Can colonic migrating motor complexes occur in mice lacking the endothelin-3 gene?

In mammals, colonic migrating motor complexes (CMMC) are a major propulsive contraction responsible for the expulsion of faecal content. Mice with a mutation of the endothelin-3 gene raised on a 129SL background strain have ~70% colonic aganglionosis, lack CMMC, and are lethal within 12 days postpartum. In contrast, endothelin-3 mutant mice raised and maintained on a C57BL6 background strain (lethal-spotted (ls/ls) mice) can live for much longer, but it is unclear whether CMMC generation is preserved in these mice also lacking the endothelin-3 gene. The aim of this study was to determine whether CMMC exist in ls/ls mouse colon and, if so, whether their existence and frequency are related to the length of aganglionosis. Spatiotemporal mapping and mechanical recordings of colonic wall movements were made from isolated whole colons obtained from wild-type and ls/ls mice. Although ls/ls mice had a megacolon, they still generated CMMC in the ganglionic segment, which on some occasions could propagate short distances into the aganglionic region. There was large variability in aganglionosis length, which showed a weak correlation with the existence or frequency of CMMC. Interestingly, CMMC propagation velocity was slower in ls/ls mice when evoked by intraluminal fluid. A myogenic motor pattern was identified in the aganglionic region that was maintained under tonic inhibition. We show that despite megacolon, ls/ls mice still generate CMMC in the ganglionic region. These offspring have sufficient propulsive motility in the ganglionic segment to live a normal murine lifespan and rarely die of bowel obstruction.

Inhibition of human drug-metabolising cytochrome P450 and UDP-glucuronosyltransferase enzyme activities in vitro by uremic toxins.

OBJECTIVE: To investigate the potential inhibitory effects of uremic toxins on the major human hepatic drug-metabolising cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) enzymes in vitro. METHODS: Benzyl alcohol, p-cresol, indoxyl sulfate, hippuric acid and a combination of the four uremic toxins were co-incubated with human liver microsomes and selective probe substrates for the major human drug-metabolising CYP and UGT enzymes. The percentage of enzyme inhibition was calculated by measuring the rates of probe metabolite formation in the absence and presence of the uremic toxins. Kinetics studies were conducted to evaluate the K i values and mechanism(s) of the inhibition of CYP2E1, CYP3A4, UGT1A1 and UGT1A9 by p-cresol. RESULTS: The individual uremic toxins inhibited CYP and UGT enzymes to a variable extent. p-Cresol was the most potent individual inhibitor, producing >50% inhibition of CYP2E1, CYP3A4, UGT1A1, UGT1A9 and UGT2B7 at a concentration of 100 µM. The greatest inhibition was observed with UGT1A9. p-Cresol was shown to be an uncompetitive inhibitor of UGT1A9, with unbound K i values of 9.1 and 2.5 µM in the absence and presence of bovine serum albumin (BSA), respectively. K i values for p-cresol inhibition of human liver microsomal CYP2E1, CYP3A4 and UGT1A1 ranged from 43 to 89 µM. A combination of the four uremic toxins produced >50% decreases in the activities of CYP1A2, CYP2C9, CYP2E1, CYP3A4, UGT1A1, UGT1A9 and UGT2B7. CONCLUSIONS: Uremic toxins may contribute to decreases in drug hepatic clearance in individuals with kidney disease by inhibition of hepatic drug-metabolising enzymes.

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

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.