Neuronal cells mature faster on polyethyleneimine coated plates than on polylysine coated plates

Lelong, Isabelle H.; Petegnief, Valérie; Rebel, G.

doi:10.1002/jnr.490320411

Cited by 49 publications

(40 citation statements)

References 46 publications

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“…Once the dissection above is complete, MEA preparation is continued by placing 100 μl of polyethyleneimine (PEI) solution into the center of each MEA. 13 In our hands, PEI solution provides less clustering of cells in the MEA than using polylysine. The dish is allowed to sit at room temperature in the laminar flow hood for 30 minutes with lids lightly resting on the MEAs to prevent evaporation.…”

Section: B Brain Dissectionmentioning

confidence: 72%

How to Culture, Record and Stimulate Neuronal Networks on Micro-electrode Arrays (MEAs)

Hales

Rolston

Potter

2010

JoVE

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For the last century, many neuroscientists around the world have dedicated their lives to understanding how neuronal networks work and why they stop working in various diseases. Studies have included neuropathological observation, fluorescent microscopy with genetic labeling, and intracellular recording in both dissociated neurons and slice preparations. This protocol discusses another technology, which involves growing dissociated neuronal cultures on micro-electrode arrays (also called multi-electrode arrays, MEAs).There are multiple advantages to using this system over other technologies. Dissociated neuronal cultures on MEAs provide a simplified model in which network activity can be manipulated with electrical stimulation sequences through the array's multiple electrodes. Because the network is small, the impact of stimulation is limited to observable areas, which is not the case in intact preparations. The cells grow in a monolayer making changes in morphology easy to monitor with various imaging techniques. Finally, cultures on MEAs can survive for over a year in vitro which removes any clear time limitations inherent with other culturing techniques. 1Our lab and others around the globe are utilizing this technology to ask important questions about neuronal networks. The purpose of this protocol is to provide the necessary information for setting up, caring for, recording from and electrically stimulating cultures on MEAs. In vitro networks provide a means for asking physiologically relevant questions at the network and cellular levels leading to a better understanding of brain function and dysfunction. Video LinkThe video component of this article can be found at https://www.jove.com/video/2056/ Protocol A. IntroductionThere are billions of neurons in the human brain. Each of these neurons can have hundreds to thousands of connections often times with many different cells. These connections harbor the signals that allow us to walk, play a piano, ride a bicycle, laugh, cry, and remember. For the last century, many neuroscientists around the world have dedicated their lives to understanding how neuronal networks work and why they stop working in various diseases.There are many tools one can use when taking on this seemingly insurmountable task. Initial studies were focused on neuropathological observation of how neural circuits are organized in the brains of humans and other animals. The advent of fluorescence microscopy and genetic labeling expanded the scope of these structural studies exploring cellular composition down to the protein and DNA level.But these experiments did not address the dynamic function of the brain, only the static arrangement. For understanding ongoing neural activity, popular techniques generally focus on examining electrophysiological activity with intracellular recording. Single neurons of dissociated cultures provide a useful reductionist model, however this technique is limited by short time intervals for recording from single cells. This model also provides limited in...

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Section: B Brain Dissectionmentioning

confidence: 72%

How to Culture, Record and Stimulate Neuronal Networks on Micro-electrode Arrays (MEAs)

Hales

Rolston

Potter

2010

JoVE

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“…Dissociated neurons were plated at a density of less than 50 neurons/mm 2 on polyethyleneimine-coated glass coverslips that were attached over a hole in the bottom of a culture dish (Whitlon and Baas, 1992). Coverslips were coated with 0.7% polyethyleneimine (PEI; Sigma; see Rüegg and Hefti, 1984;Lelong et al, 1992) after the assembly of the chambers, as described by Keith and Farmer (1993). [PEI induces the development of dendrites with compact growth cones reminiscent of those described for cortical dendrites in vivo ( Peters et al, 1991)].…”

Section: Cell Culturementioning

confidence: 99%

Glutamate modulation of dendrite outgrowth: Alterations in the distribution of dendritic microtubules

Wilson

Keith

1998

J. Neurosci. Res.

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Glutamate can both facilitate and inhibit dendrite outgrowth in vitro. The major effects of low levels of glutamate occur only on the dendrites (not the axon) of pyramidal neurons and may be important for modulating dendrite outgrowth during neuronal development in vivo. Cytoskeletal changes resulting from glutamate exposure must underlie these changes in dendrite outgrowth. In the present study, hippocampal neuron cultures were used to measure the outgrowth of both axons and immature dendrites in the presence or absence of 50 microM glutamate. Subsequently, neurons were extracted and fixed for immunofluorescent labeling of microtubules and rhodamine phalloidin labeling of microfilaments. Additionally, neurons were prepared for electron microscopy to examine dendritic microtubules at the ultrastructural level. Glutamate led to increased dendrite outgrowth in the short term (4 hr) and dendrite retraction in the long term (8 hr). After short-term glutamate exposures, no obvious morphological changes occur in either the microtubules or microfilaments. However, longer glutamate exposure causes a decrease in the number of microtubules in the distal region of retracting dendrites, and causes an increase in microtubule number in the dendritic shaft of both retracting and growing dendrites. Thus, the microtubule cytoskeleton may be involved in producing the changes in dendrite outgrowth caused by glutamate exposure.

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“…In order to follow hemocyte shape changes in condition of limited cell displacements, some experiments were repeated on polyethyleneimine-coated culture dishes. This surface treatment promotes cell adhesion (Lelong et al 1992). In these conditions, morphologic differences between round filopodia-exhibiting cells and spread granulocytes were enhanced (Fig.…”

Section: Resultsmentioning

confidence: 93%

Cell tracking and velocimetric parameters analysis as an approach to assess activity of mussel (Mytilus edulis) hemocytes in vitro

Rioult

Lebel²,

Foll

2013

Cytotechnology

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Hemocytes constitute the key element of innate immunity in bivalves, being responsible for secretion of antimicrobial peptides and release of zymogens from the prophenoloxidase system within the hemolymph compartment, reactive oxygen species production and phagocytosis. Hemocytes are found (and collected) as cells in suspension in circulating hemolymph. Hemocytes are adherent cells as well, infiltrating tissues and migrating to infected areas. In the present study, we applied an approach based on fluorescent staining and nuclei-tracking to determine migration velocity of hemocytes from the blue mussel, Mytilus edulis, in culture. Freshly collected hemocytes attached to substrate and start to move spontaneously in few minutes. Two main hemocyte morphologies can be observed: small star-shaped cells which were less motile and spread granular cells with faster migrations. Cell-tracking was combined to MTT mitochondria metabolic rate measurements in order to monitor global cell population activity over 4 days of culture. A transient peak of cell activity was recorded after 24-48 h of culture, corresponding to a speed up of cell migration. Videomicroscopy and cell tracking techniques provide new tools to characterize activity of mussel immunocytes in culture. Our analysis of hemocyte migration reveals that motility is very sensitive to cell environmental factors.

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Neuronal cells mature faster on polyethyleneimine coated plates than on polylysine coated plates

Cited by 49 publications

References 46 publications

How to Culture, Record and Stimulate Neuronal Networks on Micro-electrode Arrays (MEAs)

How to Culture, Record and Stimulate Neuronal Networks on Micro-electrode Arrays (MEAs)

Glutamate modulation of dendrite outgrowth: Alterations in the distribution of dendritic microtubules

Cell tracking and velocimetric parameters analysis as an approach to assess activity of mussel (Mytilus edulis) hemocytes in vitro

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