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This study evaluated the contribution of endothelins to changes in sensitivity to mechanical stimulation of the lower abdomen and hind paw associated with 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis. The frequency of withdrawal responses to 10 consecutive applications of von Frey probes to the lower abdomen (0.07 g) or hind paw (0.4 g) was assessed in male BALB/c mice before and after intracolonic TNBS injection (0.5 mg in 100 microL of 35% ethanol). TNBS (0.5 mg) induced referred mechanical hyperalgesia in the abdomen (response frequencies at 24 h: saline 11.0% +/- 3.1%, TNBS 48.0% +/- 6.9%) and hind paw (frequencies at 24 h: saline 12.5% +/- 4.7%, TNBS 47.1% +/- 7.1%) lasting up to 72 and 48 h, respectively. Mice receiving 1.0 or 1.5 mg TNBS assumed hunch-backed postures and became immobile during abdominal mechanical stimulation, suggestive of excessive ongoing pain. Atrasentan (ETA receptor antagonist; 10 and 30 mg/kg, i.v.) given 24 h after TNBS abolished hind paw and abdominal mechanical hyperalgesia for 2-3 h. A-192621 (ETB receptor antagonist; 20 mg/kg, i.v.) attenuated abdominal mechanical hyperalgesia at the 3 h time point only. Thus, endothelins contribute importantly to abdominal and hind paw referred mechanical hyperalgesia during TNBS-induced colitis mainly through ETA receptor-signaled mechanisms.

The relationships between changes in intracellular Ca2+ and smooth muscle tension triggered by endothelin-1 and the selective endothelin ETB receptor agonist sarafotoxin S6c, as well as their susceptibility to modification by the nonselective cyclooxygenase blocker indomethacin, were assessed in guinea pig isolated gallbladder strips. Cumulative additions of either agonist (1, 10, and 100 nM) induced simultaneous graded, strongly correlated, slowly developing, and sustained changes in tension and intracellular Ca+2 (Fura-2 technique). Sarafotoxin S6c was more effective than endothelin-1 in raising intracellular Ca2+ at 1 or 10 nM, but their abilities to cause contractions were similar at all concentrations. Indomethacin (5.6 microM) markedly inhibited the changes in both intracellular Ca2+ and tension caused by all concentrations of sarafotoxin S6c (in response to 100 nM, increases in Ca2+ fluorescence intensity and tension were inhib ited from 7.7 +/- 0.7 to 4.0 +/- 0.4% and from 460 +/- 100 to 160 +/- 40 mg, respectively) but only reduced the contraction triggered by 100 nM endothelin-1 (from 560 +/- 100 to 230 +/- 70 mg). Endothelin-1 caused greater prostacyclin release from gallbladder than sarafotoxin S6c (at 100 nM, 6-keto-PGF1alpha levels in the medium rose 4.8- and 2.8-fold, respectively; P < 0.05) and slightly increased thromboxane A2 release (1.6-fold; P < 0.05). Thus, gallbladder contractions triggered by combined ETA/ETB or selective ETB receptor stimulation (with endothelin-1 or sarafotoxin S6c, respectively) are strongly correlated with increases in intracellular Ca2+ but differentially affected by indomethacin. It remains to be assessed if this difference is because endothelin-1 triggers greater prostacyclin release than sarafotoxin S6c and (or) is due to the coupling of ETA and ETB receptors to distinct patterns of generation of cyclooxygenase-derived eicosanoids.

Changes in responsiveness of the vas deferens and urinary bladder to bradykinin (BK) receptor agonists (Tyr8-BK and des-Arg9-BK), substance P (SP), and endothelin-1 (ET-1) were assessed 8 weeks after streptozotocin (STZ)-induced diabetes. Preparations from control or STZ-treated (60 mg/kg i.p.) male rats were tested for contractile and neurogenic twitch potentiating (TP, in VD only) effects of all four agonists (1 nM to 0.3 or 3 microM). In diabetic VD, contractile effects of Tyr8-BK, des-Arg9-BK, and SP were enhanced, but ET-1 effects were unchanged. In contrast, TP by des-Arg9-BK was unaffected, that by Tyr8-BK was decreased, and those by SP and ET-1 were increased. In diabetic UB, only contractions to des-Arg9-BK and SP were enhanced. Following insulin replacement (human, 1-3 U/day s.c.), starting 1 week after STZ, TP induced by Tyr8-BK and des-Arg9-BK in VD were further inhibited, but all other changes in both preparations were reversed at least partially. Insulin treatment of nondiabetic rats, however, also affected VD (but not UB) responsiveness, such that contractions to Tyr8-BK and TP by ET-1 were increased, but TP by Tyr8-BK was decreased. Thus, STZ-induced type I diabetes causes important alterations in responsiveness of non-vascular smooth muscle tissues of the rat to BK, SP, and ET-1. Long term insulin replacement, at doses normalising glycaemia, effectively reversed most changes in VD or UB responsiveness, but it is unclear if this is truly due to blocking of STZ-induced changes, since the treatment also affected responsiveness of nondiabetic tissues.

The present study investigates the influence of endothelin (ET) related peptides (0.3-30 pmol/paw) on both phases of nociception and on edema induced by intraplantar injection of formalin (0.5% in 20 microL) in the mouse hind paw. The first phase of nociception (0-5 min after injection) was significantly potentiated by simultaneous injection of either ET-1 (10 or 30 pmol/paw) or ET-3 (10 pmol/paw), but not of the selective ET3 receptor agonist sarafotoxin S6c (SRTX-c; up to 30 pmol/paw). All three peptides potentiated the second phase (10-30 min after injection) of formalin-induced nociception (at 3-30, 1-30, and 10-30 pmol/paw for ET-1, ET-3, and SRTX-c, respectively), whereas only ET-1 (10 or 30 pmol/paw) effectively enhanced edema caused by formalin (30 min after injection). Histamine also potentiated all three responses triggered by formalin, but was 30- to 100-fold less potent than ET-1. Treatment with the mixed ETA/ETB receptor antagonist bosentan (10 mg/kg i.p., 1 h beforehand) did not influence nociceptive and edematogenic responses to formalin or their potentiation by histamine (3 nmol/paw), but did inhibit the potentiations induced by ET-1 (10 pmol/paw). Thus, ET-1 potentiates formalin-induced nociception and edema in the mouse. These actions are possibly mediated via ETB and ETA receptors, respectively, but their true identity and the mechanisms involved still remain to be fully elucidated.