686 - Smooth muscle cell depletion of piezo1 leads to impaired contractile properties in the small bowel

Publication Number: 686.6552

Geoanna Bautista, University of California, Davis, School of Medicine, Sacramento, CA, United States; Nicole R. Kushnir, University of California, Los Angeles David Geffen School of Medicine, Tujunga, CA, United States; Allison Flores, University of California, Los Angeles David Geffen School of Medicine, Mission Hills, CA, United States; Elmira Tokhtaeva, UCLA, Los Angeles, CA, United States; Steven McElroy, University of California Davis Children's Hospital, Sacramento, CA, United States; Fernando Santana, University of California, Davis, School of Medicine, Davis, CA, United States; James Dunn, Stanford University School of Medicine, Palo Alto, CA, United States; Martin Martin, UCLA Mattel Childrens Hospital, Los Angeles, CA, United States

Background: Piezo1 is a mechanosensitive cation channel expressed in various intestinal muscularis cells including the SMCs, ICC and Pdgfrα+ cells that make up the SIP syncytium modulating GI contractility. However, the role of Piezo1 in modulating intestinal contractility remain poorly understood.

Objective: Given that SMCs are the primary effectors of gut motility, we aimed to test the hypothesis that selective depletion of Piezo1 in SMCs will result in impaired contractile force generation and tonicity with increasing stretch.

Design/Methods: Using the Piezo1/Myh11-ERT2/Cre-LoxP system, we generated mice with tamoxifen (Tam) inducible-Piezo1 knockout (Piezo1ΔSMC) in the SMCs of the small bowel muscularis. Mice at 28 days of age were given Tam or oil vehicle (control), then assessed 21 days after. Isotonic force and contractility measurements were obtained on ex vivo segments of distal small bowel from adult mice at baseline and with stretch. We further assessed the length-tension relation, viscoelastic properties, and tissue stress-strain behavior following acute stretch to steady state.

Results: Ex vivo, myograph experiments showed that loss of SMC Piezo1 led to reduced amplitude (strength) and AUC (force generation), with decreased frequency but shorter duration of contractions at baseline and following acute stretch (p < 0.001). The addition of the L-type channel inhibitor, nicardipine to Piezo1ΔSMC led to a further reduction in contractile strength to near cessation. While the addition of carbachol to Piezo1ΔSMC mice was 3-fold less than the response induced in control mice. Piezo1ΔSMC mice generated a lower passive tension, or tonicity with increasing circumference, compared to control mice (p < 0.001). Piezo1ΔSMC mice demonstrated a greater total percent reduction in maximum force and consistently lower active forces at steady state (p < 0.01). Piezo1ΔSMC mice also had a more rapid rate of relaxation, particularly in the first 5 seconds of relaxation following acute stretch. Inhibitors of enteric neuronal input (TTX and L-NNA/ODQ) did not impact the relative reduction in force generated in Piezo1ΔSMC mice.

Conclusion(s): These findings demonstrate that loss of Piezo1 in SMCs leads to impaired contractile function, force generation, viscoelastic, and stress-strain properties. This collectively suggests that there may be impairments at the smooth muscle cell level that may directly and indirectly impact the contractile properties of the bowel. Furthermore, high-resolution imaging of the muscularis suggests a unique intracellular role of Piezo1 that may be particularly relevant during the neonatal period.