title: The initiation of synaptic 2-AG mobilization requires both an increased supply of diacylglycerol precursor and increased postsynaptic calcium.
abstract: On-demand postsynaptic synthesis and release of endocannabinoid lipids and subsequent binding to presynaptic CB1 receptors (CB1Rs) mediates short and long-term depression (LTD) of excitatory transmission in many brain regions. However, mechanisms involved in the synthesis of the endocannabinoid 2-arachidonoylglycerol (2-AG) by diacylglycerol lipase α (DGLα) are poorly understood. Since Gq-coupled receptor activation can stimulate production of a major DGL substrate 1-stearoyl-2-arachidonoyl-sn-glycerol (SAG) by PLCβ, we sought to determine if 2-AG biosynthesis was limited only by a lack of substrate availability, or if other pathways, such as Ca(2+) signaling, also need to be simultaneously engaged. To address this question, we loaded medium spiny neurons of the dorsolateral striatum with SAG while monitoring excitatory synaptic inputs. SAG-loading had no significant effect on evoked excitatory synaptic currents when cells were voltage-clamped at -80 mV. However, depolarization of MSNs to -50 mV revealed a SAG-loading dependent decrease in the amplitude of excitatory currents that was accompanied by an increase in paired pulse ratio, consistent with decreased glutamate release. Both effects of loading SAG at -50 mV were blocked by chelation of postsynaptic Ca(2+) using BAPTA or by bath application of tetrahydrolipstatin (THL), a DGL inhibitor. Loading of SAG into glutamatergic pyramidal neurons of the amygdala similarly inhibited excitatory synaptic inputs and increased the PPR. SAG-induced depression was absent in both regions from mice lacking CB1Rs. These data show that increasing substrate availability alone is insufficient to drive 2-AG mobilization and that DGL-dependent synaptic depression via CB1R activation requires postsynaptic Ca(2+) signals.
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA.
authors.forename: Brian C
authors.initials: BC
authors.lastname: Shonesy
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA; Department of Psychiatry, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA.
authors.forename: Danny G
authors.initials: DG
authors.lastname: Winder
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA; Department of Psychiatry, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA.
authors.forename: Sachin
authors.initials: S
authors.lastname: Patel
authors.affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt Brain Institute, Vanderbilt-Kennedy Center for Research on Human Development, Vanderbilt University Medical Center, Nashville, TN, USA. Electronic address: roger.colbran@vanderbilt.edu.
authors.forename: Roger J
authors.initials: RJ
authors.lastname: Colbran
mesh_terms: D000818:Animals; D001095:Arachidonic Acids / Q000096:biosynthesis*; D020013:Calcium Signaling; D003342:Corpus Striatum / Q000187:drug effects / Q000502:physiology*; D004075:Diglycerides / Q000378:metabolism* / Q000494:pharmacology; D063388:Endocannabinoids / Q000096:biosynthesis*; D005989:Glycerides / Q000096:biosynthesis*; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D013569:Synapses / Q000378:metabolism*
chemical_list: D001095:Arachidonic Acids; D004075:Diglycerides; D063388:Endocannabinoids; D005989:Glycerides; C075809:1-stearoyl-2-arachidonoylglycerol; C094503:glyceryl 2-arachidonate
keywords: 2-arachidonoylglycerol; Diacylglycerol lipase; Endocannabinoids