Keith Hillier

Characterization of the 5-hydroxytryptamine receptor type involved in inhibition of spontaneous activity of human isolated colonic circular muscle.

1 Experiments were carried out to characterize pharmacologically the 5‐hydroxytryptamine (5‐HT) receptor types which mediate inhibition of spontaneous contractions of the intertaenial circular muscle in human isolated colon. 2 5‐HT caused a reproducible concentration‐dependent inhibition of spontaneous contractions of the circular muscle of human colon in vitro with a mean EC50 value of 0.2 μm and 95% confidence limits of 0.1–0.5 μm. No evidence for a contractile action of 5‐HT was found. Tetrodotoxin (TTX, 1.5 μm) caused a rightward shift of the concentration‐response curve of 5‐HT with a concentration‐ratio of 2.9. 3 The inhibitory response to 5‐HT was mimicked by several indoles with the rank order of potency 5‐HT > 5‐methoxytryptamine = α‐methyl‐5‐HT > 5‐carboxamidotryptamine >> 2‐methyl‐5‐HT. 5‐Hydroxyindalpine was inactive. 4 The substituted benzamides were agonists with the following rank order of potency, 5‐HT > renzapride > zacopride > metoclopramide > cisapride. 5 The inhibitory responses to 5‐HT were not inhibited by methysergide (10 μm) or methiothepin (1 μm), which are antagonists selective for 5‐HT1‐like and 5‐HT2 receptors, nor by ondansetron (10 μ) which is an antagonist at 5‐HT3 receptors. 6 The inhibitory responses induced by 5‐HT and 5‐methoxytryptamine were competitively antagonized by a weak 5‐HT4 receptor antagonist, tropisetron, with pKB values of approximately 6. Tropisetron had no significant effect on the inhibitory response curve produced by isoprenaline (0.01–100 μm). 7 The pharmacological profile of the 5‐HT‐evoked relaxations of human colon circular muscle are consistent with activation of a 5‐HT4‐like receptor.

PROSTAGLANDIN E AND F CONCENTRATIONS IN HUMAN ENDOMETRIUM AFTER INSERTION OF INTRAUTERINE CONTRACEPTIVE DEVICE

The prostaglandin (P.G.) content of endometrium from fourteen women was examined before and 1-5 months after insertion of an intrauterine contraceptive device. A statistically significant increase in P.G.E but not P.G.F was observed. The P.G.E concentration increased in eleven to fourteen cases and fell in three. The P.G.F content increased in five cases and fell in eight.

Colonic expression of leukotriene-pathway enzymes in inflammatory bowel diseases

Background Leukotrienes derived from the 5‐lipoxygenase pathway are proinflammatory lipid mediators that possibly play a role in inflammatory bowel diseases. The expression of 5‐lipoxygenase pathway proteins has not previously been examined in colonic mucosa in inflammatory bowel disease. Results Quantitative immunohistochemical analyses showed that, compared to those of the control subjects (n = 9), colonic biopsies from patients with active inflammatory bowel disease (n = 17) had 3‐ to 7‐fold higher mean counts of cells expressing 5‐lipoxygenase (P = 0.03), 5‐lipoxygenase‐activating protein (P = 0.005), and the leukotriene A4 hydrolase (P = 0.004), which make up the biosynthetic pathway of the potent neutrophil chemotaxin leukotriene B4. Immunoexpression of the leukotriene C4 synthase was unaltered (P > 0.2). The increased representation of leukotriene B4–pathway enzymes was associated with higher counts of neutrophils (P = 0.0001), macrophages (P = 0.03), eosinophils (P = 0.0004), CD8+ T cells (P < 0.001), activated T cells (P < 0.05), and B cells (P < 0.05) but not of mast cells (P > 0.9). These eicosanoid and cellular changes were most marked in the subgroup of patients with ulcerative colitis (n = 9), and were absent in patients with quiescent disease (n = 6). The anomalies in the 5‐lipoxygenase pathway were accompanied as expected by more cells immunostaining for cytokine‐inducible COX‐2 (P = 0.004, n = 17), but this study also revealed a greater number of cells expressing COX‐1 in the samples from the patients in the ulcerative colitis subgroup (P = 0.03, n = 9). Conclusions The 5‐lipoxygenase data provide a cellular basis for increased tissue synthesis of the leukotriene B4, as reflected in the colonic mucosa and rectal dialysates of patients with active inflammatory bowel disease, which contributes to neutrophil influx and colonic injury. The COX‐1/COX‐2 data highlight the ambiguous functional role of prostanoid pathways in inflammatory bowel diseases. (Inflamm Bowel Dis 2007)

Characterization of muscarinic receptors mediating contractions of circular and longitudinal muscle of human isolated colon.

1 The effects of seven muscarinic receptor antagonists were used to characterize the receptors which mediate carbachol‐evoked contractions of intertaenial circular and taenial longitudinal muscle in human isolated colon. The effects of these antagonists were studied upon colon contractions induced by cumulatively added carbachol which had mean EC50 values of 11.7 ± 2.3 μm (n = 8) and 12.6 ± 2.3 μm (n = 8) respectively upon circular and longitudinal smooth muscle. 2 All antagonists displaced concentration‐response curves to carbachol to the right in a parallel manner. The maximum concentration of each antagonist added (30 nm–10 μm) did not significantly suppress the maximum response. 3 In circular muscle, the M3 muscarinic receptor antagonists, 4‐diphenylacetoxy‐N‐methylpiperidine methiodide (4‐DAMP), hexahydrosiladiphenidol (HHSiD) and para‐fluoro‐hexahydrosiladiphenidol (p‐F‐HHSiD) inhibited responses with pA2 values of 9.41 ± 0.23, 7.17 ± 0.07, 6.94 ± 0.18 respectively. The M2 muscarinic receptor antagonist, AF‐DX 116, the M2/M4 muscarinic receptor antagonist, himbacine, and the M1 muscarinic receptor antagonist, pirenzepine, yielded pA2 values of 7.36 ± 0.43, 7.47 ± 0.14 and 7.23 ± 0.48 respectively. The non‐selective antagonist, atropine, had a pA2 of 8.72 ± 0.28. 4 In longitudinal muscle 4‐DAMP, HHSiD, p‐F‐HHSiD, AF‐DX 116, himbacine and pirenzepine gave pA2 values of 9.09 ± 0.16, 7.45 ± 0.43, 7.44 ± 0.21, 6.44 ± 0.1, 7.54 ± 0.40, 6.87 ± 0.38 respectively. Atropine yielded a pA2 value of 8.60 ± 0.08. 5 The pharmacological profile of antagonist affinities at the muscarinic receptor population responding to muscarinic agonist‐evoked contraction is similar to that widely accepted as characterizing the activation of an M3 muscarinic receptor subtype, although pA2 values of some antagonists are lower than that seen in other investigations.

Regulation of noradrenaline overflow in rat cerebral cortex by prostaglandin E2.

1 The effects of prostaglandin E2 (PGE2), PGF2α and PGI2 and of inhibitors of prostaglandin synthesis and action, on the K+‐evoked [3H]‐noradrenaline ([3H]‐NA) overflow from rat cerebral cortex slices have been investigated. 2 PGE2 reduced, while indomethacin (a prostaglandin synthesis inhibitor) or SC 19220 (a prostaglandin receptor antagonist) increased, the evoked overflow compared with controls. 3 The inhibition of [3H]‐NA overflow by PGE2 was dose‐dependently antagonized by SC 19220. 4 The results indicate that PGE2 modulates NA release in rat cerebral cortex in vitro.

Differences in response to 5-HT4 receptor agonists and antagonists of the 5-HT4-like receptor in human colon circular smooth muscle.

1 In isolated circular smooth muscle strips of human colon 5‐hydroxytryptamine (5‐HT) produced a concentration‐related inhibition of spontaneous motility. 2 The azabicycloalkyl benzimidazolones, BIMU 8 and BIMU 1, which have 5‐HT4 receptor stimulant properties, inhibited motility with EC50 values of 0.76 μM and 3.19 μM respectively and their Emax values were not significantly different from 5‐HT (EC50, 0.13 μM). 3 The 5‐HT4 receptor antagonist, DAU 6285 (1–10 μM), displaced the 5‐HT concentration‐response curve to the right in a parallel concentration‐dependent manner without depressing the maximum. The Schild plot was linear and the slope did not differ significantly from unity giving a pA2 value of 6.32. 4 The high affinity selective 5‐HT4 receptor antagonist, GR 113808, at a concentration of 3 nM displaced the 5‐HT concentration‐response curve in a parallel manner giving an apparent pKB estimate of 8.9 ±0.24. However, higher concentrations of 10–100 nM GR 113808 did not result in a further significant displacement of the 5‐HT concentration‐response curve and there was no suppression of Emax. 5 GR 113808 (10 nM) also caused a parallel displacement of the concentration‐response curve to the 5‐HT4 receptor agonist, 5‐methoxytryptamine (5‐MeOT) giving apparent pKB values ranging from 8.3–9.3. 6 GR 113808 (3–100 nM) failed to displace 5‐HT or 5‐MeOT concentration‐response curves in tissue strips from 3 patients out of a total of 10 patients studied in whom the response to 5‐HT and 5‐MeOT was normal. 7 The 5‐HT4 receptor antagonist, SDZ 205–557 (0.3–10 μM), had no significant effect on 5‐HT‐induced inhibition of spontaneous motility. 8 The present results are discussed in the light of variability of response to GR 113808 and SDZ 205–557 in other tissues. 9 Overall, our data indicate that human colon circular smooth muscle can be regarded as a site in which 5‐HT4‐like receptors are present but it is as yet unclear whether these results are also an indication of receptor variation.

The role of nitric oxide in mediating non-adrenergic non-cholinergic relaxation in longitudinal muscle of human taenia coli

Electrical field stimulation (EFS) of isolated longitudinal muscle of human taenia coli at 4Hz produced relaxation which was abolished by tetrodotoxin but not adrenergic and cholinergic blockade (NANC-relaxation). NG-nitro L-arginine (L-NOARG; 1-100 microM), an NO synthesis inhibitor, produced a concentration-dependent partial inhibition of the NANC response; 10 microM L-NOARG inhibited EFS-induced relaxation by 48.6 +/- 5.20% and 100 microM L-NOARG by 54.2 +/- 10.1%. L-Arginine (1mM), but not D-arginine (1mM) partially reversed the inhibitory effect and this was inversely proportional to the concentration of L-NOARG used. Cumulative administration of NO (acidified sodium nitrite solution; 1-100 microM) produced a concentration-dependent relaxation of the strips. L-NOARG (1 mM) did not affect either NO or isoprenaline-induced relaxations. These results provide the first preliminary evidence that NO is partially responsible for the NANC inhibitory transmission in the longitudinal muscle of the taenia coli of human colon.

Prostaglandin-induced abortion and outcome of subsequent pregnancies: a prospective controlled study.

We analysed a prospective series of 204 pregnancies occurring in 168 women after a prostaglandin-induced abortion. The mean (±standard error of mean) interval between abortion and first subsequent conception was 10·4 ± 0·6 months; no patient reported secondary subfertility. Fifty-five of the subsequent pregnancies were terminated, 23 during the second trimester, again using prostaglandins. Of the 149 pregnancies not terminated, 127 were delivered at term, and 19 spontaneously aborted, seven during the second trimester; there was one missed abortion and two ectopic pregnancies. Morbidity in the 127 term pregnancies was infrequent; spontaneous preterm labour occurred in three patients, and four singleton infants weighed less than 2500 g at birth. There was no apparent association between morbidity in the subsequent pregnancies and the period of gestation at the time of the previous abortion, route of prostaglandin administration, or need for post-abortion curettage. The results obtained overall were very similar to a control group of 612 women consecutively admitted for delivery or abortion to the Oxford obstetrical and gynaecological units. There was, however, an increased incidence of spontaneous abortion and placenta praevia after prostaglandin-induced abortion, and the multigravidae in that group had a longer average duration of labour than the control group. Sixty-five per cent of the post-abortion pregnancies were unplanned compared with 36% of the control group.

The effect of prostaglandin E2 infusion in the fetal lamb on fetal plasma acth, prolactin and cortisol concentrations

PGE2 (2 micrograms/min) has been infused for 1h into the fetal jugular vein of 8 chronically catheterized fetuses on 13 occasions from 112 to 138 days gestation. Infusion of ethanol vehicle alone was conducted in fetuses from 111-139 days gestation. PGE2 administration produced a significant increase in fetal plasma cortisol after 30 min. No significant change was observed in fetal plasma prolactin concentration. Fetal plasma ACTH concentration was significantly elevated above resting concentration after 30 min. of PGE2 infusion. Metabolic clearance rate of PGE2 was 860 ml/min or 350 ml/kg/min. Intrauterine pressure was not changes during the infusion at any gestational age.

Synthesis of prostaglandins by the human uterine cervix in vitro during passive mechanical stretch

Arachidonic acid and prostaglandins (PGs) have a variety of effects on the human uterine cervix. In vitro, these include alterations in motility (Najak et a1 1970) and the biochemical modification of the cervix matrix proteins (Hillier & Wallis 1981; Norstrom et all981; Wilhelmsson et a1 1981). In vivo, PGs may cause a beneficial softening of the cervix at term in humans (Mackenzie 1981) and laboratory animals (Stys et a1 1981) thus aiding the processes of labour. The human cervix synthesizes PGs in vitro (Ellwood et a1 1980; Hillier & Wallis 1981) and if this occurs in vivo it may help remodel the cervical tissue structure at term. Because uterine contractility stretches the cervix and contractions not felt by the pregnant mother can occur well before the clinically identifiable onset of labour (Anderson & Turnbull 1969) we investigated the effect of passive mechanical stretching on PG synthesis by human isolated uterine cervix.