We investigated the eect of the cannabinoid agonist (+)WIN-55212-2 on human ileum longitudinal smooth muscle preparations, either electrically stimulated or contracted by carbachol. Electrical ®eld stimulation mostly activated cholinergic neurons, since atropine and tetrodotoxin (TTX), alone or coincubated, reduced twitch responses to a similar degree (85%). (+)WIN-55212-2 concentrationdependently inhibited twitch responses (IC 50 73 nM), but had no additive eect with atropine or TTX. The cannabinoid CB 1 receptor antagonist SR 141716 (pA 2 8.2), but not the CB 2 receptor antagonist, SR 144528, competitively antagonized twitch inhibition by (+)WIN-55212-2. Atropine but not (+)WIN-55212-2 or TTX prevented carbachol-induced tonic contraction.These results provide functional evidence of the existence of prejunctional cannabinoid CB 1 -receptors in the human ileum longitudinal smooth muscle. Agonist activation of these receptors prevents responses to electrical ®eld stimulation, presumably by inhibiting acetylcholine release. SR 141716 is a potent and competitive antagonist of cannabinoid CB 1 receptors naturally expressed in the human gut.

The role played by the mechanical tissue stress in supporting lymph formation and propulsion in thoracic tissues was studied in deeply anesthetized rats (n = 13) during spontaneous breathing or mechanical ventilation. After arterial and venous catheterization and insertion of an intratracheal cannula, fluorescent dextrans were injected intrapleurally to serve as lymphatic markers. After 2 h, the fluorescent intercostal lymphatics were identified, and the hydraulic pressure in lymphatic vessels (P lymph) and adjacent interstitial space (P int) was measured using micropuncture. During spontaneous breathing, end-expiratory P lymph and corresponding P int were -2.5 +/- 1.1 (SE) and 3.1 +/- 0.7 mmHg (P < 0.01), which dropped to -21.1 +/- 1.3 and -12.2 +/- 1.3 mmHg, respectively, at end inspiration. During mechanical ventilation with air at zero end-expiratory alveolar pressure, P lymph and P int were essentially unchanged at end expiration, but, at variance with spontaneous breathing, they increased at end inspiration to 28.1 +/- 7.9 and 28.2 +/- 6.3 mmHg, respectively. The hydraulic transmural pressure gradient (DeltaP tm = P lymph - P int) was in favor of lymph formation throughout the whole respiratory cycle (DeltaP tm = -6.8 +/- 1.2 mmHg) during spontaneous breathing but not during mechanical ventilation (DeltaP tm = -1.1 +/- 1.8 mmHg). Therefore, data suggest that local tissue stress associated with the active contraction of respiratory muscles is required to support an efficient lymphatic drainage from the thoracic tissues.

The data revealed a great functional complexity of the diaphragmatic lymphatic network and suggested that cardiogenic oscillations may play an important role in promoting lymph formation and propulsion from interstitial tissues with subatmospheric tissue pressure.

Background and aims: Diverticulosis is a common disease of not completely defined pathogenesis. Motor abnormalities of the intestinal wall have been frequently described but very little is known about their mechanisms. We investigated in vitro the neural response of colonic longitudinal muscle strips from patients undergoing surgery for complicated diverticular disease (diverticulitis). Methods: The neural contractile response to electrical field stimulation of longitudinal muscle strips from the colon of patients undergoing surgery for colonic cancer or diverticulitis was challenged by different receptor agonists and antagonists. Results: Contractions of colonic strips from healthy controls and diverticulitis specimens were abolished by atropine. The b adrenergic agonist (2) isoprenaline and the tachykinin NK 1 receptor antagonist SR140333 had similar potency in reducing the electrical twitch response in controls and diseased tissues, while the cannabinoid receptor agonist (+)WIN 55,212-2 was 100 times more potent in inhibiting contractions in controls (IC50 42 nmol/l) than in diverticulitis strips. SR141716, a selective antagonist of the cannabinoid CB 1 receptor, had no intrinsic activity in control preparations but potentiated the neural twitch in diseased tissues by up to 196% in a concentration dependent manner. SR141716 inhibited (+)WIN 55,212-2 induced relaxation in control strips but had no efficacy on (+)WIN 55,212-2 responses in strips from diverticular disease patients. Colonic levels of the endogenous ligand of cannabinoid and vanilloid TRPV1 receptors anandamide were more than twice those of control tissues (54 v 27 pmol/g tissue). The axonal conduction blocker tetrodotoxin had opposite effects in the two preparations, completely inhibiting the contractions of control strips but potentiating those in diverticular preparations, an effect selectively inhibited by SR140333. Conclusions: Neural control of colon motility is profoundly altered in patients with diverticulitis. Their raised levels of anandamide, apparent desensitisation of the presynaptic neural cannabinoid CB 1 receptor, and the SR141716 induced intrinsic response, suggest that endocannabinoids may be involved in the pathophysiology of complications of colonic diverticular disease.

In seven anesthetized rabbits we measured the size, shape, and density of lymphatic stomata on the peritoneal and pleural sides of the diaphragm. The diaphragm was fixed in situ and processed for scanning electron microscopy. Results are from 2,902 peritoneal and 3,086 pleural fields (each 1,620 microns 2) randomly chosen from the various specimens. Stomata were seen in 9% of the fields examined, and in 30% of the cases they appeared grouped in clusters with 2-14 stomata/field. Stoma density was 250 +/- 242 and 72 +/- 57 (SD) stomata/mm2 on peritoneal and pleural sides, respectively, and it was similar over the muscular and tendinous portion of the two surfaces. The maximum diameter ranged from less than 1 to approximately 30 microns, with an average value of 1.2 +/- 3.1 micron. The ratio of the maximum to the minimum diameter and the surface area averaged 2 +/- 1.4 and 0.7 +/- 2.4 micron 2, respectively. The maximum and minimum diameter and surface area values followed a lognormal frequency distribution, suggesting that stomata geometry is affected by diaphragmatic tension.

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1 Human in vitro preparations of transverse or distal colonic circular smooth muscle were potently and dose-dependently contracted by neurokinin A (EC 50 , 4.9 nM), the tachykinin NK 2 -receptor selective agonist (SR 144743) and N,N-demethyl (SR 144782) metabolites, were used to challenge agonist responses, as appropriate. SR 144190 and the metabolites all potently and competitively antagonized the response to [b-Ala 8 ]NKA (4 ± 10), with similar potency (Schild plot pA 2 values 9.4, 9.4 and 9.3, slope=1). SR 48968 antagonism was not competitive: the Schild plot slope was biphasic with a high (X intercept*9.3) and a low (X intercept 8.4, slope 1.6) a nity site. Co-incubation of SR 48968 (10, 100 nM) and SR 144782 (10 nM) produced additive e ects; in this experimental condition, SR 48968 apparent a nity (pK B ) was 8.2. In addition, SR 144782 (0.1 mM) antagonized responses to neurokinin A, substance P and neurokinin B, with pK B consistent with its a nity for tachykinin NK 2 -receptors. The potent and selective NK 1 and NK 3 -receptor antagonists, SR 140333 and SR 142801 (both 0.1 mM), failed to inhibit contractions induced by SP or NKB. 4 In conclusion, the in vitro mechanical responses of circular smooth muscle preparations from human colon are strongly consistent with the presence of non-neuronal tachykinin NK 2 -receptors, but not tachykinin NK 1 -or NK 3 -receptors. Our ®ndings with SR 48968 suggest the existence of two tachykinin NK 2 -receptor subtypes, that it seems to distinguish, unlike SR 144190 and its metabolites. However, the precise nature of SR 48968 allotopic antagonism remains to be elucidated, since allosteric e ects at the tachykinin NK 2 -receptor might well account for the complexity of the observed interaction.

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