The endocannabinoid (eCB) system comprised of lipophylic signaling molecules, postsynaptic production enzymes, and presynaptic G-protein coupled receptors (GPCR) has recently emerged as a key mediator in a broad range of neuronal plasticity. Initial interest in the system stemmed from, delta(9)-tetrahydrocannabinol (THC) the main psychoactive ingredient in the illicit drug marijuana which elicits various effects on the central nervous system (CNS) through interactions with the eCB system. Further study described a nearly ubiquitous yet tunable system that may underlie many CNS functions. Studying the eCB system offers insight into the basic science of synaptic transmission but more importantly the mechanism by which postsynaptic determinants may influence their synaptic inputs to adapt to changing CNS conditions. The objective of these studies was to determine the properties of THC that dictate its relationship to the intact eCB system and to characterize how specific members of the homer family of postsynaptic scaffolding proteins impact differing forms of eCB-mediated plasticity.
Activation of the cannabinoid receptor-1 (CB1R) by THC or eCBs leads to several presynaptic consequences, but most importantly to inhibition of voltage-gated calcium channels (VGCC) and thus a reduction of action potential induced neurotransmitter release. In the first study presented here, we described a voltage-mediated switch by which THC acts as an agonist of CB1R or as an antagonist to effects at CB1R mediated by eCBs. Using patch clamp electrophysiology to study excitatory synaptic transmission we discovered that the rate at which the presynaptic neuron is depolarized has drastic effects on THC mediated inhibition. Excitatory postsynaptic currents (EPSCs) evoked at 0.1 Hz were suppressed by THC, however THC did not effect EPSCs evoke at 0.5 Hz. Experiments using other CB1R agonists Win55212,2 (Win-2) and 2-Arachydonylglycerol (2-AG) did not demonstrate sensitivity to stimulus rate. THC application at 0.5 Hz stimulation antagonized the inhibition produced by Win-2. THC but not Win-2 inhibited influx of calcium through VGCCs in a rate dependent manner. Win-2 but not THC shifted the voltage-dependent activation of tail currents. THC but not Win-2 mediated inhibition of VGCCs displayed mild prepulse facilitation. THC antagonized depolarization-induced suppression of excitation (DSE). These findings illustrate the complex interplay between THC and the eCB system and suggest that behavioral responses to THC may stem from a combination of both CB1R activation and inhibition of eCB action at CB1R.
Postsynaptic induction of the eCB system and subsequent production of eCBs may be achieved via several interrelated molecular mechanisms that focus around glutamatergic ionotropic and metabotropic signaling and postsynaptic intercellular Ca2+ concentration ([Ca2+]i). Homer, a family of postsynaptic scaffolding proteins, is present in the hippocampus and has been found to organize many of the structural and mechanistic proteins necessary for several forms of synaptic plasticity. In the second study we describe a homer isoform-specific switch in eCB plasticity that primes neurons for ionotropic mediated production of eCBs but shifts them away from metabotropic eCB production. Transfection of hippocampal cultures with homer 1a (H1a) enhances DSE. Transfection of cultures with H1a inhibits metabotropic suppression of excitation (MSE). In combination experiments on the same neuron with DSE followed by MSE, DSE is enhanced and MSE is inhibited in H1a expressing neurons. Brain derived neurotrophic factor (BDNF) induces formation of H1a mRNA and leads to functional expression of H1a which dissociates homer 1c-GFP puncta. BDNF mediates an enhancement of DSE and inhibition of MSE similar to that found in H1a transfected neurons. These findings identify H1a as a crucial mediator of eCB signaling that may be induced by BDNF and can lead to an enhancement of depolarization mediated eCB production. The second study delineated how postsynaptic homer proteins can influence differential paths to activating eCB mediated synaptic plasticity.
The physiological manifestations of eCB signaling have tentacles that extend from neurotransmitter release to higher brain function through the regulation of Ca2+ channels and induction of synaptic plasticity. These studies are directed toward understanding the way in which THC interacts with the eCB system, and the postsynaptic scaffolding protein complement which induces changes to eCB-mediated plasticity. Delineating these concepts may lead to better understanding of higher brain function and will offer new targets for therapeutic development.
University of Minnesota Ph.D. dissertation. October 2009. Major: Pharmacology. Advisor: Stanley A Thyer. 1 computer file (PDF); appendix A. Ill. (some col.)
Roloff, Alan Mathhew.
Modulation of hippocampal endocannabinoid plasticity by homer proteins and Delta(9)-tetrahydrocannabinol..
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