Identification of the endocannabinoid working as a retrograde messenger and its release mechanism

最近的研究表明，内源性大麻素作为一种逆行信使，在大脑突触传递的活动依赖性调节中发挥重要作用。内源性大麻素的释放是由突触后去极化或I组代谢型谷氨酸受体（I-mGluRs）的激活诱导的。释放的内源性大麻素然后激活突触前CB 1大麻素受体并抑制递质释放。然而，内源性大麻素产生的机制尚未完全确定。在这项研究中，我们使用培养的大鼠海马神经元，并记录大麻素敏感的抑制性突触后电流（IPSC），它可以被用作释放的内源性大麻素的生物传感器。结果表明：1.在三种内源性大麻素（anandamide，2-AG，noladin ether）中，2-AG对IPSC的抑制作用最强且可逆; 2.内源性大麻素的释放是由M1/M3受体的突触后激活引起的 ...更多信息 以及I-mGluRs。PLCβ1基因敲除小鼠神经元的去极化诱导的内源性大麻素释放依赖于Ca^<2+>的升高，PLC抑制剂ET-18可抑制这种去极化诱导的内源性大麻素释放，但PLCβ1或PLCδ基因敲除小鼠不能消除这种释放。两者都不能诱导内源性大麻素释放，同时处理，内源性大麻素释放。这种类型的释放也从PLCβ1基因敲除小鼠中消除。利用TRPC 6通道作为PLC活性的生物传感器，我们证明了去极化和受体激活对内源性大麻素释放的协同作用可以通过受体驱动PLCβ激活的Ca^2+依赖性来解释，这些结果表明，突触后激活（去极化或受体激活）刺激PLC并通过随后的DAG脂肪酶的酶活性诱导2-AG的产生。在不同的刺激条件下，所涉及的PLC的同工酶可能不同。少

Recent studies have revealed that endocannabinoids work as a retrograde messenger and play an important role in activity-dependent modulations of synaptic transmission in the brain. The endocannabinoid release is induced by postsynaptic depolarization or activation of group I metabotropic glutamate receptors (I-mGluRs). The released endocannabinoid then activates presynaptic CB1 cannabinoid receptors and suppresses the transmitter release. The mechanisms of endocannabinoid production, however, have not been fully determined. In this study, we used rat cultured hippocampal neurons and recorded cannabinoid-sensitive inhibitory postsynaptic currents (IPSCs), which can be used as biosensor of released endocannabinoids. We obtained the following results.1.Among three candidates of endocannabinoids (anandamide,2-AG, noladin ether),2-AG was the most effective and reversible in suppressing IPSCs.2.The endocannabinoid release was induced by postsynaptic activation, of M1/M3 muscarinic receptors … More as well as I-mGluRs. This receptor-driven endocannabinoid release was eliminated from the neurons prepared from PLCβ1-knockout mice.3.The depolarization-induced endocannabinoid release, which is dependent on Ca^<2+> elevation, was suppressed by the PLC inhibitor ET-18,but not eliminated from PLCβ1- or PLCδ-knockout mice.4.When weak depolarization and weak activation of I-mGluRs or M1/M3 muscarinic receptors, both of which can not induce endocannabinoid release, were simultaneously treated, endocannabinoids were released. This type of release was also eliminated from PLCβ1-knockout mice. Using TRPC6 channels as biosensor of PLC activity, we demonstrated that the synergistic effect of depolarization and receptor activation on endocannabinoid release can be explained by the Ca^<2+>-dependence of receptor-driven PLCβ activation.These results suggest that the postsynatpic activation (depolarization or receptor activation) stimulates PLCs and induces 2-AG production through the subsequent enzymatic activity by DAG lipase. The isozymes of PLCs involved might be different in different stimulation conditions. Less