Combined integrin activation and intracellular cAMP cause Rho GTPase dependent growth cone collapse on laminin-1

Lemons, Michele L.; Condic, Maureen L.

doi:10.1016/j.expneurol.2006.06.008

Cited by 37 publications

(70 citation statements)

References 58 publications

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“…Surprisingly, different concentrations of the same molecule can have dramatic effects on the internal state of neurons as well as their cell surface composition. For example, intracellular levels of cAMP and surface levels of integrins are significantly different in neurons plated on these two substrata 7,8 . Additional studies have shown that other molecules, including fibronectin and chondroitin sulfate proteoglycans, impact the expression of cell surface molecules and neuronal motility [7][8][9][10][11] .…”

Section: Introductionmentioning

confidence: 99%

“…For example, intracellular levels of cAMP and surface levels of integrins are significantly different in neurons plated on these two substrata 7,8 . Additional studies have shown that other molecules, including fibronectin and chondroitin sulfate proteoglycans, impact the expression of cell surface molecules and neuronal motility [7][8][9][10][11] . In addition, soluble molecules such as neurotrophins and neurotropins also impact cell membrane composition and neuronal motility [12][13][14][15][16] and can be easily and accurately manipulated in a cell culture model.…”

Section: Introductionmentioning

confidence: 99%

“…This procedure has been used to make significant breakthroughs in neurobiology, including axon outgrowth 5,7,8,10,11,[16][17][18][19][20][21] and was modified from a procedure designed to isolate ganglion cells 22 . There are several advantages to this approach.…”

Section: Introductionmentioning

confidence: 99%

“…The procedure for double immunocytochemistry against neural cell adhesion molecule (NCAM) and β1 integrins is provided. Data generated from these immunocytochemical methods have been used to examine the spatial patterning and intensity of several molecules in cultured neurons 8,16 . .…”

Section: Introductionmentioning

confidence: 99%

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Isolation and Culture of Dissociated Sensory Neurons From Chick Embryos

Powell

Vinod

Lemons

2014

JoVE

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Neurons are multifaceted cells that carry information essential for a variety of functions including sensation, motor movement, learning, and memory. Studying neurons in vivo can be challenging due to their complexity, their varied and dynamic environments, and technical limitations. For these reasons, studying neurons in vitro can prove beneficial to unravel the complex mysteries of neurons. The well-defined nature of cell culture models provides detailed control over environmental conditions and variables. Here we describe how to isolate, dissociate, and culture primary neurons from chick embryos. This technique is rapid, inexpensive, and generates robustly growing sensory neurons. The procedure consistently produces cultures that are highly enriched for neurons and has very few non-neuronal cells (less than 5%). Primary neurons do not adhere well to untreated glass or tissue culture plastic, therefore detailed procedures to create two distinct, well-defined laminin-containing substrata for neuronal plating are described. Cultured neurons are highly amenable to multiple cellular and molecular techniques, including co-immunoprecipitation, live cell imagining, RNAi, and immunocytochemistry. Procedures for double immunocytochemistry on these cultured neurons have been optimized and described here. Video LinkThe video component of this article can be found at

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Isolation and Culture of Dissociated Sensory Neurons From Chick Embryos

Powell

Vinod

Lemons

2014

JoVE

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“…Their functions are regulated by a conformational change in the extracellular domain from a low-affinity to a high-affinity state (also called "inside-out " signaling or integrin activation) and "outside-in " signaling (the signaling cascades propagated intracellularly following integrin-ligand binding). Enhancing ligand-binding affinity via integrin activation promotes neurite outgrowth from cultured CNS and PNS neurons (Ivins et al, 2000;Lein et al, 2000;Lemons and Condic, 2006;Tan et al, 2011). Crucially, integrin activation also enhances the ability of neurons to extend axons in the presence of inhibitory molecules such as amino-Nogo and the chondroitin sulfate proteoglycan (CSPG) aggrecan, which can both inactivate axonal integrins (Hu and Strittmatter, 2008;Tan et al, 2011) and are highly expressed after a CNS injury (Liu et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Kindlin-1 Enhances Axon Growth on Inhibitory Chondroitin Sulfate Proteoglycans and Promotes Sensory Axon Regeneration

Tan¹,

Andrews²,

Kwok³

et al. 2012

J. Neurosci.

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Growing and regenerating axons need to interact with the molecules in the extracellular matrix as they traverse through their environment. An important group of receptors that serve this function is the integrin superfamily of cell surface receptors, which are evolutionarily conserved ␣␤ heterodimeric transmembrane proteins. The function of integrins is controlled by regulating the affinity for ligands (also called "integrin activation"). Previous results have shown that CNS inhibitory molecules inactivate axonal integrins, while enhancing integrin activation can promote axon growth from neurons cultured on inhibitory substrates. We tested two related molecules, kindlin-1 and kindlin-2 (Fermitin family members 1 and 2), that can activate ␤1, ␤2, and ␤3 integrins, for their effects on integrin signaling and integrin-mediated axon growth in rat sensory neurons. We determined that kindlin-2, but not kindlin-1, is endogenously expressed in the nervous system. Knocking down kindlin-2 levels in cultured sensory neurons impaired their ability to extend axons, but this was partially rescued by kindlin-1 expression. Overexpression of kindlin-1, but not kindlin-2, in cultured neurons increased axon growth on an inhibitory aggrecan substrate. This was found to be associated with enhanced integrin activation and signaling within the axons. Additionally, in an in vivo rat dorsal root injury model, transduction of dorsal root ganglion neurons to express kindlin-1 promoted axon regeneration across the dorsal root entry zone and into the spinal cord. These animals demonstrated improved recovery of thermal sensation following injury. Our results therefore suggest that kindlin-1 is a potential tool for improving axon regeneration after nervous system lesions.

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Schwann cell migration is integrin‐dependent and inhibited by astrocyte‐produced aggrecan

Afshari

Kwok

White

et al. 2010

Glia

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Schwann cells transplantation has considerable promise in spinal cord trauma to bridge the site of injury and for remyelination in demyelinating conditions. They support axonal regeneration and sprouting by secreting growth factors and providing a permissive surface and matrix molecules while shielding axons from the inhibitory environment of the central nervous system. However, following transplantation Schwann cells show limited migratory ability and they are unable to intermingle with the host astrocytes. This in turn leads to formation of a sharp boundary and an abrupt transition between the Schwann cell graft and the host tissue astrocytes, therefore preventing regenerating axons from exiting the graft. The objective of this study was to identify inhibitory elements on astrocytes involved in restricting Schwann cell migration. Using in vitro assays of cell migration, we show that aggrecan produced by astrocytes is involved in the inhibition of Schwann cell motility on astrocytic monolayers. Knockdown of this proteoglycan in astrocytes using RNAi or digestion of glycosaminglycan chains on aggrecan improves Schwann cell migration. We further show aggrecan mediates its effect by disruption of integrin function in Schwann cells, and that the inhibitory effects of aggrecan can overcome by activation of Schwann cell integrins.

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Combined integrin activation and intracellular cAMP cause Rho GTPase dependent growth cone collapse on laminin-1

Cited by 37 publications

References 58 publications

Isolation and Culture of Dissociated Sensory Neurons From Chick Embryos

Isolation and Culture of Dissociated Sensory Neurons From Chick Embryos

Kindlin-1 Enhances Axon Growth on Inhibitory Chondroitin Sulfate Proteoglycans and Promotes Sensory Axon Regeneration

Schwann cell migration is integrin‐dependent and inhibited by astrocyte‐produced aggrecan

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