The effect of motilin on the rectum in healthy volunteers

Abstract

Aims

The role of motilin in the regulation of upper gastrointestinal (GI) motility is well defined. However, little is known about the effects on the distal GI tract. To investigate the effect of exogenous motilin on rectal function, barostat measurements in the rectum were performed and lower abdominal symptoms were scored.

Methods

Eight fasted, healthy volunteers were infused intravenously with synthetic motilin or placebo over 90 min in a double-blind, randomized, cross-over design. Rectum volume was measured with a barostat device during constant pressure and during isobaric distensions. Lower abdominal symptoms were scored by visual analogue scales. Plasma motilin concentrations were measured by radioimmunoassay.

Results

Baseline rectum volumes were similar between treatments: 185 ± 62 mL (motilin) and 136 ± 41 mL (placebo). During the constant pressure procedure, motilin increased rectum volume [area under the effect curve (AUEC)] by 6%[95% confidence interval (CI) −3, 16] of baseline, compared with placebo. During isobaric distensions motilin increased rectum volume (AUEC) by 43 mL (95% CI 0.4, 85; P < 0.05) and compliance by 10 mL mmHg⁻¹ (95% CI 0.3, 20; P < 0.05) relative to placebo. Motilin did not induce changes in the sensation of rectal feelings.

Conclusion

Exogenous motilin increased rectal compliance in healthy volunteers, without affecting rectal sensations.

Introduction

Motilin is a 22-amino acid peptide that is produced and released by enterochromaffin cells in the proximal small intestine [1]. It is well known that motilin stimulates proximal gastrointestinal (GI) motility [2, 3]. However, limited information is available about the effect of motilin on the distal GI tract. It has been reported that motilin receptors are present in human colon [4]. Previous in vitro experiments on the rabbit colon have shown the stimulatory response to motilin and the motilin agonist, erythromycin [5, 6]. In vitro studies have also shown that motilin and erythromycin induce contractions of smooth muscle strips in humans [7, 8]. In addition, in vivo, motilin and erythromycin increase the motor activity of the human colon [9]. At present, no knowledge is available on the presence of motilin receptors in the human rectum. The role of motilin on rectum motility in humans is not clear as the reported stimulatory effects of motilin on the rectum result from in vitro animal research only [6].

Irritable bowel syndrome (IBS) is a functional bowel disorder in which abdominal discomfort or pain is associated with features of disordered defecation or a change in bowel habit [10]. As alterations in motilin levels are present in IBS patients, it has been suggested that disturbances in the regulatory function of motilin may be associated with this ill-defined clinical entity [11–13].

In the light of potential future therapeutic applications of motilin agonists or antagonists in lower GI motility disorders such as IBS, a study was performed to assess the effects of motilin on the distal GI tract in humans.

This study investigated the effects of motilin on rectal volume during constant pressure and rectal compliance during isobaric distensions. The rectum was selected for barostat measurements, as this is a suitable and relatively noninvasive approach to measure distal GI function and provides an initial idea on the role of motilin in distal GI function in humans.

Methods

Experimental design

The study was carried out using a double-blind, randomized, placebo-controlled, two-way, cross-over design. The Medical Ethics Committee of the Leiden University Medical Centre (LUMC) approved the study protocol. The study was performed according to Good Clinical Practice and International Conference of Harmonization guidelines.

Subjects

Eight healthy volunteers (6F/2M; mean age 29 years; range 21–43 years) participated in the study after having given written informed consent. Subjects were eligible if they were healthy assessed by a medical screening. They were excluded if they had a history of GI symptoms, abdominal surgery, used medication or had positive Helicobacter pylori serology.

Barostat

An electronic barostat device (Synectics Visceral Stimulator; Synectics Medical, Stockholm, Sweden) was used to measure volume changes in the rectum. A polyethylene bag (1000 mL maximum) was tied to the end of a 19-French multilumen catheter. The catheter was connected to a barostat device [14]. Pressure (mmHg) and volume (mL) were constantly monitored and recorded on a personal computer with dedicated software that corrects volume for air compressibility and temperature (Polygram for Windows, SVS module; Synectics Medical).

Study days

Each subject participated on two study days, separated by at least 7 days, during which placebo (NaCl 0.9%) or motilin (4 pmol kg⁻¹ min⁻¹) was administered intravenously over 90 min. The motilin used in this study was synthetic human motilin manufactured by the American Peptide Co. Inc. (Sunnyvale, CA, USA) and prepared for human use by Clinalfa AG (Läufelfingen, Switzerland). After an overnight fast of at least 10 h, the subjects reported to the LUMC in the morning of each study day. Two cannulas were inserted in contralateral forearm veins, one for blood sampling and one for the administration of motilin or placebo. Before introduction of the rectal barostat catheter, the rectum was cleansed with a water enema [14]. The catheter was inserted in the rectum with the distal part of the bag at 2 cm proximal from the anal sphincter; this position was checked by digital examination. The subjects were positioned in a 10° Trendelenburg position to reduce gravitational effects on the barostat recording [15].

After determination of the minimal distending pressure (MDP; pressure level needed to overcome intra-abdominal pressure) [16], a constant pressure procedure was started at a pressure of 3 mmHg above MDP. Following a 20-min baseline recording period, motilin or placebo infusion was started for a period of 90 min. The constant pressure procedure continued for 45 min after the start of the infusion. Thereafter, following a resting period of 20 min, a stepwise pressure distension procedure was performed. Isobaric distensions were performed in 1-mmHg steps, every 60 s from 0 mmHg to a maximum of 30 mmHg. The procedure stopped if a bag volume of 800 mL was reached or if the subject could not tolerate further distension. During the distension procedure, at every second pressure increment, the participants were asked to fill out perception questionnaires [100 mm horizontal visual analogue scales (VAS)][17]. Feelings of urge to defecate (rectal filling) and lower abdominal discomfort were scored.

After approximately 4 h, subjects were offered lunch and discharged from the unit.

Sample handling and motilin assay

Blood sampling for motilin assay (5 mL; Na/EDTA-coated tubes) was performed predose (t = −1 min) and at t = 10, 20, 60 and 90 min. Blood samples were collected on ice, were centrifuged for 10 min at 4 °C (2000 g) and separated plasma was stored at −20 °C until analysis. Assay for motilin was done with a sensitive (limit of detection = 10 pmol L⁻¹) and specific radioimmunoassay [18].

Data analysis

Motilin pharmacokinetics

The pharmacokinetic parameters for exogenous motilin were estimated, taking into account the endogenous motilin levels, as described previously [2]. A continuous endogenous motilin production was assumed and the rate of production was estimated in a one-compartment model. The modelling was done using NONMEM V software providing individual nonlinear regression estimates for the parameters (NONMEM Project Group, UCSF, CA, USA).

Rectum volume during the constant pressure procedure

Rectum volume was measured continuously and average values over 5-min epochs were analysed. Baseline rectum volume was calculated as the average volume of the 5-min period preceding the start of the infusion. This baseline volume was set at 100%. Subsequent volumes were calculated as percentage relative to this baseline, as time-averaged area under the effect curve (AUEC), and as maximum change in rectum volume (maximum effect; E_max) [19].

Rectum volume during the distension procedure

Rectum volume was measured continuously and average values over the last 30 s of each pressure level were analysed. Rectum volumes as time-averaged AUEC, and volumes at highest pressure level were calculated.

Compliance of the rectum was quantified from the distension data, by smoothing the distension volume/pressure curve (Figure 2) using a sigmoid E_max model. The maximum slope was used as a measure of compliance. Sigmoid E_max modelling was performed using WinNonlin V3.1 (Pharsight Corp., Mountain View, California, USA).

Figure 2.

Rectum volume (mL) (mean ± SD) during isobaric distensions (n = 8) during motilin (•) and placebo (○).

Abdominal sensation

Perception scores on 100 mm VAS were calculated in mm. Perception scores were calculated as time-average AUEC of the values at every second pressure increment.

Results are expressed as means ± SD. Differences between treatments were used to test for the presence of a motilin effect using paired Student t-tests and are reported with 95% confidence interval (CI). P-values < 0.05 were considered significant. All analyses were performed using SPSS for Windows V10.07 (SPSS, Inc., Chicago, IL, USA).

Results

The procedure was well tolerated by each subject and there were no withdrawals.

Motilin pharmacokinetics

Baseline plasma motilin concentrations were similar between treatments: 64 ± 34 (placebo) and 56 ± 28 pmol L⁻¹ (motilin). The pharmacokinetic model used to describe the motilin kinetics estimated an endogenous motilin concentration of 53 ± 29 pmol L⁻¹ for the motilin experiment, which corresponded well with the average motilin concentration of 57 ± 30 pmol L⁻¹ during placebo. During the 90-min infusion of motilin, plasma motilin rose to a steady-state concentration [C_ss; calculated as C_ss= endogenous level + (infusion rate/clearance)] of 494 ± 136 pmol L⁻¹. Ninety-five percent of the steady-state concentration was reached within 39 ± 9 min. The estimated motilin clearance was 625 ± 93 mL min⁻¹, with a volume of distribution of 8.0 ± 1.2 L and an elimination half-life of 9.1 ± 2.0 min.

Rectum volume during constant pressure procedure

MDPs and baseline rectum volumes were similar between treatments (Table 1). Compared with placebo, motilin slightly increased averaged rectum volume (AUEC) by 6% (95% CI −3, 16) and maximal rectum volume (E_max) by 5% (95% CI −8, 18) from baseline (Figure 1).

Figure 1.

Baseline-corrected rectum volume (%) (mean ± SD) during the constant pressure procedure [minimal distending pressure (MDP) + 3 mmHg] (n = 8), during motilin (•) and placebo (○).

Table 1.

Rectum parameters (mean ± SD) during motilin and placebo administration, during the minimal distending pressure (MDP), constant pressure and distension procedure (n = 8), and differences between treatments with the corresponding 95% confidence interval (95% CI)

	Motilin	Placebo	Difference (95% CI)
MDP (mmHg)	12.1 ± 1.9	11.5 ± 2.3	0.6 (−0.7, 2.0)
Constant pressure procedure
Baseline rectum volume (mL)	185 ± 62	136 ± 41	49 (−21, 118)
Average rectum volume (AUEC)(% from baseline)	106 ± 13	100 ± 16	6 (−3, 16)
Maximal rectum volume (Emax)(% from baseline)	115 ± 12	110 ± 11	5 (−8, 18)
Distension procedure
Average rectum volume (AUEC) (mL)	161 ± 55	118 ± 43	43 (0.4, 85)*
Rectum volume at highest pressure (mL)	349 ± 98	297 ± 88	51 (−9, 112)
Rectum compliance (mL mmHg−1)	37 ± 16	27 ± 11	10 (0.3, 20)*

^*P < 0.05.

Rectum volume during distension procedure

Distension of the rectum with increasing pressure levels resulted in increasing rectum volumes during either treatment. However, during motilin administration the rectum was able to distend more than during placebo (Figure 2), which was reflected by significant increases in both rectum volume and compliance. Motilin increased mean rectum volume (AUEC) by 43 mL (95% CI 0.4, 85; P < 0.05) and the volume at highest pressure level by 51 mL (95% CI −9, 112), compared with placebo. Relative to placebo treatment, rectum compliance increased by 10 mL mmHg⁻¹ (95% CI 0.3, 20; P < 0.05) during motilin treatment (Table 1).

Abdominal sensation

The stepwise pressure increments resulted in increasing perception scores for feelings of urge to defecate and lower abdominal discomfort, but this did not differ between the treatments (Figure 3a,b).

Figure 3.

(a) Visual analogue scales (VAS) urge (mm) (mean ± SD) during motilin (•) and placebo (○) infusion. (b) VAS discomfort (mm) (mean ± SD) during motilin (•) and placebo (○) infusion.

Discussion

This study is the first to demonstrate that motilin has effects on the human rectum. It was found that motilin induced a slight but significant relaxation of the rectum in healthy volunteers. Despite the effect on rectal compliance, no effects on rectal sensation scores were detected. This is in line with previous studies in fasted healthy volunteers, where motilin was shown to affect motoric endpoints while not affecting subjective abdominal sensations [2, 20].

Stimulatory action of motilin on the proximal GI tract has been well documented. For instance, motilin reportedly increases proximal gastric tone [21] and antrum contraction frequency [2] and induces phase 3 activity of the migrating motor complex [3]. To a lesser extent, stimulatory actions of motilin on distal parts of the GI tract have been described. Recently, the presence of motilin receptors has been described for the human colon [4] and contractile effects of motilin on the circular smooth muscle of human colon have been reported [8, 9, 22]. However, no data are available about the presence of motilin receptors on the human rectum, nor about its effect on this organ. The present study shows that motilin increased rectal volume, but the effect reached significance only during the distension procedure. The disparity between the two barostat procedures may be explained by the observation that motilin plasma concentrations did not reach steady-state during the first procedure, while the distension procedure was performed during steady-state motilin concentration. However, we have previously shown that with the presently employed infusion regimen, motilin effects on the stomach may already be assessed before steady state of motilin has been reached [2].

The present data may indicate the presence of functional motilin receptors in the human rectum. However, even if these receptors are not present the findings of this study can be reconciled with the available evidence of motilin effects. For instance, it is well known that motor activity of the colon increases after meal ingestion. This gastrocolic reflex which is the colonic response to eating is often followed by defecation [23]. During the reflex, contractions of the colon propel intestinal contents into the rectum. The rectum has to accommodate these contents, thereby acting as a storage organ. Furthermore, an adaptive response of the rectum to distension exists, reflected by a volume increase in response to rectal distension [24]. Therefore, the relaxation of the rectum as demonstrated in the current study may reflect the peristaltic reflex whereby the rectum relaxes in response to contractions of the colon.

It is not uncommon that compounds have different and sometimes even opposing effects on GI motility. For example, neurotensin has regional differences in its effects on GI smooth muscle [25]. Also, for erythromycin, stimulatory effects [8, 9], no effect [26] and even inhibitory effects [27] on GI motility have been described. The underlying mechanism of motilin stimulating colonic activity and increasing rectal compliance can not be deduced from the current study. Probably, motilin does not influence GI motility by a single mechanism but by several pathways, such as direct action on motilin receptors on GI smooth muscle and indirectly via numerous neuronal pathways resulting in a variety of effects. Further mechanistic studies are necessary to clarify the mechanism by which the effect is established.

The present study may be considered not to provide information about the role of motilin in regulating rectal compliance under physiological conditions, as the infusion resulted in motilin concentrations approximately 10-fold higher than baseline motilin concentrations. However, as motilin is synthesized and released in the GI tract and as it is likely that motilin is metabolized hepatically, it is conceivable that peripheral motilin levels do not represent the concentration at the site of action. Thus, the attained high peripheral motilin concentrations can be of physiological relevance at the site of action (in the gut). This is supported further by the fact that contractions in the distal stomach were attained by these concentrations and the amount of contractions was well within the physiological range [2].

It is well known that IBS is associated with intestinal motility disturbances [28–30] and that alteration in motilin levels are present in IBS patients [13, 31]; increased motilin levels have been found in diarrhoea-predominant IBS, whereas decreased levels are present in constipation-predominant IBS [12, 32]. The observation of decreased motilin concentrations in constipation-predominant IBS patients coincides with data showing decreased rectal compliance in these patients [33], which corresponds well with the present finding of motilin increasing rectal compliance. It can thus be speculated that motilin plays a role in IBS, and that interference with this regulatory mechanism may provide new treatment opportunities for this complex disease entity.

In conclusion, motilin increased rectal compliance without affecting lower abdominal sensation. As motilin seems to be involved in the regulation of lower GI motility, motilin agonists or antagonists may be of therapeutic relevance in lower abdominal motility disorders such as IBS, chronic constipation or acute diarrhoea.