Dynamic monitoring of single-terminal norepinephrine transporter rate in the rodent cardiovascular system: a novel fluorescence imaging method

Research output: Contribution to journalArticlepeer-review

Standard

Harvard

APA

Vancouver

Author

Bibtex

@article{1a8c081bd0a34b0ebed11a94d574a0e9,
title = "Dynamic monitoring of single-terminal norepinephrine transporter rate in the rodent cardiovascular system: a novel fluorescence imaging method",
abstract = "Here, we validate the use of a novel fluorescent norepinephrine transporter (NET) substrate for dynamic measurements of transporter function in rodent cardiovascular tissue; this technique avoids the use of radiotracers and provides single-terminal resolution.Rodent (Wistar rats and C57BL/6 mice) hearts and mesenteric arteries (MA) were isolated, loaded with NET substrate Neurotransmitter Transporter Uptake Assay (NTUA) ex vivo and imaged with confocal microscopy.NTUA labelled noradrenergic nerve terminals in all four chambers of the heart and on the surface of MA. In all tissues, a temperature-dependent, stable linear increase in intra-terminal fluorescence upon NTUA exposure was observed; this was abolished by NET inhibitor desipramine (1 μM) and reversed by indirectly-acting sympathomimetic amine tyramine (10 μM). NET reuptake rates were similar across the mouse cardiac chambers. In both species, cardiac NET activity was significantly greater than in MA (by 62 ± 29% (mouse) and 21 ± 16% (rat)). We also show that mouse NET reuptake rate was twice as fast as that in the rat (for example, in the heart, by 94 ± 30%). Finally, NET reuptake rate in the mouse heart was attenuated with muscarinic agonist carbachol (10 μM) thus demonstrating the potential for parasympathetic regulation of norepinephrine clearance.Our data provide the first demonstration of monitoring intra-terminal NET function in rodent cardiovascular tissue. This straightforward method allows dynamic measurements of transporter rate in response to varying physiological conditions and drug treatments; this offers the potential to study new mechanisms of sympathetic dysfunction associated with cardiovascular disease.",
keywords = "Norepinephrine transporter kinetics, fluorescence imaging, sympathetic, rodent, cardiovascular system, Cardiovascular system, Fluorescence imaging, Rodent, Sympathetic",
author = "Lily Cao and Andy Holmes and Janice Marshall and Larissa Fabritz and Keith Brain",
note = "Copyright {\textcopyright} 2019 The Authors. Published by Elsevier B.V. All rights reserved.",
year = "2020",
month = jan,
doi = "10.1016/j.autneu.2019.102611",
language = "English",
volume = "223",
journal = "Autonomic Neuroscience",
issn = "1566-0702",
publisher = "Elsevier",

}

RIS

TY - JOUR

T1 - Dynamic monitoring of single-terminal norepinephrine transporter rate in the rodent cardiovascular system: a novel fluorescence imaging method

AU - Cao, Lily

AU - Holmes, Andy

AU - Marshall, Janice

AU - Fabritz, Larissa

AU - Brain, Keith

N1 - Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.

PY - 2020/1

Y1 - 2020/1

N2 - Here, we validate the use of a novel fluorescent norepinephrine transporter (NET) substrate for dynamic measurements of transporter function in rodent cardiovascular tissue; this technique avoids the use of radiotracers and provides single-terminal resolution.Rodent (Wistar rats and C57BL/6 mice) hearts and mesenteric arteries (MA) were isolated, loaded with NET substrate Neurotransmitter Transporter Uptake Assay (NTUA) ex vivo and imaged with confocal microscopy.NTUA labelled noradrenergic nerve terminals in all four chambers of the heart and on the surface of MA. In all tissues, a temperature-dependent, stable linear increase in intra-terminal fluorescence upon NTUA exposure was observed; this was abolished by NET inhibitor desipramine (1 μM) and reversed by indirectly-acting sympathomimetic amine tyramine (10 μM). NET reuptake rates were similar across the mouse cardiac chambers. In both species, cardiac NET activity was significantly greater than in MA (by 62 ± 29% (mouse) and 21 ± 16% (rat)). We also show that mouse NET reuptake rate was twice as fast as that in the rat (for example, in the heart, by 94 ± 30%). Finally, NET reuptake rate in the mouse heart was attenuated with muscarinic agonist carbachol (10 μM) thus demonstrating the potential for parasympathetic regulation of norepinephrine clearance.Our data provide the first demonstration of monitoring intra-terminal NET function in rodent cardiovascular tissue. This straightforward method allows dynamic measurements of transporter rate in response to varying physiological conditions and drug treatments; this offers the potential to study new mechanisms of sympathetic dysfunction associated with cardiovascular disease.

AB - Here, we validate the use of a novel fluorescent norepinephrine transporter (NET) substrate for dynamic measurements of transporter function in rodent cardiovascular tissue; this technique avoids the use of radiotracers and provides single-terminal resolution.Rodent (Wistar rats and C57BL/6 mice) hearts and mesenteric arteries (MA) were isolated, loaded with NET substrate Neurotransmitter Transporter Uptake Assay (NTUA) ex vivo and imaged with confocal microscopy.NTUA labelled noradrenergic nerve terminals in all four chambers of the heart and on the surface of MA. In all tissues, a temperature-dependent, stable linear increase in intra-terminal fluorescence upon NTUA exposure was observed; this was abolished by NET inhibitor desipramine (1 μM) and reversed by indirectly-acting sympathomimetic amine tyramine (10 μM). NET reuptake rates were similar across the mouse cardiac chambers. In both species, cardiac NET activity was significantly greater than in MA (by 62 ± 29% (mouse) and 21 ± 16% (rat)). We also show that mouse NET reuptake rate was twice as fast as that in the rat (for example, in the heart, by 94 ± 30%). Finally, NET reuptake rate in the mouse heart was attenuated with muscarinic agonist carbachol (10 μM) thus demonstrating the potential for parasympathetic regulation of norepinephrine clearance.Our data provide the first demonstration of monitoring intra-terminal NET function in rodent cardiovascular tissue. This straightforward method allows dynamic measurements of transporter rate in response to varying physiological conditions and drug treatments; this offers the potential to study new mechanisms of sympathetic dysfunction associated with cardiovascular disease.

KW - Norepinephrine transporter kinetics

KW - fluorescence imaging

KW - sympathetic

KW - rodent

KW - cardiovascular system

KW - Cardiovascular system

KW - Fluorescence imaging

KW - Rodent

KW - Sympathetic

UR - http://www.scopus.com/inward/record.url?scp=85077770768&partnerID=8YFLogxK

U2 - 10.1016/j.autneu.2019.102611

DO - 10.1016/j.autneu.2019.102611

M3 - Article

C2 - 31901784

VL - 223

JO - Autonomic Neuroscience

JF - Autonomic Neuroscience

SN - 1566-0702

M1 - 102611

ER -