INHIBITION BY CANNABINOID RECEPTOR AGONISTS OF ACETYLCHOLINE-RELEASE FROM THE GUINEA-PIG MYENTERIC PLEXUS

1 The dose-related inhibition of the twitch responses of the myenteric plexus-longitudinal muscle preparation of the guinea-pig small intest ine by cannabinoid (CB) agonists, (+)-WIN 55212 and CP 55940 during st imulation at 0.1 Hz with supramaximal voltage was confirmed. These ago nists inhibited acetylcholine (ACh) release in the presence of physost igmine (7.7 mu M) thus indicating a prejunctional site of action. 2 In hibition of twitch responses and ACh release by CB agonists was revers ed by the CB1-selective cannabinoid receptor antagonist, SR141716A. Do se-response curves to (+)-WIN 55212 and CP 55940 were shifted to the r ight, with no reduction of maximal response, by pretreatment with SR14 1716A (31.6-1000 nM), but not its vehicle, Tween 80 (1 mu M) However, at very high concentrations (25-400 mu M), Tween 80 itself caused a do se-related inhibition of the twitch response which was significantly r educed in the presence of SR141716A (1 mu M). The opioid receptor anta gonist, naloxone (1 mu M) had no significant effect on the inhibition by CP 55940 of the twitch response. 3 (+)-WIN 55212, CP 55940 and Twee n 80 (50 mu M) had no effect on responses to exogenous ACh, confirming that their actions were prejunctional. SR141716A (1 mu M) did not inc rease the sensitivity of the longitudinal muscle to either ACh or hist amine, but inhibited the responses to high doses of ACh. 4 The (-)-ena ntiomer of WIN 55212, was approximately 300 times less active thin the (+) enantiomer in inhibiting the twitch response, had no CB1 antagoni st activity against the active isomer and did not inhibit the release of ACh in the presence of physostigmine. 5 The dissociation constant ( K-D) values for SR 141716A against the inhibitory effect of (+)-WIN 55 212 and CP 55940 on the twitch response were 12.07 nM (95% confidence intervals 8.55 and 20.83) and 6.44 nM (95% confidence intervals 4.70 a nd 10.24), respectively. In experiments in which the release of ACh wa s inhibited by (+)-WIN 55212, the K-D values were 9.21 nM and 10.53 nM at SR141716A concentrations of 31.6 nM and 100 nM, respectively. The K-D values for the antagonism by naloxone of the inhibition of the twi tch responses and the inhibition of ACh release by normorphine in this preparation were found to be 2.38+/-0.69 nM and 2.00+/-0.9 nM, respec tively. 6 During maximal inhibition of ACh release by (+)-WIN 55212, t he addition of normorphine (400 nM) caused a further significant decre ase in ACh output. 7 SR141716A alone produced a significant increase i n ACh release in both the absence and presence of exogenous cannabinoi d drugs, hence we conclude that it has a presynaptic site of action. W e also conclude that SR141716A acts either by antagonizing the effect of an endogenous CB1 receptor agonist or by having an inverse agonist effect at these receptors.