Detection of local ATP release from activated platelets  using cell surface-attached firefly luciferase

Beigi, Reza; Kobatake, Eiry; Aizawa, Masuo; Dubyak, George R.

doi:10.1152/ajpcell.1999.276.1.c267

Cited by 282 publications

(217 citation statements)

References 28 publications

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“…This technique allows the detection of ten to 20 times higher peak concentrations of ATP. Thus, it has been possible to obtain a much more accurate estimate of the ATP concentrations near the plasma membrane [30,31]. It has also provided convincing evidence that the actual concentrations of extracellular ATP during stimulation lie well within the range of nucleotide affinities of P2 receptors.…”

Section: The Luciferin-luciferase Techniquementioning

confidence: 94%

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ATP release from non-excitable cells

Praetorius

Leipziger

2009

Purinergic Signalling

207

229

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All cells release nucleotides and are in one way or another involved in local autocrine and paracrine regulation of organ function via stimulation of purinergic receptors. Significant technical advances have been made in recent years to quantify more precisely resting and stimulated adenosine triphosphate (ATP) concentrations in close proximity to the plasma membrane. These technical advances are reviewed here. However, the mechanisms by which cells release ATP continue to be enigmatic. The current state of knowledge on different suggested mechanisms is also reviewed. Current evidence suggests that two separate regulated modes of ATP release co-exist in nonexcitable cells: (1) a conductive pore which in several systems has been found to be the channel pannexin 1 and (2) vesicular release. Modes of stimulation of ATP release are reviewed and indicate that both subtle mechanical stimulation and agonist-triggered release play pivotal roles. The mechano-sensor for ATP release is not yet defined.

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Section: The Luciferin-luciferase Techniquementioning

confidence: 94%

“…This problem has been tackled by binding firefly luciferase to surface proteins. In this way, the detecting enzyme is placed in close proximity to the plasma membrane [31,32]. The technique exploits the strong protein binding property of bacterial protein A to IgG immunoglobulins.…”

Section: The Luciferin-luciferase Techniquementioning

confidence: 99%

ATP release from non-excitable cells

Praetorius

Leipziger

2009

Purinergic Signalling

207

229

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“…(4) A previous method in-volved the localization of a reporter to the outside surface via an affinity process, in this case using a protein A-luciferase chimera and coating the cell surface with IgG. (83) In both cases the addition of luciferin to the medium, in the presence of ATP, causes photon emission. Pellegatti's and colleagues' approach (4) is particularly attractive to the analysis of ATP dynamics in cell aggregates, although they discuss some limitations in the use of the expressible indicator, notably a low affinity allowing measurements only above 5-10M.…”

Section: Atp In Vivomentioning

confidence: 99%

Windows to cell function and dysfunction: Signatures written in the boundary layers

Smith¹,

Collis²,

Messerli³

2010

BioEssays

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The medium surrounding cells either in culture or in tissues contains a chemical mix varying with cell state. As solutes move in and out of the cytoplasmic compartment they set up characteristic signatures in the cellular boundary layers. These layers are complex physical and chemical environments whose profiles both reflect cell physiology and provide conduits for intercellular messaging. Here we review some of the most relevant characteristics of the extracellular/intercellular space. Our initial focus is primarily with cultured cells but we extend our consideration to the far more complex environment of tissues and discuss how chemical signatures in the boundary layer can or may affect cell function. Critical to the entire essay are the methods used, or being developed, to monitor chemical profiles in the boundary layers. We review recent developments in ultramicro electrochemical sensors and tailored optical reporters suitable for the task 20 in hand. IntroductionThe advent of electron tomography with 3 dimensional image reconstruction has revealed a fundamental beauty and complexity behind the structure of the living cell. Notable contributions have come from several imaging laboratories but one of the most compelling can be found in Marsh et al.(1) From such reconstructions the complexity within a cell is quite staggering and in the 4 th dimension of time this entire structure is in motion. The question that becomes extremely interesting is how molecular and ionic signals are regulated and move within these matrices, where chemical diffusion will be significantly altered. (2) However, complexity, both structural and functional, does not stop at the plasma membrane but extends outwards 30 into the extracellular/intercellular space (EICS: Fig.1). The chemical dynamics within this space are hypothesized to play key roles in cancer, spreading depression, epilepsy and sleep disorders. (3)(4)(5)(6) The purpose of this essay is to consider what we can learn from this space and the methods for following the dynamics of chemical movement within the diffusive boundary layers that comprise an integral part of a cell. We will start by noting the physical and chemical features involved.

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“…These high concentration requirements for ATP raise the issue about whether these levels are achievable during physiological regulation of the P2X 7 receptor. In the microenvironment of cell compartments it is possible that local release of ATP from adjacent cells (e.g., platelets [28]) or even autocrine release induced by activating agents such as LPS [29,30] may provide ATP concentrations that approach the 1-5 mM range. However, much remains to be uncovered about how the P2X 7 receptors work under normal physiological conditions.…”

Section: Macrophages Less Responsive Than Monocytes To P2x 7 Agonistsmentioning

confidence: 99%

P2X7 receptor and macrophage function

Wewers

Sarkar

2009

Purinergic Signalling

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Macrophages are unique innate immune cells that play an integral role in the defense of the host by virtue of their ability to recognize, engulf, and kill pathogens while sending out danger signals via cytokines to recruit and activate inflammatory cells. It is becoming increasingly clear that purinergic signaling events are essential components of the macrophage response to pathogen challenges and disorders such as sepsis may be, at least in part, regulated by these important sensors. The activation of the P2X 7 receptor is a powerful event in the regulation of the caspase-1 inflammasome. We provide evidence that the inflammasome activation requires "priming" of macrophages prior to ATP activation of the P2X 7 R. Inhibition of the inflammasome activation by the tyrosine kinase inhibitor, AG126, suggests regulation by phosphorylation. Finally, the P2X 7 R may also be activated by other elements of the host response such as the antimicrobial peptide LL-37, which adds a new, physiologically relevant agonist to the P2X 7 R pathway. Therapeutic approaches to inflammation and sepsis will certainly be enhanced by an increased understanding of how purinergic receptors modulate the inflammasomes.

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Detection of local ATP release from activated platelets using cell surface-attached firefly luciferase

Cited by 282 publications

References 28 publications

ATP release from non-excitable cells

ATP release from non-excitable cells

Windows to cell function and dysfunction: Signatures written in the boundary layers

P2X7 receptor and macrophage function

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