Neurites of PC12 and chick dorsal root ganglion neurons behave as viscoelastic solids in response to applied forces. This passive behavior can be modeled with three mechanical elements; a relatively stiff, undamped spring in series with a Voight element composed of a less stiff spring in parallel with a dashpot. In response to applied tensions greater than 100 microdynes, PC12 cells show lengthening behavior distinct from and in addition to the passive viscoelastic response. We interpret this as "towed growth" (Bray, D. 1984. Dev. Biol. 102:379-389) because the neurites can become twice as long without obvious thinning of the neurite and because in two cases neurite tensions fell below original rest tensions, a result that cannot be obtained with passive viscoelastic elements. The rate of towed growth showed a linear dependence of growth rate with applied tensions in 8 of 12 PC12 neurites exposed to applied tension greater than 100 microdynes. Both PC12 and chick sensory neurons showed evidence of retraction when neurite tensions were suddenly diminished. This response was measured as tension recovery after slackening in chick sensory neurites. In 62% of the cases, tension recovery exceeded and sometimes doubled the preexperimental steady-state tension. Our data indicate that this response is active tension generation by the neurite shaft. We conclude that neurite length is regulated by axial tension in both elongation and retraction. Our data suggest a three-way controller: above some tension set point, the neurite is stimulated to elongate. Below some different, lower tension threshold the neurite is stimulated to retract. Between these two tension thresholds, the neurite responds passively as a viscoelastic solid.

Neurites of chick sensory neurons in culture were attached by their growth cones to glass needles of known compliance and were subjected to increasing tensions as steps of constant force; each step lasted 30-60 min and was 25-50 mu dyn greater than the previous step. After correcting for elastic stretching, neurite elongation rate increased in proportion to tension magnitude greater than a tension threshold. The value of the tension threshold required for growth varied between 25 and 560 mu dyn, with most between 50 and 150 mu dyn. The growth sensitivity of neurites to tension was surprisingly high: an increase in tension of 1 mu dyn increased the elongation rate an average of about 1.5 microns/hr. The linear relationship between growth rate and tension provides a simple control mechanism for axons to accommodate tissue expansion in growing animals that consistently maintains a moderate rest tension on axons. Styrene microspheres treated with polyethyleneimine were used to label the surface of neurites in order to determine the site and pattern of surface addition during the experimental "towed growth" regime. New membrane is added interstitially throughout the neurite, but different regions of neurite vary widely in the amount of new membrane added. This contrasts with membrane addition specifically at the distal end in growth-cone-mediated growth. The different sites for membrane addition in growth mediated by towing and by the growth cone indicate that the membrane addition process is sensitive to the mode of growth. We confirmed the finding of Bray (1984) that neurites can be initiated de novo by application of tension to the cell margin of chick sensory neurons.(ABSTRACT TRUNCATED AT 250 WORDS)

Whether the axonal framework is stationary or moves is a central debate in cell biology. To better understand this problem, we developed a mathematical model that incorporates force generation at the growth cone, the viscoelastic properties of the axon, and adhesions between the axon and substrate. Using force-calibrated needles to apply and measure forces at the growth cone, we used docked mitochondria as markers to monitor movement of the axonal framework. We found coherent axonal transport that decreased away from the growth cone. Based on the velocity profiles of movement and the force applied at the growth cone, and by varying the attachment of the axonal shaft to the coverslip, we estimate values for the axial viscosity of the axon (3 x 10(6) +/- 2.4 x 10(6) Pa.s) and the friction coefficient for laminin/polyornithine-based adhesions along the axon (9.6 x 10(3) +/- 7.5 x 10(3) Pa.s). Our model suggests that whether axons elongate by tip growth or stretching depends on the level of force generation at the growth cone, the viscosity of the axon, and the level of adhesions along the axon.

Here we asked whether applied mechanical tension would stimulate undifferentiated minor processes of cultured hippocampal neurons to become axons and whether tension could induce a second axon in an already polarized neuron. Experimental tension applied to minor processes produced extensions that demonstrated axonal character, regardless of the presence of an existing axon. Towed neurites showed a high rate of spontaneous growth cone advance and could continue to grow out for 1–3 d after towing. The developmental course of experimental neurites was found to be similar to that of unmanipulated spontaneous axons. Furthermore, the experimentally elongated neurites showed compartmentation of the axonal markers dephospho-tau and L-1 in towed outgrowth after 24 h. Extension of a second axon from an already polarized neuron does not lead to the loss of the spontaneous axon either immediately or after longer term growth. In addition, we were able to initiate neurites de novo that subsequently acquired axonal character even though spontaneous growth cone advance began while the towed neurite was still no longer than its sibling processes. This suggests that tension rather than the achievement of a critical neurite length determined axonal specification.

Mechanical tension is a particularly effective stimulus for axonal elongation, but little is known about how it leads to the formation of new axon. To better understand this process, we examined the movement of axonal branch points, beads bound to the axon, and docked mitochondria while monitoring axonal width. We found these markers moved in a pattern that suggests elongation occurs by viscoelastic stretching and volume addition along the axon. To test the coupling between "lengthening" and "growth," we measured axonal width while forcing axons to grow and then pause by controlling the tension applied to the growth cone or to the cell body. We found axons thinned during high rates of elongation and thickened when the growth cones were stationary. These findings suggest that forces cause lengthening because they stretch the axon and that growth occurs, in a loosely coupled step, by volume addition along the axon.

Important new theories in science often ignite heated debates. If they do not, they are probably of little significance. Thus a strong argument in support of the importance of the tensegrity model of cell and tissue architecture first proposed almost 20 years ago (23, 24) is the large number of public and private criticisms that have been mounted against this theory. Demonstration of the ability of the tensegrity model to explain complex mechanical behaviors in viruses, nuclei, cells, tissues, and organs in animals as well as in insects and plants (reviewed in Refs. 4, 5, 7, 10, 17, 20-24, 26, 30, 32, 42) has led to a drastic reduction in the number of these confrontations. Nevertheless, some intransigent critics remain. However, their remaining objections are limited in scope and largely result, I believe, from an overly concrete definition of what tensegrity is and how it can be applied. My purpose here, at the request of the Editor and Associate Editors of this journal, is to present the argument in support of the tensegrity model and to respond to some of these remaining concerns. The tensegrity model states that cells, tissues, and other biological structures at smaller and larger size scales in the hierarchy of life gain their shape stability and their ability to exhibit integrated mechanical behavior through use of the structural principles of tensegrity architecture (5, 20, 22-24). The term, "tensegrity" (contraction of "tensional integrity") was first created by the architect R. Buckminster Fuller, who first explored use of this form of structural stabilization as early as 1927 in his plan for the Wichita Dymaxion house, which minimized weight by separating compression members from tension members (31). To create this cylindrical building, Fuller proposed to set a central mast in the earth as a vertical compression strut and to suspend from it multiple circular floors (horizontal wheels) using tension cables. Tensile guy wires that linked the mast to surrounding anchors in the ground provided the balancing tension necessary to stabilize the entire structure. "Fuller called this special discontinuous-compression, continuous-tension

During development, neurons send out axonal processes that can reach lengths hundreds of times longer than the diameter of their cell bodies. Recent studies indicate that en masse microtubule translocation is a significant mechanism underlying axonal elongation, but how cellular forces drive this process is unknown. Cytoplasmic dynein generates forces on microtubules in axons to power their movement through 'stop-and-go' transport, but whether these forces influence the bulk translocation of long microtubules embedded in the cytoskeletal meshwork has not been tested. Here, we use both function-blocking antibodies targeted to the dynein intermediate chain and the pharmacological dynein inhibitor ciliobrevin D to ask whether dynein forces contribute to en bloc cytoskeleton translocation. By tracking docked mitochondria as fiducial markers for bulk cytoskeleton movements, we find that translocation is reduced after dynein disruption. We then directly measure net force generation after dynein disruption and find a dramatic increase in axonal tension. Taken together, these data indicate that dynein generates forces that push the cytoskeletal meshwork forward en masse during axonal elongation.

Abstract. The growth cone must push its substrate rearward via some traction force in order to propel itself forward. To determine which growth cone behaviors produce traction force, we observed chick sensory growth cones under conditions in which force production was accommodated by movement of obstacles in the environment, namely, neurites of other sensory neurons or glass fibers. The movements of these obstacles occurred via three, different, stereotyped growth cone behaviors: (a) filopodial contractions, (b) smooth rearward movement on the dorsal surface of the growth cone, and (c) interactions with ruffling lamellipodia. More than 70% of the obstacle movements were caused by filopodial contractions in which the obstacle attached at the extreme distal end of a filopodium and moved only as the filopodium changed its extension. Filopodial contractions were characterized by frequent changes of obstacle velocity and direction. Contraction of a single filopodium is estimated to exert 50-90/~dyn of force, which can account for the pull exerted by chick sensory growth cones. Importantly, all five cases of growth cones growing over the top of obstacle neurites (i.e., geometry that mimics the usual growth cone/substrate interaction), were of the filopodial contraction type. Some 25 % of obstacle movements occurred by a smooth backward movement along the top surface of growth cones. Both the appearance and rate of movements were similar to that reported for retrograde flow of cortical actin near the dorsal growth cone surface. Although these retrograde flow movements also exerted enough force to account for growth cone pulling, we did not observe such movements on ventral growth cone surfaces. Occasionally obstacles were moved by interaction with ruffling lamellipodia. However, we obtained no evidence for attachment of the obstacles to ruffling lamellipodia or for directed obstacle movements by this mechanism. These data suggest that chick sensory growth cones move forward by contractile activity of filopodia, i.e., isometric contraction on a rigid substrate. Our data argue against retrograde flow of actin producing traction force.