Luisa Pinto

Cannabinoid CB1‐receptor mediated regulation of gastrointestinal motility in mice in a model of intestinal inflammation

We have studied the effect of cannabinoid agonists (CP 55,940 and cannabinol) on intestinal motility in a model of intestinal inflammation (induced by oral croton oil in mice) and measured cannabinoid receptor expression, endocannabinoids (anandamide and 2‐arachidonylglycerol) and anandamide amidohydrolase activity both in physiological and pathophysiological states. CP 55,940 (0.03 – 10 nmol mouse−1) and cannabinol (10 – 3000 nmol mouse−1) were more active in delaying intestinal motility in croton oil‐treated mice than in control mice. These inhibitory effects were counteracted by the selective cannabinoid CB1 receptor antagonist SR141716A (16 nmol mouse−1). SR141716A (1 – 300 nmol mouse−1), administered alone, increased intestinal motility to the same extent in both control and croton oil‐treated mice Croton oil‐induced intestinal inflammation was associated with an increased expression of CB1 receptor, an unprecedented example of up‐regulation of cannabinoid receptors during inflammation. High levels of anandamide and 2‐arachidonylglycerol were detected in the small intestine, although no differences were observed between control and croton oil‐treated mice; by contrast anandamide amidohydrolase activity increased 2 fold in the inflamed small intestine. It is concluded that inflammation of the gut increases the potency of cannabinoid agonists possibly by ‘up‐regulating’ CB1 receptor expression; in addition, endocannabinoids, whose turnover is increased in inflamed gut, might tonically inhibit intestinal motility.

Phytochemical compounds involved in the anti-inflammatory effect of propolis extract

Two ethanolic propolis extracts (EPE) with and without the caffeic acid phenethyl ester (CAPE), CAPE and galangin (major components of propolis) were investigated for anti-inflammatory activity in rats using carrageenin foot oedema, carrageenin pleurisy and adjuvant arthritis. In our experiments, EPE with CAPE and CAPE alone significantly inhibited carrageenin oedema, carrageenin pleurisy and adjuvant arthritis. In contrast EPE without CAPE and galangin did not exhibit anti-inflammatory effects in acute and chronic inflammation. These results suggest that the anti-inflammatory activity of propolis is due to CAPE.

An endogenous cannabinoid tone attenuates cholera toxin-induced fluid accumulation in mice.

BACKGROUND & AIMS Cholera toxin (CT) is the most recognizable enterotoxin causing secretory diarrhea, a major cause of infant morbidity and mortality throughout the world. In this study, we investigated the role of the endogenous cannabinoid system (i.e., the cannabinoid receptors and their endogenous ligands) in CT-induced fluid accumulation in the mouse small intestine. METHODS Fluid accumulation was evaluated by enteropooling; endocannabinoid levels were measured by isotope-dilution gas chromatography mass spectrometry; CB(1) receptors were localized by immunohistochemistry and their messenger RNA (mRNA) levels were quantified by reverse-transcription polymerase chain reaction (PCR). RESULTS Oral administration of CT to mice resulted in an increase in fluid accumulation in the small intestine and in increased levels of the endogenous cannabinoid, anandamide, and increased expression of the cannabinoid CB(1) receptor mRNA. The cannabinoid receptor agonist CP55,940 and the selective cannabinoid CB(1) receptor agonist arachidonoyl-chloro-ethanolamide inhibited CT-induced fluid accumulation, and this effect was counteracted by the CB(1) receptor antagonist SR141716A, but not by the CB(2) receptor antagonist SR144528. SR141716A, per se, but not the vanilloid VR1 receptor antagonist capsazepine, enhanced fluid accumulation induced by CT, whereas the selective inhibitor of anandamide cellular uptake, VDM11, prevented CT-induced fluid accumulation. CONCLUSIONS These results indicate that CT, along with enhanced intestinal secretion, causes overstimulation of endocannabinoid signaling with an antisecretory role in the small intestine.

Phytotherapy and quality of herbal medicines.

The extensive use of plants as medicines has pointed out that herbal medicines are not as safe as frequently claimed. Therefore, it can be harmful to take herbal medicines without being aware of their potential adverse effects. A comprehensive surveillance system for monitoring the adverse effects of herbal medicines is now essential. Also knowledge of the correlation existing between the marker compounds and other ingredients that occur in plants is a necessary requirement for ensuring the efficacy and quality of the herbal medicines.

The role of cannabinoid receptors in intestinal motility, defaecation and diarrhoea in rats

We have studied the effects of the cannabinoid receptor agonists (R)-(+)[2,3-dihydro-5-methyl-3-(4-morpholinylmethyl)pyrrolo[1,2, 3-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone (WIN 55,212-2, 0. 3-5 mg/kg, i.p.) and (-)-cis-3-[2-hydroxy-4-(1, 1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl)cyclohexanol) (CP 55,940, 0.03-1 mg/kg, i.p.), the cannabinoid CB(1) receptor antagonist (N-piperidin-1-yl)-5-(4-chlorophenyl)-1-2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (SR141716A, 0. 3-5 mg/kg, i.p.) and the cannabinoid CB(2) receptor antagonist N-[-(1S)-endo-1,3,3-trimethyl bicyclo [2.2.1] heptan-2-yl]-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazo le- 3-carboxamide (SR144528, 1 mg/kg, i.p.) on intestinal motility, defaecation and castor-oil (1 ml/100 g rat, orally)-induced diarrhoea in the rat. SR141716A, but not SR144528, increased defaecation and upper gastrointestinal transit, while WIN 55,212-2 and CP 55,940 decreased upper gastrointestinal transit but not defaecation. WIN 55,212-3 (5 mg/kg), the less active enantiomer of WIN 55,212-2, was without effect. A per se non-effective dose of SR141716A (0.3 mg/kg), but not of SR144528 (1 mg/kg) or the opioid receptor antagonist, naloxone (2 mg/kg i.p.), counteracted the inhibitory effect of both WIN 55,212-2 (1 mg/kg) and CP 55,940 (0.1 mg/kg) on gastrointestinal motility. WIN 55,212-2 did not modify castor-oil-induced diarrhoea, while CP 55,940 produced a transient delay in castor-oil-induced diarrhoea at the highest dose tested (1 mg/kg), an effect counteracted by SR141715A (5 mg/kg). These results suggest that (i) intestinal motility and defaecation could be tonically inhibited by the endogenous cannabinoid system, (ii) exogenous activation of cannabinoid CB(1) receptors produces a reduction in intestinal motility in the upper gastrointestinal tract but not in defaecation, (iii) endogenous or exogenous activation of cannabinoid CB(2) receptors does not affect defaecation or intestinal motility and (iv) the cannabinoid receptor agonist, CP 55, 940, possesses a weak and transient antidiarrhoeal effect while the cannabinoid receptor agonist, WIN 55,212-2, does not possess antidiarrhoeal activity.

Central and peripheral cannabinoid modulation of gastrointestinal transit in physiological states or during the diarrhoea induced by croton oil

We have evaluated the effect of cannabinoid drugs, administered intraperitoneally (i.p.) or intracerebroventricularly (i.c.v.) on upper gastrointestinal transit in control and in croton oil‐treated mice. The cannabinoid agonists, WIN 55,212‐2 (2–239 nmol mouse−1) and cannabinol (24–4027 nmol mouse−1), decreased while the CB1 antagonist SR141716A (2–539 nmol mouse−1) increased transit in control mice. WIN 55,212‐2, cannabinol and SR141716A had lower ED50 values when administered i.c.v., than when administered i.p. The CB2 antagonist SR144528 (52 nmol mouse−1, i.p.) was without effect. During croton oil (0.01 ml mouse−1, p.o.)‐induced diarrhoea, the ED50 values of i.p.‐injected WIN 55,212‐2 and cannabinol (but not SR141716A) were significantly decreased (compared to control mice). However, the ED50 values of WIN 55,212‐2 were similar after i.p. or i.c.v. administration. The inhibitory effects of WIN 55,212‐2 and cannabinol were counteracted by SR141716A (16 nmol mouse−1, i.p.) but not by SR144528 (52 nmol mouse−1, i.p.) both in control and croton‐oil treated mice. Ganglionic blockade with hexamethonium (69 nmol mouse−1, i.p.) did not modify the inhibitory effect of i.p.‐injected cannabinoid agonists either in control or in croton‐oil treated mice. The lower ED50 values of cannabinoid drugs after i.c.v. administration suggest a central (CB1) site of action. However, a peripheral site of action is suggested by the lack of effect of hexamethonium. In addition, croton oil‐induced diarrhoea enhances the effect of cannabinoid agonists by a peripheral mechanism.

The endocannabinoid system and the molecular basis of paralytic ileus in mice

The endocannabinoid system (i.e., the cannabinoid receptors and their endogenous ligands) plays an important role in the physiological control of intestinal motility. However, its participation in intestinal pathological states is still poorly understood. In the present study, we investigated the possible role of the endocannabinoid system in the pathogenesis of paralytic ileus, a pathological state consisting of decreased intestinal motility following peritonitis, surgery, or other noxious situations. Ileus was induced by i.p. administration of acetic acid, and gastrointestinal propulsion was assessed by the charcoal method. Endocannabinoid levels were measured by isotope‐dilution gas chromatography‐mass spectrometry, whereas cannabinoid CB1 receptors were identified by immunohistochemistry. Acetic acid administration inhibited gastrointestinal transit (ileus), and this effect was accompanied by increased levels of the endocannabinoid anandamide compared with control mice and by overexpression of CB1 receptors in myenteric nerves. Furthermore, acetic acid‐induced ileus was alleviated by the CB1 receptor antagonist SR141716A and worsened by VDM11, a selective inhibitor of anandamide cellular uptake (and hence inactivation). From these findings, it can be concluded that the intestinal hypomotility typical of paralytic ileus is due, at least in part, to the enhancement of anandamide levels and CB1 expression during this condition, and that selective, nonpsychotropic CB1 receptor antagonists could represent new drugs to treat this disorder.

In-vivo and in-vitro anti-inflammatory effect of echinacea purpurea and Hypericum perforatum

Echinacea purpurea (L.) Moench and Hypericum perforatum (L.) were evaluated for their anti‐inflammatory activity against carrageenan‐induced paw oedema in mice. Each drug was administered orally to mice at 30 and 100 mg kg−1, twice daily. Only the higher dose significantly inhibited, time dependently, the formation of oedema, evaluated as area under the curve (echinacea P < 0.01; hypericum P < 0.05). Western blot analysis showed that in‐vivo treatment with these extracts could modulate lipopolysaccharide (LPS) and interferon‐γ induced cyclooxygenase‐2 (COX‐2) and inducible nitric oxide synthase (iNOS) expression in peritoneal macrophages. In particular, treatment with 100 mg kg−1 hypericum inhibited both iNOS and COX‐2 expression, whereas treatment with 100 mg kg−1 echinacea down‐regulated only COX‐2 expression. The present study suggests that the anti‐inflammatory effect of these extracts could be in part related to their modulation of COX‐2 expression.

Effect of vanilloid drugs on gastrointestinal transit in mice

We have studied the effect of capsaicin, piperine and anandamide, drugs which activate vanilloid receptors and capsazepine, a vanilloid receptor antagonist, on upper gastrointestinal motility in mice. Piperine (0.5 – 20 mg kg−1 i.p.) and anandamide (0.5 – 20 mg kg−1 i.p.), dose‐dependently delayed gastrointestinal motility, while capsaicin (up to 3 mg kg−1 i.p.) was without effect. Capsazepine (15 mg kg−1 i.p.) neither per se affected gastrointestinal motility nor did it counteract the inhibitory effect of both piperine (10 mg kg−1) and anandamide (10 mg kg−1). A per se non effective dose of SR141716A (0.3 mg kg−1 i.p.), a cannabinoid CB1 receptor antagonist, counteracted the inhibitory effect of anandamide (10 mg kg−1) but not of piperine (10 mg kg−1). By contrast, the inhibitory effect of piperine (10 mg kg−1) but not of anandamide (10 mg kg−1) was strongly attenuated in capsaicin (75 mg kg−1 in total, s.c.)‐treated mice. Pretreatment of mice with NG‐nitro‐L‐arginine methyl ester (25 mg kg−1 i.p.), yohimbine (1 mg kg−1, i.p.), naloxone (2 mg kg−1 i.p.), or hexamethonium (1 mg kg−1 i.p.) did not modify the inhibitory effect of both piperine (10 mg kg−1) and anandamide (10 mg kg−1). The present study indicates that the vanilloid ligands anandamide and piperine, but not capsaicin, can reduce upper gastrointestinal motility. The effect of piperine involves capsaicin‐sensitive neurones, but not vanilloid receptors, while the effect of anandamide involves cannabinoid CB1, but not vanilloid receptors.

Aloe and its therapeutic use

Aloe is one of the few medicinal plants that has maintained its popularity for a long period of time. Aloe latex is used for its laxative effect and should be distinguished from aloe gel, used both in cosmetics and ointments for skin ailments. Aloe whole leaf is another preparation used internally as a drink in a wide range of human diseases including cancer, AIDS, ulcerative colitis and other disturbances. The concomitant use of honey may make the aloe whole leaf therapy more palatable and efficient.