Effects of endogenous cannabinoid anandamide on excitation–contraction coupling in rat ventricular myocytes

A role for anandamide (N-arachidonoyl ethanolamide; AEA), a major endocannabinoid, in the cardiovascular system in various pathological conditions has been reported in earlier reports. In the present study, the effects of AEA on contractility, Ca²⁺ signaling, and action potential (AP) characteristics were investigated in rat ventricular myocytes. Video edge detection was used to measure myocyte shortening. Intracellular Ca²⁺ was measured in cells loaded with the fluorescent indicator fura-2 AM. AEA (1 μM) caused a significant decrease in the amplitudes of electrically evoked myocyte shortening and Ca²⁺ transients. However, the amplitudes of caffeine-evoked Ca²⁺ transients and the rate of recovery of electrically evoked Ca²⁺ transients following caffeine application were not altered. Biochemical studies in sarcoplasmic reticulum (SR) vesicles from rat ventricles indicated that AEA affected Ca²⁺-uptake and Ca²⁺-ATPase activity in a biphasic manner. [³H]-ryanodine binding and passive Ca²⁺ release from SR vesicles were not altered by 10 μM AEA. Whole-cell patch-clamp technique was employed to investigate the effect of AEA on the characteristics of APs. AEA (1 μM) significantly decreased the duration of AP. The effect of AEA on myocyte shortening and AP characteristics was not altered in the presence of pertussis toxin (PTX, 2 μg/ml for 4 h), AM251 and SR141716 (cannabinoid type 1 receptor antagonists; 0.3 μM) or AM630 and SR 144528 (cannabinoid type 2 receptor antagonists; 0.3 μM). The results suggest that AEA depresses ventricular myocyte contractility by decreasing the action potential duration (APD) in a manner independent of CB1 and CB2 receptors.     

The alteration of rabbit skeletal sarcoplasmic reticulum function by N-acylethanolamine, a lipid associated with myocardial infarction

The ability of N-acylethanolamines (pharmacologically active lipid metabolites which accumulate in canine myocardium during experimentally induced infarctions) to alter Ca2+ fluxes in a biological membrane system was studied using sarcoplasmic reticulum vesicles prepared from rabbit skeletal muscle. The effects of two N-acylethanolamines, the N-oleyl and N-lauryl derivatives, were compared to those of the lipophilic drugs, dibucaine and propranolol. The rate and extent of Ca2+ sequestration, Ca2+-Mg2+-ATPase activity and retention time of Ca2+ by the vesicles were all stimulated at low concentrations of the four compounds studied and inhibited at higher concentrations. The stoichiometry between Ca2+-pumping rates and ATPase activity was partially "uncoupled" indicating that both the calcium pump and the membrane permeability were affected by the drugs. However, although all four compounds exhibited the same qualitative behavior, the effects of the two N-acylethanolamines were more pronounced than dibucaine and propranolol and occurred at much lower concentrations. These results suggest that the N-acylethanolamines may have important physiological effects in the myocardium and, at least at lower concentrations, stimulate myocardial contractility by increasing the rate of calcium flux across the sarcoplasmic reticulum.