Two distinct calcium signaling behaviors are present in interstitial cells of Cajal of the gastroesophageal junction

Abstract

Background/Objectives: The lower esophageal sphincter (LES) is located within the gastroesophageal junction (GEJ) and generates tone to prevent gastric reflux into the esophagus. Previously, we demonstrated that the generation and maintenance of tone in the LES clasp muscle involves intramuscular interstitial cells of Cajal (ICC-IM) (Drumm et al., 2022; PMID: 35229888). The aim of the current study was to further characterize Ca2+ signaling in ICC-IM within the GEJ. We hypothesize that Ca2+ release from the endoplasmic reticulum (ER) activates Ca2+-activated Cl- channels (ANO1) in ICC leading to depolarization that is conducted to smooth muscle cells (SMCs) via gap junctions; this in turn leads to Ca2+ influx via L-type Ca2+ channels (CavL) and the development of tone. Methods: Confocal spinning disc microscopy was used to image Ca2+ transients in the LES and distal esophagus of mice expressing GCaMP6f in ICC (Kit-GCaMP6f). Muscles of the GEJ imaged with the submucosal surface upwards at 20x. Ani9 (3mM), nifedipine (1mM) and CPA (10mM) were used to inhibit ANO1, CavL and SERCA, respectively. Pinacidil (10mM) was used to activate KATP channels. Results: ICC-IM within the GEJ displayed two types of Ca2+ signaling behaviors. Ca2+ transients in ICC-IM of the LES clasp muscle were stochastic, arising from multiple firing sites within the same cell. These Ca2+ transients were not synchronized within the same cell or between adjacent cells. In contrast, two Ca2+ signaling patterns were observed in ICC-IM of the distal esophagus where smooth muscle transitions to skeletal muscle. One ICC-IM population (Type I) shared the Ca2+ signaling behaviors of ICC-IM in the LES clasp. Another ICC population (Type II) exhibited rhythmic whole-cell Ca2+ transients that were synchronized between adjacent Type II ICC. Whole cell Ca2+ transients in Type II ICC were abolished by Ani9, nifedipine or pinacidil, revealing underlying Ca2+ transients similar to those observed in Type I ICC. In contrast, Ca2+ transients in Type I ICC were resistant to Ani9, nifedipine and pinacidil. Ca2+ transients were abolished in both types of ICC by removal of extracellular Ca2+, however the activity in Type I ICC persisted longer than that of Type II ICC. Discussion/Conclusions: This study demonstrates two distinct Ca2+ signaling behaviors in ICC-IM of the GEJ with differing pharmacology. This is similar to what has been previously described within the internal anal sphincter (IAS) (Hannigan et al., 2020; PMID: 32587396). In the IAS, it has been suggested that Type II ICC-IM generate slow waves however their role in the GEJ remains unclear as slow waves have not been described within this region of the mouse. Thus, the functional role of ICC-IM in the GEJ, particularly Type II ICC, warrants further investigation. NIH DK129528 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.