Abstract
Interstitial cells of Cajal (ICC) are pacemaker cells that generate electrical slow waves in gastrointestinal (GI) smooth muscles. Slow waves organize basic motor patterns, such as peristalsis and segmentation in the GI tract. Slow waves depend upon activation of Ca2+-activated Cl– channels (CaCC) encoded by Ano1. Slow waves consist of an upstroke depolarization and a sustained plateau potential that is the main factor leading to excitation-contraction coupling. The plateau phase can last for seconds in some regions of the GI tract. How elevated Ca2+ is maintained throughout the duration of slow waves, which is necessary for sustained activation of CaCC, is unknown. Modeling has suggested a role for Na+/Ca2+ exchanger (NCX) in regulating CaCC currents in ICC, so we tested this idea on murine intestinal ICC. ICC of small and large intestine express NCX isoforms. NCX3 is closely associated with ANO1 in ICC, as shown by immunoprecipitation and proximity ligation assays (PLA). KB-R7943, an inhibitor of NCX, increased CaCC current in ICC, suggesting that NCX, acting in Ca2+ exit mode, helps to regulate basal [Ca2+]i in these cells. Shifting NCX into Ca2+ entry mode by replacing extracellular Na+ with Li+ increased spontaneous transient inward currents (STICs), due to activation of CaCC. Stepping ICC from −80 to −40 mV activated slow wave currents that were reduced in amplitude and duration by NCX inhibitors, KB-R7943 and SN-6, and enhanced by increasing the NCX driving force. SN-6 reduced the duration of clustered Ca2+ transients that underlie the activation of CaCC and the plateau phase of slow waves. Our results suggest that NCX participates in slow waves as modeling has predicted. Dynamic changes in membrane potential and ionic gradients during slow waves appear to flip the directionality of NCX, facilitating removal of Ca2+ during the inter-slow wave interval and providing Ca2+ for sustained activation of ANO1 during the slow wave plateau phase.
Highlights
Interstitial cells of Cajal provide the pacemaker activity responsible for electrical slow waves in gastrointestinal (GI) muscles (Ward et al, 1994; Huizinga et al, 1995; Dickens et al, 1999; Zhu et al, 2009)
We found that NCX3 is associated with ANO1 channels, and we evaluated the function of Na+/Ca2+ exchanger (NCX) in activating and sustaining activation of Ca2+-activated Cl− channels (CaCC) during slow wave currents using the patch clamp techniques on single Interstitial cells of Cajal (ICC) and imaging of Ca2+ transients in intact networks of pacemaker ICC
We showed that ICC collected by fluorescence-activated cell sorting (FACS), as performed in this study, showed minimal expression of Pdgfra, Myh11 and Uchl1 and elevated levels of Kit transcripts (ICC marker) relative to the unsorted cells, suggesting that sorted ICC were highly purified by FACS (Peri et al, 2013)
Summary
Interstitial cells of Cajal provide the pacemaker activity responsible for electrical slow waves in gastrointestinal (GI) muscles (Ward et al, 1994; Huizinga et al, 1995; Dickens et al, 1999; Zhu et al, 2009). A major conductance responsible for slow wave currents in ICC is due to Ca2+-activated Cl− channels (CaCC) encoded by Ano (Hirst et al, 2002; Hwang et al, 2009; Zhu et al, 2009). The plateau phase of slow waves has been attributed to clusters of Ca2+ transients in ICC (Drumm et al, 2017) that cause sustained activation of CaCC (Hirst et al, 2002; Kito and Suzuki, 2003), but the mechanism(s) responsible for sustaining Ca2+ release events during the plateau phase are unknown
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
