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Journal Club Theme of July 1 2008: Mechanics in Neuronal Development

The biological world is part of the physical world around us and obeys its laws. In particular, physical interactions can be as important in determining tissue or cell fate as biochemical stimuli, and they may also contribute to pathological conditions. The application of cell biomechanics contributes to an understanding of many processes that take place in our body such as cell movement, cell division, phagocytosis, and cellular contractility. Therefore, biomechanical investigations contribute to our understanding of the normal functioning of living organisms, help to predict changes which arise due to alterations of their environment, and maybe also to propose methods of artificial intervention.

Most biomechanical studies have so far focused on systems where mechanics obviously plays an important role, such as the locomotor system, the cardiovascular system, and the lung. However, even if not that obvious, biomechanical aspects may also play an important role in the central nervous system (CNS).

Our nervous system consists of two basic cell types, neurons and glial cells. Neurons, which transmit and process information, extend cell processes, typically several dendrites and one axon. The cues determining which neuronal process becomes the axon have long been unclear. In a groundbreaking study from almost 25 years ago Dennis Bray showed that axonal growth can be initiated de novo by the application of mechanical tension, a process he termed “towed growth”:

Bray, D. (1984). "Axonal growth in response to experimentally applied mechanical tension." Dev Biol 102(2): 379-89.

How tension may influence tissue formation in the CNS is described in the second paper:

Van Essen, D. C. (1997). A tension-based theory of morphogenesis and compact wiring in the central nervous system. Nature 385:313-318.

In the first part of this very interesting article the author briefly reviews experimental data on passive and active mechanical properties of neurons, mainly found in the lab of Steve Heidemann, and subsequently develops a theory that explains the morphogenesis of the CNS, e.g., the specific folding of the brain or the development of the retinal fovea, by tension and hydrostatic pressure.

The third paper in this Journal Club presents another key study in the field of neuromechanics. The group of Paul Janmey could show that not only active tension but also passive material properties of the cells’ environment may influence their growth and function:

Georges, P. C., W. J. Miller, D. F. Meaney, E. S. Sawyer, and P. A. Janmey. (2006). Matrices with compliance comparable to that of brain tissue select neuronal over glial growth in mixed cortical cultures. Biophys J 90:3012-3018.

The authors show that neurons are capable of sensing the compliance of their environment in vitro and prefer relatively compliant substrates. Interestingly, glial cells in vitro grow better on stiffer materials. Thus, the cells in the CNS seem to be able to feel and to respond to the mechanical properties of their environment.
These data point towards an important contribution of mechanics to the development of the CNS and also to certain pathological processes.

Future research promises to reveal increasingly interesting facts about the influence of mechanics on CNS physiology and pathology, and it seems very likely that this growing knowledge may be exploited for example in the design of new neural implants and in the treatment of injuries to the fragile nervous tissue.

Mike Ciavarella's picture

 
 

 

Nano-opto-mechanical characterization of neuron membrane mechanics under cellular growth and differentiation

Ashwini Gopal1, Zhiquan Luo2, Jae Young Lee3, Karthik Kumar1, Bin Li2, Kazunori Hoshino1, Christine Schmidt3, Paul S. Ho2 and Xiaojing Zhang1 Contact Information

(1) 
Department
of Biomedical Engineering, The University of Texas at Austin, 1
University Station, ENS 12, Austin, TX 78712-0238, USA

(2) 
Department of Mechanical Engineering, University of Texas at Austin, Austin, TX 78758, USA

(3) 
Department of Chemical Engineering, University of Texas at Austin, Austin, TX 78712, USA

Published online: 16 May 2008

Abstract  We
designed and fabricated silicon probe with nanophotonic force sensor to
directly stimulate neurons (PC12) and measured its effect on neurite
initiation and elongation. A single-layer pitch-variable diffractive
nanogratings was fabricated on silicon nitride probe using e-beam
lithography, reactive ion etching and wet-etching techniques. The
nanogratings consist of flexure folding beams suspended between two
parallel cantilevers of known stiffness. The probe displacement,
therefore the force, can be measured through grating transmission
spectrum. We measured the mechanical membrane characteristics of PC12
cells using the force sensors with displacement range of 10 μm and
force sensitivity 8 μN/μm. Young’s moduli of 425 ± 30 Pa are measured
with membrane deflection of 1% for PC12 cells cultured on
polydimethylsiloxane (PDMS) substrate coated with collagen or laminin
in Ham’s F-12K medium. In a series of measurements, we have also
observed stimulation of directed neurite contraction up to 6 μm on
extended probing for a time period of 30 min. This method is applicable
to measure central neurons mechanics under subtle tensions for studies
on development and morphogenesis. The close synergy between the
nano-photonic measurements and neurological verification can improve
our understanding of the effect of external conditions on the
mechanical properties of cells during growth and differentiation.

Keywords  Mechanotransduction - Cytomechanics - PC12 - Cell
membrane - Growth - Differentiation - Nanogratings - Micro-electro-mechanical
systems (MEMS) - Force sensor

Contact Information
Xiaojing Zhang
Email: John.Zhang@engr.utexas.edu

Mike Ciavarella's picture

Cellular control lies in the balance of forces - all 8 versions »
ME Chicurel, CS Chen, DE Ingber - Current Opinion in Cell Biology, 1998 - Elsevier
Addresses Departments of Surgery and Pathology, Children's Hospital and Harvard
Medical School, Enders 1007, 300 Longwood Avenue, Boston, MA 02115, USA *e-mail:
ingber@al.tch.harvard.edu Current Opinion in Cell Biology 1998, 10:232-239 ...
Cited by 261 - Related Articles - Web Search - BL Direct

 

The structural and mechanical complexity of cell-growth control - »
S Huang, DE Ingber - Nature Cell Biology, 1999 - nature.com
Over the past decade, enormous advances have been made in our understanding of
the molecules that mediate the control of cell proliferation. Soluble mitogens,
insoluble extracellular matrix (ECM) molecules, cell-surface growth-factor ...
Cited by 238 - Related Articles - Web Search - BL Direct

 

Tensegrity II. How structural networks influence cellular information processing networks
DE Ingber - Journal of Cell Science, 2003 - jcs.biologists.org
The major challenge in biology today is biocomplexity: the need to explain how
cell and tissue behaviors emerge from collective interactions within complex
molecular networks. Part I of this two-part article, described a mechanical ...
Cited by 198 - Related Articles - Web Search - BL Direct

Tensegrity I. Cell structure and hierarchical systems biology - all 3 versions »
DE Ingber - Journal of Cell Science, 2003 - jcs.biologists.org
In 1993, a Commentary in this journal described how a simple mechanical model of
cell structure based on tensegrity architecture can help to explain how cell
shape, movement and cytoskeletal mechanics are controlled, as well as how ...
Cited by 174 - Related Articles - Web Search - BL Direct

Cell surface area regulation and membrane tension.
CE Morris, U Homann - J Membr Biol, 2001 - ncbi.nlm.nih.gov
The beautifully orchestrated regulation of cell shape and volume are central
themes in cell biology and physiology. Though it is less well recognized, cell
surface area regulation also constitutes a distinct task for cells. ...
Cited by 118 - Related Articles - Web Search - BL Direct

Forces on adhesive contacts affect cell function - all 4 versions »
CG Galbraith, MP Sheetz - Current Opinion in Cell Biology, 1998 - Elsevier
Cellular forces acting on the adhesive contacts made with the extracellular
matrix (ECM) contribute significantly to cell shape, viability, signal
transduction and motility. In the past two years, research has determined ...
Cited by 116 - Related Articles - Web Search - BL Direct

 

 

 

 

 

MichelleLOyen's picture

From the discussions I've heard at recent conferences, the tensegrity idea for cells is fairly controversial.

MichelleLOyen's picture

Kristian,

do you think this question of 2D vs 3D experimental conditions for cell-matrix interaction experiments is potentially important in the CNS context?

It's definitely important.

There are a couple of excellent studies on the behaviour of cells on 2D substrates of different compliance. Additionally to the one mentioned above I want to point out here another paper from the group of Paul Janmey by Flanagan et al. (2002). In this paper it is shown that neurons grow better and extend more branches on compliant substrates.

Complementary, there is a paper from Ravi Bellamkonda's lab in which they have investigated the behaviour of neurons in 3D gels. In Balgude et al. (2001) it is shown that neurite extension rates decrease with increasing matrix stiffness. However, this effect might also be due to a decrease in meshsize of the gels with increasing stiffness.

I think currently there is a need of investigations in both 2D and 3D. The comparison of these studies might allow deeper insights into cellular mechanisms including mechanosensitivity and -responsiveness.

Ideally, studies would be conducted in 3D matrices with properties similar to those found in vivo. But I guess there is still a long road to travel...

 

The Gopal et al. paper is very interesting. There is another paper by Lu YB et al. (2006) in which the mechanical properties of acutely isolated neurons and glial cells from hippocampus and retina are assessed by atomic force microscopy. The values in this paper are very similar to those from Gopal and colleagues.

Migration of neurons along compliant radial glial cells 

Interestingly, glial cells, which grow better on stiffer subtrates, are very compliant, while neurons, which prefer compliant substrates, are considerably stiffer. There are interesting data about the growth and migration patterns of neurons in the developing CNS. Growing axons and migrating neurons in the premature cortex are both known to follow radial glial cells. The compliance of those glial cells has been suggested in this paper to constitute a (mechanical) guidance cue for neurons.

 

Mike Ciavarella's picture

with an elasticity problem, just define the problem, and we can make this experiment of finding a solution collaboratively.

The ultimate goal of cell mechanics is probably to measure the mechanical properties of individual cells (or even parts of them) in situ.  This measurement is currently (to my knowledge) not possible. And not to make it too easy: I'd be mostly interested in cells of the CNS, which is surrounded by pretty stiff "envelops" (the brain by the skull, the spinal cord by vertebrae, and the retina by the sclera). If you'd find a way how to measure these cells, that'd be awesome. Is this enough of a challenge?Smile

Mike Ciavarella's picture

I agree that measurement is the ultimate need, and you seem not the only one around to think it. However, I am not expert.  I can only do a search for you.

Maybe some progress exists however:check these, including a patent!  Let me know if these were trivial results:

 

Correlation between heart valve interstitial cell stiffness and ...

Ultimately, it would be most appropriate to measure cellular stiffness in situ; however, mechanical testing of intact cells is much more complex, ...

Tensile Properties and Local Stiffness of Cells

cell stiffness was significantly higher in the cells of contractile phenotype than ... on the AFM measurement. of local stiffness of living cells in situ. ...

i1ntegrating cell's stiffness. measurements. and. penetration. The stiffness ..... [28] M. Nakajima, F. Arai, and T. Fukuda, "In-situ measurement ofYoung's ... 

Method and system for measurement of mechanical properties of ...

Diagnosis of In Situ Changes in Cell Structure and Function ..... Taxol (15 μM) increased cell stiffness by 80% in cancer cells. ...

Science Links Japan | Tensile Properties of Cultured Aortic Smooth ...

Abstract;We established a quasi-in situ tensile test to measure the tensile properties ... that cell stiffness is overestimated when cells are trypsinized. ...

Fluorescence-Imaged Microdeformation of the Outer Hair Cell ...

Key words: cochlea; inner ear; hearing; cytoskeleton; cell stiffness; .... the respective staining protocols were performed in situ (described below). ...

Veeco's BioScope II AFM Aids Cell Biology Researchers at ...

13 mar 2007 ... a cell's stiffness and the stiffness of the matrix it sits on, ... of the BioScope II enables novel in-situ techniques for measuring ...

MRS Website : Meeting Scene - Day 1

Subra Suresh, currently Professor and soon to be Dean of Engineering at MIT (USA), presented a talk on his research while epitomizes the present conference ...

The biomechanics toolbox: experimental approaches for living cells ..


Measure ligand-receptor unbinding forces [18,19]. spectroscopy. Microneedle. MN. Qualitative cell stiffness during migration [20,21]. Optical tweezers ...

2007 APS March Meeting
For both cell lines, we probe cell stiffness measured by magnetic ...... Germany; Massachusetts Institute of Technology, USA, A. MICOULET, S. SURESH, MIT, ...

 

Mike Ciavarella's picture

Challenging if you want more interest and help from imechanica ...

I'm sorry about being abroad and having no internet access for the last couple of days. That was unfortunately not to avoid.

Thanks for the papers, some of them are quite interesting, even if I don't see a solution yet for the (experimental) problem I had mentioned before.

Another problem, which I'd like to come up with, is the extraction of mechanical material properties of let's say an isotropic material that is in close connection to another isotropic material of comparable compliance. (Ultimately, I'd really like to know how to extract mechanical cell properties from cells that are cultured on compliant materials and probed with an AFM...)

Is this a more appropriate challenge? 

Mike Ciavarella's picture

 

Kristian

the general area now you are touching is "material identification" and  "inverse problems" --the literature is so vast on this that I do not even attempt a review. Sorry!  However, I don't remember where I read this, but somewhere Drucker, one of the founders of plasticity, said that there will never be enough mathematical functions to describe material behaviour.  I don't know the exact quote, but the meaning is clear.  Material behaviour is very rich, already with plasticity of metals, let alone with cells as you are doing!  So indeed, very challenging, good luck.

 Mike

Dear Mike,

I was afraid you'd say something like this... If you have any idea how to approach this problem I'd be very greatful.

Thanks for your help.

Kristian 

Mike Ciavarella's picture

Maybe you should ask Michelle Oyen --- she is your Editor and she is in Cambridge. I remember she sent me very nice papers on viscoelasticity using indentation.

If you have a substrate, you simply have different material properties. But I am sure you can apply the same principles, only the parameters to identify increase.  So you need more data to start from.

To be able to recognize "independent" measurements, you need some test so they are not correlated.   Give me more info but it is not my field!  Also, you set 2 lines and you expect me to write a paper?  We should set this on a equal basis, so I also set you a problem and we work together?  ;)

Thanks for your help. I never said I expect you to write a paper...

I'm already in contact with Michelle. Let's see what we can do together... 

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