Epithelial Mechanosensitive Ion Channel Piezo2 Contributes to Pressure‐Induced Epithelial Chloride Secretion in Mouse Colon

Abstract

BACKGROUNDMechanical stimulation of the gastrointestinal (GI) epithelium causes changes in epithelial ion transport by an unclear mechanism. We recently discovered the mechanosensitive ion channel Piezo2 is specifically expressed by the enteroendocrine cells (EECs) and is necessary for EEC mechanosensitivity. Further, in mouse small bowel, the pharmacologic inhibition or genetic knockout of the epithelial Piezo2 led to an inhibition of the pressure‐induced short‐circuit current (Isc) in the Ussing chamber. These results suggested that EEC Piezo2 plays a role in pressure‐induced epithelial ion transport, but the mechanism and whether these responses are region‐specific remains uknown.AIMDetermine whether epithelial Piezo2‐mediated pressure‐induced Isc is due to Clsecretion in the mouse colon.METHODSFull‐thickness mid‐colon preparations of VilCre/+::Piezo2f/f (Piezo2CKO) or Vil+/+::Piezo2f/f (Piezo2WT) mice at 0.3 cm2 were mounted in 4mL Ussing chambers with normal Krebs solution. The epithelial side was intermittently pressure‐clamped at a range of pressures (5, 10, 20, 30, 40, 60 mmHg) for 10s each. Forskolin (FSK, 10μM) was applied at the end of the experiments, and FSK response was used to normalize the pressure‐induced responses (ΔIsc/ΔIsc,FSK).RESULTSPressure applied to the mucosa of Piezo2CKO and Piezo2WTcolons caused a pressure‐dependent increase in Isc. Responses in Piezo2CKO colons were smaller than Piezo2WT at 40mmHg (0.41±0.1 vs. 0.76±0.1) and 60mmHg (0.51±0.1 vs. 0.87±0.1) (n=9–11, P<0.05, two‐way ANOVA). To determine the nature of the pressure‐induced Isc we removed Cl− from the Krebs solutions. Cl‐removal resulted in XX. We found a nearloss in the mechanically induced ΔIsc in both Piezo2WT tissues and Piezo2CKO: 5mmHg (− 88.85±0.1% and −87.44±0.1% Piezo2WT), 10mmHg (−86.46±0.1% Piezo2CKO and − 90.30±0.1% Piezo2WT), 20mmHg (−88.50±0.01% Piezo2CKO and −89.86±0.1% Piezo2WT), 40mmHg (−91.78±0.1% Piezo2CKO and −91.35±0.04% Piezo2WT), and 60mmHg (−93.93±0.04% Piezo2CKO and −92.08±0.04% Piezo2WT (n= 5–10, P <0.05, unpaired t‐test). Replacement of Cl− led to full recovery of the mechanically induced ΔIsc in both Piezo2WT and Piezo2CKO tissues when tested at 30 mmHg (Piezo2WT: 2.51±1.4 vs. 73.52±33.8, n=6 and 3, P <0.05, unpaired t‐test), (Piezo2CKO: 1.80±1.0 vs. 45.14±12.4, n=5 and 4, P <0.05, unpaired t‐test).CONCLUSIONSHydrostatic pressure applied to the colon epithelium leads to an increase in epithelial secretion that depends chloride and Piezo2 contributes to this mechanosensitive chloride secretion.Support or Funding InformationSupported by AGA RSA, NIH DK106456, NIH DK119683, NIH DK123549