Control of intrinsic pacemaker frequency and velocity of colonic migrating motor complexes in mouse

Abstract

The mechanisms that control the frequency and propagation velocity of colonic migrating motor complexes (CMMCs) in mammals are poorly understood. Previous in vitro studies on whole mouse colon have shown that CMMCs occur frequently (~every 1-3 min) when the colon is devoid of all fecal content. Consequently, these studies have concluded that the generation of CMMCs and the frequency which they occur does not require the presence of fecal content in the lumen. However, in these studies, stimuli have always been unavoidably applied to these empty colonic preparations, facilitating recordings of CMMC activity. We tested whether CMMCs still occur in empty whole colonic preparations, but when conventional recording methods are not used. To test this, we used video imaging, but did not utilize standard recording methods. In whole isolated colons containing multiple endogenous fecal pellets, CMMCs occurred frequently (1.9 ± 0.1/min) and propagated at 2.2 ± 0.2 mm/s. Surprisingly, when these preparations had expelled all content, CMMCs were absent in 11/24 preparations. In the remaining preparations, CMMCs occurred rarely (0.18 ± 0.02/min) and at reduced velocities (0.71 ± 0.1 mm/s), with reduced extent of propagation. When conventional recording techniques were then applied to these empty preparations, CMMC frequency significantly increased, as did the extent of propagation and velocity. We show that in contrast to popular belief, CMMCs either do not occur when the colon is free of luminal contents, or, they occur at significantly lower frequencies. We believe that previous in vitro studies on empty segments of whole mouse colon have consistently demonstrated CMMCs at high frequencies because conventional recording techniques stimulate the colon. This study shows that CMMCs are normally absent, or infrequent in an empty colon, but their frequency increases substantially when fecal content is present, or, if in vitro techniques are used that stimulate the intestine.

Keywords: colon; enteric nervous system; migrating motor complex; myenteric plexus; peristalsis.

Figure 1

Different types of preparations used to record CMMC activity in isolated whole segments of mouse colon. (A) Preparation 1 is a novel preparation, where video recordings are made from the empty whole colon, with only a single pin inserted at proximal and distal end to anchor the preparation to the colon. No force transducer clips pinch the bowel. (B) Shows preparation 2, where the colon has force transducer clips attached to the serosal surface, but no resting tension is applied to the colon. (C) Preparation 3, shows the colon tied to an infusion bath with suture thread. (D) Preparation 4 shows the oral and anal ends of the colon tied (as in preparation 3), but this preparation is also pierced with hooks and attached to force transducer under 1 gm resting tension.

Figure 2

Spatiotemporal maps of colonic circumferential wall diameter in 4 separate mice, showing propulsion of natural pellets in an aboral direction along the full length of isolated colon. (A,D) are diagrammatic representations of the preparations of colon that contain endogenous fecal pellets and were used to record changes in circumferential diameter by a video camera mounted directly above the preparation. These were then converted to the D-maps as shown in (B,C,E,F). CMMCs occur which induce pellet movement, with some pellets being propelled significant distances aborally. (E) Shows that after a single pellet has been expelled, the colon is empty and becomes relatively silent (indicated from the point at the downward arrow on the map).

Figure 3

Spatiotemporal maps of colonic wall diameter showing CMMCs in 4 separate empty colonic preparations. (A,D) are diagrammatic representations of the empty preparations, with the video camera mounted directly above each preparation. (B,C) Show infrequent CMMCs contractions propagating at a slower velocity, that typically do not propagate over the full length of colon. (E) Shows the absence of contractions in an empty colon. (F) has no obvious CMMCs present.

Figure 4

(B,D,F) are three separate recordings, from the same segment of whole mouse colon, using different recording methodologies. (A) Shows a diagrammatic representation of the preparation from which video recordings were made from a whole colon that contained multiple fecal pellets. (B) Shows the D-map from the same preparation, with CMMCs present that propel a number of pellets aborally. (C) Shows the preparation from which video recordings were made from the same segment of colon, but devoid of all fecal pellets. (D) Shows a D-map from the same preparation of colon as in (B) but the colon is now devoid of all fecal pellets. CMMCs are considerably less frequent. (E) Shows a diagram of the preparation but now when isometric force transducers are attached to the circular muscle at three sites along the colon, with 1 gm resting tension imposed. (F) Shows that under these recording conditions, CMMCs are now regularly recorded which propagated from oral, to mid, to distal colon.

Figure 5

There was a statistically significant reduction in the number of CMMCs that occurred per minute in full segments of colon (that lacked luminal contents) compared with the same segments that contained multiple endogenous pellets. (A) empty preparations which had hooks applied and 1 gm tension imposed induced a significant increase in CMMC frequency (compare Empty data set with Hooked data). “Tied” refers to when the colon is cannulated at the oral and anal ends with suture thread and “clipped” refers to when the preparations have stainless steel clips to pinch the bowel wall (e.g., Figure 1B). (B) the extent of propagation of individual CMMCs was significantly increased preparations were hooked and resting tension imposed, compared with the same preparations empty. (C) There was a statistically significant reduction in the propagation velocity of CMMCs between full and empty preparations. When these same preparations were then stimulated with one of the two conventional techniques, tied or hooked, CMMC velocity significantly increased.