| 1. |
- Lovric, Jelena, 1980, et al.
(författare)
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Analysis of liposome model systems by time-of-flight secondary ion mass spectrometry
- 2014
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Ingår i: Surface and Interface Analysis: 19th International Conference on Secondary Ion Mass Spectrometry (SIMS). - : Wiley. - 0142-2421 .- 1096-9918.
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Konferensbidrag (refereegranskat)abstract
- Time-of-flight secondary ion mass spectrometry (ToF-SIMS) is an important technique for studying chemical composition of micrometer scale objects because of its high spatial resolution imaging capabilities and chemical specificity. In this work, we focus on the application of ToF-SIMS to gain insight into the chemistry of micrometer size liposomes as a potential model for neurotransmitter vesicles. Two models of giant liposomes were analyzed: histamine and aqueous two-phase system-containing liposomes. Characterization of the internal structure of single fixed liposomes was carried out both with the Bi-3(+) and C-60(+) ion sources. The depth profiling capability of ToF-SIMS was used to investigate the liposome interior.
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| 2. |
- Li, Xianchan, 1982, et al.
(författare)
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Mechanistic Aspects of Vesicle Opening during Analysis with Vesicle Impact Electrochemical Cytometry
- 2017
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 89:17, s. 9416-9423
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Tidskriftsartikel (refereegranskat)abstract
- Vesicle impact electrochemical cytometry (VIEC) has been used to quantify the vesicular transmitter content in mammalian vesicles. In the present study, we studied the mechanism of VIEC by quantifying the catecholamine content in single vesicles isolated from pheochromocytoma (PC12) cells. These vesicles contain about one tenth of the catecholamine compared with adrenal chromaffin vesicles. The existence of a prespike foot for many events suggests the formation of an initial transiently stable pore at the beginning of vesicle rupture. Increasing the detection temperature from 6 to 30 degrees C increases the possibility of vesicle rupture on the electrode, implying that there is a temperature-dependent process that facilitates electroporation. Natively larger vesicles are shown to rupture earlier and more frequently than smaller ones in VIEC. Likewise, manipulating vesicle content and size with drugs leads to similar trends. These data support the hypothesis that electroporation is the primary force for pore opening in VIEC. We further hypothesize that a critical step for initiating vesicle opening by electroporation is diffusion of membrane proteins away from the membrane region of contact with the electrode to allow closer contact, increasing the lateral potential field and thus facilitating electroporation.
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| 3. |
- Li, Xianchan, 1982, et al.
(författare)
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Nanopore Opening at Flat and Nanotip Conical Electrodes during Vesicle Impact Electrochemical Cytometry
- 2018
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Ingår i: ACS Nano. - : American Chemical Society (ACS). - 1936-0851 .- 1936-086X. ; 12:3, s. 3010-3019
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Tidskriftsartikel (refereegranskat)abstract
- The oxidation of catecholamine at a microelectrode, following its release from individual vesicles, allows interrogation of the content of single nanometer vesicles with vesicle impact electrochemical cytometry (VIEC). Previous to this development, there were no methods available to quantify the chemical load of single vesicles. However, accurate quantification of the content is hampered by uncertainty in the proportion of substituent molecules reaching the electrode surface (collection efficiency). In this work, we use quantitative modeling to calculate this collection efficiency. For all vesicles except those at the very edge of the electrode, modeling shows that ∼100% oxidation efficiency is achieved when employing a 33 μm diameter disk microelectrode for VIEC, independent of the location of the vesicle release pore. We use this to experimentally determine a precise distribution of catecholamine in individual vesicles extracted from PC12 cells. In contrast, we calculate that when a nanotip conical electrode (∼4 μm length, ∼1.5 μm diameter at the base) is employed, as in intracellular VIEC (IVIEC), the current-time response depends strongly on the position of the catecholamine-releasing pore in the vesicle membrane. When vesicle release occurs with the pore opening occurring far from the electrode, lower currents and partial oxidation (∼75%) of the catecholamine are predicted, as compared to higher currents and ∼100% oxidation, when the pore is close to/at the electrode surface. As close agreement is observed between the experimentally measured vesicular content in intracellular and extracted vesicles from the same cell line using nanotip and disk electrodes, respectively, we conclude that pores open at the electrode surface. Not only does this suggest that electroporation of the vesicle membrane is the primary driving force for catecholamine release from vesicles at polarized electrodes, but it also indicates that IVIEC with nanotip electrodes can directly assess vesicular content without correction. © 2018 American Chemical Society.
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| 4. |
- Li, Xianchan, 1982, et al.
(författare)
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Quantitative Chemical Measurements of Vesicular Transmitters with Electrochemical Cytometry
- 2016
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Ingår i: Accounts of Chemical Research. - : American Chemical Society (ACS). - 0001-4842 .- 1520-4898. ; 49:10, s. 2347-2354
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Tidskriftsartikel (refereegranskat)abstract
- Electrochemical cytometry adds a new dimension to our ability to study the chemistry and chemical storage of transmitter molecules stored in nanometer vesicles. The approach involves the adsorption and subsequent rupture of vesicles on an electrode surface during which the electroactive contents are quantitatively oxidized (or reduced). The measured current allows us to count the number of molecules in the vesicles using Faraday's law and to correlate this-to the amount of molecules released when single exocytosis events take place at communicating cells. The original format for this method involved a capillary electrophoresis separation step to singly address each vesicle, but we have more recently discovered that cellular vesicles tend to adsorb to carbon electrodes and spontaneously as well as stochastically rupture to give mostly single vesicle events. This approach, called impact electrochemical cytometry, even though the impact is perhaps not the important part of this process, has been studied and the vesicle rupture appears to be at the interface between the vesicle and the electrode and is probably driven by electroporation. The pore size and rate of content electrolysis are a function of the pore diameter and the presence of a protein core in the vesicles. In model liposomes with no protein, events appear extremely rapidly as the soft nanoparticles impact the electrode and the contents are oxidized. It appears that the proteins decorating the surface of the vesicle are important in maintaining a gap from the electrode and when this gap is closed electroporation takes place. Models of the event response times suggest the pores formed are small enough so we can carry out these measurements at nanotip electrodes and we have used this to quantify the vesicle content in living cells in a mode we call intracellular impact electrochemical cytometry. The development of electrochemical cytometry allows comparison between vesicle content and vesicular release and we have found that only part of the vesicle content is released in typical exocytotic cases measured by amperometry. This has led to the novel hypothesis that most exocytosis from dense core vesicles is via mechanism where vesicles fuse with the cell membrane, some content is released and then close again to be reloaded and reused. It leaves open the possibility that cells regulate release during individual events. This might be important in learning and memory and be a nonreceptor pharmaceutical target for brain related disorders. Indeed, the concept of the chemo-brain observed in cisplatin-treated cancer patients appears to be at least in part the result of changing the fraction of transmitter released and we have been able to show this by using the combined amperometric measurement of release and electrochemical cytometry at model cells.
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| 5. |
- Li, Xianchan, 1982, et al.
(författare)
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Quantitative Measurement of Transmitters in Individual Vesicles in the Cytoplasm of Single Cells with Nanotip Electrodes
- 2015
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Ingår i: Angewandte Chemie International Edition. - : Wiley. - 1433-7851 .- 1521-3773. ; 54:41, s. 11978-11982
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Tidskriftsartikel (refereegranskat)abstract
- The quantification of vesicular transmitter content is important for studying the mechanisms of neurotransmission and malfunction in disease, and yet it is incredibly difficult to measure the tiny amounts of neurotransmitters in the attoliter volume of a single vesicle, especially in the cell environment. We introduce a novel method, intracellular vesicle electrochemical cytometry. A nanotip conical carbon-fiber microelectrode was used to electrochemically measure the total content of electroactive neurotransmitters in individual nanoscale vesicles in single PC12 cells as these vesicles lysed on the electrode inside the living cell. The results demonstrate that only a fraction of the quantal neurotransmitter content is released during exocytosis. These data support the intriguing hypothesis that the vesicle does not open all the way during the normal exocytosis process, thus resulting in incomplete expulsion of the vesicular contents.
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| 6. |
- Li, Xianchan, 1982, et al.
(författare)
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Single cell amperometry reveals curcuminoids modulate the release of neurotransmitters during exocytosis from PC12 cells
- 2016
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Ingår i: Journal of Electroanalytical Chemistry. - : Elsevier BV. - 1572-6657. ; 781, s. 30-35
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Tidskriftsartikel (refereegranskat)abstract
- We used single cell amperometry to examine whether curcumin and bisdemethoxycurcumin (BDMC), substances that are suggested to affect learning and memory, can modulate monoamine release from PC12 cells. Our results indicate both curcumin and BDMC need long-term treatment (72 h in this study) to influence exocytosis effectively. By analyzing the parameters calculated from single exocytosis events, it can be concluded that curcumin and BDMC affect exocytosis through different mechanisms. Curcumin accelerates the event dynamics with no significant change of the monoamine amount released from single exocytotic events, whereas BDMC attenuates the amount from single exocytotic event with no significant change of the event dynamics. This comparison of the effect of curcumin and BDMC on exocytosis at the single cell level brings insight into their different mechanisms, which might lead to different biological actions. The effect of curcumin and BDMC on the opening and closing of the exocytotic fusion pore were also investigated. These results might be helpful for understanding the improvement of learning and memory and the anti-depression properties of curcuminoids.
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| 7. |
- Li, Xianchan, 1982, et al.
(författare)
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Using Single-Cell Amperometry To Reveal How Cisplatin Treatment Modulates the Release of Catecholamine Transmitters during Exocytosis
- 2016
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Ingår i: Angewandte Chemie-International Edition. - : Wiley. - 1433-7851 .- 1521-3773. ; 55:31, s. 9041-9044
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Tidskriftsartikel (refereegranskat)abstract
- The pretreatment of cultured pheochromocytoma (PC12) cells with cis-diamminedichloroplatinum (cisplatin), an anti-cancer drug, influences the exocytotic ability of the cells in a dose-dependent manner. Low concentrations of cisplatin stimulate catecholamine release whereas high concentrations inhibit it. Single-cell amperometry reflects that 2 mm cisplatin treatment increases the frequency of exocytotic events and reduces their duration, whereas 100 mm cisplatin treatment decreases the frequency of exocytotic events and increases their duration. Furthermore, the stability of the initial fusion pore that is formed in the lipid membrane during exocytosis is also regulated differentially by different cisplatin concentrations. This study thus suggests that cisplatin influences exocytosis by multiple mechanisms.
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| 8. |
- Majdi, Soodabeh, 1980, et al.
(författare)
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Selected recent in vivo studies on chemical measurements in invertebrates
- 2015
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Ingår i: Analyst. - : Royal Society of Chemistry (RSC). - 0003-2654 .- 1364-5528. ; 140:11, s. 3676-3686
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Tidskriftsartikel (refereegranskat)abstract
- In vivo measurements of neurotransmitters and related compounds have provided a better understanding of the chemical interactions that are a major part in functioning of brains. In addition, a great deal of technology has been developed to measure chemical species in other areas of living organisms. A key part of this work has been sampling technologies as well as direct measurements in vivo. This is extremely important when sampling from the smallest animal systems. Yet, very small invertebrate systems are excellent models and often have better defined and more easily manipulated genetics. This review focuses on in vivo measurements, electrochemical methods, fluorescence techniques, and sampling and is further narrowed to work over approximately the last three years. Rapid developments of in vivo studies in these model systems should aid in finding solutions to biological and bioanalytical challenges related to human physiological functions and neurodegenerative diseases.
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| 9. |
- Mohammadi, Amir Saeid, et al.
(författare)
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Mass Spectrometry Imaging Suggests That Cisplatin Affects Exocytotic Release by Alteration of Cell Membrane Lipids
- 2018
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 90:14, s. 8509-8516
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Tidskriftsartikel (refereegranskat)abstract
- We used time-of-flight secondary ion mass spectrometry (TOFSIMS) imaging to investigate the effect of cisplatin, the first member of the platinum-based anticancer drugs, on the membrane lipid composition of model cells to see if lipid changes might be involved in the changes in exocytosis observed. Platinum-based anticancer drugs have been reported to affect neurotransmitter release resulting in what is called the "chemobrain"; however, the mechanism for the influence is not yet understood. TOF-SIMS imaging was carried out using a high energy 40 keV (CO2)(6000)(+) gas cluster ion beam with improved sensitivity for intact lipids in biological samples. Principal components analysis showed that cisplatin treatment of PC12 cells significantly affects the abundance of different lipids and their derivatives, particularly phosphatidylcholine and cholesterol, which are diminished. Treatment of cells with 2 mu M and 100 mu M cisplatin showed similar effects on induced lipid changes. Lipid content alterations caused by cisplatin treatment at the cell surface are associated with the molecular and bimolecular signaling pathways of cisplatin-induced apoptosis of cells. We suggest that lipid alterations measured by TOF-SIMS are involved, at least in part, in the regulation of exocytosis by cisplatin.
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| 10. |
- Phan, Nhu TN, 1981, et al.
(författare)
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Measuring synaptic vesicles using cellular electrochemistry and nanoscale molecular imaging
- 2017
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Ingår i: Nature Reviews Chemistry. - : Springer Science and Business Media LLC. - 2397-3358. ; 1
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Tidskriftsartikel (refereegranskat)abstract
- The synaptic vesicle, a cellular compartment tens to hundreds of nanometres in size, is a main player in the process of exocytosis for neuronal communication. Understanding the regulatory mechanism of neurotransmission and neurological disorders requires the quantification of chemicals transmitted between cells. These challenging single vesicle measurements can be performed using analytical techniques described in this Review. In vivo amperometry at living cells can be used to quantify the amount of neurotransmitter released from a vesicle. By contrast, intracellular vesicle impact electrochemical cytometry allows the amount of molecules to be determined inside single vesicles. Although the dominant mode of exocytosis from vesicles is still under debate, several experiments point to the importance of partial release modes. Making use of fluorescent or isotopically labelled probes enables super-resolution optical and mass spectrometric imaging of molecular composition and activity of single vesicles. Correlating results from these nanoscopic techniques with those from electrochemistry has proved advantageous in understanding the relationship between vesicle structure and function. © 2017 Macmillan Publishers Limited.
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