Effects of substance P on spontaneous contraction in the flexure region of the guinea pig colon.

Effects of Substance P on spontaneous contractions of the circular muscle of the flexure region of guinea pig colon were studied by mechanical tension recording. Substance P (3 nM-10 nM) produced tonic contraction associated with phasic activities but the contraction was found stronger at higher concentration. Verapamil (3 microM), a voltage dependent L-type Ca(2+) channel blocker completely blocked the spontaneous activities and also Substance P induced contraction. These results suggest that Substance P produce contraction by Ca(2+) influx and the Ca(2+) influx occurs by activating verapamil sensitive Ca(2+) channel.

Special smooth muscle cells along the submucosal surface of the guinea pig colon with reference to its spontaneous contractions.

Antiperistalses occur from the flexure region of the guinea pig colon. We previously demonstrated that the circular muscle at the mesenteric border of the flexure region produced spontaneous regular contractions and found special smooth muscle cells believed to be pacemakers along the submucosal surface of the circular muscle layer. In this study, we revealed bipolar- and multipolar-type special smooth muscle cells along the submucosal surface of the muscle layer. Their slender cell processes contacted each other and formed a cellular network. Caveolae, filament structures expressing smooth muscle actin, vimentin, some desmin, and basal lamina were prominent features. The special smooth muscle cells corresponded to c-Kit-immunopositive cells and so-called interstitial cells or interstitial cells of Cajal in other reports. Their population was larger in the flexure region and the proximal colon than in the distal colon. The circular muscle layer at the flexure region was thicker than in other regions. The contraction in the flexure region showed the highest frequency and regularity. The dense population of special smooth muscle cells at the flexure region and thicker muscle layer may make the mechanical contraction more regular. The antiperistalsis from the flexure region could be explained in relation to the highest frequency of the pulsating contraction.

Caffeine and ryanodine may act on the plasma membrane of the circular muscle at the flexure region in the guinea-pig colon.

The actions of caffeine and ryanodine on the spontaneous rhythmic activities in the guinea-pig colon were studied by mechanical tension recording. Caffeine reduced the amplitude of the spontaneous rhythmic activity at low concentration (0.3 mM-1 mM). At high concentration (3-10 mM), it induced a phasic transient contraction. The spontaneous rhythmic activity and a phasic contraction induced by caffeine, were blocked by verapamil (3 microM) or by removal of external Ca2+. Ryanodine affected neither resting tension nor frequency of spontaneous activity at 1 microM. However in the circular muscle strips pretreated with ryanodine, a sustained contraction was initiated after the removal of caffeine (10 mM). Continuous Ca2+ influx was necessary for spontaneous rhythmic activities and a phasic transient contraction, because it was abolished completely by the removal of external Ca2+, Verapamil (3 microM), a voltage gated L-type Ca2+ channels blocker, inhibited the spontaneous rhythmic activities and also inhibited phasic transit contraction followed by a sustained contraction induced by 10 mM caffeine. Our results suggest that caffeine may produce a sustained contraction by activating verapamil sensitive Ca2+ channel. In the muscle pretreated with both caffeine and ryanodine, continuous Ca2+ influx may occur also through verapamil sensitive pathway.

A smooth muscle nodule producing 10-12 cycle/min regular contractions at the mesenteric border of the pacemaker area in the guinea-pig colon.

At the boundary between the proximal and distal divisions of the colon in the guinea-pig is a ring-like section which rhythmically contracts. HUKUHARA and his co-researchers demonstrated that antiperistaltic movements in the proximal colon start from this ring-like section, the so-called pacemaker area. Tissue specimens, 0.1-0.3 mm in width/height x 4-7 mm in length, were prepared from various parts of this area. Significantly, in the circular muscle at the mesenteric border, a nodular structure spontaneously producing 10-12 cycle/min regular mechanical contractions was found. Moreover, histological investigations after physiological recording revealed that the presence of the inner-most and/or outermost portions of the circular muscle coat was not necessary for these spontaneous activities. Champy-Maillet (ZIO) staining showed that smooth muscle cells in this spontaneously contracting nodule were heavily innervated. Transmission electron microscopy showed that the smooth muscle tissue of this particular area was characterized by scanty interstitial elements such as fibroblasts. Plasma membranes of adjacent smooth muscle cells were frequently in direct contact with each other, forming many gap junctions. Scanning electron microscopy in the specimen prepared using a NaOH-maceration method revealed fine three-dimensional relationships between nerve terminals and smooth muscle cells. The nodular structure described in this paper may provide a useful experimental model for the investigation of colonic motility and its neural control.

Nitrinergic nerves controlling pacemaker activities of the inner sublayer (P-layer) in the canine proximal colon circular muscles.

The inner sublayer (P-layer) of the circular muscle coat in the canine proximal colon has been known to produce spontaneous mechanical contractions associated with characteristic electrical activities called slow waves. We recorded the mechanical activities of tissue preparations from this P-layer. Normal Krebs solution (K+; 6 mM) was used as the perfusate. Elevation of extracellular K+ concentrations in the range of 12 mM and 36 mM induced intensified phasic contractions. Administration of an NO-synthase inhibitor, N omega-nitro-arginine methyl ester (L-NAME, 50 microM), enhanced both the spontaneous mechanical rhythms and high extracellular K(+)-induced contractions. Administration of the substrate for NO synthases, L-arginine (400 microM) remarkably suppressed the effects of L-NAME on the amplitude of the spontaneous rhythms and on responses to extracellular high K+. Histological structures of nerves in the P-layer were investigated by an NADPH (nicotinamide adenine dinucleotide phosphate)-diaphorase technique and by the immunohistochemistry of NO-synthases, since NO-producing (nitrinergic) nerves usually, if not always, show a histochemical NADPH-diaphorase positive reaction in formaldehyde-fixed specimens, and since features of ganglia and nerve strands in the outer subdivision of the submucosal plexus (plexus submucosus externus; or so-called Henle's plexus) together with the delicate network of nerve terminal varicosities within the P-layer were clearly visualized by this method. The topographical arrangement of nitrinergic nerves supported the view that they produce nitric oxide (NO), being one of the major chemical mediators of the neural control of the spontaneous rhythms in the P-layer.