FFA3 activation inhibits nicotine‐induced secretion and motility in rat proximal colon
Authors: Kaji Izumi , Akiba Yasutada , Furuyama Takafumi , Kaunitz Jonathan D 
Published: November 2023

Short‐chain fatty acids (SCFA) are microbial fermentation products that are absorbed from the colon. Although luminal SCFA stimulate colonic secretion and motility, we have recently reported that activation of a SCFA receptor termed FFA3, expressed in cholinergic nerves, suppresses cholinergic‐mediated transepithelial anion secretion. Since myenteric neurons are FFA3‐positive, we further investigated how activation of neurally‐expressed FFA3 affects colonic motility.We used the selective FFA3 agonists (N‐[2‐methylphenyl]‐[4‐furan‐3‐yl]‐2‐methyl‐5‐oxo‐1,4,5,6,7,8‐hexahydro‐quinoline‐3‐carboxamide; MQC) and AR420626 to activate neural FFA3. Short‐circuit current (Isc) was measured to determine anion secretion in Ussing‐chamber. Circular muscle contractions were measured with isolated muscle strips without mucosa and submucosa. Muscle contractions were analyzed by wave analysis according to the parameters: basal tone, peak frequency, peak force, and half‐width (HW) of peak. The effect of FFA3 agonists on defecation in vivo was investigated with an established exogenous serotonin‐induced defecation model.In Ussing chambered mucosa‐submucosa preparations, pretreatment with MQC (100 μM) or AR420626 (10 μM) suppressed the Isc increase induced by carbachol (CCh, 10 μM) or nicotine (100 μM) by 65%, but did not affect the response to bethanechol (100 μM). The inhibitory effect of FFA3 activation on CCh‐evoked Isc was reversed by pretreatment with the Gi/o inhibitor pertussis toxin (500 ng/mL).Circular muscle strips exhibited periodic spontaneous contractions that were not altered by the application of MQC or AR420626. The addition of nicotine (10 μM) into the organ bath transiently relaxed the muscle, inhibited by the NO synthase inhibitor L‐NAME. In the presence of L‐NAME and the cyclooxygenase inhibitor indomethacin, nicotine (100 μM) induced large amplitude contractions. Pretreatment with FFA3 agonists inhibited nicotine‐induced transient relaxation and contraction, but had no effect on bethanechol‐induced contractions, suggesting that FFA3 agonists had no direct effect on smooth muscle contraction. Pretreatment with tetrodotoxin did not alter the nicotine‐evoked contraction or the inhibitory effect of FFA3 agonists, whereas atropine or hexamethonium inhibited nicotine‐evoked contractions as did FFA3 agonists. The effect of AR420626 was more potent than the effect of MQC. Nicotine‐evoked contraction was abolished by AR420626 (10 μM) and reversed by pretreatment with pertussis toxin, suggesting that FFA3 activation suppresses nicotinic neural activity in the myenteric neurons, consistent with a FFA3‐mediated secretory pathway.In conscious rats, serotonin (10 mg/kg, i.p.) treatment significantly increased the volume of fecal output, compared with the non‐treated group, although frank diarrhea was not present. Pretreatment with AR420626 (0.1 mg/kg, i.p.) significantly suppressed serotonin‐induced fecal output. These results are suggesting that FFA3 agonists may be useful therapeutically for diarrheal disorders due to their highly targeted and localized anti‐cholinergic effects.Support or Funding InformationSupported by VA Merit Review and NIH R01 DK54221