Transient Receptor Potential Melastatin 4 Channel Regulation by Inositol Trisphosphate Receptor in Human Urinary Bladder Smooth Muscle

Abstract

Overactive bladder (OAB) is a highly prevalent and debilitating condition that lacks an effective therapeutic treatment. The contraction and relaxation of urinary bladder smooth muscle (UBSM) facilitates the voiding and storage of urine, the two main functions of the urinary bladder. Some forms of OAB have been directly linked to UBSM dysfunction. Thus, efforts to uncover unknown regulatory mechanisms in UBSM function are urgently needed. Among the potential novel targets for OAB, emerging evidence points to key roles for transient receptor potential melastatin 4 (TRPM4) channels in UBSM excitability and contractility. However, the cellular mechanisms regulating TRPM4 channel activity in human UBSM has been virtually unexplored. As a potential cellular mechanism regulating TRPM4 channel activity in UBSM, we have directed our attention to the inositol trisphosphate (IP3) receptors (IP3Rs), which facilitate the release of Ca2+ from the sarcoplasmic reticulum (SR). Since the TRPM4 channels are activated by Ca2+, IP3R‐mediated Ca2+ release of the SR represents a potential Ca2+ source for TRPM4 channel activation. In accordance with the approved IRB Pro00045232 of the Medical University of South Carolina, we used clinically‐characterized human UBSM tissues from 6 donor patients to investigate the molecular and functional interactions of the TRPM4 channels and IP3Rs in human UBSM. Using in situ proximity ligation assay (PLA) and amphotericin‐B perforated patch‐clamp electrophysiology, we sought to test the hypothesis that TRPM4 channels are tightly associated with the IP3Rs and are activated by IP3R‐mediated Ca2+ release of the SR in human UBSM. As determined by in situ PLA, we demonstrated colocalization of the TRPM4 channels and IP3Rs in human UBSM cells. As the TRPM4 channels and IP3Rs must be located within close apposition to functionally interact, these findings support the concept of a TRPM4‐IP3R regulatory mechanism in human UBSM function. To investigate IP3R regulation of TRPM4 channel activity, we sought to determine the consequences of IP3R pharmacological inhibition on TRPM4 channel‐mediated transient inward cation currents (TICCs). In freshly‐isolated human UBSM cells, the selective IP3R inhibitor xestospongin C significantly decreased TICCs. These findings suggest that the SR IP3Rs have a key role in mediating the Ca2+‐dependent activation of TRPM4 channels in human UBSM. In conclusion, this study provides novel insight into the cellular mechanisms regulating TRPM4 channel activity in human UBSM function. Our study reveals that the TRPM4 channels and IP3Rs are spatially and functionally coupled in human UBSM, information that is critical for further evaluating the potential role of the TRPM4 channels as novel therapeutic targets for OAB.Support or Funding InformationSupported by NIH grant R01‐DK106964 to Georgi V. Petkov and F31‐DK104528 to Aaron Provence.