Ca2+ transients in ICC-MY define the basis for the dominance of the corpus in gastric pacemaking

Changes in interstitial cells and gastric excitability in a mouse model of sleeve gastrectomy

“…We also recorded an additional site (site #3) between site#1 and site#2 in imaging experiments ( Fig 9 ). To reduce motion artifacts from gastric contractions, the L-type calcium channel, nicardipine (1 μM), was used throughout the imaging experiments as previously described [ 38 ]. Under control conditions, gastric antral muscles from sham-operated mice generated subcellular Ca 2+ transients that originated from numerous distinct firing sites in ICC-MY networks.…”

“…Male C57BL mice, aged 8 weeks, and with a body weight between 28–30 grams were used for VSG. For Ca 2+ -imaging experiments Kit CreERT2Ejb1 /+ ( Kit CreERT2 ) mice were obtained from Dr Dieter Saur (Technical University Munich, Germany) [ 37 ], and crossed with GCaMP6f-floxed mice to generate an inducible Cre strain as previously described [ 38 ]. Inducible Cre mice were injected with tamoxifen (Tam; intraperitoneal injection) at 8 weeks of age (2 mg of TAM for three consecutive days), to induce activation of the Cre recombinase and expression of the cell-specific (ICC) GCaMP6f Ca 2+ -sensor.…”

“…Preparations were then visualized and imaged using a spinning-disk confocal microscope (CSU-W1 spinning disk; Yokogawa Electric Corporation, Tokyo, Japan) mounted to an upright Eclipse FN1 microscope equipped with 40X 0.8 NA CFI Fluor lens (Nikon Instruments Inc., NY, USA). Image sequences and movies of Ca 2+ activity were collected at 33 fps using MetaMorph acquisition software (Molecular devices, CA, USA) as previously described [ 38 ]. Movies were converted to stacked TIFF images (tagged image file format), and image processing and analysis were done using a custom software (Volumetry G8a, G.W.H.).…”

New open-source software for subcellular segmentation and analysis of spatiotemporal fluorescence signals using deep learning

“…GCaMP6f-floxed mice (B6;129S- Gt(ROSA)26Sor tm95.1(CAG-GCaMP6f)Hze /J) were acquired from Jackson Laboratories (Bar Harbor, MN, USA) and crossed with Kit-Cre mice (c-Kit +/Cre−ERT2 ), provided by Dr. Dieter Saur (Technical University Munich, Munich, Germany). Kit-Cre-GCaMP6f mice were injected with tamoxifen at 6–8 weeks of age (2 mg for three consecutive days), as previously described ( Baker et al., 2021a , 2021b ). 15 days after tamoxifen injection, Kit-Cre-GCaMP6f mice were anaesthetized by isoflurane inhalation (Baxter, Deerfield, IL, USA) and sacrificed by cervical dislocation.…”

“…Ca 2+ STMaps are widely used for the analysis and quantification of voltage and Ca 2+ dynamics parameters that retain Ca 2+ event information as a function of space occupied over time ( Cheng et al., 1996 ; Colman et al., 2017 ; Hennig et al., 1999 ; Lee et al., 2009 ; Lentle and Hulls, 2018 ; Roome and Kuhn, 2018 ; Waadt et al., 2017 ). As a result, STMaps can provide a platform to effectively extract quantifiable cellular Ca 2+ data ( Baker et al., 2021a , 2021b ; Drumm et al., 2014 ; Fedigan et al., 2017 ; Sancho et al., 2017 ; Sergeant et al., 2006 ). Extracting dynamic fluorescent data can be a challenging task, as the information is most often manually defined using single-line pixel measurement or region of interest (ROI).…”