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Of Mice and MAGL (Monoacylglycerol Lipase)
Bach, S; Bombinski, T; Daniels, M; Gross, D; Hogg, T; Martin, T; McMurray, D; Naber, E; Perez, N; Schulman, A; Tucker, S; Andera-Cato, S; Arnold, A;
Blumberg, A; Bord, M; Feiertag, A; Greaves, M; Her, A; Kennedy, E; Orozco, C; Rice, C; Rodgers, A; Sauer, A; Schubert, J; Tubbs, C; Wray, T
Teacher Advisor: G. Vogt - Brown Deer High School 8060 N. 60th, Brown Deer, WI 53223
Mentors: L. Shrestha, C. Hillard, Ph.D. Director of the Neuroscience Research Center
Medical College of Wisconsin
Abstract
Figure 4: Inhibition of MAGL Hinders Hydrolysis of 2-AG
Figure 2: Endocannabinoid Signaling System
About 18.8 million American adults suffer depressive disorders that may
occur with anxiety and substance abuse. Tetrahydrocannabinol (THC), a
compound in marijuana, is a cannabinoid chemical that binds to and
activates cannabinoid receptors (CB1) in the pre-synaptic cell membrane
as part of neuron-to-neuron transmission in the endocannabinoid system
(ECS). Glutamate in the pre-synaptic cell is released and binds to the
post-synaptic cell triggering the synthesis and release of 2arachidonoylglycerol (2-AG). 2-AG returns to the pre-synaptic cell binding
to and activating CB1 receptors. THC mimics 2-AG action, and is used to
study the ECS retrograde signaling system and its effect on appetite and
mood. A protein from the pre-synaptic cell, monoacylglycerol lipase
(MAGL), hydrolyzes 2-AG into arachidonic acid (AA) and glycerol
controlling 2-AG levels. When MAGL is hyperactive, too much 2-AG
degrades, causing 2-AG shortage, prompting depression, anxiety, and
other neurodegenerative diseases. Hypoactive MAGL activity creates an
excess of 2-AG, causing obesity, diabetes, and addictive behaviors. The
Brown Deer Students Modeling a Research Topic Team, in alliance with
MSOE, built a MAGL model using a 3D printer. Study of MAGL hydrolysis
sites may provide the key to regulating MAGL’s enzymatic activity leading
to therapies that will prevent neurodegenerative disorders.
Step 5:
MAGL hydrolyzes 2-AG
into arachidonic acid
and glycerol.
(See Figure 1)
PRE-SYNAPTIC CELL
Step 1:
Glutamate is
released from the
pre-synaptic cell,
and binds to the
post-synaptic
cell.
MAGL
Blockade of 2-Arachidonoylglycerol Hydrolysis by Selective Monoacylglycerol Lipase Inhibitor 4-Nitrophenyl 4-(Dibenzo[d[ [,13]dioxol-5yl(hydroxy)methyl)piperidine-1carboxylate (JXL184) Enhances Retrograde Endocannabinoid Signaling Bin Pan, Wei Wang, Jonathan Z. Long,
Dalong Sun, Cecilia J Hillard, Benjamin F. Cravatt, Qing-sang Liu
Electrodes placed in neurons from a slice of rat brain were used to detect
the excitatory post-synaptic currents (EPSC) that run through the postsynaptic neurons when glutamate reacts with its receptors. These data
were plotted on the y-axis of Figure 4. Small pulses were used to release
glutamate from the pre-synaptic cell but not enough to trigger the
synthesis of 2-AG. The baseline result between -50 and 0 seconds is at
100% of the EPSC amplitude because there is no 2-AG present. At time
0, the electrical pulses were vastly increased for a few milliseconds,
triggering 2-AG synthesis. The presence of 2-AG decreased the release
of glutamate reflected in a decrease in EPSC amplitude. The control line
returned to 100% after a few seconds due to MAGL breaking down 2-AG.
The other two curves demonstrate the inhibitory effect of JZL184 on
MAGL activity; blocking MAGL delays the breakdown of 2-AG and delays
the return time to control EPSC amplitude.
Step 4:
SYNAPTIC CLEFT 2-AG binds to CB1
receptor on the presynaptic cell inhibiting
glutamate release.
Introduction
Approximately 80% of the 18.8 million American adults suffering from the
serious neurological disorder known as depression are not receiving any
treatment. Elucidation of neural signaling mechanisms in the brain may
lead to more effective treatments. Specific brain cells called neurons
transmit messages from pre-synaptic neurons to post-synaptic neurons
across a gap called the synapse using neurotransmitter chemicals. One
potential target of drug treatment is the regulation of the enzyme
monoacylglycerol lipase (MAGL), found in the endocannabinoid system,
shown in Figure 2. MAGL is an enzyme found in the pre-synaptic cell that
hydrolyzes 2-AG:
•Increase in MAGL activity decreases 2-AG concentrations resulting in
depression and anxiety.
•Decrease in MAGL activity increases 2-AG concentrations producing
anti-depressant effects.
Inhibition of MAGL will increase 2-AG, therefore making MAGL a good
target for drug therapy of depression.
Step 2:
The binding of
glutamate to the
post-synaptic cellPOST-SYNAPTIC
CELL
Step
3:
triggers the
2-AG travels back to
synthesis and
the pre-synaptic cell
release of 2-AG.
and binds to CB1
Figure 5: Inhibition of MAGL Calms Mouse Behavior
16
14
Marbles Buried
receptors.
POST-SYNAPTIC CELL
(A Review on the Monoacylglycerol Lipase: At the Interface Between Fat and Endocannabinoid Signaling G. Labar, J. Wouters,
D.M. Lambert “Current Medicinal Chemistry, 2010, 17, 2588-2607”)
http://depressiontreatmenthelp.org/depression-statistics.php
12
10
8
6
4
2
0
Vehicle
Figure 1: MAGL Hydrolyzes 2-AG to Glycerol and
Arachidonic Acid
16
40
JZL-184 (mg/kg)
MAGL
Asp-239
Catalytic Triad
His-269
3hju.pdb
MAGL
2
Figure 3: Monoacylglycerol Lipase,
in model form
Ser-122
Ser-122
Research has been conducted using mice to study their marble burying
behavior. Mice normally demonstrate anxious behavior when foreign
objects are placed in their cage. When marbles were introduced the mice
responded by frantically trying to bury them. The y-axis demonstrates the
number of marbles buried with a vehicle and increasing concentrations of
JZL184. The vehicle includes the experimental protocol used to inject the
mice. The data shown in Figure 5 illustrates that injection of JZL184
reduces the anxious behavior of the mice causing them to bury fewer
marbles.
Kinsey SG, et al, Inhibition of endocannabinoid catabolic enzymes elicits anxiolytic-like effects in the marble burying assay,
Pharmacol Biochem Behav (2010), doi:10.1016/j.pbb.2010.12.002
Biological Significance
JZL184 has an inhibitory effect on MAGL activity. Regulation of MAGL
activity could effectively treat anxiety and depression disorders in human.
SMART Teams are supported by the National Institutes of Health (NIH)- National Center for Research Resources-Science Education Partnership Award (NCRR-SEPA), and an NIH CTSA Award to the Medical College of Wisconsin.