Direct optogenetic stimulation of smooth muscle cells to control gastric contractility

Vogt, Markus; Schulz, Benjamin; Wagdi, Ahmed; Lebert, Jan; Belle, Gijsbert J. van; Christoph, Jan; Bruegmann, Tobias; Patejdl, Robert

doi:10.7150/thno.53883

Cited by 15 publications

(13 citation statements)

References 83 publications

(46 reference statements)

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“…Stable neurogenic contractions could be elicited over more than three hours of incubation time when sera of healthy volunteers were used. The magnitude of the obtained EFS responses was lower than high-K + or CCh contractions, the relation between the different responses being comparable to that we observed in murine fundus preparations in a recent study using the same setup and equivalent EFS-parameters 50 . The most likely explanation is that EFS-pulses, inevitably exciting all kinds of intramural nerve fibers, activate the coincident release of excitatory and inhibitory transmitters which leads to dampened response compared to purely excitatory transmitter actions exerted by CCh or direct SMC-depolarizations by K + .…”

Section: Discussionsupporting

confidence: 87%

A novel ex vivo model for critical illness neuromyopathy using freshly resected human colon smooth muscle

Patejdl

Klawitter

Walter

et al. 2021

Sci Rep

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Patients suffering from critical illness are at risk to develop critical illness neuromyopathy (CINM). The underlying pathophysiology is complex and controversial. A central question is whether soluble serum factors are involved in the pathogenesis of CINM. In this study, smooth muscle preparations obtained from the colon of patients undergoing elective surgery were used to investigate the effects of serum from critically ill patients. At the time of blood draw, CINM was assessed by clinical rating and electrophysiology. Muscle strips were incubated with serum of healthy controls or patients in organ baths and isometric force was measured. Fifteen samples from healthy controls and 98 from patients were studied. Ratios of responses to electric field stimulation (EFS) before and after incubation were 118% for serum from controls and 51% and 62% with serum from critically ill patients obtained at day 3 and 10 of critical illness, respectively (p = 0.003, One-Way-ANOVA). Responses to carbachol and high-K+ were equal between these groups. Ratios of post/pre-EFS responses correlated with less severe CINM. These results support the existence of pathogenic, i.e. neurotoxic factors in the serum of critically ill patients. Using human colon smooth muscle as a bioassay may facilitate their future molecular identification.

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Section: Discussionsupporting

confidence: 87%

A novel ex vivo model for critical illness neuromyopathy using freshly resected human colon smooth muscle

Patejdl

Klawitter

Walter

et al. 2021

Sci Rep

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“…Importantly, having an optogenetic receptor at hand which is commonly expressed in humans provides a unique opportunity for translational approaches because ectopic expression most likely does not trigger immune responses 66 , 67 . Increasing activity of G q signaling with hOPN5 expression and illumination might help to increase the efficiency of optogenetic strategies 32 , specifically those targeting smooth muscle contractility 68 , 69 . We could not detect any side effects by sole overexpression of hOPN5 in cells and organs which excludes dark activity of the receptor or alterations of the native G q signaling cascade.…”

Section: Discussionmentioning

confidence: 99%

Selective optogenetic control of Gq signaling using human Neuropsin

et al. 2022

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Gq proteins are universally important for signal transduction in mammalian cells. The underlying kinetics and transformation from extracellular stimuli into intracellular signaling, however could not be investigated in detail so far. Here we present the human Neuropsin (hOPN5) for specific and repetitive manipulation of Gq signaling in vitro and in vivo with high spatio-temporal resolution. Properties and G protein specificity of hOPN5 are characterized by UV light induced IP3 generation, Ca2+ transients and inhibition of GIRK channel activity in HEK cells. In adult hearts from a transgenic animal model, light increases the spontaneous beating rate. In addition, we demonstrate light induced contractions in the small intestine, which are not detectable after pharmacological Gq protein block. All-optical high-throughput screening for TRPC6 inhibitors is more specific and sensitive than conventional pharmacological screening. Thus, we demonstrate specific Gq signaling of hOPN5 and unveil its potential for optogenetic applications.

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“…The high processing speeds allow us to immediately assess the quality of the videos in between recordings. We previously used the Farnebäck GPU algorithm to track motion and compensate for motion artifacts in calcium imaging data of isolated stomach strips ( 36 ). The high-resolution calcium imaging data was acquired at a spatial resolution of 1, 280 × 1, 080 pixels and acquisition speeds of 50 fps using a Zyla camera (Andor, Oxford Instruments, UK).…”

Section: Resultsmentioning

confidence: 99%

“…The experimental imaging data processed and analyzed in this study consists of voltage-sensitive optical mapping videos of a mouse and a rabbit heart imaged ex vivo with a Basler acA720-520um camera (720 × 540 pixels full resolution), voltage-sensitive optical mapping videos of a rabbit heart imaged ex vivo with a Brainvision Scimedia MiCAM N256 camera (256 × 256 pixels full resolution) and an IDS μEye UI-3060CP-M-GL camera (1, 936 × 1, 216 pixels full resolution), voltage-sensitive optical mapping videos of a pig heart imaged ex vivo with a Photometrics Teledyne Evolve 128 camera (128 × 128 pixels full resolution), and calcium- and voltage-sensitive optical mapping videos of a cardiomyocyte culture, differentiated from human induced pluripotent stem cells (iPSCs), imaged in vitro with an IDS μEye UI-3060CP-M-GL camera (1, 936 × 1, 216 pixels full resolution). In addition, we discuss calcium-sensitive optical mapping data of an isolated stomach strip imaged in vitro with an Oxford Instruments Andor camera (1, 280 × 1, 080 pixels full resolution) which we analyzed in another study with our methodology ( 36 ). All tissues contract and deform strongly as they were imaged in the absence of pharmacological excitation-contraction uncoupling agents.…”

Section: Methodsmentioning

confidence: 99%

“…While brightness constancy is almost never perfectly fulfilled in most other real-life scenarios as well, during optical mapping the brightness changes can become particularly severe as they are an intended feature and central part of the measurement: the stronger the brightness changes, the better the measured signal. In previous studies, refer to review by Nesmith et al ( 31 ), the numerical tracking was either facilitated by attaching physical markers to the tissue ( 18 , 32 , 33 ) or performed entirely without markers solely tracking the tissue's natural features ( 6 , 15 , 16 , 19 , 20 , 34 – 36 ). Particularly in the latter case, when solely the fluorescing tissue is imaged, brightness changes can lead to tracking artifacts ( 16 ), and it is, therefore, critical to carefully consider these brightness changes when assessing the performance of numerical motion tracking algorithms.…”

Section: Introductionmentioning

confidence: 99%

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Real-Time Optical Mapping of Contracting Cardiac Tissues With GPU-Accelerated Numerical Motion Tracking

Lebert

Ravi

Kensah

et al. 2022

Front. Cardiovasc. Med.

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Optical mapping of action potentials or calcium transients in contracting cardiac tissues are challenging because of the severe sensitivity of the measurements to motion. The measurements rely on the accurate numerical tracking and analysis of fluorescence changes emitted by the tissue as it moves, and inaccurate or no tracking can produce motion artifacts and lead to imprecise measurements that can prohibit the analysis of the data. Recently, it was demonstrated that numerical motion-tracking and -stabilization can effectively inhibit motion artifacts, allowing highly detailed simultaneous measurements of electrophysiological phenomena and tissue mechanics. However, the field of electromechanical optical mapping is still young and under development. To date, the technique is only used by a few laboratories, the processing of the video data is time-consuming and performed offline post-acquisition as it is associated with a considerable demand for computing power. In addition, a systematic review of numerical motion tracking algorithms applicable to optical mapping data is lacking. To address these issues, we evaluated 5 open-source numerical motion-tracking algorithms implemented on a graphics processing unit (GPU) and compared their performance when tracking and compensating motion and measuring optical traces in voltage- or calcium-sensitive optical mapping videos of contracting cardiac tissues. Using GPU-accelerated numerical motion tracking, the processing times necessary to analyze optical mapping videos become substantially reduced. We demonstrate that it is possible to track and stabilize motion and create motion-compensated optical maps in real-time with low-resolution (128 x 128 pixels) and high resolution (800 x 800 pixels) optical mapping videos acquired at 500 and 40 fps, respectively. We evaluated the tracking accuracies and motion-stabilization capabilities of the GPU-based algorithms on synthetic optical mapping videos, determined their sensitivity to fluorescence signals and noise, and demonstrate the efficacy of the Farnebäck algorithm with recordings of contracting human cardiac cell cultures and beating hearts from 3 different species (mouse, rabbit, pig) imaged with 4 different high-speed cameras. GPU-accelerated processing provides a substantial increase in processing speed, which could open the path for more widespread use of numerical motion tracking and stabilization algorithms during routine optical mapping studies.

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Direct optogenetic stimulation of smooth muscle cells to control gastric contractility

Cited by 15 publications

References 83 publications

A novel ex vivo model for critical illness neuromyopathy using freshly resected human colon smooth muscle

A novel ex vivo model for critical illness neuromyopathy using freshly resected human colon smooth muscle

Selective optogenetic control of Gq signaling using human Neuropsin

Real-Time Optical Mapping of Contracting Cardiac Tissues With GPU-Accelerated Numerical Motion Tracking

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