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AbstractWe investigated the involvement of adenosine in ischemic preconditioning (IPC) by the unspecific antagonist, 8‐phenyltheophylline (8‐PT). Anesthetized Wistar rats were treated as follows: 1. nonischemic controls, 2. ischemic controls: 60 min of clamping of the common hepatic artery followed by 60 min reperfusion, 3. IPC: 10 min ischemia followed by 15 min reperfusion, prior to the identical ischemia‐reperfusion (IR) period as in group 2, 4. 8‐PT + IPC: 8‐PT 10 mg/kg i. v. was given 10 min prior to the identical procedure as in group 3. The peripheral liver blood flow was monitored by laser‐Doppler flowmetry. Blood alanine aminotransferase (ALT) was analyzed once every 60 min. IPC significantly reduced impairment of liver blood flow, as well as ALT increase during reperfusion. This effect was abolished by pretreatment with 8‐PT. Adenosine appears to be a crucial effector in IPC. Clinical studies need to be undertaken to explore a possible effect of IPC in liver transplantation.

Relaxations of segments of rat distal colon were elicited by hypertonic solutions of potassium (K(+); final concentration, 20.8 or 50.8 mM). The initial part of the response to K(+) was antagonized by the nerve blocker tetrodotoxin. This effect could, moreover, be significantly antagonized by apamin (a blocker of K(+) channels), reactive blue 2 (a P(2y)-purinoceptor antagonist), N(G)-nitro-L-arginine (an inhibitor of NO synthase), 1H-[1,2,4]- oxadiazolo[4,3-a]quinoxaline-1-one (ODQ; an inhibitor of soluble guanylyl cyclase), or N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide (H-89; an inhibitor of cAMP-dependent protein kinase). Sodium nitroprusside (a donor of NO) and vasoactive intestinal peptide (VIP) both relaxed the tissues. The response to sodium nitroprusside was abolished by ODQ and unaffected by H-89, and that to VIP was partially inhibited by VIP(10-28) (a VIP receptor antagonist), ODQ, or H-89. When combining reactive blue 2 and N(G)-nitro-L-arginine, the response to 50.8 mM K(+) was reduced by approximately 70% and was abolished by the concomitant administration of these antagonists and VIP(10-28). ATP, NO, and VIP may, thus, be inhibitory neurotransmitters in rat distal colon.

Peroperative manometry was performed in 12 patients operated on with endoanal proctectomy and a hand-sewn pouch-anal anastomosis and in 12 in whom proctectomy was performed entirely from above, with the ileal pouch stapled to the top of the anal canal. Results from both groups showed that division of the superior rectal artery reduced the median (95 per cent confidence interval (c.i.)) resting anal pressure from 77.5 (69.9–83.3) mmHg to 64.5 (55.2–70.0) mmHg (P< 0.01). Complete rectal mobilization to the pelvic floor decreased resting pressure by an additional 22 per cent, to a median of 50.0 (95 per cent c.i. 40.1–53.5) mmHg (P< 0.01). After completion of anastomosis, irrespective of the operative technique used, a further decline in median pressure to 35.0 (95 per cent c.i. 26.0–47.7) mmHg could be demonstrated (P < 0.05). This study indicates that anal sphincter pressure is reduced to a similar extent after hand-sewn and stapled anastomoses. Injury to the autonomic nervous supply to the anal sphincter mechanism might be the major cause for this reduction.

A method to study electrically induced distal colonic motility in the rat in vivois reported. The animals were anaesthetized with methohexital and chloralose and were artificially ventilated. Motility of a segment (2 cm) of the distal colon was monitored as volume changes of an intraluminal balloon, introduced via the anus. Local electrical stimulation of the wall of the segment was achieved by means of a bipolar electrode folded around the gut. Stimulations produced reproducible contractile responses in a frequency dependent fashion. Stimulation characteristics resembled those of other autonomic neuro‐effector systems. The adrenergic neuron‐blocker, guanethidine, significantly lowered colonic tone, but had no other effects on spontaneous or electrically induced motility. Atropine significantly lowered colonic tone. After the administration of this compound the electrically induced contractions were significantly smaller with a shorter duration and, furthermore, appeared upon the cessation of stimulation (‘off’ or ‘rebound’ contraction). Following the administration of tetrodotoxin (TTX, given close i.a. via a cannula with its tip in distal aorta) basal colonic tone and the number of spontaneously occurring contractions increased. The amplitude and duration of the electrically induced responses were significantly attenuated and, furthermore, appeared as ‘rebound’ contractions which were preceded by a relaxation. Such TTX‐resistant responses may be myogenic, but a neurogenic origin cannot be excluded. The present study showed that local electrical stimulation of the distal colon elicits cholinergic contractions, but also atropine‐ and TTX‐resistant motor responses.

Background: Nitric oxide (NO) mediates inhibition of gastrointestinal smooth-muscle cells via non-adrenergic, non-cholinergic (NANC) nervous pathways. The effect of NO on the absorption and secretion by the mucosa of the gastrointestinal and hepatobiliary tracts is less well known. The aim of this study was to evaluate the effects of a pharmacologic blockade of NO synthase on sphincter of Oddi activity, gallbladder function, and bile secretion and to demonstrate the presence of NO synthase-positive neurons in this region.Methods: Experiments were conducted on anesthetized cats after blockage of noradrenergic and cholinergic neurotransmission. Flow resistance in the sphincter of Oddi, gallbladder fluid absorption and motility, bile outflow from the liver, and bile salt secretion were registered.Results: Flow resistance exerted by the sphincter of Oddi increased dose-dependently in response to the NO synthase blocker NG-nitro-L-arginine. The increase in flow resistance was reversed stereospecifically by L-arginine, the substrate for NO synthesis. No significant effects on bile secretion, gallbladder fluid transport, or gallbladder motility were observed. NO synthase-positive neurons were identified close to the sphincter of Oddi and in the gallbladder mucosa.Conclusions: This tonically active inhibitory NANC innervation of the sphincter of Oddi may be important in the physiologic regulation of the bile duct pressure.

Anticholinergic drugs are used on an empirical basis for treatment of functional disturbances after restorative proctocolectomy, but their mode of action on ileal pouch performance is mainly unknown. We studied the acute effects of atropine or benzilonium on pouch characteristics and anal sphincter function in 20 patients with a pelvic pouch. Pouch volume was increased by 27% by atropine at distension with 20 cm H2O (p < 0.01). Benzilonium tended to have a similar effect, but the changes did not reach statistical significance (p = 0.06). Pouch contractility, as reflected by volume fluctuations and pressure changes during distension, was almost abolished by both drugs. Sensory thresholds for sense of filling and, particularly, urge were raised. Resting anal pressure was slightly lowered, whereas no significant effect was found on maximal squeeze pressure. In conclusion, anticholinergics appear to have specific properties of action on small-intestinal reservoirs, constituting possible explanations for the empirically observed beneficial effects of anticholinergic treatment of functional disturbances after restorative proctocolectomy.

Lange S. Delbro DS. Jennische E. Evans blue permeation of intestinal mucosa in the rat. Scand J Gastroenterol 1994;29:38-46.The azo dye Evans blue (EB; molecular weight, 960.83) is widely used as an indicator of increased capillary permeability. In the present study, however, rat gut absorption of EB was investigated after dye instillation in either the small or large intestine. During a brief period of ether anaesthesia, EB was injected either into jejunal loops with a challenge period of 30 or 60 min or into a proximal and a distal colon loop with a challenge period of 30, 60, or 120 min. After the rats had been killed the intestinal specimens were washed with 6 mM acetylcysteine dissolved in phosphate-buffered saline, which efficiently cleared the tissues of mucus, and thus of EB trapped in mucus. Only EB absorbed by the gut wall remained to be estimated, and this absorption was found to be both dose- and time-dependent in the jejunum and the colon. After instillation in the colon, but not in jejunum, EB could be detected in the blood. EB absorption from the jejunum remained unaffected by the addition of either ouabain (1 mM) or lidocaine (0.38 mM). Either of these compounds inhibited EB uptake in the proximal part of the colon, while enhancing it in the distal part. Fluorescence microscopy showed penetration into the intestinal wall to be a prerequisite for EB to become fluorescent, and EB fluorescence increased with time. It is proposed that EB is transported over the mucosa by the paracellular route and that the amount of absorbed EB reflects epithelial permeability differently in different parts of the gastrointestinal tract. The results suggest that active mechanisms also contribute to the EB uptake in the large intestine. A particularly noteworthy finding was the existence of a functional heterogeneity between the proximal and distal colon.

In anaesthetized rats treated with guanethidine, the pylorus was ligated. A catheter was inserted into the stomach via the mouth and anchored by a ligature around the cervical oesophagus. The catheter was used for instillation into the stomach of a fixed volume (9 ml) of either isotonic or 1 M NaCl and for the recording of intragastric pressure. Neurogenic modulation of gastric motility caused by either solution was analysed by studying the effect of the systemic administration of the ganglionic blocker hexamethonium on gastric tone and on phasic motor activity. Gastric motility was not significantly changed by hexamethonium in the animals that had been subjected to isotonic NaCl. After intragastric treatment with 1 M NaCl, however, hexamethonium caused increased gastric tone and decreased phasic motor activity. An analysis of these results suggests that 1 M NaCl activated vagal non-adrenergic, non-cholinergic relaxatory and cholinergic excitatory motor neurons, respectively. These chemically induced modulations of gastric motility were dependent on the spinal cord and, most probably, were due to activation of the mucosal sensory endings of the splanchnic nerves, leading to reflex spinovagal motor adjustments.

Nitric oxide (NO) has emerged as one of several important intraovarian regulatory factors. In particular, NO has been implicated in the processes of ovulation and atresia-related apoptosis. The aim of the present study was to investigate the presence and distribution of the NO-generating nitric oxide synthase (NOS) enzymes in the ovary during follicular development, ovulation and luteal formation of the equine chorionic gonadotrophin (ECG)/human chorionic gonadotrophin (HCG)-primed rat. NADPH diaphorase activity was used as a histochemical marker for NOS within the ovary. Diaphorase reactivity was most abundant in the stroma (S) of the ovary and in the theca (T) layer of the follicle. In luteinized ovaries, weaker diaphorase reactivity was present within the corpora lutea (CL). Two different isoforms of NOS, the constitutively expressed endothelial NOS (eNOS) and the inducible isoform of NOS (iNOS), were immunolocalized in ovaries of immature rats and in ECG/HCG-primed rats during the periovulatory period from HCG injection until 2 days after ovulation. In addition, ovarian concentrations of eNOS and iNOS were quantified by immunoblotting. Immunoblotting with a monoclonal anti-eNOS antibody demonstrated the presence of eNOS mainly in the residual ovary (ROV) during the periovulatory period. In luteinized ovaries, higher concentrations of eNOS were seen in CL, while those in the ROV at this stage were lower than in the periovulatory ovary. Immature ovaries contained diminutive amounts of eNOS, detectable mostly in the ROV compartment. In contrast, iNOS was barely detectable during follicular development to the preovulatory stage. A slight elevation of iNOS was observed in the granulosa cells at 6 h after the HCG injection. The levels of iNOS during the luteal phase were also low. Immunohistochemical analysis using polyclonal eNOS and iNOS antibodies revealed the localization of these two isoforms primarily in the S and the T of the periovulatory ovary. In luteinized ovaries, positive immunoreactivity was also seen within the CL. With a monoclonal antibody against eNOS, intense immunoreactivity was observed in the S, T and within CL. There was a particularly strong staining in blood vessels. These data demonstrate the presence of an intraovarian NO-generating system. The localization of this system to the S, T and CL suggests a role for NO in the ovulatory process and in the regulation of CL function.

The physiological and clinical effects of loperamide treatment versus placebo were investigated in a randomized, double-blind, crossover study in patients operated with restorative proctocolectomy. Sixteen patients operated with endoanal mucosectomy and a handsewn ileal pouch-anal anastomosis and 14 patients operated with abdominal proctocolectomy and stapling of the pouch to the top of the anal canal were studied. While loperamide treatment increased resting anal pressure in both groups of patients by approximately 20% (P<0.05), squeeze pressure was not affected. Loperamide did not affect pouch volume or contractility. Sensory thresholds and the recto/pouch-anal inhibitory reflex were not influenced by loperamide treatment. Clinical function was improved, with a reduced bowel frequency and an improved nighttime continence, with less soiling (P<0.05) as well as need to wear a protective pad.

Background: The relaxatory effect of acetylcholine was investigated on the feline internal anal sphincter (IAS), in vitro. Results: Acetylcholine (10, 30, 100, and 1000 μM) caused a concentration-dependent relaxation of the same magnitude in strips from the proximal and distal IAS. The antagonist of nitric oxide synthase, N$oM-nitro-L-arginine (L-NNA; 1, 10, and 100 μM), in a concentration-dependent and stereospecific manner, blocked the acetylcholine-induced relaxation, leaving a residual response of 10-30%. The blocking effect of L-NNA (100 μM) could not be shown in tissues that had been incubated with the substrate for nitric oxide synthase, L-arginine (1 mM). Conclusions: The present results suggest that the acetylcholine-induced relaxation of the IAS to a major extent is due to an activation of nitrergic, inhibitory motor neurons to the IAS.

Recto-anal motility response to bladder distension was studied under general anaesthesia in 12 patients undergoing intestinal resection for Crohn's disease of the small intestine or colonic cancer. The effect of epidural anaesthesia on anal tone and on the motility response to bladder distension was studied in six of these patients. An anal pressure increase on bladder distension was observed in all individuals. No motility response was noted in the rectum. The anal pressure response to bladder distension was abolished by epidural anaesthesia. It was concluded that anal pressure in man under general anaesthesia was tonically influenced by the thoracolumbar sympathetic outflow. An excitatory vesico-anal reflex was demonstrated. It appears as this reflex is mediated via the spinal cord.

1. The effects of electrical stimulation of the peripheral end of the cervical vagal nerve on jejunal motility were investigated in anaesthetized cats, pretreated with guanethidine, with sectioned splanchnic nerves and ligated adrenal vessels. Motility was monitored as volume changes of an intraluminal balloon. 2. Vagal stimulation elicited frequency‐dependent hypermotility with a short latency. Relaxatory events were also observed, which could indicate the presence of a non‐adrenergic inhibitory pathway. 3. After atropine treatment, contractions and relaxations could still be elicited. The former were compared to cholinergic contractions and showed a lower maximal amplitude and a longer latency to onset. Moreover, they were antagonized by 80‐100% by the opioid receptor antagonist, naloxone. 4. Vagal stimulation after hemicholinium, given in order to deplete the preganglionic acetylcholine content, elicited naloxone‐sensitive contractions. This suggests that a subpopulation of the vagal preganglionic fibres is non‐cholinergic. 5. Isolation of the balloon‐containing segment did not qualitatively alter the responses, indicating that the vagal fibres reach the small intestine via the paravascular mesenteric nerves. 6. It is concluded that cholinergic and non‐adrenergic, non‐cholinergic (NANC) contractions, as well as relaxations, could be elicited by efferent vagal stimulation. The NANC contractions seem to result from the activation of opioid receptors causing disinhibition of a tonic neurogenic restraint on the gut muscle.

The intestinal vasodilation evoked by mechanical mucosal stimulation or by transmural electrical field stimulation was abolished by close i.a. injection of Apamin, a polypeptide originally isolated from bee venom. Apamin also blocked the vasodilatation induced by close i.a. infusion of vasoactive intestinal polypeptide (VIP). It is suggested that Apamin is a VIP receptor antagonist.