Artificial cells: Unique insights into exocytosis using liposomes and lipid nanotubes

Cans, Ann-Sofie; Wittenberg, Nathan J.; Karlsson, Roger; Sombers, Leslie A.; Karlsson, Mattias; Orwar, Owe; Ewing, Andrew G.

doi:10.1073/pnas.232702599

Cited by 129 publications

(122 citation statements)

References 33 publications

Supporting

Mentioning

117

Contrasting

Order By: Relevance

“…It can be noted that the size of these spots in diameter is close to the size of vesicles reported in the literature ͑ranging from 50 nm to 1 m in diameter͒. 33 To acquire temporal and spatially resolved information, fluorescence intensity for bright spots ͑five regions of interest selected͒ was plotted against time ͓Fig. 5͑a͔͒.…”

Section: A Fluorescence Imaging Evaluationsupporting

confidence: 63%

Release monitoring of single cells on a microfluidic device coupled with fluorescence microscopy and electrochemistry

et al. 2010

View full text Add to dashboard Cite

A method for monitoring the biological exocytotic phenomena on a microfluidic system was proposed. A microfluidic device coupled with functionalities of fluorescence imaging and amperometric detection has been developed to enable the real-time monitoring of the exocytotic events. Exocytotic release of single SH-SY5Y neuroblastoma cells was studied. By staining the cells located on integrated microelectrodes with naphthalene-2,3-dicarboxaldehyde, punctuate fluorescence consistent with localization of neurotransmitters stored in vesicles was obtained. The stimulated exocytotic release was successfully observed at the surface of SH-SY5Y cells without refitting the commercial inverted fluorescence microscope. Spatially and temporally resolved exocytotic events from single cells on a microfluidic device were visualized in real time using fluorescence microscopy and were amperometrically recorded by the electrochemical system simultaneously. This coupled technique is simple and is hoped to provide new insights into the mechanisms responsible for the kinetics of exocytosis.

show abstract

Section: A Fluorescence Imaging Evaluationsupporting

confidence: 63%

Release monitoring of single cells on a microfluidic device coupled with fluorescence microscopy and electrochemistry

et al. 2010

View full text Add to dashboard Cite

show abstract

“…On the other hand, this should also allow the formation of wider exocytotic fusion pores by relaxing at a molecular level the important local curvature strains created into lipid nanotubes or toroid junctions connecting cell and vesicle membranes. 42,43 Conversely, at the low concentrations used in this work SDS has been shown to have no effect on the SNAREs machineries that lead to initial fusion pores formation. 51,52 Hence, eliciting exocytosis by SDS at these low concentrations should not tamper at all with the formation of calcium-induced SNAREs complexes that initiates the critical process leading to the opening of the initial exocytotic fusion pores.…”

Section: Resultsmentioning

confidence: 68%

How “Full” is “Full Fusion” during Exocytosis from Dense Core Vesicles? Effect of SDS on “Quantal” Release and Final Fusion Pore Size

Ren

Lin

et al. 2016

J. Electrochem. Soc.

View full text Add to dashboard Cite

In this work, release was elicited from PC12 using sufficiently small concentrations of sodium dodecyl sulfate (SDS) to perturb normal release mechanism in an attempt to reveal concealed information while keeping physiologically compatible conditions. Amperometry was used to monitor and quantify the released fluxes and kinetics in individual vesicular events. This showed that stimulating release with SDS 350 μM leads to a doubling of the quantity of catecholamine cations released per event and much larger release fluxes as compared to controls (release elicited with K + 105 mM under same conditions) or SDS 250 μM. These quantitative measurements confirm our previous theoretical model and reports based on ex situ cytometric experiments on isolated PC12 vesicles, which established that release is far from being total under normal exocytotic conditions. Secondly, this establishes that the maximal size of fusion pores at the end of the "full fusion" phase is limited by some contraption unrelated to the membrane. Indeed, the present results are entirely consistent with the fact that SDS 350 μM allows the fusion pore to expand to a double size (ca. 28 nm radius) compared to controls and SDS 250 μM (ca. 14 nm radius). The considerable importance of the mechanism of vesicular exocytosis in biology and medicine [1][2][3][4][5][6] has stimulated an increasing number of reports. This high importance is evidenced, for example, by the award of two recent Nobel prizes in physiology and medicine bearing on these issues, the latest one being awarded in 2013 to James E. Rothman, Randy W. Schekman, and Thomas C. Südhof. [3][4][5][6] In neurons and endocrine cells neurotransmitters are stored and transported inside vesicles to be finally delivered at specific release sites of the outer cell membrane.7 Vesicles and cells membranes connect there through activation of SNAREs complexes following intracellular gated calcium ions influxes to form a nanometric fusion pore through which neurotransmitters can diffuse away.7-9 Most of the biochemical cellular processes regulating the formation and the intracellular traffic of these vesicles -as well as the corresponding biomolecular machineries involved -are now well established and documented. However, albeit this recognition, and after more than 20 years of important intensive and thoughtful works the crucial ultimate stages of the process which govern vesicular exocytotic events, viz., the very delivery of neurotransmitters in the extracellular space, are not fully understood beyond those relative to the transient initial fusion pore through which only a modest amount of neurotransmitters may be released 10-12 whose dimensions (ca. 1.2 nm radius) and fast dynamics (Kiss and Run) have been characterized by patch-clamp, [13][14][15][16][17][18] and very recently through amperometry. 19In endocrine cells, the main releasing stage involves a fast expansion of the initial fusion pore, generally thought to lead to a full incorporation of the former vesicle membrane into the cell one, 20 throug...

show abstract

“…Note that this is bound to remain true even when the vesicle-cell membrane connection is not tore-like as drawn in Figure 2 but involves for example a bilipidic tube connection as has been suggested recently. [11] Indeed, provided that the diffusion coefficient in such intermediate structure remains comparable to that in the extracellular fluid and that the length of the bilipidic tube remains at most comparable to the vesicle radius, the kinetics of release will be exclusively commanded by the rate of release from the spherical vesicle.…”

Section: Introductionmentioning

confidence: 99%

Reconstruction of Aperture Functions during Full Fusion in Vesicular Exocytosis of Neurotransmitters

2010

View full text Add to dashboard Cite

Individual vesicular exocytosis of adrenaline by dense core vesicles in chromaffin cells is considered here as a paradigm of many situations encountered in biology, nanosciences and drug delivery in which a spherical container releases in the external environment through gradual uncovering of its surface. A procedure for extracting the aperture (opening) function of a biological vesicle fusing with a cell membrane from the released molecular flux of neurotransmitter as monitored by amperometry has been devised based on semi-analytical expressions derived in a former work [C. Amatore, A. I. Oleinick, I. Svir, ChemPhysChem 2009, 10, DOI: 10.1002/cphc.200900646]. This precise analysis shows that in the absence of direct information about the radius of the vesicle or about the concentration of the adrenaline cation stored by the vesicle matrix, current spikes do not contain enough information to determine the maximum aperture angle. Yet, a statistical analysis establishes that this maximum aperture angle is most probably less than a few tens of degrees, which suggests that full fusion is a very improbable event.

show abstract

Artificial cells: Unique insights into exocytosis using liposomes and lipid nanotubes

Cited by 129 publications

References 33 publications

Release monitoring of single cells on a microfluidic device coupled with fluorescence microscopy and electrochemistry

Release monitoring of single cells on a microfluidic device coupled with fluorescence microscopy and electrochemistry

How “Full” is “Full Fusion” during Exocytosis from Dense Core Vesicles? Effect of SDS on “Quantal” Release and Final Fusion Pore Size

Reconstruction of Aperture Functions during Full Fusion in Vesicular Exocytosis of Neurotransmitters

Contact Info

Product

Resources

About