Capillary zone electrophoresis with laser-induced fluorescence detection for the determination of nanomolar concentrations of noradrenaline and dopamine: application to brain microdialyzate analysis

Robert, F.; Bert, Lionel; Denoroy, Luc; Renaud, Bernard

doi:10.1021/ac00107a013

Cited by 114 publications

(44 citation statements)

References 28 publications

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“…Such a mechanism, however, may have limitations for uphill transport. Basal cytoplasmic concentration of catecholamines is 1-2 M in pheochromocytoma cells or giant snail neurons (44, 45) whereas basal levels of extracellular catecholamines are typically 5-50 nM (46,47). Because uncoupled mechanisms only support a 10:1 ratio at Ϫ60 mV, it is unlikely FIG.…”

Section: Resultsmentioning

confidence: 99%

Patch-clamp and amperometric recordings from norepinephrine transporters: Channel activity and voltage-dependent uptake

Galli

Blakely²,

DeFelice³

1998

Proc. Natl. Acad. Sci. U.S.A.

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Transporters for the biogenic amines dopamine, norepinephrine, epinephrine and serotonin are largely responsible for transmitter inactivation after release. They also serve as high-affinity targets for a number of clinically relevant psychoactive agents, including antidepressants, cocaine, and amphetamines. Despite their prominent role in neurotransmitter inactivation and drug responses, we lack a clear understanding of the permeation pathway or regulation mechanisms at the single transporter level. The resolution of radiotracer-based flux techniques limits the opportunities to dissect these problems. Here we combine patch-clamp recording techniques with microamperometry to record the transporter-mediated flux of norepinephrine across isolated membrane patches. These data reveal voltage-dependent norepinephrine flux that correlates temporally with antidepressant-sensitive transporter currents in the same patch. Furthermore, we resolve unitary flux events linked with bursts of transporter channel openings. These findings indicate that norepinephrine transporters are capable of transporting neurotransmitter across the membrane in discrete shots containing hundreds of molecules. Amperometry is used widely to study neurotransmitter distribution and kinetics in the nervous system and to detect transmitter release during vesicular exocytosis. Of interest regarding the present application is the use of amperometry on inside-out patches with synchronous recording of flux and current. Thus, our results further demonstrate a powerful method to assess transporter function and regulation.Transporter-mediated clearance of neurotransmitters after vesicular release represents a crucial mechanism for temporal and spatial regulation of chemical signaling (1-5). Transporters for the biogenic amines dopamine, norepinephrine (NE), and serotonin (5HT) have particular significance as targets for clinically relevant psychoactive agents including, cocaine, antidepressants, and amphetamines (6). Cocaine and antidepressants are transporter antagonists that act with varying degrees of specificity to enhance synaptic concentrations of amines by limiting clearance (2, 6-8). Amphetamines enhance transporter mediated efflux in concert with a depletion of vesicular amine stores (8, 9). The localization of cocaine and antidepressant-sensitive transporters to the presynaptic neuron extends their influence to homeostatic relationships for transmitter biosynthesis and release.To date, the mechanisms of plasma membrane amine transport have been inferred primarily through transmitter-based flux measurements on populations of cells of isolated membrane fractions (10, 11). These assays predict the transport of amines to be coupled energetically to the inward movement of Na and Cl ions. In the case of 5HT transporters, this flux is coupled to the counter transport of K ions (11, 12), rendering 5HT transporters theoretically electroneutral. In the case of the NE transporter, the coupling stoichiometry predicted from cell and membrane vesicle flu...

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Section: Resultsmentioning

confidence: 99%

Patch-clamp and amperometric recordings from norepinephrine transporters: Channel activity and voltage-dependent uptake

Galli

Blakely²,

DeFelice³

1998

Proc. Natl. Acad. Sci. U.S.A.

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“…Several detection techniques in CE, including LIF [20][21][22], UV [23,24], MS [25], EC [26][27][28], and fluorescence [29], have been developed to determine catecholamines. Although LIF detection has the advantage of high sensitivity, the disadvantages are expensive instrumentation and complicated approach.…”

Section: Introductionmentioning

confidence: 99%

Determination of catecholamines by CE with direct chemiluminescence detection

Liu

Wang

et al. 2007

Electrophoresis

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A rapid and sensitive method to detect three catecholamines, isoprenaline, epinephrine, and dopamine, by CE coupled with direct luminol-potassium periodate chemiluminescence (CL) detection is described. The conditions for CE separation and CL reaction were systematically optimized. Under the optimum conditions, the baseline separation of three catecholamines was achieved within 6.5 min. The LODs obtained in standard solution were 5.3 6 10 -8 mol/L for isoprenaline, 4.7 6 10 -8 mol/L for epinephrine, and 1.5 6 10 -7 mol/L for dopamine. The RSD of the migration time and peak area were less than 1.8 and 3.6% (n = 5), respectively. The present method was applied to the determination of the dopamine in urine samples of cigarette smokers and nonsmokers. The results obtained indicate that there is a close relationship between the content of dopamine in human urine and the amount of cigarettes smoked daily; the level of dopamine in smokers is higher than in nonsmokers.

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“…In microdialysates, the CA determination by CZE-LIFD has been performed in low concentration, small volume samples derivatized with NAD to give highly fluorescent compounds. Robert et al [136] and Bert et al [137] have accomplished very reliable and sensitive determinations of CA. NE detection limit was 5 Â 10 À9 M in rat medial frontal cortex microdialysates [136].…”

Section: Ce-lifd Coupled To Microdialysis Studies Neuroactive Substamentioning

confidence: 99%

“…Robert et al [136] and Bert et al [137] have accomplished very reliable and sensitive determinations of CA. NE detection limit was 5 Â 10 À9 M in rat medial frontal cortex microdialysates [136]. NE detection with derivatization on-line of 2-min microdialysates reached high microdialysis temporal resolution [50].…”

Section: Ce-lifd Coupled To Microdialysis Studies Neuroactive Substamentioning

confidence: 99%

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

Páez

Hernández

2001

Biopharm & Drug Disp

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Capillary electrophoresis (CE) is a high-efficiency analytical technique that has had a great impact as a tool in biomedical research, clinical and forensic practice in the last ten years. Only in one of the applications, the DNA analysis, it has had an explosive exponential growth in the last few years. This impact is expressed in an enormous amount of CE articles and many reviews. The CE advantages with respect to other analytical techniques: the required very small sample volume, rapid analysis, great resolution power and low costs, have made this technique ideal for the analysis of a numerous endogenous and exogenous substances present in biological fluids. The different modes of CE have been coupled to different detection techniques such as UV-absorbance, electrochemical, mass spectrometry and laser-induced fluorescence detection (LIFD) to detect different nature and molecular size separated analytes. This review focuses mostly on the applications of CE-LIFD, to measure drugs and endogenous neuroactive substances such as amino acids and monoamines, especially in microdialysis samples from experimental animals and humans. CE-LIFD trends are discussed: automated faster analysis with capillary array systems, resolution power improvement, higher detection sensitivity, and CE systems miniaturization for extremely small sample volume, in order to make CE easier and affordable to the lab bench or the clinical bed.

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Capillary zone electrophoresis with laser-induced fluorescence detection for the determination of nanomolar concentrations of noradrenaline and dopamine: application to brain microdialyzate analysis

Cited by 114 publications

References 28 publications

Patch-clamp and amperometric recordings from norepinephrine transporters: Channel activity and voltage-dependent uptake

Patch-clamp and amperometric recordings from norepinephrine transporters: Channel activity and voltage-dependent uptake

Determination of catecholamines by CE with direct chemiluminescence detection

Biomedical applications of capillary electrophoresis with laser‐induced fluorescence detection

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