Peripheral peptide YY inhibits propulsive colonic motor function through Y2 receptor in conscious mice

Abstract

Peptide YY (PYY) antisecretory effect on intestinal epithelia is well established, whereas less is known about its actions to influence colonic motility in conscious animals. We characterized changes in basal function and stimulated colonic motor function induced by PYY-related peptides in conscious mice. PYY(3-36), PYY, and neuropeptide Y (NPY) (8 nmol/kg) injected intraperitoneally inhibited fecal pellet output (FPO) per hour during novel environment stress by 90%, 63%, and 57%, respectively, whereas the Y(1)-preferring agonists, [Pro(34)]PYY and [Leu(31),Pro(34)]NPY, had no effect. Corticotrophin-releasing factor 2 receptor antagonist did not alter PYY(3-36) inhibitory action. PYY and PYY(3-36) significantly reduced restraint-stimulated defecation, and PYY(3-36) inhibited high-amplitude distal colonic contractions in restrained conscious mice for 1 h, by intraluminal pressure with the use of a microtransducer. PYY suppression of intraperitoneal 5-hydroxytryptophan induced FPO and diarrhea was blocked by the Y(2) antagonist, BIIE0246, injected intraperitoneally and mimicked by PYY(3-36), but not [Leu(31),Pro(34)]NPY. PYY(3-36) also inhibited bethanechol-stimulated FPO and diarrhea. PYY(3-36) inhibited basal FPO during nocturnal feeding period and light phase in fasted/refed mice for 2-3 h, whereas the reduction of food intake lasted for only 1 h. PYY(3-36) delayed gastric emptying after fasting-refeeding by 48% and distal colonic transit time by 104%, whereas [Leu(31),Pro(34)]NPY had no effect. In the proximal and distal colon, higher Y(2) mRNA expression was detected in the mucosa than in muscle layers, and Y(2) immunoreactivity was located in nerve terminals around myenteric neurons. These data established that PYY/PYY(3-36) potently inhibits basal and stress/serotonin/cholinergic-stimulated propulsive colonic motor function in conscious mice, likely via Y(2) receptors.

Fig. 1.

Intraperitoneal peptide YY (PYY) and PYY_3–36 inhibit fecal pellet output (FPO) response to novelty environmental stress for 1 h in mice. A: novelty increased defecation in naive mice (n = 16) exposed singly to a novel environment (box) compared with trained mice (n = 16) accustomed singly to the box for 1 h/day for 3–4 days prior. B: dose-response of PYY_3–36 on novelty stress-induced defecation. PYY_3–36 was injected intraperitoneally 10 min before the stress (n = 7–8/group). C: effect of intraperitoneal injection of neuropeptide Y (NPY)/PYY agonists with different Y receptor selectivity on novelty stress-induced defecation. Peptides (8.0 nmol/kg ip) or saline were injected 10 min before the stress. FPO was monitored every 15 min for 1 h. Data are means ± SE of n = 7–13/group. *P < 0.05 vs. saline at the corresponding time.

Fig. 2.

Corticotrophin-releasing factor 2 is not involved in PYY_3–36 inhibition of 1-h novelty stress-induced defecation. Astressin₂-B (Ast₂B, 30 μg/kg ip) or vehicle was injected 10 min before intraperitoneal injection of PYY_3–36 (8 nmol/kg) or saline, and mice were exposed to novelty stress 10 min later. Data are means ± SE of n = 6–9/group. *P < 0.05 vs. vehicle-saline; #P < 0.05 vs. Ast₂B-saline at the corresponding time.

Fig. 3.

Intraperitoneal PYY and PYY_3–36 inhibit 1-h restraint stress-induced FPO in mice. Peptides (8 nmol/kg) and saline (Sal) were injected 10 min before restraint stress (RS) or nonstress (NS) (mice were placed in the monitor box for 1 h, in which they were acquainted). Data are means ± SE of n = 7–9/group. *P < 0.05 vs. Sal-NS; #P < 0.05 vs. PYY-NS or PYY_3–36-NS.

Fig. 4.

Intraperitoneal PYY_3–36 reduces distal colonic intraluminal pressure using a noninvasive miniature pressure transducer inserted into the distal colon in restrained conscious mice. After intraperitoneal injection of PYY_3–36 (8 nmol/kg, n = 9) or saline (n = 6), mice were briefly anesthetized for insertion of the pressure transducer, and distal intracolonic pressure was recorded 10 min later for 1 h. A: representative raw trace during the 1-h recording in mice injected with intraperitoneal saline and PYY_3–36. B: time course of the phasic component of intraluminal pressure trace of area under curve (pAUC; mmHg × min) over 1-h recording. C: mean of distal colonic contractions (number/h) with high amplitude (>25 mmHg) in the first 15 min and in 15–60 min. D: mean of distal colonic contractions (number/h) with low amplitude (10–25 mmHg) in the first 15 min and in 15–60 min. F: correlation between 1-h distal colonic contractions (number/h) with high amplitude (>25 mmHg) and FPO. Data are means ± SE. *P < 0.05 vs. saline; #P < 0.05 vs. 0–15 min of the same treatement.

Fig. 5.

Intraperitoneal PYY-induced suppression of colonic secretory motor response to 5-hydroxytryptophan (5-HTP) through Y₂ receptor in conscious mice. A: 30-min FPO after PYY, Y₁ agonist, [Leu31,Pro34]NPY, and Y₂ agonist, PYY_3–36 (8 nmol/kg), or saline were injected intraperitoneally 10 min before 5-HTP (10 mg/kg ip); n = 7–10/group. *P < 0.05 vs. saline-saline; #P < 0.05 vs. saline-5-HTP. B: diarrhea score from the same mice in A. *P < 0.05 vs. saline-saline; #P < 0.05 vs. saline-5-HTP. C: PYY-induced suppression of 30-min stimulated defecation induced by intraperitoneal 5-HTP in mice, with reversal by the Y₂ antagonist, BIIE0246. BIIE0246 (5 mg/kg) or vehicle was injected intraperitoneally at 20 min before and PYY_3–36 (8 nmol/kg) 10 min before 5-HTP (10 mg/kg) or saline in mice (n = 6–10/group). Data are means ± SE *P < 0.05 vs. vehicle-saline-5-HTP; #P < 0.05 vs. vehicle-saline-5-HTP.

Fig. 6.

Dose-related action of intraperitoneal PYY_3–36 on bethanechol-induced stimulation of FPO (A) and diarrhea (B) in mice. PYY_3–36 (8 and 25 nmol/kg) or saline was injected intraperitoneally 10 min before intraperitoneal bethanechol chloride (5 mg/kg) or saline. FPO was monitored every 15 min for 1 h. Data are means ± SE of n = 7/group. *P < 0.05 vs. saline-saline; ^#P < 0.05 vs. saline-bethanechol.

Fig. 7.

Time course of PYY_3–36-induced inhibition of food intake (A and B) and fecal output (C and D) monitored simultaneously hourly for 4 h in fasted/refed mice (A and C) or in response to dark phase in freely fed mice (B and D). Data are means ± SE of n = 8/group. *P < 0.05 vs. saline.

Fig. 8.

Intraperitoneal PYY_3–36 inhibits gastric emptying of solid food (A), distal colonic transit (B), and defecation (C) monitored simultaneously in fasted/refed mice. Peptides, Y₁ agonist [Leu³¹,Pro³⁴]NPY (Y₁), PYY_3–36 (8 nmol/kg), or saline was injected intraperitoneally after 1-h refeeding in mice fasted overnight. Data are means ± SE of n = 7–8/group; *P < 0.05 vs. saline.

Fig. 9.

Expression of Y₂ receptor at the gene and protein levels in proximal and distal colon of naive mice. A: representative gel images of Y₂ mRNA (423 bp) detected by RT-PCR in both proximal colon (pC) and distal colon (dC) of mice. S16 is the housekeeping gene (309 bp). B: semiquantitative analysis of Y₂ mRNA gel bands performed by NIH imaging. M, mucosal layer; S, layers including smooth muscles and enteric plexi; n = 4; *P < 0.05 vs. pC-M and dC-M, respectively; #P < 0.05 vs. pC-S. C: primary antibody control by preabsorption with immunogenic peptide in myenteric plexus of the distal colon. D and E: representative photomicrographs of Y₂ immunoreactivity in the distal colon submucosal (D) and myenteric (E) plexus. Scale bar = 50 μm.