Thien P Dinh

University of California, Irvine, Irvine, CA, United States

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Publications (7)46.99 Total impact

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    ABSTRACT: The endogenous cannabinoid 2-arachidonoylglycerol (2-AG) is produced by neurons and other cells in a stimulus-dependent manner and undergoes rapid biological inactivation through transport into cells and catalytic hydrolysis. The enzymatic pathways responsible for 2-AG degradation are only partially understood. We have shown previously that overexpression of monoacylglycerol lipase (MGL), a cytosolic serine hydrolase that cleaves 1- and 2-monoacylglycerols to fatty acid and glycerol, reduces stimulus-dependent 2-AG accumulation in primary cultures of rat brain neurons. We report here that RNA interference-mediated silencing of MGL expression greatly enhances 2-AG accumulation in HeLa cells. After stimulation with the calcium ionophore ionomycin, 2-AG levels in MGL-silenced cells were comparable with those found in cells in which 2-AG degradation had been blocked using methyl arachidonyl fluorophosphonate, a nonselective inhibitor of 2-AG hydrolysis. The results indicate that MGL plays an important role in the degradation of endogenous 2-AG in intact HeLa cells. Furthermore, immunodepletion experiments show that MGL accounts for at least 50% of the total 2-AG-hydrolyzing activity in soluble fractions of rat brain, suggesting that this enzyme also contributes to 2-AG deactivation in the central nervous system.
    Molecular Pharmacology 12/2004; 66(5):1260-4. · 4.41 Impact Factor
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    Lisa Walter, Thien Dinh, Nephi Stella
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    ABSTRACT: The cytoplasm of neural cells contain millimolar amounts of ATP, which flood the extracellular space after injury, activating purinergic receptors expressed by glial cells and increasing gliotransmitter production. These gliotransmitters, which are thought to orchestrate neuroinflammation, remain widely uncharacterized. Recently, we showed that microglial cells produce 2-arachidonoylglycerol (2-AG), an endocannabinoid known to prevent the propagation of harmful neuroinflammation, and that ATP increases this production by threefold at 2.5 min (Witting et al., 2004). Here we show that ATP increases 2-AG production from mouse astrocytes in culture, a response that is more rapid (i.e., significant within 10 sec) and pronounced (i.e., 60-fold increase at 2.5 min) than any stimulus-induced increase in endocannabinoid production reported thus far. Increased 2-AG production from astrocytes requires millimolar amounts of ATP, activation of purinergic P2X7 receptors, sustained rise in intracellular calcium, and diacylglycerol lipase activity. Furthermore, we show that astrocytes express monoacylglycerol lipase (MGL), the main hydrolyzing enzyme of 2-AG, the pharmacological inhibition of which potentiates the ATP-induced 2-AG production (up to 113-fold of basal 2-AG production at 2.5 min). Our results show that ATP greatly increases, and MGL limits, 2-AG production from astrocytes. We propose that 2-AG may function as a gliotransmitter, with MGL inhibitors potentiating this production and possibly restraining the propagation of harmful neuroinflammation.
    Journal of Neuroscience 10/2004; 24(37):8068-74. · 6.91 Impact Factor
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    ABSTRACT: Fatty acid amide hydrolase (FAAH) and monoglyceride lipase (MGL) catalyse the hydrolysis of the endocannabinoids anandamide and 2-arachidonoyl glycerol. We investigated their ultrastructural distribution in brain areas where the localization and effects of cannabinoid receptor activation are known. In the hippocampus, FAAH was present in somata and dendrites of principal cells, but not in interneurons. It was located mostly on the membrane surface of intracellular organelles known to store Ca(2+) (e.g. mitochondria, smooth endoplasmic reticulum), less frequently on the somatic or dendritic plasma membrane. MGL immunoreactivity was found in axon terminals of granule cells, CA3 pyramidal cells and some interneurons. In the cerebellum, Purkinje cells and their dendrites are intensively immunoreactive for FAAH, together with a sparse axon plexus at the border of the Purkinje cell/granule cell layers. Immunostaining for MGL was complementary, the axons in the molecular layer were intensively labelled leaving the Purkinje cell dendrites blank. FAAH distribution in the amygdala was similar to that of the CB(1) cannabinoid receptor: evident signal in neuronal somata and proximal dendrites in the basolateral nucleus, and hardly any labelling in the central nucleus. MGL staining was restricted to axons in the neuropil, with similar relative signal intensities seen for FAAH in different nuclei. Thus, FAAH is primarily a postsynaptic enzyme, whereas MGL is presynaptic. FAAH is associated with membranes of cytoplasmic organelles. The differential compartmentalization of the two enzymes suggests that anandamide and 2-AG signalling may subserve functional roles that are spatially segregated at least at the stage of metabolism.
    European Journal of Neuroscience 08/2004; 20(2):441-58. · 3.75 Impact Factor
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    ABSTRACT: The control of endocannabinoid actions on cortical neurons by a putative carrier-mediated uptake is still poorly understood at the level of synaptic transmission. We investigated the effect of an endocannabinoid, 2-arachidonoyl glycerol (2-AG), on inhibitory postsynaptic currents (IPSCs) in hippocampal slices under physiological conditions, and when uptake was altered by a selective blocker or lower temperature. Bath application of 2-AG (20 micro m) caused a 40% reduction in the amplitude of IPSCs evoked in the perisomatic region of CA1 pyramidal neurons at room temperature; this effect could be blocked by a specific CB(1) receptor antagonist, AM251. By contrast, a smaller (20%) but significant suppression of inhibitory transmission was found by 2-AG at 33-35 degrees C. This reduced blocking effect at physiological temperature could be brought back to 40% by coapplying the endocannabinoid uptake blocker, AM404 (10 or 20 micro m) with 2-AG. In parallel experiments, we measured [(3)H]2-AG uptake at different temperatures in primary cultures prepared from cortical neurons. These data confirmed a striking inhibition of [(3)H]2-AG uptake at room temperature compared with values observed at 37 degrees C. Uptake could be significantly modified by anandamide, 2-AG and AM404, suggesting a common transporter for the two endocannabinoids. These findings together demonstrate the presence of an effective endocannabinoid uptake in cortical neurons, which could considerably alter the spatial and temporal constraints of endocannabinoid signalling at physiological temperature, and which may critically change the interpretation of findings at room temperature.
    European Journal of Neuroscience 07/2004; 19(11):2991-6. · 3.75 Impact Factor
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    ABSTRACT: Febrile (fever-induced) seizures are the most common form of childhood seizures, affecting 3%-5% of infants and young children. Here we show that the activity-dependent, retrograde inhibition of GABA release by endogenous cannabinoids is persistently enhanced in the rat hippocampus following a single episode of experimental prolonged febrile seizures during early postnatal development. The potentiation of endocannabinoid signaling results from an increase in the number of presynaptic cannabinoid type 1 receptors associated with cholecystokinin-containing perisomatic inhibitory inputs, without an effect on the endocannabinoid-mediated inhibition of glutamate release. These results demonstrate a selective, long-term increase in the gain of endocannabinoid-mediated retrograde signaling at GABAergic synapses in a model of a human neurological disease.
    Neuron 09/2003; 39(4):599-611. · 15.77 Impact Factor
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    ABSTRACT: 2-Arachidonoylglycerol (2-AG) is a naturally occurring monoglyceride that activates cannabinoid receptors and meets several key requisites of an endogenous cannabinoid substance. It is present in the brain (where its levels are 170-folds higher than those of anandamide), is produced by neurons in an activity- and calcium-dependent manner, and is rapidly eliminated. The mechanism of 2-AG inactivation is not completely understood, but is thought to involve carrier-mediated transport into cells followed by enzymatic hydrolysis. We examined the possible role of the serine hydrolase, monoglyceride lipase (MGL), in brain 2-AG inactivation. We identified by homology screening a cDNA sequence encoding for a 303-amino acid protein, which conferred MGL activity upon transfection to COS-7 cells. Northern blot and in situ hybridization analyses revealed that MGL mRNA is unevenly present in the rat brain, with highest levels in regions where CB1 cannabinoid receptors are also expressed (hippocampus, cortex, anterior thalamus and cerebellum). Immunohistochemical studies in the hippocampus showed that MGL distribution has striking laminar specificity, suggesting a presynaptic localization of the enzyme. Adenovirus-mediated transfer of MGL cDNA into rat cortical neurons increased the degradation of endogenously produced 2-AG in these cells, whereas no such effect was observed on anandamide degradation. These results indicate that hydrolysis via MGL may be a primary route of 2-AG inactivation in intact neuronal cells.
    Chemistry and Physics of Lipids 01/2003; 121(1-2):149-58. · 2.59 Impact Factor
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    ABSTRACT: The endogenous cannabinoids (endocannabinoids) are lipid molecules that may mediate retrograde signaling at central synapses and other forms of short-range neuronal communication. The monoglyceride 2-arachidonoylglycerol (2-AG) meets several criteria of an endocannabinoid substance: (i) it activates cannabinoid receptors; (ii) it is produced by neurons in an activity-dependent manner; and (iii) it is rapidly eliminated. 2-AG inactivation is only partially understood, but it may occur by transport into cells and enzymatic hydrolysis. Here we tested the hypothesis that monoglyceride lipase (MGL), a serine hydrolase that converts monoglycerides to fatty acid and glycerol, participates in 2-AG inactivation. We cloned MGL by homology from a rat brain cDNA library. Its cDNA sequence encoded for a 303-aa protein with a calculated molecular weight of 33,367 daltons. Northern blot and in situ hybridization analyses revealed that MGL mRNA is heterogeneously expressed in the rat brain, with highest levels in regions where CB(1) cannabinoid receptors are also present (hippocampus, cortex, anterior thalamus, and cerebellum). Immunohistochemical studies in the hippocampus showed that MGL distribution has striking laminar specificity, suggesting a presynaptic localization of the enzyme. Adenovirus-mediated transfer of MGL cDNA into rat cortical neurons increased MGL expression and attenuated N-methyl-D-aspartate/carbachol-induced 2-AG accumulation in these cells. No such effect was observed on the accumulation of anandamide, another endocannabinoid lipid. The results suggest that hydrolysis by means of MGL is a primary mechanism for 2-AG inactivation in intact neurons.
    Proceedings of the National Academy of Sciences 09/2002; 99(16):10819-24. · 9.81 Impact Factor