 2009) In addition to this, mice null forseveral laminin chains, such as a1 and a5, are embryonic lethal (Alpy et al., 2005;Miner et al., 1998) This lethality of disrupting laminin expression is one of the rea-sons why the function of laminins in adult neurogenesis has not been as extensivelystudied However, the major laminin receptors, integrins, and dystroglycan havebeen studied in the adult stem cell niche and are discussed later Interestingly, thecells within the adult niche expressing the highest levels of laminin receptor integrina6b1 are those located closest to the laminin-rich vasculature, the area of neurogen-esis within the niche (Shen et al., 2008) As in the embryo, reduction of laminin con-taining fractones disrupts neurogenesis in a mouse model of autism (Mercier et al.,

2011) How laminin has this effect on adult neurogenesis is not fully understood, butwork in vitro and in the embryo suggests that laminin provides multiple signals.Adult NSCs can be maintained in vitro on laminin (Pollard et al., 2006), which

can enhance NSC proliferation (Flanagan et al., 2008; Hall et al., 2008)

Hippocam-pal NSPCs also showed increased self-renewal and reduced differentiation when

plated on laminin (Imbeault et al., 2009) More recently, the addition of a

laminin-rich ECM has been shown to be sufficient to promote the generation of

cor-tical neurons from iPS cells, forming complex 3D structures (Lancaster et al., 2013)

10 CHONDROITIN SULFATE PROTEOGLYCANS IN THE NSC

COMPARTMENT

It has already been pointed out that the chondroitin sulfate proteoglycan (CSPG)

DSD-1-PG/phosphacan is highly enriched in the adult NSC niche of the SVZ

(Gates et al., 1995) Proteoglycans consist of a core protein and at least one

cova-lently attached GAG side chain One distinguishes proteoglycan subclasses

accord-ing to the constitutaccord-ing carbohydrate polymers, namely, chondroitin sulfate, dermatan

sulfate, keratan sulfate, and heparan sulfate A large variety of proteoglycans have

been described in neural tissues, which influence the biology of NSCs by their

GAG chains, O- or N-linked carbohydrate epitopes, or their core proteins Tissue

fractionation studies performed with rat brain revealed that heparan sulfate

proteo-glycans (HSPGs) are tightly associated with cell membranes, whereas CSPGs are

mainly recovered in detergent-free salt extracts The lectican family comprises

bre-vican, neurocan, versican, and aggrecan and represents the major population of

CSPGs in the CNS (Bandtlow and Zimmermann, 2000) Versican occurs in four

iso-forms (V0–V3) and is expressed in mature oligodendrocytes (Zimmermann and

Dours-Zimmermann, 2008) CSPGs including the lecticans are secreted into the

cul-ture medium of neurospheres, the culcul-ture model of NSPCs (Ida et al., 2006; Kabos

et al., 2004) This motivated the development of strategies and methods to study the

biological roles of CSPGs in the NSC niche (Sirko et al., 2010a) Embryonic radial

glial cells similar to adult neurogenic niches express DSD-1-PG/phosphacan, a

sol-uble CSPG of the mouse CNS and homologue rat phosphacan (Faissner et al., 2006)

The core protein of DSD-1-PG/phosphacan carries chondroitin sulfate chains of the

CS-A and CS-C subtype, a keratan sulfate recognized by the MAb 3H1, and the

DSD-1 epitope The latter is recognized by the MAb 473HD and depends on a chain

length of seven disaccharides, the sulfation of the carbohydrate backbone, and the

presence of CS-D dimers (Clement et al., 1998; Ito et al., 2005) The antibody

473HD can be used to enrich for neurosphere-forming units, and the DSD-1 epitope

is enriched on radial glial surfaces and hence can be considered a novel marker of

NSCs (von Holst et al., 2006) When the DSD-1 epitope and chondroitin sulfates

al-together are removed by the enzyme chondroitinase ABC, the growth behavior of

neurospheres is modified; the proliferation rate of NSCs is reduced in vitro and

in vivo; and the differentiation of neurons is impaired, while the emergence of

astro-cytes is favored (Sirko et al., 2007; von Holst et al., 2006) In this setting, the enzyme

treatment selectively interferes with FGF2 signaling (Sirko et al., 2010b) In view of

the importance of sulfation for the definition of GAG microdomains, including the

DSD-1 epitope, the synthetic machinery of sulfation was examined The relevantsulfotransferases could be detected in radial glia, neurospheres, and the adult neuro-genic niche (Akita et al., 2008) Suppression of sulfation by chlorate results in a dose-dependent reduction of neurosphere number that cannot be rescued by the addition ofpurified GAG chains In spinal cord progenitors, the addition of chlorate results inaltered cell cycle progression and an increased rate of neurogenesis paralleled by adelay of neuronal maturation (Karus et al., 2012) Taken together, these results sug-gest that the CSPGs intervene in important signaling processes in the NSC niche,partly via complex GAG microdomains that serve as docking sites for biologicallyactive molecules (Purushothaman et al., 2012).

11 MEMBRANE-BASED PART-TIME CSPGs

The secreted and glycosylated core protein of phosphacan is a splice variant of theentire extracellular region of the largest isoform of the transmembrane receptor pro-tein tyrosine phosphatase-beta (RPTP-b/z) The large and a short RPTP-b/z receptorvariant possess a transmembrane domain and two cytoplasmic phosphotyrosinephosphatase modules The small receptor variant is devoid of the GAG attachmentregion and hence not exposing chondroitin sulfate, whereas the large receptor canoccur as a part-time proteoglycan (Faissner et al., 2006) RPTP-b/z isoforms are dif-ferentially expressed in NSPCs, glial precursor cells, radial glia, Golgi cells, and as-trocytes of different developmental stages and regions of the CNS (Lamprianou andHarroch, 2006) Within the ECM, DSD-1PG/phosphacan interacts with various li-gands, for example, the ECM glycoprotein tenascin-C (Garwood et al., 2001).The RPTP-b/z serves as a receptor of the cytokines midkine and pleiotrophin andpresumably further, yet to be identified ligands (Mohebiany et al., 2013) Beyondthe GAGs, the core proteins of RPTP-b/z are derivatized with N- or O-linked gly-cans, for example, the HNK-1 epitope or LewisX-type glycans, the latter acting

as markers for NSCs (Hennen and Faissner, 2012; Hennen et al., 2011) Themembrane-based NG2 is expressed in the oligodendrocyte lineage and in pericytes

in the CNS and expressed either as glycoprotein or as CSPG There, it has been plicated in the regulation of cell proliferation and migration (Nishiyama et al., 2009;Trotter et al., 2010)

im-12 HSPGs IN THE NSC COMPARTMENT

Heparan sulfate consists of repeating subunits ofN-acetylglucosamine and nic acid.N-acetylglucosamine and glucuronic acid can be sulfated at the C6 positionand the C2 position, respectively The spatial pattern of sulfates creates chargedmicrodomains that serve as docking sites for distinct proteins For example, specificbinding motifs have been identified for FGF2, PDGF, or antithrombin (Sarrazin

glucuro-et al., 2011) HSPGs comprise the transmembrane syndecans and the GPI-linked

glypicans HSPGs have been attributed many functions, including ligand–receptor

clustering; storage and presentation of morphogens, cytokines, and chemokines;

bar-rier and basement membrane formation; and regulation of cell adhesion and motility

(Sarrazin et al., 2011) Glypicans are enriched in the ventricular zone of the

devel-oping CNS In the adult complexes of laminin glycoproteins, nidogen, collagen IV,

and HSPGs occur as so-called fractones in the SVZ Fractones bind FGF2 and may

thereby influence the proliferation behavior of NSPCs localized in their vicinity

(Kerever et al., 2007) HSPGs are expressed in various phases of CNS development,

and roles in the regulation of progenitor proliferation and in axon growth and

guid-ance have been suggested (Maeda et al., 2011) These ideas are now being tested by

conditional knockout of the glycosyltransferase Ext1, a key enzyme of heparan

sulfate synthesis (Yamaguchi et al., 2010)

13 ECM RECEPTORS IN NSCs AND GLIAL PROGENITORS

The integrins represent an important family of heterodimeric ECM receptors and are

the major receptor for laminins within the CNS (Gardiner, 2011), although several

integrin receptors have also been described for tenascin-C, for example, Itga1b8 or

Itgavb3 (Joester and Faissner, 2001) The beta1-integrin subunit is highly enriched

in neural progenitor cells; high expression levels are seen on neurospheres and within

the VZ of the developing cortex (Campos et al., 2004; Hall et al., 2006)

Beta1-integrins contribute substantially to NSC maintenance, and disruption of integrin

func-tion at either the pial basement membrane, via genetic ablafunc-tion, or ventricular surface,

via blocking antibodies or disintegrins, promotes process detachment, apoptosis, and

altered neurogenesis (Fietz et al., 2010; Graus-Porta et al., 2001; Loulier et al., 2009;

Radakovits et al., 2009) suggesting a structural role of integrins within the niche

Recent studies show increasing evidence for a role of integrin signaling in the

regulation of embryonic NSC behavior Transcriptome analysis of the different

ger-minal zones of the developing cortex revealed high levels of ECM and integrin

ex-pression within the areas of NSC self-renewal, consistent with findings in vitro

showing loss of beta1-integrin reduced neurosphere proliferation (Fietz et al.,

2012; Leone et al., 2005) More recently, it has been reported that the activation

of the Itgavb3 stimulates the expansion of the Tbr2-positive intermediate progenitors

in the mouse telencephalon (Stenzel et al., 2014), suggesting the increased area of

integrin expression within the outer SVZ of the human brain may play a role in

cor-tical expansion (Fietz et al., 2010, 2012)

In the adult niches, integrins are also important for the regulation of neurogenesis

Beta8 integrin-null mice have multiple defects within the SVZ, including disrupted

architecture and perturbed migration of neuroblasts, increased apoptosis, and

re-duced proliferation (Mobley et al., 2009) Beta1-integrin is highly expressed within

the adult SVZ, where interestingly it was not expressed in the quiescent NSCs but

upregulated upon their activation (Kazanis et al., 2010) Blocking antibodies against

beta1-integrin increased proliferation and migration of precursors (Kazanis et al.,

2010), an effect opposite to the beta8 integrin-null mouse This is most likely due tothe different ligands of the two integrins, laminin for beta1 and TGF-b for beta8,leading to opposing downstream signals As in the embryo, there is increasing evi-dence that integrin signaling as well as adhesion plays an important role in the reg-ulation of NSCs Loss of integrin-linked kinase (ILK) from NSPCs in both the SVZand the SGZ enhanced proliferation via increased activation of JNK (Porcheri et al.,

2014) Integrin signaling can also interact with other major neurogenic pathwayssuch as Notch, Wnt, and various growth factors (Alam et al., 2007; Arora et al.,2012; Brizzi et al., 2012; Campos et al., 2006; Rallis et al., 2010), predicting a morecomplicated role of integrin signaling within the niche that is yet to be discovered.There are many other ECM receptors expressed on NSCs aside from integrins,including the laminin receptor dystroglycan During development, dystroglycan isexpressed on the end feet of radial glia and is required for basement membrane in-tegrity, regulation of proliferation, and the correct lamination of the cortex (Myshrall

et al., 2012; Satz et al., 2010; Schroder et al., 2007) In the adult, dystroglycan in glialcells stabilizes the glia limitans and promotes neuronal migration and axonal path-finding (Satz et al., 2010; Wright et al., 2012)

These ECM receptors are also expressed on the OPC stem cell population WithinOPCs, dystroglycan and beta1-integrin promote filopodial formation and processbranching (Eyermann et al., 2012), and dystroglycan can promote OPC differentia-tion via IGF-1 and myelination of mature oligodendrocytes (Colognato et al., 2007;Galvin et al., 2010) Integrins have been shown to regulate different stages of OPCdevelopment (Colognato et al., 2004, 2007) Integrin avb3 can also promote OPCproliferation (Blaschuk et al., 2000), and beta1-integrin has also been shown to have

an instructive role on OPC differentiation and myelination Expression of a dominantnegative beta1-integrin containing only a functional intracellular part but lackingligand binding extracellular domains increased the threshold axon diameter for mye-lination and reduced myelination efficiency of oligodendrocytes (Camara et al.,

2009) This reduction in myelination was thought to occur via reduced MAPKactivity (Lee et al., 2006), and further studies showed that activation of integrin sig-naling initiated the translation of MBP via the mRNA-binding protein hnRNP-K(Laursen et al., 2011) Laminins, the major ligands of beta1-integrin and dystrogly-can, are also important in glial development Laminin a2 is required for the gener-ation and maturation of OPCs in the postnatal brain (Buttery and ffrench-Constant,1999; Relucio et al., 2012), which in turn requires ILK (Chun et al., 2003) Lamininalso plays a role in oligogliogenesis, with an increase in OPCs produced when NSCswere plated onto laminin (Sypecka et al., 2009)

14 CONCLUSIONS

While the studies of ECM molecules in the CNS niches reviewed above have shownsignificant expression of tenascins, laminins, and proteoglycans, it is clear that thefunction of these molecules remains elusive This likely reflects the complex com-binatorial associations in vivo, with perturbation of single molecules therefore

having limited effects Additionally, other ECM families require investigation For

example, the gene expression of human neurogenic regions (Fietz et al., 2012)

showed high levels of collagens, and these are attractive candidates for further work

Given the importance of understanding and manipulating adult NSCs for

regenera-tive neurology, an expansion of work in this area is clearly warranted

Outstanding Questions

• What are the composition and variability of the ECM in the NSC niches?

• Does the niche ECM change with age?

• What is the stoichiometric and the spatial organization of niche ECM?

• How is the niche ECM regulated by cytokines?

• Which transcription factors regulate the niche ECM?

• How does the niche ECM respond to CNS lesions and degenerative processes?

• Which components of the niche are required for maintenance of stemness?

• Which components of the niche regulate differentiation events?

• Which receptor systems are involved in regulating the response to niche ECM?

• Which signaling mechanisms are controlled by these receptors?

• Is it possible to design artificial niches for NSCs?

ACKNOWLEDGMENTS

We acknowledge grant support by the Stem Cell Network North Rhine Westphalia, the

German Research Foundation (DFG: SPP 1109, Fa 159/16-1, GRK 736, and GSC 98/1),

the German Ministry of Education, Research and Technology (BMBF 01GN0503), and the

Ruhr University (President’s special programme call 2008) We are thankful to Dr T Czopka

for the generation oftenascin-C riboprobes and Melina Terhufen and Nina Kornblum for the

experimental procedures

REFERENCES

Akita, K., von Holst, A., Furukawa, Y., Mikami, T., Sugahara, K., Faissner, A., 2008

Expres-sion of multiple chondroitin/dermatan sulfotransferases in the neurogenic regions of the

embryonic and adult central nervous system implies that complex chondroitin sulfates

have a role in neural stem cell maintenance Stem Cells 26, 798–809

Alam, N., Goel, H.L., Zarif, M.J., Butterfield, J.E., Perkins, H.M., Sansoucy, B.G., Sawyer, T.K.,

Languino, L.R., 2007 The integrin-growth factor receptor duet J Cell Physiol

213, 649–653

Alpy, F., Jivkov, I., Sorokin, L., Klein, A., Arnold, C., Huss, Y., Kedinger, M.,

Simon-Assmann, P., Lefebvre, O., 2005 Generation of a conditionally null allele of the laminin

alpha1 gene Genesis 43, 59–70

Anthony, T.E., Klein, C., Fishell, G., Heintz, N., 2004 Radial glia serve as neuronal

progen-itors in all regions of the central nervous system Neuron 41, 881–890

Arora, N., Mainali, D., Smith, E.A., 2012 Unraveling the role of membrane proteins Notch,

Pvr, and EGFR in altering integrin diffusion and clustering Anal Bioanal Chem

404, 2339–2348

Aumailley, M., Bruckner-Tuderman, L., Carter, W.G., Deutzmann, R., Edgar, D., Ekblom, P.,Engel, J., Engvall, E., Hohenester, E., Jones, J.C., Kleinman, H.K., Marinkovich, M.P.,Martin, G.R., Mayer, U., Meneguzzi, G., Miner, J.H., Miyazaki, K., Patarroyo, M.,Paulsson, M., Quaranta, V., Sanes, J.R., Sasaki, T., Sekiguchi, K., Sorokin, L.M., Talts, J.F.,Tryggvason, K., Uitto, J., Virtanen, I., von der Mark, K., Wewer, U.M., Yamada, Y.,Yurchenco, P.D., 2005 A simplified laminin nomenclature Matrix Biol 24, 326–332.Bandtlow, C.E., Zimmermann, D.R., 2000 Proteoglycans in the developing brain: new con-ceptual insights for old proteins Physiol Rev 80, 1267–1290.

Barros, C.S., Franco, S.J., Muller, U., 2011 Extracellular matrix: functions in the nervous tem Cold Spring Harb Persp Biol 3, a005108

sys-Baumgartner, S., Martin, D., Hagios, C., Chiquet-Ehrismann, R., 1994 Tenm, a Drosophilagene related to tenascin, is a new pair-rule gene EMBO J 13, 3728–3740

Blaschuk, K.L., Frost, E.E., ffrench-Constant, C., 2000 The regulation of proliferation anddifferentiation in oligodendrocyte progenitor cells by alphaV integrins Development

J Cell Biol 185, 699–712

Campos, L.S., Leone, D.P., Relvas, J.B., Brakebusch, C., Fassler, R., Suter, U., Constant, C., 2004 Beta1 integrins activate a MAPK signalling pathway in neural stemcells that contributes to their maintenance Development 131, 3433–3444

ffrench-Campos, L.S., Decker, L., Taylor, V., Skarnes, W., 2006 Notch, epidermal growth factor ceptor, and beta1-integrin pathways are coordinated in neural stem cells J Biol Chem

Clement, A.M., Nadanaka, S., Masayama, K., Mandl, C., Sugahara, K., Faissner, A., 1998 TheDSD-1 carbohydrate epitope depends on sulfation, correlates with chondroitin sulfate

D motifs, and is sufficient to promote neurite outgrowth J Biol Chem

273, 28444–28453

Colognato, H., Ramachandrappa, S., Olsen, I.M., ffrench-Constant, C., 2004 Integrins directSrc family kinases to regulate distinct phases of oligodendrocyte development J CellBiol 167, 365–375

Colognato, H., Galvin, J., Wang, Z., Relucio, J., Nguyen, T., Harrison, D., Yurchenco, P.D.,ffrench-Constant, C., 2007 Identification of dystroglycan as a second laminin receptor inoligodendrocytes, with a role in myelination Development 134, 1723–1736

Copertino, D.W., Jenkinson, S., Jones, F.S., Edelman, G.M., 1995 Structural and functional

similarities between the promoters for mouse tenascin and chicken cytotactin Proc Natl

Acad Sci U S A 92, 2131–2135

Copertino, D.W., Edelman, G.M., Jones, F.S., 1997 Multiple promoter elements differentially

regulate the expression of the mouse tenascin gene Proc Natl Acad Sci U S A

94, 1846–1851

Couturier, L., Mazouni, K., Schweisguth, F., 2013 Numb localizes at endosomes and controls

the endosomal sorting of notch after asymmetric division in Drosophila Curr Biol

23, 588–593

Craig, A.M., Kang, Y., 2007 Neurexin-neuroligin signaling in synapse development Curr

Opin Neurobiol 17, 43–52

Czopka, T., Von Holst, A., Schmidt, G., ffrench-Constant, C., Faissner, A., 2009 Tenascin

C and tenascin R similarly prevent the formation of myelin membranes in a

RhoA-dependent manner, but antagonistically regulate the expression of myelin basic protein

via a separate pathway Glia 57, 1790–1801

Czopka, T., von Holst, A., ffrench-Constant, C., Faissner, A., 2010 Regulatory mechanisms

that mediate tenascin C-dependent inhibition of oligodendrocyte precursor differentiation

J Neurosci 30, 12310–12322

Dityatev, A., Seidenbecher, C.I., Schachner, M., 2010 Compartmentalization from the

out-side: the extracellular matrix and functional microdomains in the brain Trends Neurosci

33, 503–512

Dobbertin, A., Czvitkovich, S., Theocharidis, U., Garwood, J., Andrews, M.R., Properzi, F.,

Lin, R., Fawcett, J.W., Faissner, A., 2010 Analysis of combinatorial variability reveals

selective accumulation of the fibronectin type III domains B and D of tenascin-C in injured

brain Exp Neurol 225, 60–73

Doetsch, F., Petreanu, L., Caille, I., Garcia-Verdugo, J.M., Alvarez-Buylla, A., 2002 EGF

converts transit-amplifying neurogenic precursors in the adult brain into multipotent stem

cells Neuron 36, 1021–1034

Drago, J., Nurcombe, V., Bartlett, P.F., 1991 Laminin through its long arm E8 fragment

pro-motes the proliferation and differentiation of murine neuroepithelial cells in vitro Exp

Cell Res 192, 256–265

Eyermann, C., Czaplinski, K., Colognato, H., 2012 Dystroglycan promotes filopodial formation

and process branching in differentiating oligodendroglia J Neurochem 120, 928–947

Faissner, A., 1997 Glial derived extracellular matrix components: important roles in axon

growth and guidance Neuroscientist 3, 371–380

Faissner, A., Kruse, J., 1990 J1/tenascin is a repulsive substrate for central nervous system

neurons Neuron 5, 627–637

Faissner, A., Steindler, D., 1995 Boundaries and inhibitory molecules in developing neural

tissues Glia 13, 233–254

Faissner, A., Heck, N., Dobbertin, A., Garwood, J., 2006 DSD-1-proteoglycan/phosphacan

and receptor protein tyrosine phosphatase-beta isoforms during development and

regen-eration of neural tissues Adv Exp Med Biol 557, 25–53

Ffrench-Constant, C., Raff, M.C., 1986 Proliferating bipotential glial progenitor cells in adult

rat optic nerve Nature 319, 499–502

Fietz, S.A., Huttner, W.B., 2011 Cortical progenitor expansion, self-renewal and

neurogenesis-a polarized perspective Curr Opin Neurobiol 21, 23–35

Fietz, S.A., Kelava, I., Vogt, J., Wilsch-Brauninger, M., Stenzel, D., Fish, J.L., Corbeil, D.,Riehn, A., Distler, W., Nitsch, R., Huttner, W.B., 2010 OSVZ progenitors of humanand ferret neocortex are epithelial-like and expand by integrin signaling Nat Neurosci.

13, 690–699

Fietz, S.A., Lachmann, R., Brandl, H., Kircher, M., Samusik, N., Schroder, R.,Lakshmanaperumal, N., Henry, I., Vogt, J., Riehn, A., Distler, W., Nitsch, R.,Enard, W., Paabo, S., Huttner, W.B., 2012 Transcriptomes of germinal zones of humanand mouse fetal neocortex suggest a role of extracellular matrix in progenitor self-renewal.Proc Natl Acad Sci U S A 109, 11836–11841

Flanagan, L.A., Lu, J., Wang, L., Marchenko, S.A., Jeon, N.L., Lee, A.P., Monuki, E.S., 2008.Unique dielectric properties distinguish stem cells and their differentiated progeny StemCells 26, 656–665

Freeman, M.R., 2010 Specification and morphogenesis of astrocytes Science 330, 774–778.Galvin, J., Eyermann, C., Colognato, H., 2010 Dystroglycan modulates the ability of insulin-like growth factor-1 to promote oligodendrocyte differentiation J Neurosci Res

88, 3295–3307

Garcion, E., Faissner, A., ffrench-Constant, C., 2001 Knockout mice reveal a contribution ofthe extracellular matrix molecule tenascin-C to neural precursor proliferation and migra-tion Development 128, 2485–2496

Garcion, E., Halilagic, A., Faissner, A., ffrench-Constant, C., 2004 Generation of an mental niche for neural stem cell development by the extracellular matrix molecule tenas-cin C Development 131, 3423–3432

environ-Gardiner, N.J., 2011 Integrins and the extracellular matrix: key mediators of development andregeneration of the sensory nervous system Dev Neurobiol 71, 1054–1072

Garthe, A., Kempermann, G., 2013 An old test for new neurons: refining the Morris watermaze to study the functional relevance of adult hippocampal neurogenesis Front Neu-rosci 7, 63

Garwood, J., Rigato, F., Heck, N., Faissner, A., 2001 Tenascin glycoproteins and the mentary ligand DSD-1-PG/phosphacan-structuring the neural extracellular matrix duringdevelopment and repair Restor Neurol Neurosci 19, 51–64

comple-Garwood, J., Garcion, E., Dobbertin, A., Heck, N., Calco, V., ffrench-Constant, C.,Faissner, A., 2004 The extracellular matrix glycoprotein tenascin-C is expressed byoligodendrocyte precursor cells and required for the regulation of maturation rate,survival and responsiveness to platelet-derived growth factor Eur J Neurosci

20, 2524–2540

Garwood, J., Theocharidis, U., Calco, V., Dobbertin, A., Faissner, A., 2012 Existence oftenascin-C isoforms in rat that contain the alternatively spliced AD1 domain are develop-mentally regulated during hippocampal development Cell Mol Neurobiol 32, 279–287.Gates, M.A., Thomas, L.B., Howard, E.M., Laywell, E.D., Sajin, B., Faissner, A., Gotz, B.,Silver, J., Steindler, D.A., 1995 Cell and molecular analysis of the developing and adultmouse subventricular zone of the cerebral hemispheres J Comp Neurol 361, 249–266.Gherzi, R., Briata, P., Boncinelli, E., Ponassi, M., Querze, G., Viti, F., Corte, G., Zardi, L.,

1997 The human homeodomain protein OTX2 binds to the human tenascin-C promoterand trans-represses its activity in transfected cells DNA Cell Biol 16, 559–567.Gotz, M., Huttner, W.B., 2005 The cell biology of neurogenesis Nat Rev Mol Cell Biol

6, 777–788

Gotz, M., Stoykova, A., Gruss, P., 1998 Pax6 controls radial glia differentiation in the cerebralcortex Neuron 21, 1031–1044

Graus-Porta, D., Blaess, S., Senften, M., Littlewood-Evans, A., Damsky, C., Huang, Z.,

Orban, P., Klein, R., Schittny, J.C., Muller, U., 2001 Beta1-class integrins regulate the

development of laminae and folia in the cerebral and cerebellar cortex Neuron

31, 367–379

Halfter, W., Dong, S., Yip, Y.P., Willem, M., Mayer, U., 2002 A critical function of the pial

basement membrane in cortical histogenesis J Neurosci 22, 6029–6040

Hall, P.E., Lathia, J.D., Miller, N.G., Caldwell, M.A., ffrench-Constant, C., 2006 Integrins are

markers of human neural stem cells Stem Cells 24, 2078–2084

Hall, P.E., Lathia, J.D., Caldwell, M.A., ffrench-Constant, C., 2008 Laminin enhances the

growth of human neural stem cells in defined culture media BMC Neurosci 9, 71

Hansen, D.V., Lui, J.H., Parker, P.R., Kriegstein, A.R., 2010 Neurogenic radial glia in the

outer subventricular zone of human neocortex Nature 464, 554–561

Hartfuss, E., Galli, R., Heins, N., Gotz, M., 2001 Characterization of CNS precursor subtypes

and radial glia Dev Biol 229, 15–30

Hennen, E., Faissner, A., 2012 LewisX: a neural stem cell specific glycan? Int J Biochem

Cell Biol 44, 830–833

Hennen, E., Czopka, T., Faissner, A., 2011 Structurally distinct LewisX glycans distinguish

subpopulations of neural stem/progenitor cells J Biol Chem 286, 16321–16331

Holm, P.C., Mader, M.T., Haubst, N., Wizenmann, A., Sigvardsson, M., Gotz, M., 2007

Loss-and gain-of-function analyses reveal targets of Pax6 in the developing mouse

telenceph-alon Mol Cell Neurosci 34, 99–119

Husmann, K., Faissner, A., Schachner, M., 1992 Tenascin promotes cerebellar granule cell

migration and neurite outgrowth by different domains in the fibronectin type III repeats

J Cell Biol 116, 1475–1486

Ida, M., Shuo, T., Hirano, K., Tokita, Y., Nakanishi, K., Matsui, F., Aono, S., Fujita, H.,

Fujiwara, Y., Kaji, T., Oohira, A., 2006 Identification and functions of chondroitin sulfate

in the milieu of neural stem cells J Biol Chem 281, 5982–5991

Ihrie, R.A., Alvarez-Buylla, A., 2011 Lake-front property: a unique germinal niche by the

lateral ventricles of the adult brain Neuron 70, 674–686

Imbeault, S., Gauvin, L.G., Toeg, H.D., Pettit, A., Sorbara, C.D., Migahed, L., DesRoches, R.,

Menzies, A.S., Nishii, K., Paul, D.L., Simon, A.M., Bennett, S.A., 2009 The extracellular

matrix controls gap junction protein expression and function in postnatal hippocampal

neural progenitor cells BMC Neurosci 10, 13

Ito, Y., Hikino, M., Yajima, Y., Mikami, T., Sirko, S., von Holst, A., Faissner, A., Fukui, S.,

Sugahara, K., 2005 Structural characterization of the epitopes of the monoclonal

anti-bodies 473HD, CS-56, and MO-225 specific for chondroitin sulfate D-type using the

oligosaccharide library Glycobiology 15, 593–603

Jankovski, A., Sotelo, C., 1996 Subventricular zone-olfactory bulb migratory pathway in the

adult mouse: cellular composition and specificity as determined by heterochronic and

het-erotopic transplantation J Comp Neurol 371, 376–396

Joester, A., Faissner, A., 1999 Evidence for combinatorial variability of tenascin-C isoforms

and developmental regulation in the mouse central nervous system J Biol Chem

274, 17144–17151

Joester, A., Faissner, A., 2001 The structure and function of tenascins in the nervous system

Matrix Biol 20, 13–22

Jones, F.S., Jones, P.L., 2000 The tenascin family of ECM glycoproteins: structure, function,

and regulation during embryonic development and tissue remodeling Dev Dyn

218, 235–259