VasoTracker, a Low-Cost and Open Source Pressure Myograph System for Vascular Physiology

Lawton, Penelope F.; Lee, Matthew D.; Saunter, Christopher D.; Girkin, John M.; McCarron, John G.; Wilson, Calum

doi:10.3389/fphys.2019.00099

Cited by 33 publications

(25 citation statements)

References 73 publications

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“…In some experiments, Ca 2+ signals were calibrated using Ca 2þ Images were acquired at 10 or 5 Hz (consistent within each experimental protocol) using an upright fluorescence microscope equipped with a 16× objective lens (0.8 NA; Nikon, Tokyo, Japan) and large format (1,024 × 1,024; 13-μm pixels) back-illuminated electronmultiplying charge-coupled device camera (iXon 888; Andor, Belfast, UK) and stored for offline analysis. An edge-detection algorithm (Lawton et al, 2019) was used to track the width of arteries in image recordings. The algorithm extracts an intensity profile along a scanline orientated perpendicular to the longitudinal axis of the vessel.…”

Section: Extraction and Analysis Of Endothelial Ca 2+ Signalsmentioning

confidence: 99%

“…Diameter measurements of pressurized arteries were performed using the VasoTracker open source pressure myograph system (system (RRID:SCR_017233; Lawton et al, 2019). Mesenteric arteries were dissected free from surrounding tissue, cleaned of connective tissue, and mounted on similar sized glass pipettes in a VasoTracker pressure myograph chamber.…”

Section: Pressure Myographymentioning

confidence: 99%

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Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors

Heathcote

Lee

Zhang

et al. 2019

British J Pharmacology

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Background and Purpose The TRPV4 ion channels are Ca 2+ permeable, non‐selective cation channels that mediate large, but highly localized, Ca 2+ signals in the endothelium. The mechanisms that permit highly localized Ca 2+ changes to evoke cell‐wide activity are incompletely understood. Here, we tested the hypothesis that TRPV4‐mediated Ca 2+ influx activates Ca 2+ release from internal Ca 2+ stores to generate widespread effects. Experimental Approach Ca 2+ signals in large numbers (~100) of endothelial cells in intact arteries were imaged and analysed separately. Key Results Responses to the TRPV4 channel agonist GSK1016790A were heterogeneous across the endothelium. In activated cells, Ca 2+ responses comprised localized Ca 2+ changes leading to slow, persistent, global increases in Ca 2+ followed by large propagating Ca 2+ waves that moved within and between cells. To examine the mechanisms underlying each component, we developed methods to separate slow persistent Ca 2+ rise from the propagating Ca 2+ waves in each cell. TRPV4‐mediated Ca 2+ entry was required for the slow persistent global rise and propagating Ca 2+ signals. The propagating waves were inhibited by depleting internal Ca 2+ stores, inhibiting PLC or blocking IP 3 receptors. Ca 2+ release from stores was tightly controlled by TRPV4‐mediated Ca 2+ influx and ceased when influx was terminated. Furthermore, Ca 2+ release from internal stores was essential for TRPV4‐mediated control of vascular tone. Conclusions and Implications Ca 2+ influx via TRPV4 channels is amplified by Ca 2+ ‐induced Ca 2+ release acting at IP 3 receptors to generate propagating Ca 2+ waves and provide a large‐scale endothelial communication system. TRPV4‐mediated control of vascular tone requires Ca 2+ release from the internal store.

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Section: Extraction and Analysis Of Endothelial Ca 2+ Signalsmentioning

confidence: 99%

Section: Pressure Myographymentioning

confidence: 99%

Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors

Heathcote

Lee

Zhang

et al. 2019

British J Pharmacology

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“…Outside diameter recording was used simultaneously with servo-null micropressure measurement to test and validate our method; however, we also confirmed that inner diameter tracking gave comparable results. We used outside over inner diameter tracking because outside diameter tracking can easily be achieved using various already available, open-source tools: VasoTracker (Lawton et al 2019), ImageJ, etc. In addition to reporting novel findings on valve function/dysfunction in lymphatic valves from Foxc2 Cre/+ and Prox1-CreER T2 ;Cx43 fx/fx ;Cx37 −/− mice, the significance of this work comes in making available a more accessible technique for quantitative measurement of lymphatic valve function.…”

Section: Discussionmentioning

confidence: 99%

Simplified method to quantify valve back‐leak uncovers severe mesenteric lymphatic valve dysfunction in mice deficient in connexins 43 and 37

Castorena‐Gonzalez

Srinivasan

King

et al. 2020

The Journal of Physiology

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Key points Lymphatic valve defects are one of the major causes of lymph transport dysfunction; however, there are no accessible methods for quantitatively assessing valve function. This report describes a novel technique for quantifying lymphatic valve back‐leak. Postnatal endothelial‐specific deletion of connexin 43 (Cx43) in connexin 37 null (Cx37−/−) mice results in rapid regression of valve leaflets and severe valve dysfunction. This method can also be used for assessing the function of venous and lymphatic valves from various species, including humans. Abstract The lymphatic system relies on robust, spontaneous contractions of collecting lymphatic vessels and one‐way secondary lymphatic valves to efficiently move lymph forward. Secondary valves prevent reflux and allow for the generation of propulsive pressure during each contraction cycle. Lymphatic valve defects are one of the major causes of lymph transport dysfunction. Genetic mutations in multiple genes have been associated with the development of primary lymphoedema in humans; and many of the same mutations in mice result in valve defects that subsequently lead to chylous ascites or chylothorax. At present the only experimental technique for the quantitative assessment of lymphatic valve function utilizes the servo‐null micropressure system, which is highly accurate and precise, but relatively inaccessible and difficult to use. We developed a novel, simplified alternative method for quantifying valve function and determining the degree of pressure back‐leak through an intact valve in pressurized, single‐valve segments of isolated lymphatic vessels. With this diameter‐based method, the competence of each lymphatic valve is challenged over a physiological range of pressures (e.g. 0.5–10cmH2O) and pressure back‐leak is extrapolated from calibrated, pressure‐driven changes in diameter upstream from the valve. Using mesenteric lymphatic vessels from C57BL/6J, Ub‐CreERT2;Rasa1fx/fx, Foxc2Cre/+, Lyve1‐Cre;Cx43fx/fx, and Prox1‐CreERT2;Cx43fx/fx;Cx37−/− mice, we tested our method on lymphatic valves displaying a wide range of dysfunction, from fully competent to completely incompetent. Our results were validated by simultaneous direct measurement of pressure back‐leak using a servo‐null micropressure system. Our diameter‐based technique can be used to quantify valve function in isolated lymphatic valves from a variety of species. This method also revealed that haplodeficiency in Foxc2 (Foxc2Cre/+) is not sufficient to cause significant valve dysfunction; however, postnatal endothelial‐specific deletion of Cx43 in Cx37−/− mice results in rapid regression of valve leaflets and severe valve dysfunction.

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“…First-order mesenteric vessels were selected as a widely used model of vascular and endothelial function since there is a wealth of published background information to aid experimental design and data interpretation. For pressure myograph experiments, we used the open source pressure myograph system, VasoTracker [49] and recorded outer vessel diameter in response to various treatments at 70 mmHg and 37°C. For Ca 2+ imaging experiments, arteries were opened longitudinally, and endothelial cells were preferentially loaded with the Ca 2+ indicator, Cal-520/AM (5 μM).…”

Section: Experimental Techniquesmentioning

confidence: 99%

Disrupted endothelial cell heterogeneity and network organization impair vascular function in prediabetic obesity

et al. 2020

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Background: Obesity is a major risk factor for diabetes and cardiovascular diseases such as hypertension, heart failure, and stroke. Impaired endothelial function occurs in the earliest stages of obesity and underlies vascular alterations that give rise to cardiovascular disease. However, the mechanisms that link weight gain to endothelial dysfunction are ill-defined. Increasing evidence suggests that endothelial cells are not a population of uniform cells but are highly heterogeneous and are organized as a communicating multicellular network that controls vascular function. Purpose: To investigate the hypothesis that disrupted endothelial heterogeneity and network-level organization contribute to impaired vascular reactivity in obesity. Methods and results: To study obesity-related vascular function without complications associated with diabetes, a state of prediabetic obesity was induced in rats. Small artery diameter recordings confirmed nitric-oxide mediated vasodilator responses were dependent on increases in endothelial calcium levels and were impaired in obese animals. Single-photon imaging revealed a linear relationship between blood vessel relaxation and population-wide calcium responses. Obesity did not alter the slope of this relationship, but impaired calcium responses in the endothelial cell network. The network comprised structural and functional components. The structural architecture, a hexagonal lattice network of connected cells, was unchanged in obesity. The functional network contained sub-populations of clustered specialized agonist-sensing cells from which signals were communicated through the network. In obesity there were fewer but larger clusters of sensory cells and communication path lengths between clusters increased. Communication between neighboring cells was unaltered in obesity. Altered network organization resulted in impaired, population-level calcium signaling and deficient endothelial control of vascular tone. Conclusions: The distribution of cells in the endothelial network is critical in determining overall vascular response. Altered cell heterogeneity and arrangement in obesity decreases endothelial function and provides a novel framework for understanding compromised endothelial function in cardiovascular disease.

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VasoTracker, a Low-Cost and Open Source Pressure Myograph System for Vascular Physiology

Cited by 33 publications

References 73 publications

Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors

Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors

Simplified method to quantify valve back‐leak uncovers severe mesenteric lymphatic valve dysfunction in mice deficient in connexins 43 and 37

Disrupted endothelial cell heterogeneity and network organization impair vascular function in prediabetic obesity

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VasoTracker, a Low-Cost and Open Source Pressure Myograph System for Vascular Physiology

Cited by 33 publications

References 73 publications

Endothelial TRPV4 channels modulate vascular tone by Ca2+‐induced Ca2+ release at inositol 1,4,5‐trisphosphate receptors

Endothelial TRPV4 channels modulate vascular tone by Ca2+‐induced Ca2+ release at inositol 1,4,5‐trisphosphate receptors

Simplified method to quantify valve back‐leak uncovers severe mesenteric lymphatic valve dysfunction in mice deficient in connexins 43 and 37

Disrupted endothelial cell heterogeneity and network organization impair vascular function in prediabetic obesity

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Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors

Endothelial TRPV4 channels modulate vascular tone by Ca²⁺‐induced Ca²⁺ release at inositol 1,4,5‐trisphosphate receptors