Biomedical subjects

Areles Molleman

Publications and source records attributed to Areles Molleman.

8 recordsLinked to original sources

PubMed2006

Cannabinoid signalling in TNF-alpha induced IL-8 release.

The molecular events mediating the immunomodulatory properties of cannabinoids have remained largely unresolved. We have therefore investigated the molecular mechanism(s) through which R-(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl] pyrrolo[1,2,3-de]-1,4-benzoxazinyl]-(1-napthanlenyl) methanone (WIN55212-2) modulate production of interleukin-8 (IL-8) in HT-29 cells. Release of IL-8 induced by tumor necrosis factor-alpha (TNF-alpha) was determined by enzyme-linked immunosorbent assay (ELISA). Changes in expression of inhibitory kappa B (IkappaB) were monitored by Western blotting and activation of nuclear factor-kappa B (NF-kappaB) was determined in electrophoretic mobility shift assay (EMSAs). TNF-alpha induced release of IL-8 was inhibited by WIN55212-2 which also blocked the degradation of IkappaB-alpha and activation of NF-kappaB induced by TNF-alpha. These data provide strong evidence that WIN55212-2 may modulate IL-8 release by negatively regulating the signaling cascade leading to the activation of NF-kappaB. These findings highlight a potential mechanism for the immunomodulatory properties of cannabinoids and contribute towards acquiring a clear understanding of the role of cannabinoids in inflammation.

Benzoxazines↗

PubMed2005

Cannabinoid signalling.

After their discovery, the two known cannabinoid receptors, CB(1) and CB(2), have been the focus of research into the cellular signalling mechanisms of cannabinoids. The initial assessment, mainly derived from expression studies, was that cannabinoids, via G(i/o) proteins, negatively modulate cyclic AMP levels, and activate inward rectifying K(+) channels. Recent findings have complicated this assessment on different levels: (1) cannabinoids include a wide range of compounds with varying profiles of affinity and efficacy at the known CB receptors, and these profiles do not necessarily match their biological activity; (2) CB receptors appear to be intrinsically active and possibly coupled to more than one type of G protein; (3) CB receptor signalling mechanisms are diverse and dependent on the system studied; (4) cannabinoids have other targets than CB receptors. The aim of this mini review is to discuss the current literature regarding CB receptor signalling pathways. These include regulation of adenylyl cyclase, MAP kinase, intracellular Ca(2+), and ion channels. In addition, actions of cannabinoids that are not mediated by CB(1) or CB(2) receptors are discussed.

Adenylyl Cyclases↗

PubMed2005

The responses to manipulation of extracellular and intracellular calcium are altered in the streptozotocin-diabetic rat colon and ileum.

Studies were performed to see if alterations in Ca2+ homeostasis underlie the gastrointestinal motility complications seen in many diabetic patients. Experiments were performed on colonic and ileal tissues taken from streptozotocin-induced diabetic and control rats. Diabetes caused alterations in the responses of the tissues to Ca2+ manipulation but these differed between the colon and ileum. In the colon a small but not significant increase in contractile responses to CaCl2 was observed in diabetic tissues, whereas the responses of the ileum were depressed relative to those of the controls. In contrast, responses of the diabetic ileum to the Ca2+ channel agonist Bay K8644 were greater than those of the controls, whilst the agonist failed to contract the colon. Similarly, the Ca2+-ATPase inhibitors, thapsigargin and cyclopiazonic acid, produced contractions which were greater in diabetic ileal tissues. Thus, alterations in the responses of the diabetic gut to Ca2+ manipulation are complex, and also tissue-specific.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed2005

Arachidonic acid mediates non-capacitative calcium entry evoked by CB1-cannabinoid receptor activation in DDT1 MF-2 smooth muscle cells.

Cannabinoid CB1-receptor stimulation in DDT1 MF-2 smooth muscle cells induces a rise in [Ca2+]i, which is dependent on extracellular Ca2+ and modulated by thapsigargin-sensitive stores, suggesting capacitative Ca2+ entry (CCE), and by MAP kinase. Non-capacitative Ca2+ entry (NCCE) stimulated by arachidonic acid (AA) partly mediates histamine H1-receptor-evoked increases in [Ca2+]i in DDT1 MF-2 cells. In the current study, both Ca2+ entry mechanisms and a possible link between MAP kinase activation and increasing [Ca2+]i were investigated. In the whole-cell patch clamp configuration, the CB-receptor agonist CP 55, 940 evoked a transient, Ca2+-dependent K+ current, which was not blocked by the inhibitors of CCE, 2-APB, and SKF 96365. AA, but not its metabolites, evoked a transient outward current and inhibited the response to CP 55,940 in a concentration-dependent manner. CP 55,940 induced a concentration-dependent release of AA, which was inhibited by the CB1 antagonist SR 141716. The non-selective Ca2+ channel blockers La3+ and Gd3+ inhibited the CP 55,940-induced current at concentrations that had no effect on thapsigargin-evoked CCE. La3+ also inhibited the AA-induced current. CP 55,940-induced AA release was abolished by Gd3+ and by phospholipase A2 inhibition using quinacrine; this compound also inhibited the outward current. The CP 55,940-induced AA release was strongly reduced by the MAP kinase inhibitor PD 98059. The data suggest that in DDT1 MF-2 cells, AA is an integral component of the CB1 receptor signaling pathway, upstream of NCCE and, via PLA2, downstream of MAP kinase.

Arachidonic Acid↗

PubMed2003

Pharmacological characterisation of cannabinoid receptors inhibiting interleukin 2 release from human peripheral blood mononuclear cells.

The effects of a range of cannabinoid receptor agonists and antagonists on phytohaemagglutinin-induced secretion of interleukin-2 from human peripheral blood mononuclear cells were investigated. The nonselective cannabinoid receptor agonist WIN55212-2 ((R)-(+)-[2,3-dihydro-5-methyl-3-[4-morpholinylmethyl]pyrrolo[1,2,3-de]1,4-benzoxazin-6-yl](1-naphthyl) methanone mesylate) and the selective cannabinoid CB(2) receptor agonist JWH 015 ((2-methyl-1-propyl-1H-indol-3-yl)-1-napthalenylmethanone) inhibited phytohaemagglutinin (10 microg/ml)-induced release of interleukin-2 in a concentration-dependent manner (IC(1/2max), WIN55212-2=8.8 x 10(-7) M, 95% confidence limits (C.L.)=2.2 x 10(-7)-3.5 x 10(-6) M; JWH 015=1.8 x 10(-6) M, 95% C.L.=1.2 x 10(-6)-2.9 x 10(-6) M, n=5). The nonselective cannabinoid receptor agonists CP55,940 ((-)-3-[2-hydroxy-4-(1,1-dimethyl-hepthyl)-phenyl]4-[3-hydroxypropyl]cyclo-hexan-1-ol), Delta(9)-tetrahydrocannabinol and the selective cannabinoid CB(1) receptor agonist ACEA (arachidonoyl-2-chloroethylamide) had no significant (P>0.05) inhibitory effect on phytohaemagglutinin-induced release of interleukin-2. Dexamethasone significantly (P<0.05) inhibited phytohaemagglutinin-induced release of interleukin-2 in a concentration-dependent manner (IC(1/2max)=1.3 x 10(-8) M, 95% C.L.=1.4 x 10(-9)-3.2 x 10(-8) M). The cannabinoid CB(1) receptor antagonist SR141716A (N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride) (10(-6) M) did not antagonise the inhibitory effect of WIN55212-2 whereas the cannabinoid CB(2) receptor antagonist SR144528 (N-(1,S)-endo-1,3,3-trimethyl bicyclo(2,2,1)heptan-2-yl)-5-(4-chloro-3-methylphenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide) antagonised the inhibitory effect of WIN55212-2 (pA(2)=6.3+/-0.1, n=5). In addition, CP55,940 (10(-6) M) and Delta(9)-tetrahydrocannabinol (10(-6) M) also antagonised the inhibitory effects of WIN55212-2 (pA(2)=6.1+/-0.1, n=5 and pA(2)=6.9+/-0.2, n=5). In summary, WIN55,212-2 and JWH 015 inhibited interleukin-2 release from human peripheral blood mononuclear cells via the cannabinoid CB(2) receptor. In contrast, CP55,940 and Delta(9)-tetrahydrocannabinol behaved as partial agonists/antagonists in these cells.

Arachidonic Acids↗

PubMed2003

Inhibition of interleukin-8 release in the human colonic epithelial cell line HT-29 by cannabinoids.

We have investigated the effects of cannabinoid agonists and antagonists on tumour necrosis factor-alpha (TNF-alpha)-induced secretion of interleukin-8 from the colonic epithelial cell line, HT-29. The cannabinoid receptor agonists [(-)-3-[2-hydroxy-4-(1,1-dimethyl-heptyl)-phenyl]4-[3-hydroxypropyl]cyclo-hexan-1-ol] (CP55,940); Delta-9-tetrahydrocannabinol; [R(+)-[2,3-dihydro-5-methyl-3-[(morpholinyl) methyl] pyrrolo[1,2,3-de]1,4-benzoxazin-6-yl](1-naphthyl) methanone mesylate] (WIN55,212-2) and 1-propyl-2-methyl-3-naphthoyl-indole (JWH 015) inhibited TNF-alpha induced release of interleukin-8 in a concentration-dependent manner. The less active enantiomer of WIN55212-2, [S(-)-[2,3-dihydro-5-methyl-3-[(morpholinyl)methyl]pyrrolo[1,2,3-de]1,4-benzoxazin-6-yl](1-naphthyl) methanone mesylate (WIN55212-3), and the cannabinoid CB(1) receptor agonist arachidonoyl-2-chloroethylamide (ACEA) had no significant effect on TNF-alpha-induced release of interleukin-8. The cannabinoid CB(1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1,4-pyrazole-3-carboxamide hydrochloride (SR141716A; 10(-6) M) antagonised the inhibitory effect of CP55,940 (pA(2)=8.3+/-0.2, n=6) but did not antagonise the inhibitory effects of WIN55212-2 and JWH 015. The cannabinoid CB(2) receptor antagonist N-(1,S)-endo1,3,3-trimethylbicyclo(2,2,1)heptan-2-yl)-5(4-chloro-3-methyl-phenyl)-1-(4-methylbenzyl)-pyrazole-3-carboxamide (SR144528; 10(-6) M) antagonised the inhibitory effects of CP55,940 (pA(2)=8.2+/-0.8, n=6), WIN55212-2 (pA(2)=7.1+/-0.3, n=6) and JWH 015 (pA(2)=7.6+/-0.3, n=6), respectively. Western immunoblotting of HT-29 cell lysates revealed a protein with a size that is consistent with the presence of cannabinoid CB(2) receptors. We conclude that in HT-29 cells, TNF-alpha-induced interleukin-8 release is inhibited by cannabinoids through activation of cannabinoid CB(2) receptors.

Arachidonic Acids↗

PubMed2002

Modulation of the release of endogenous gamma-aminobutyric acid by cannabinoids in the guinea pig ileum.

Interactions between cannabinoid CB(1) and GABA receptors and ligands were investigated in the myenteric plexus-longitudinal muscle of the guinea pig ileum. Electrically evoked contractions of the myenteric plexus-longitudinal muscle were inhibited by the cannabinoid receptor agonist CP55,940 ((-)-cis-3-[2-Hydroxy-4-(1,1-dimethylheptyl) phenyl]-trans-4-(3-hydroxypropyl) cyclohexanol), the GABA(B) receptor agonist, baclofen (4-amino-3-(chlorophenyl) butanoic acid), or exogenous GABA. Electrically evoked contractions of the myenteric plexus-longitudinal muscle were also inhibited by the addition of the GABA releasing agent ethylenediamine. CP55,940 (1 nM) or the endogenous cannabinoid anandamide (arachidonyl ethanolamide, 1 microM) reduced the inhibition produced by ethylenediamine, while in contrast, anandamide (10 microM) significantly increased the inhibition produced by ethylenediamine. The results suggest that while there is no interaction between cannabinoid CB(1) and GABA(B) receptors in the myenteric plexus-longitudinal muscle of the guinea pig, cannabinoid CB(1) receptor stimulation reduces the ethylenediamine-evoked GABA release. In addition, anandamide at higher concentrations also potentiates the inhibitory effect of ethylenediamine at least partly by stimulating vanilloid receptors.

Animals↗

PubMed2002

High-conductance chloride channels generate pacemaker currents in interstitial cells of Cajal.

BACKGROUND & AIMS: Interstitial cells of Cajal (ICCs) are responsible for slow, wave-driven, rhythmic, peristaltic motor patterns in the gastrointestinal tract. The aim was to identify and characterize the ion channels that generate the underlying pacemaker activity. METHODS: Single ion channel recordings were obtained from nonenzymatically isolated ICCs and studied by using the cell attached and inside-out configurations of the patch clamp technique. RESULTS: A high-conductance chloride channel was observed in ICCs that was spontaneously and rhythmically active at the same frequency as the rhythmic inward currents defining ICC pacemaker activity, 20-30 cycles/min at room temperature. Main conductance levels occurred between 122-144 pS and between 185-216 pS. Periodicity in the channel opening coincided with periodicity in membrane potential change, hence, at the single channel level, chloride channels were seen to be associated with the generation of rhythmic changes in membrane potential. CONCLUSIONS: ICCs harbor high-conductance chloride channels that participate in the generation of pacemaker activity and may become a target for pharmacologic treatment of gut motor disorders.

Animals↗