Multiple Forms of Endocannabinoid and Endovanilloid Signaling Regulate the Tonic Control of GABA Release

Abstract

PersistentCB 1 cannabinoidreceptoractivitylimitsneurotransmitterreleaseatvarioussynapsesthroughoutthebrain.However,itisnotfullyunderstood how constitutively active CB 1 receptors, tonic endocannabinoid signaling, and its regulation by multiple serine hydrolases contribute to the synapsespecific calibration of neurotransmitter release probability. To address this question at perisomatic and dendritic GABAergic synapses in the mouse hippocampus,weusedacombinationofpairedwhole-cellpatch-clamprecording,liquidchromatography/tandemmassspectrometry,stochasticoptical reconstruction microscopy super-resolution imaging, and immunogold electron microscopy. Significance StatementTonic cannabinoid signaling plays a critical role in the regulation of synaptic transmission. However, the mechanistic details of how persistent CB 1 cannabinoid receptor activity inhibits neurotransmitter release have remained elusive. Therefore, electrophysiological recordings, lipid measurements, and super-resolution imaging were combined to elucidate those signaling molecules and mechanisms that underlie tonic cannabinoid signaling. The findings indicate that constitutive CB 1 activity has pivotal function in the tonic control of hippocampal GABA release. Moreover, the endocannabinoid 2-arachidonoylglycerol (2-AG) is continuously generated postsynaptically, but its synaptic effect is regulated strictly by presynaptic monoacylglycerol lipase activity. Finally, anandamide signaling antagonizes tonic 2-AG signaling via activation of postsynaptic transient receptor potential vanilloid TRPV1 receptors. This unexpected mechanistic diversity may be necessary to fine-tune GABA release probability under various physiological and pathophysiological conditions.

Cannabidiol as a Potential Treatment for Anxiety Disorders

“…The BeCB systemî ncludes AEA and 2-arachidonoylglycerol; their respective degradative enzymes fatty acid amide hydroxylase (FAAH) and monoacylglycerol lipase; the CB 1 R and related CB 2 receptor (the latter expressed mainly in the periphery); as well as several other receptors activated by eCBs, including the TRPV1 receptor, peroxisome proliferator-activated receptor-γ, and G protein-coupled 55 receptor, which functionally interact with CB 1 R signaling (reviewed in [21,24]). Interactions with the TRPV1 receptor, in particular, appear to be critical in regulating the extent to which eCB release leads to inhibition or facilitation of presynaptic neurotransmitter release [25]. The TRPV1 receptor is a postsynaptic cation channel that underlies sensation of noxious heat in the periphery, with capsacin (hot chili) as an exogenous ligand.…”

Correlated confocal and super-resolution imaging by VividSTORM

“…recording 5,20 . To obtain high-resolution STORM images of our proteins of interest, we have built on a procedure, which was previously used for STORM microscopy in fixed brain sections 21 , and we have modified this procedure for free-floating immunostaining to maximize the sensitivity and specificity of antibody binding.…”

“…To address this issue, we have implemented the analysis algorithms used earlier 5,20 in VividSTORM, a stand-alone software with graphical user interface (GUI), which can readily open, visualize and analyze corresponding conventional optical microscopy images and molecule lists. By following the analysis workflow presented earlier 5 , the localization points (LPs) in the STORM molecule list belonging to the labeled subcellular structures on the overlaid confocal image are selected.…”

“…By following the analysis workflow presented earlier 5 , the localization points (LPs) in the STORM molecule list belonging to the labeled subcellular structures on the overlaid confocal image are selected. On these data sets, a number of analysis functions can be run, in order to measure the number and density of LPs, intermolecular distance distribution, density of LPs along 3D surfaces, clustering and internalization 20 .…”

“…1) for correlated confocal and STORM superresolution microscopy, which enables quantitative molecular imaging at subcellular domains of specific cells at the nanoscale level. The protocol follows the experimental procedure that we have described earlier 5,20 . Importantly, this protocol allows the study of any target protein in a cell typespecific manner in association with physiological and pathophysiological processes.…”