Identification of functional intramuscular rectal mechanoreceptors in aganglionic rectal smooth muscle from piebald lethal mice.

The mechanosensitive endings of low-threshold, slowly adapting pelvic afferents that innervate the rectum have been previously identified as rectal intraganglionic laminar endings (rIGLEs) that lie within myenteric ganglia. We tested whether the aganglionic rectum of piebald-lethal (s(l)/s(l)) mice lacks rIGLEs and whether this could explain impaired distension-evoked reflexes from this region. Extracellular recordings were made from fine rectal nerves in C57BL/6 wild-type and s(l)/s(l) mice, combined with anterograde labeling. In C57BL/6 mice, graded circumferential stretch applied to the rectum activated graded increases in firing of slowly adapting rectal mechanoreceptors. In s(l)/s(l) mice, graded stretch of the aganglionic rectum activated similar graded increases in rectal afferent firing. Stretch-sensitive afferents responded at low mechanical thresholds and fired more intensely at noxious levels of stretch. They could also be activated by probing their receptive fields with von Frey hairs and by muscle contraction. Anterograde labeling from recorded rectal nerves identified the mechanoreceptors of muscular afferents in the aganglionic rectal smooth muscle. A population of afferents were also recorded in both C57BL/6 and s(l)/s(l) mice that were activated by von Frey hair probing, but not stretch. In summary, the aganglionic rectum is innervated by a population of stretch-sensitive rectal afferent mechanoreceptor which develops and functions in the absence of any enteric ganglia. These results suggest that in patients with Hirschsprung's disease the inability to activate extrinsic distension reflexes from the aganglionic rectum is unlikely to be due to the absence of stretch-sensitive extrinsic mechanoreceptors.

Anatomic modifications in the enteric nervous system of piebald mice and physiological consequences to colonic motor activity.

It has been assumed that in piebald lethal mice that develop megacolon, impaired colonic motor activity is restricted to the aganglionic distal colon. Peristaltic mechanical recordings, immunohistochemistry, and quantitative PCR were used to investigate whether regions of the colon, other than the aganglionic segment, may also show anatomical modifications and dysfunctional colonic motor activity. Contrary to expectations, colonic migrating motor complexes (MMCs) were absent along the whole colon of piebald lethal homozygote mice and severely impaired in heterozygote siblings. Aganglionosis was detected not only in the distal colon of piebald homozygote lethal mice (mean length: 20.4 +/- 2.1 mm) but also surprisingly in their heterozygote siblings (mean length: 12.4 +/- 1.1 mm). Unlike homozygote lethal mice, piebald heterozygotes showed no signs of megacolon. Interestingly, mRNA expression for PGP 9.5 was also dramatically reduced (by 71-99%) throughout the entire small and large bowel in both homozygote lethal and heterozygous littermates (by 67-87%). Histochemical staining confirmed a significant reduction in myenteric ganglia along the whole colon. In summary, the piebald mutation in homozygote lethal and heterozygote siblings is associated with dramatic reductions in myenteric ganglia throughout the entire colon and not limited to the distal colon as originally thought. Functionally, this results in an absence or severe impairment of colonic MMC activity in both piebald homozygote lethal and heterozygote siblings, respectively. The observation that piebald heterozygotes have an aganglionic distal colon (mean length: 12 mm) but live a normal murine life span without megacolon suggests that aganglionosis >12 mm and the complete absence of colonic MMCs may be required before any symptoms of megacolon arise.