Colonic motility is the product of myogenic, neurogenic and endocrinogenic factors that regulate the excitability of smooth muscle cells (SMCs). The traditional concept of ?myogenic? has been revised to include regulatory inputs from interstitial cells (interstitial cells of Cajal (ICC) and PDGFRa+ cells). These cells are electrically coupled to SMCs, forming the SIP syncytium. While much has been learned about ICC in other organs of the GI tract, very little is known about the 4 types of ICC in colonic muscles. We developed reporter strains and mice with exclusive expression of optogenetic Ca2+ sensors in ICC. Using these mice it is possible to isolate specific types of ICC for physiological and molecular studies and to image Ca2+ transients in ICC in situ. Our preliminary data show that all ICC in the colon employ brief Ca2+ entry and release events (i.e. Ca2+ transients) to activate Ca2+-activated Cl- channels (CaCC), encoded by Ano1, in ICC. Activation of CaCC initiates inward current, and this has a depolarizing or excitatory impact on the SIP syncytium. Ca2+ transients occur spontaneously in colonic ICC, but they are also regulated by neural and hormonal inputs. Our overarching hypothesis is that Ca2+ transients in ICC are the ?myogenic? mechanism that establishes patterning of contractions in colonic muscles. Neural and hormonal modulation of ICC Ca2+ transients establish organ level mixing and propulsive motility. We will address the following questions to investigate this hypothesis: 1. What specific Ca2+ handling behaviors are manifest in the 4 classes of colonic ICC? 2. Is the behavior of ICC affected by neural and hormonal inputs and during colonic motility behaviors such as colonic migrating motor complexes (CMMC)? 3. What is the relationship between Ca2+ transients in ICC and the electrical and mechanical events that generate colonic motility? Ca2+ transients in ICC of GCaMP6f-Kit mice will be imaged in situ using confocal microscopy while recording movements and intracellular electrical activity. Preliminary data show that basal electrical and contractile patterning in colonic muscles are disrupted by ANO1 channel antagonists. These data demonstrate the key importance of ICC in colonic motor activity, because ICC express ANO1 exclusively in GI muscles. Colonic dysmotilities have been associated with reduced populations of ICC. Therefore, we will utilize animal models with reduced ICC to explore how deficiencies in these cells affect local propagating and mixing contractions and propulsive contractions, such as CMMC. Ca2+ transients will also be characterized in animals with reduced ICC to determine how loss of these cells affects spontaneous Ca2+ signaling and neural and hormonal regulation. This study will be the first comprehensive evaluation of the pacemaker activity of colonic ICC, and the first to show that ICC are responsible for developing basal motor patterning in colonic motility. Completion of the specific aims will provide understanding of why ICC loss has deleterious effects on colonic motility and provide new techniques for evaluating the heath/function of specific types of ICC and the efficacy of therapeutic agents designed to improve ICC function in colonic muscles.

Public Health Relevance

Intrinsic and extrinsic nerves organize appropriate movements of the GI tract, however the patterning of motility begins with intrinsic myogenic behaviors. We are investigating the role of interstitial cells of Cajal (ICC) that generate pacemaker activity and transduce responses to motor neurons. Spontaneous and stimulated release of Ca2+ is fundamental to the behavior of ICC because increases in Ca2+ activate Cl- channels and initiates the basic patterns of contractile activity of colonic muscles. All contractile patterns are blocked by inhibitors of the Cl- channels that are expressed exclusively in ICC, demonstrating the gateway functions of ICC in organizing colonic motility. Mechanisms responsible for Ca2+ transients in colonic ICC and regulation of these events by enteric motor neurons will be determined. Loss of ICC has been associated with major motility disorders of the colon, and this project will shed light on why colonic motility becomes defective when ICC are disrupted.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK120759-03
Application #
10130518
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Shea-Donohue, Terez
Project Start
2019-03-01
Project End
2023-02-28
Budget Start
2021-03-01
Budget End
2022-02-28
Support Year
3
Fiscal Year
2021
Total Cost
Indirect Cost
Name
University of Nevada Reno
Department
Physiology
Type
Schools of Medicine
DUNS #
146515460
City
Reno
State
NV
Country
United States
Zip Code
89557