Enhanced anandamide degradation is associated with neuronal apoptosis induced by the HIV-1 coat glycoprotein gp120 in the rat neocortex

Human immunodeficiency virus type-1 coat glycoprotein gp120 causes delayed apoptosis in rat brain neocortex. Here, we investigated the possible role of the endocannabinoid system in this process. It is shown that gp120 causes a time-dependent increase in the activity and immunoreactivity of the anandamide (AEA)-hydrolyzing enzyme fatty acid amide hydrolase (FAAH), paralleled by increased activity of the AEA membrane transporter and decreased endogenous levels of AEA. The AEA-synthesizing phospholipase D and the AEA-binding receptors were not affected by gp120. None of the changes induced by gp120 in the cortex were induced by bovine serum albumin, nor were they observed in the hippocampus of the same animals. Also, the activity of 5-lipoxygenase, which generates AEA derivatives able to inhibit FAAH, decreased down to approximately 25% of the control activity upon gp120 treatment, due to reduced protein level ( approximately 45%). In addition, the FAAH inhibitor methyl-arachidonoyl fluorophosphonate significantly reduced gp120-induced apoptosis in rat brain neocortex, whereas selective blockers of AEA membrane transporter or of AEA-binding receptors were ineffective. Taken together, these results suggest that gp120, by activating FAAH, decreases endogenous levels of AEA, and the latter effect seems instrumental in the execution of delayed neuronal apoptosis in the brain neocortex of rats.     

Symptom-related changes of endocannabinoid and palmitoylethanolamide levels in brain areas of R6/2 mice, a transgenic model of Huntington's disease

Previous studies have shown an impairment of the endocannabinoid system in experimental models of Huntington's disease. In transgenic R6/2 mice, created by inserting exon 1 of the human IT15 mutant gene into the mouse, and exhibiting 150 CAG repeats as well as signs of HD, a progressive decline of CB(1) receptor expression and an abnormal sensitivity to CB(1) receptor stimulation have been reported. Here, by using isotope-dilution liquid chromatography-mass spectrometry, we investigated whether the levels of three endogenous neuroprotective substances, the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG), and palmitoylethanolamide (PEA), are altered in different brain areas of transgenic R6/2 versus wild-type (WT) mice at two different disease phases, i.e. in pre-symptomatic (4.5 weeks) or overtly symptomatic (10 weeks) R6/2 mice versus age-matched WT mice (n=4/group). Except for a approximately 25% decrease in 2-AG levels in the cortex, no significant changes in endocannabinoid and PEA levels were observed in pre-symptomatic R6/2 versus WT mice. By contrast, in symptomatic R6/2 mice the levels of all three compounds were significantly (approximately 30-60%) decreased in the striatum, whereas little changes were observed in the hippocampus, and a approximately 28% decrease of 2-AG levels, accompanied by a approximately 50% increase of AEA levels, was found in the cortex. These findings show that endocannabinoid levels change in a disease phase- and region-specific way in the brain of R6/2 mice and indicate that an impaired endocannabinoid system is a hallmark of symptomatic HD, thus suggesting that drugs inhibiting endocannabinoid degradation might be used to treat this disease.     

Endocannabinoids,hormone-cytokine networks and human fertility

Anandamide (N -arachidonoylethanolamine, AEA) is a major endocannabinoid, shown to impair mouse pregnancy and embryo development and to induce apoptosis in blastocysts. Here, we review the roles of AEA, of the AEA-binding cannabinoid (CB) receptors, of the selective AEA membrane transporter (AMT), and of the AEA-hydrolyzing enzyme fatty acid amide hydrolase (FAAH), in human gestation. In particular, we discuss the interplay between the endocannabinoid system and the hormone-cytokine array involved in the control of human pregnancy, showing that the endocannabinoids take part in the immunological adaptation occurring during early pregnancy. In this line, we discuss the critical role of FAAH in human peripheral lymphocytes, showing that the expression of this enzyme is regulated by progesterone, Th1 and Th2 cytokines, which also regulate fertility. Moreover, we show that AEA and the other endocannabinoid, 2-arachidonoylglycerol, inhibit the release of the fertility-promoting cytokine leukemia inhibitory factor from human lymphocytes. Taken together, low FAAH and consistently high blood levels of AEA, but not CB receptors or AMT, can be early (<8 weeks of gestation) markers of spontaneous abortion, potentially useful as diagnostic tools for large-scale, routine monitoring of gestation in humans.     

Levodopa treatment reverses endocannabinoid system abnormalities in experimental parkinsonism

Cannabinoid receptors and their endogenous ligands are potent inhibitors of neurotransmitter release in the brain. Here, we show that in a rat model of Parkinson's disease induced by unilateral nigral lesion with 6-hydroxydopamine (6-OHDA), the striatal levels of the endocannabinoid anandamide (AEA) were increased, while the activity of its membrane transporter and hydrolase (fatty-acid amide hydrolase, FAAH) were decreased. These changes were not observed in the cerebellum of the same animals. Moreover, the frequency and amplitude of glutamate-mediated spontaneous excitatory post-synaptic currents were augmented in striatal spiny neurones recorded from parkinsonian rats. Remarkably, the anomalies in the endocannabinoid system, as well as those in glutamatergic activity, were completely reversed by chronic treatment of parkinsonian rats with levodopa, and the pharmacological inhibition of FAAH restored a normal glutamatergic activity in 6-OHDA-lesioned animals. Thus, the increased striatal levels of AEA may reflect a compensatory mechanism trying to counteract the abnormal corticostriatal glutamatergic drive in parkinsonian rats. However, this mechanism seems to be unsuccessful, since spontaneous excitatory activity is still higher in these animals. Taken together, these data show that anomalies in the endocannabinoid system induced by experimental parkinsonism are restricted to the striatum and can be reversed by chronic levodopa treatment, and suggest that inhibition of FAAH might represent a possible target to decrease the abnormal cortical glutamatergic drive in Parkinson's disease.     

The postmortal accumulation of brain N-arachidonylethanolamine (anandamide) is dependent upon fatty acid amide hydrolase activity

N-arachidonylethanolamine (AEA) accumulates during brain injury and postmortem. Because fatty acid amide hydrolase (FAAH) regulates brain AEA content, the purpose of this study was to determine its role in the postmortal accumulation of AEA using FAAH null mice. As expected, AEA content in immediately frozen brain tissue was significantly greater in FAAH-deficient (FAAH-/-) than in wild-type mice. However, AEA content was significantly lower in brains from FAAH-/- mice at 5 and 24 h postmortem. Similarly, wild-type mice treated in vivo with a FAAH inhibitor (URB532) had significantly lower brain AEA content 24 h postmortem compared with controls. These data indicate that FAAH contributes significantly to the postmortal accumulation of AEA. In contrast, the accumulations of two other N-acylethanolamines, N-oleoylethanolamine (OEA) and N-palmitoylethanolamine (PEA), were not reduced at 24 h postmortem in either the FAAH-/- mice or mice treated with URB532. FAAH-/- mice accumulated significantly less ethanolamine at 24 h postmortem compared with wild-type mice, suggesting that FAAH activity plays a role in the accumulation of ethanolamine postmortem. These data demonstrate that FAAH activity differentially affects AEA and OEA/PEA contents postmortem and suggest that AEA formation specifically occurs via an ethanolamine-dependent route postmortem.     

The endocannabinoid system in human keratinocytes. Evidence that anandamide inhibits epidermal differentiation through CB1 receptor-dependent inhibition of protein kinase C,activation protein-1, and transglutaminase

Anandamide (AEA), a prominent member of the endogenous ligands of cannabinoid receptors (endocannabinoids), is known to affect several functions of brain and peripheral tissues. A potential role for AEA in skin pathophysiology has been proposed, yet its molecular basis remains unknown. Here we report unprecedented evidence that spontaneously immortalized human keratinocytes (HaCaT) and normal human epidermal keratinocytes (NHEK) have the biochemical machinery to bind and metabolize AEA, i.e. a functional type-1 cannabinoid receptor (CB1R), a selective AEA membrane transporter (AMT), an AEA-degrading fatty acid amide hydrolase (FAAH), and an AEA-synthesizing phospholipase D (PLD). We show that, unlike CB1R and PLD, the activity of AMT and the activity and expression of FAAH increase while the endogenous levels of AEA decrease in HaCaT and NHEK cells induced to differentiate in vitro by 12-O-tetradecanoylphorbol 13-acetate (TPA) plus calcium. We also show that exogenous AEA inhibits the formation of cornified envelopes, a hallmark of keratinocyte differentiation, in HaCaT and NHEK cells treated with TPA plus calcium, through a CB1R-dependent reduction of transglutaminase and protein kinase C activity. Moreover, transient expression in HaCaT cells of the chloramphenicol acetyltransferase reporter gene under control of the loricrin promoter, which contained a wild-type or mutated activating protein-1 (AP-1) site, showed that AEA inhibited AP-1 in a CB1R-dependent manner. Taken together, these data demonstrate that human keratinocytes partake in the peripheral endocannabinoid system and show a novel signaling mechanism of CB1 receptors, which may have important implications in epidermal differentiation and skin development.     

Methylation and acetylation of 15-hydroxyanandamide modulate its interaction with the endocannabinoid system

The biological activity of endocannabinoids like anandamide (AEA) and 2-arachidonoylglycerol (2-AG) is subjected in vivo to a “metabolic control”, exerted mainly by catabolic enzymes. AEA is inactivated by fatty acid amide hydrolase (FAAH), that is inhibited competitively by hydroxyanandamides (HAEAs) generated from AEA by lipoxygenase activity. Among these derivatives, 15-HAEA has been shown to be an effective (K_i ∼0.6 μM) FAAH inhibitor, that blocks also type-1 cannabinoid receptor (CB1R) but not other components of the “endocannabinoid system (ECS)”, like the AEA transporter (AMT) or CB2R. Here, we extended the study of the effect of 15-HAEA on the AEA synthetase (NAPE-PLD) and the AEA-binding vanilloid receptor (TRPV1), showing that 15-HAEA activates the former (up to ∼140% of controls) and inhibits the latter protein (down to ∼70%). We also show that 15-HAEA halves the synthesis of 2-AG and almost doubles the transport of this compound across the membrane. In addition, we synthesized methyl and acetyl derivatives of 15-HAEA (15-MeOAEA and 15-AcOAEA, respectively), in order to check their ability to modulate FAAH and the other ECS elements. In fact, methylation and acetylation are common biochemical reactions in the cellular environment. We show that 15-MeOAEA, unlike 15-AcOAEA, is still a powerful competitive inhibitor of FAAH (K_i ∼0.7 μM), and that both derivatives have negligible interactions with the other proteins of ECS. Therefore, 15-MeOAEA is a FAAH inhibitor more selective than 15-HAEA. Further molecular dynamics analysis gave clues to the molecular requirements for the interaction of 15-HAEA and 15-MeOAEA with FAAH.     

Comparison of anandamide transport in FAAH wild-type and knockout neurons: evidence for contributions by both FAAH and the CB1 receptor to anandamide uptake

The cellular inactivation of the endogenous cannabinoid (endocannabinoid) anandamide (AEA) represents a controversial and intensely investigated subject. This process has been proposed to involve two proteins, a transporter that promotes the cellular uptake of AEA and fatty acid amide hydrolase (FAAH), which hydrolyzes AEA to arachidonic acid. However, whereas the role of FAAH in AEA metabolism is well-characterized, the identity of the putative AEA transporter remains enigmatic. Indeed, the indirect pharmacological evidence used to support the existence of an AEA transporter has been suggested also to be compatible with a model in which AEA uptake is driven by simple diffusion coupled to FAAH metabolism. Here, we have directly addressed the contribution of FAAH to AEA uptake by examining this process in neuronal preparations from FAAH(-/-) mice and in the presence of the uptake inhibitor UCM707. The results of these studies reveal that (i) care should be taken to avoid the presence of artifacts when studying the cellular uptake of lipophilic molecules like AEA, (ii) FAAH significantly contributes to AEA uptake, especially with longer incubation times, and (iii) a UCM707-sensitive protein(s) distinct from FAAH also participates in AEA uptake. Interestingly, the FAAH-independent component of AEA transport was significantly reduced by pretreatment of neurons with the cannabinoid receptor 1 (CB1) antagonist SR141716A. Collectively, these results indicate that the protein-dependent uptake of AEA is largely mediated by known constituents of the endocannabinoid system (FAAH and the CB1 receptor), although a partial contribution of an additional UCM707-sensitive protein is also suggested.     

An anatomical and temporal portrait of physiological substrates for fatty acid amide hydrolase

Fatty acid amide hydrolase (FAAH) regulates amidated lipid transmitters, including the endocannabinoid anandamide and its N-acyl ethanolamine (NAE) congeners and transient receptor potential channel agonists N-acyl taurines (NATs). Using both the FAAH inhibitor PF-3845 and FAAH(-/-) mice, we present a global analysis of changes in NAE and NAT metabolism caused by FAAH disruption in central and peripheral tissues. Elevations in anandamide (and other NAEs) were tissue dependent, with the most dramatic changes occurring in brain, testis, and liver of PF-3845-treated or FAAH(-/-) mice. Polyunsaturated NATs accumulated to very high amounts in the liver, kidney, and plasma of these animals. The NAT profile in brain tissue was markedly different and punctuated by significant increases in long-chain NATs found exclusively in FAAH(-/-), but not in PF-3845-treated animals. Suspecting that this difference might reflect a slow pathway for NAT biosynthesis, we treated mice chronically with PF-3845 for 6 days and observed robust elevations in brain NATs. These studies, taken together, define the anatomical and temporal features of FAAH-mediated NAE and NAT metabolism, which are complemented and probably influenced by kinetically distinguishable biosynthetic pathways that produce these lipids in vivo.     

Endocannabinoid metabolism in the absence of fatty acid amide hydrolase (FAAH): discovery of phosphorylcholine derivatives of N-acyl ethanolamines

Lipid transmitters are tightly regulated by a balance of biosynthetic and degradative enzymes. Termination of the activity of the N-acyl ethanolamine (NAE) class of lipid-signaling molecules, including the endocannabinoid anandamide (AEA), is principally mediated by the integral membrane enzyme fatty acid amide hydrolase (FAAH) in vivo. FAAH(-/-) mice are highly sensitized to the pharmacological effects of AEA; however, these animals eventually recover from AEA treatment, implying the existence of alternative routes for NAE metabolism. Here, we have pursued the characterization of these pathways by profiling the metabolome of FAAH(-/-) mice treated with AEA. Multiple AEA-induced metabolites were observed in brains from FAAH(-/-) mice, including a major product with a mass shift of +165 Da (m/z 513). The structure of this product was determined to be O-phosphorylcholine (PC)-AEA. Analysis of untreated mice identified PC-NAEs as endogenous constituents of the central nervous system (CNS) that were highly elevated in FAAH(-/-) animals. PC-NAEs were very poor substrates for FAAH; however, a vanadate-sensitive enzymatic activity was detected in brain membranes that converted PC-NAEs back to their parent NAEs. The choline-specific phosphodiesterase NPP6 was identified as a candidate enzyme responsible for this activity. These data indicate the presence of a complete metabolic pathway for the production and degradation of PC-NAEs in the CNS that constitutes an alternative route for endocannabinoid metabolism.     

Levels, metabolism, and pharmacological activity of anandamide in CB(1) cannabinoid receptor knockout mice: evidence for non-CB(1), non-CB(2) receptor-mediated actions of anandamide in mouse brain

Anandamide [arachidonylethanolamide (AEA)] appears to be an endogenous agonist of brain cannabinoid receptors (CB(1)), yet some of the neurobehavioral effects of this compound in mice are unaffected by a selective CB(1) antagonist. We studied the levels, pharmacological actions, and degradation of AEA in transgenic mice lacking the CB(1) gene. We quantified AEA and the other endocannabinoid, 2-arachidonoyl glycerol, in six brain regions and the spinal cord by isotope-dilution liquid chromatography-mass spectrometry. The distribution of endocannabinoids and their inactivating enzyme, fatty acid amide hydrolase, were found to overlap with CB(1) distribution only in part. In CB(1) knockout homozygotes (CB(1)-/-), the hippocampus and, to a lesser extent, the striatum exhibited lower AEA levels as compared with wild-type (CB(1)+/+) controls. These data suggest a ligand/receptor relationship between AEA and CB(1) in these two brain regions, where tonic activation of the receptor may tightly regulate the biosynthesis of its endogenous ligand. 2-Arachidonoyl glycerol levels and fatty acid amide hydrolase activity were unchanged in CB(1)-/- with respect to CB(1)+/+ mice in all regions. AEA and Delta(9)-tetrahydrocannabinol (THC) were tested in CB(1)-/- mice for their capability of inducing analgesia and catalepsy and decreasing spontaneous activity. The effects of AEA, unlike THC, were not decreased in CB(1)-/- mice. AEA, but not THC, stimulated GTPgammaS binding in brain membranes from CB(1)-/- mice, and this stimulation was insensitive to CB(1) and CB(2) antagonists. We suggest that non-CB(1), non-CB(2) G protein-coupled receptors might mediate in mice some of the neuro-behavioral actions of AEA.     

Critical enzymes involved in endocannabinoid metabolism

Investigations of the pathways involved in the metabolism of endocannabinoids have grown exponentially in recent years following the discovery of cannabinoid receptors (CB) and their endogenous ligands, such as anandamide (AEA) and 2-arachidonoylglycerol (2-AG). The in vivo biosynthesis of AEA has been shown to occur through several pathways mediated by N-acylphosphatidylethanolamide-phospholipase D (NAPE-PLD), a secretory PLA(2) and PLC. 2-AG, a second endocannabinoid is generated through the action of selective enzymes such as phosphatidic acid phsophohydrolase, diacylglycerol lipase (DAGL), phosphoinositide-specific PLC (PI-PLC) and lyso-PLC. A putative membrane transporter or facilitated diffusion is involved in the cellular uptake or release of endocannabinoids. AEA is metabolized by fatty acid amidohydrolase (FAAH) and 2-AG is metabolized by both FAAH and monoacylglycerol lipase (MAGL). The author presents an integrative overview of current research on the enzymes involved in the metabolism of endocannabinoids and discusses possible therapeutic interventions for various diseases, including addiction.     

Lipopolysaccharide induces anandamide synthesis in macrophages via CD14/MAPK/phosphoinositide 3-kinase/NF-kappaB independently of platelet-activating factor

Macrophage-derived endocannabinoids have been implicated in endotoxin (lipopolysaccharide (LPS))-induced hypotension, but the endocannabinoid involved and the mechanism of its regulation by LPS are unknown. In RAW264.7 mouse macrophages, LPS (10 ng/ml) increases anandamide (AEA) levels >10-fold via CD14-, NF-kappaB-, and p44/42-dependent, platelet-activating factor-independent activation of the AEA biosynthetic enzymes, N-acyltransferase and phospholipase D. LPS also induces the AEA-degrading enzyme fatty acid amidohydrolase (FAAH), and inhibition of FAAH activity potentiates, whereas actinomycin D or cycloheximide blocks the LPS-induced increase in AEA levels and N-acyltransferase and phospholipase D activities. In contrast, cellular levels of the endocannabinoid 2-arachidonoylglycerol (2-AG) are unaffected by LPS but increased by platelet-activating factor. LPS similarly induces AEA, but not 2-AG, in mouse peritoneal macrophages where basal AEA levels are higher, and the LPS-stimulated increase in AEA is potentiated in cells from FAAH-/- as compared with FAAH+/+ mice. Intravenous administration of 107 LPS-treated mouse macrophages to anesthetized rats elicits hypotension, which is much greater in response to FAAH-/- than FAAH+/+ cells and is susceptible to inhibition by SR141716, a cannabinoid CB1 receptor antagonist. We conclude that AEA and 2-AG synthesis are differentially regulated in macrophages, and AEA rather than 2-AG is a major contributor to LPS-induced hypotension.     

Recent advances in the synthesis of endocannabinoid related ligands

The chemical strategies used for the synthesis of various ligands related to the endocannabinoid system namely anandamide (AEA), 2-arachidonylglycerol (2-Ara-Gl), CB1/(vanilloid receptors) VR1, anandamide membrane transporter (AMT) and fatty acid amide hydrolase (FAAH) are described in this review. In general, the chemical synthesis of analogs with changes in the head group of AEA was quite straightforward involving the conversion of an acid to an amide or an ester. Analogs which had modifications in the end pentyl chain were more difficult to synthesize and required multistep synthetic sequences to prepare the target compounds. A facile total synthesis of 2-Ara-Gl was reported and an HPLC procedure for its identification and quantification was developed, but because of the instability of 2-Ara-Gl another synthesis was developed so that it can be stored as the more stable phenylboronate ester. Similarly the chemical synthesis of various ligands in the remaining areas of CB1/VR1, AMT and FAAH are described. A summary of the present state of knowledge about the SAR in each area is presented to help in the design and synthesis of novel ligands for the future.     

Anandamide and 2-arachidonoylglycerol inhibit fatty acid amide hydrolase by activating the lipoxygenase pathway of the arachidonate cascade

Treatment of intact human neuroblastoma CHP100 cells with anandamide (arachidonoylethanolamide, AEA) or 2-arachidonoylglycerol (2-AG) inhibits intracellular fatty acid amide hydrolase (FAAH). This effect was not associated with covalent modifications of FAAH, since specific inhibitors of farnesyltransferase, kinases, phosphatases, glycosyltransferase or nitric oxide synthase were ineffective. Electrophoretic analysis of (33)P-labelled proteins, Western blot with anti-phosphotyrosine antibodies, and glycan analysis of cellular proteins confirmed the absence of covalent modifications of FAAH. The inhibition by AEA was paralleled by an increased arachidonate release, which was not observed upon treatment of cells with linoleoylethanolamide, palmitoylethanolamide, or oleoylethanolamide. Moreover, cell treatment with AEA or 2-AG increased the activity of cyclooxygenase and 5-lipoxygenase, and the hydro(pero)xides generated from arachidonate by lipoxygenase were shown to inhibit FAAH, with inhibition constants in the low micromolar range. Consistently, inhibitors of 5-lipoxygenase, but not those of cyclooxygenase, significantly counteracted the inhibition of FAAH by AEA or 2-AG.     

Structure-activity relationships among N-arachidonylethanolamine (Anandamide) head group analogues for the anandamide transporter

Two putative endocannabinoids, N-arachidonylethanolamine (AEA) and 2-arachidonylglycerol, are inactivated by removal from the extracellular environment by a process that has the features of protein-mediated facilitated diffusion. We have synthesized and studied 22 N-linked analogues of arachidonylamide for the purpose of increasing our understanding of the structural requirements for the binding of ligands to the AEA transporter. We have also determined the affinities of these analogues for both the CB(1) cannabinoid receptor and fatty acid amide hydrolase (FAAH). We have identified several structural features that enhance binding to the AEA transporter in cerebellar granule cells. We have confirmed the findings of others that replacing the ethanolamine head group with 4-hydroxybenzyl results in a high-affinity ligand for the transporter. However, we find that the same molecule is also a competitive inhibitor of FAAH. Similarly, replacement of the ethanolamine of AEA with 3-pyridinyl also results in a high-affinity inhibitor of both the transporter and FAAH. We conclude that the structural requirements for ligand binding to the CB(1) receptor and binding to the transporter are very different; however, the transporter and FAAH share most, but not all, structural requirements.     

Lack of CB1 receptor activity impairs serotonergic negative feedback

Serotonergic and endocannabinoid systems are important substrates for the control of emotional behaviour and growing evidence show an involvement in the pathophysiology of mood disorders. In the present study, the absence of the activity of the CB₁ cannabinoid receptor impaired serotonergic negative feedback in mice. Thus, in vivo microdialysis experiments revealed increased basal 5-HT extracellular levels and attenuated fluoxetine-induced increase of 5-HT extracellular levels in the prefrontal cortex of CB₁ knockout compared with wild-type mice. These observations could be related to the significant reduction in the 5-HT transporter binding site density detected in frontal cortex and hippocampus of CB₁ knockout mice. The lack of CB₁ receptor also altered some 5-HT receptors related to the 5-HT feedback. Extracellular recordings in the dorsal raphe nucleus (DRN) revealed that the genetic and pharmacological blockade of CB₁ receptor induced a 5-HT_1A autoreceptor functional desensitization. In situ hybridization studies showed a reduction in the expression of the 5-HT_2C receptor within several brain areas related to the control of the emotional responses, such as the DRN, the nucleus accumbens and the paraventricular nucleus of the hypothalamus, whereas an over-expression was observed in the CA3 area of the ventral hippocampus. These results reveal that the lack of CB₁ receptor induces a facilitation of the activity of serotonergic neurons in the DRN by altering different components of the 5-HT feedback as well as an increase in 5-HT extracellular levels in the prefrontal cortex in mice.     

Exogenous Nerve Growth Factor Increases the Activity of High-Affinity Choline Uptake and Choline Acetyltransferase in Brain of Fisher 344 Male Rats

The objective of this study was to determine the effect of age and chronic intracerebral administration of nerve growth factor (NGF) on the activity of the presynaptic cholinergic neuronal markers hemicholinium-sensitive high-affinity choline uptake (HACU) and choline acetyltransferase (ChAT) in the brain of Fisher 344 male rats. In 24-month-old rats, a substantial decrease in ChAT activity (30%) was measured in striatum, and decreases in HACU were found in frontal cortex (28%) and hippocampus (23%) compared with 4-month-old controls. Cholinergic neurons in brain of both young adult and aged rats responded to administration of exogenous NGF by increased expression of both phenotypes. In 4-month-old animals, NGF treatment at 1.2 micron/day resulted in increased activities of both ChAT and HACU in striatum (175 and 170%, respectively), frontal cortex (133 and 125%), and hippocampus (137 and 125%) compared with untreated and vehicle-treated 4-month-old animals; vehicle treatment had no effect on the activity of either marker. In 24-month-old animals treated with NGF for 2 weeks, ChAT activity was increased in striatum (179%), frontal cortex (134%), and hippocampus (119%) compared with 24-month-old control animals. Synaptosomal HACU in 24-month-old rats was increased in striatum (151%) and frontal cortex (128%) after 2 weeks of NGF treatment, but hippocampal HACU was not significantly different from control values.(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of fatty acid amide hydrolase inhibition in nicotine reward and dependence

The endogenous cannabinoid anandamide (AEA) exerts the majority of its effects at CB₁ and CB₂ receptors and is degraded by fatty acid amide hydrolase (FAAH). FAAH KO mice and animals treated with FAAH inhibitors are impaired in their ability to hydrolyze AEA and other non-cannabinoid lipid signaling molecules, such as oleoylethanolamide (OEA) and palmitoylethanolamide (PEA). AEA and these other substrates activate non-cannabinoid receptor systems, including TRPV1 and PPAR-α receptors. In this mini review, we describe the functional consequences of FAAH inhibition on nicotine reward and dependence as well as the underlying endocannabinoid and non-cannabinoid receptor systems mediating these effects. Interestingly, FAAH inhibition seems to mediate nicotine dependence differently in mice and rats. Indeed, pharmacological and genetic FAAH disruption in mice enhances nicotine reward and withdrawal. However, in rats, pharmacological blockade of FAAH significantly inhibits nicotine reward and has no effect in nicotine withdrawal. Studies suggest that non-cannabinoid mechanisms may play a role in these species differences.