Characterization of maternal motivation in the lactating rat: Contrasts between early and late postpartum responses

We previously assessed the motivational properties of pups relative to those of cocaine in parturient female rats (dams) across the postpartum period and demonstrated that the larger subset of dams in early postpartum (PPD8) preferred the pup-associated chamber, whereas the majority of dams tested in late postpartum (PPD16) preferred the cocaine-associated chamber [Mattson, B.J., Williams, S., Rosenblatt, J.S., Morrell, J.I. 2001. Comparison of two positive reinforcing stimuli: pups and cocaine throughout the postpartum period. Behav. Neurosci., 115, 683-694; Seip, K.M., Morrell, J.I. 2007. Increasing the incentive salience of cocaine challenges preference for pup- over cocaine-associated stimuli during early postpartum: place preference and locomotor analyses in the lactating female rat. Psychopharmacology 194, 309-319]. The present study uses a dual-choice conditioned place preference to ask how the progression of the postpartum period, including natural pup development, influences maternal motivation for pups. Preferences for cued chambers associated with pups that were age-matched to the postpartum stage of the dam in contrast to a stimulus with little incentive salience were higher during the early than the late postpartum, suggesting that the incentive salience of pups diminishes as the postpartum period progresses. Preferences of the early postpartum dams deprived of pups for 15 min, 2, 6, 12 or 22 hrs prior to conditioning and testing did not differ statistically but there was a trend of more pup preference after 22 hr deprivation; pup age was not an important factor in early postpartum. In marked contrast, late postpartum dams only exhibited robust pup-associated place preference when they were conditioned with young (4-7 day-old) pups or after a 22 hr period of deprivation from contemporaneous pups. Together these results suggest that both forces are at work in the mother-pup dyad, changes in the pups as they develop and changes in the physiological and endocrine state of the female as she progresses through the postpartum period.     

Effects of rTMS on Hippocampal Endocannabinoids and Depressive-like Behaviors in Adolescent Rats

Depression is a common mental disorder in adolescents, with a prevalence rate of 5.6%. Current anti-depressive options for adolescents are limited: psychological intervention and conventional antidepressants have low efficacy, a delayed onset of action and increased possibility of suicidal risk. Repetitive transcranial magnetic stimulation (rTMS) as an effective and noninvasive physical therapy for adult depression has been investigated in recent years. However, whether it also produces similar effects on juvenile depression and the underlying mechanism are not clearly understood. In this study, chronic unpredictable mild stress (CMS) was applied to 3-week-old male Sprague Dawley rats for 21 days. Then rTMS was performed for seven consecutive days, and the anti-depressive effects were evaluated by behavioral tests including the sucrose preference test (SPT), the forced swimming test (FST), and the novelty suppressed feeding test (NSF). Expression of hippocampal cannabinoid type I receptor (CB1R), 2-arachidonoylglycerol (2-AG) and relative synthetase and degradative enzymes-diacylglycerol lipase (DAGL) and monoacylglycerol lipase (MAGL) were also investigated. The behavioral parameters were also observed after the administration of the selective CB1 receptor antagonist AM251. The results showed that CMS induced a significant decrease in sucrose preference, a significant increase of immobility time in the FST, and an increased latency to feed in the NSF. In addition, reduced hippocampal CB1 receptor, 2-AG level and increased MAGL protein expression level were also observed in CMS rats. Meanwhile, rTMS treatment upregulated 2-AG level in the hippocampus and ameliorated depressive-like behaviors. The anti-depressive effect of rTMS was attenuated by AM251, a specific CB1R antagonist that was administered 30 min before the onset of rTMS by either intraperitoneal administration or hippocampal microinjection. These results indicate that rTMS can be used as an antidepressive therapy for juvenile depression at least partly mediated by increasing hippocampal 2-AG and CB1 receptor expression levels.     

Is Self‐Grooming by Male Prairie Voles a Predictor of Mate Choice?

Self-grooming by mammals is a form of scent dissemination in which individuals anoint themselves with salivary, anogenital, and other body odours. Self-grooming has been proposed to be a sexually selected trait favoured in reproductive competition and sexual attraction. We tested the hypothesis that females would show a mating preference for males that self-groomed more than a reproductive competitor that groomed less. In mate-choice experiments in which females had a choice of two tethered males, non pair-bonded females did not choose males based on their frequency of self-grooming. In a second experiment in which pair-bonded females in postpartum oestrus had access to their current mate and two strange males, strange males groomed significantly more than pair-bonded mates, yet attained the fewest copulations. Non pair-bonded females and pair-bonded males and females groomed significantly less often than did non pair-bonded males. Self-grooming behaviour was consistent with the sexual attraction hypothesis, but the frequency of self-grooming did not increase a male's mating success. We conclude that the frequency and time spent self grooming are not good predictors of mating success.     

Lid domain plasticity and lipid flexibility modulate enzyme specificity in human monoacylglycerol lipase

Human monoacylglycerol lipase (MAGL) is a membrane-interacting enzyme that generates pro-inflammatory signaling molecules. For this reason, MAGL inhibition is a promising strategy to treat pain, cancer, and neuroinflammatory diseases. MAGL can hydrolyze monoacylglycerols bearing an acyl chain of different lengths and degrees of unsaturation, cleaving primarily the endocannabinoid 2-arachidonoylglycerol. Importantly, the enzymatic binding site of MAGL is confined by a 75-amino-acid-long, flexible cap domain, named ‘lid domain’, which is structurally similar to that found in several other lipases. However, it is unclear how lid domain plasticity affects catalysis in MAGL. By integrating extensive molecular dynamics simulations and free-energy calculations with mutagenesis and kinetic experiments, we here define a lid-domain-mediated mechanism for substrate selection and binding in MAGL catalysis. In particular, we clarify the key role of Phe159 and Ile179, two conserved residues within the lid domain, in regulating substrate specificity in MAGL. We conclude by proposing that other structurally related lipases may share this lid-domain-mediated mechanism for substrate specificity.     

Development of terphenyl-2-methyloxazol-5(4H)-one derivatives as selective reversible MAGL inhibitors

Monoacylglycerol lipase is a serine hydrolase that plays a major role in the degradation of the endocannabinoid neurotransmitter 2-arachidonoylglycerol. A wide number of MAGL inhibitors are reported in literature; however, many of them are characterised by an irreversible mechanism of action and this behavior determines an unwanted chronic MAGL inactivation, which acquires a functional antagonism of the endocannabinoid system. The possible use of reversible MAGL inhibitors has only recently been explored, due to the lack of known compounds possessing efficient reversible inhibitory activities. In this work, we report a new series of terphenyl-2-methyloxazol-5(4H)-one derivatives characterised by a reversible MAGL-inhibition mechanism. Among them, compound 20b showed to be a potent MAGL reversible inhibitor (IC₅₀?=?348?nM) with a good MAGL/FAAH selectivity. Furthermore, this compound showed antiproliferative activities against two different cancer cell lines that overexpress MAGL.     

Relationship between the preference for sake (Japanese rice wine) and the movements of metabolic parameters coinciding with sake intake

Selection of foods by animals largely depends on their physiological condition. In this paper, we reported how physiological changes in rats after ingestion of sake (Japanese rice wine) affected their preference for different kinds of sake. Rats could discriminate among various kinds of sakes in a two-bottle choice test, even after adjustment of the glucose concentration or alcohol concentration of the sake, suggesting that the choice of a sake by animals was based on more than one ingredient. To identify effect of a rat's physiological condition on the selection of sake, we monitored the levels of blood glucose, nonesterified fatty acids (NEFA), and ketone bodies in serum in mice after forced intragastric ingestion of sake that had previously been offered to the rats in a two-bottle choice test. Blood glucose levels in mice were not different between the rats fed the palatable and unpalatable sakes, and the NEFA level and ketone level were high in rats fed the unpalatable sake. To further clarify the relationship between physiological condition and preference for sake, rats were offered eight kinds of Junmai-syu, which is made with only rice and no sub-ingredient. Rats could still discriminate among Junmai-syu. Furthermore, with two exceptions, the preference for a sake was significantly correlated with the ketone level. The order in the ratio of blood insulin level to blood glucagon level, which is an indicator of metabolism, was correlated with the order in preference to sake. These results suggest that some physiological factors besides oral stimulation also are important factors in the selection of a sake.     

Identification and characterization of a new reversible MAGL inhibitor

Monoacylglycerol lipase is a serine hydrolase that play a major role in the degradation of 2-arachidonoylglycerol, an endocannabinoid neurotransmitter implicated in several physiological processes. Recent studies have shown the possible role of MAGL inhibitors as anti-inflammatory, anti-nociceptive and anti-cancer agents. The use of irreversible MAGL inhibitors determined an unwanted chronic MAGL inactivation, which acquires a functional antagonism function of the endocannabinoid system. However, the application of reversible MAGL inhibitors has not yet been explored, mainly due to the scarcity of known compounds possessing efficient reversible inhibitory activities. In this study we reported the first virtual screening analysis for the identification of reversible MAGL inhibitors. Among the screened compounds, the (4-(4-chlorobenzoyl)piperidin-1-yl)(4-methoxyphenyl)methanone (CL6a) is a promising reversible MAGL inhibitor lead (K_i = 8.6 μM), which may be used for the future development of a new class of MAGL inhibitors. Furthermore, the results demonstrate the validity of the methodologies that we followed, encouraging additional screenings of other commercial databases.     

4-Aryliden-2-methyloxazol-5(4H)-one as a new scaffold for selective reversible MAGL inhibitors

This study reports on a preliminary structure–activity relationship exploration of 4-aryliden-2-methyloxazol-5(4H)-one-based compounds as MAGL/FAAH inhibitors. Our results highlight that this scaffold may serve for the development of selective MAGL inhibitors. A 69-fold selectivity against MAGL over FAAH was achieved for compound 16b (MAGL and FAAH IC₅₀?=?1.6 and 111?µM, respectively). Furthermore, the best compound behaved as a reversible ligand and showed promising antiproliferative activity in cancer cells.     

Piperazine and piperidine carboxamides and carbamates as inhibitors of fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL)

The key hydrolytic enzymes of the endocannabinoid system, fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL), are potential targets for various therapeutic applications. In this paper, we present more extensively the results of our previous work on piperazine and piperidine carboxamides and carbamates as FAAH and MAGL inhibitors. The best compounds of these series function as potent and selective MAGL/FAAH inhibitors or as dual FAAH/MAGL inhibitors at nanomolar concentrations. This study revealed that MAGL inhibitors should comprise leaving-groups with a conjugate acid pK_a of 8–10, while diverse leaving groups are tolerated for FAAH inhibitors.     

Docking based virtual screening and molecular dynamics study to identify potential monoacylglycerol lipase inhibitors

Monoacylglycerol lipase (MAGL) is one of the key enzymes of the endocannabinoid system (ECS). It hydrolyzes one of the major endocannabinoid, 2-arachidonoylglycerol (2-AG), an endogenous full agonist at G protein coupled cannabinoid receptors CB1 and CB2. Numerous studies showed that MGL inhibitors are potentially useful for the treatment of pain, inflammation, cancer and CNS disorders. These provocative findings suggested that pharmacological inhibition of MAGL function may confer significant therapeutic benefits. In this study, we presented hybrid ligand and structure-based approaches to obtain a novel set of virtual leads as MAGL inhibitors. The constraints used in this study, were Glide score, binding free energy estimates and ADME properties to screen the ZINC database, containing approximately 21 million compounds. A total of seven virtual hits were obtained, which showed significant binding affinity towards MAGL protein. Ligand, ZINC24092691 was employed in complex form with the protein MAGL, for molecular dynamics simulation study, because of its excellent glide score, binding free energy and ADME properties. The RMSD of ZINC24092691 was observed to stay at 0.1 nm (1 Å) in most of the trajectories, which further confirmed its ability to inhibit the protein MAGL. The hits were then evaluated for their ability to inhibit human MAGL. The compound ZINC24092691 displayed the noteworthy inhibitory activity reducing MAGL activity to 21.15% at 100 nM concentration, with an IC₅₀ value of 10 nM.     

Lipoprotein lipase in rat heart--II. Influence of apolipoproteins and nutritional factors on tri-, di- and monoacylglycerol lipase activities in post-heparin effluents

1. The in vitro effects of serum and apolipoproteins (apo), and the influence of the nutritional state of the animals were compared on triacylglycerol lipase (TAGL), diacylglycerol lipase (DAGL) and monoacylglycerol lipase (MAGL) activities in post-heparin effluent from rat heart. 2. Serum and apoC-II stimulated DAGL and MAGL 3-fold less than TAGL, the activity that measures lipoprotein lipase (LPL). 3. The preexisting nutritional state of the heart, that strongly modulated LPL, did not influence DAGL and MAGL. 4. ApoA-I, apoC-I, apoC-III1 and apoC-III2 did not stimulate LPL and counteracted its stimulation by apoC-II; MAGL, and not DAGL, was inhibited by apoA-I and apoC-I, an effect reversed by apoC-II. 5. TAGL, DAGL and MAGL appeared to act as a single physiological unit, although differing in functional details; MAGL displayed the greatest dissimilarity.     

Preference for own versus conspecific pups by inbred and outbred rats

Communal care of young may occur when related females live in close proximity to each other. The degree of relatedness between females may affect discrimination of own from conspecific young. Retrieval tests were used to examine preference for own versus conspecific pups by inbred and outbred rats. Dams were given retrieval tests in a neutral arena every other day from day 1 through 10 postpartum. Outbred, but not inbred, dams showed a significant preference for their own pups. Outbred dams also retrieved pups more slowly than did inbred dams. Differences in the behavior of dams and/ or similarity of cues presented by pups may be responsible for the lack of preference for own pups shown by inbred dams. The results of this study suggest that indiscriminate care of young should occur when closely related females share a nest.     

Monoacylglycerol lipase promotes progression of hepatocellular carcinoma via NF-κB-mediated epithelial-mesenchymal transition

Background

Monoacylglycerol lipase (MAGL), a critical lipolytic enzyme, has emerged as a key regulator of tumor progression, yet its biological function and clinical significance in hepatocellular carcinoma (HCC) is still unknown.

Methods

In this study, we used a tissue microarray containing samples from 170 HCC patients to evaluate the expression of MAGL and its correlation with other clinicopathologic characteristics. In addition, we investigated the regulating effects of MAGL on various HCC lines. Finally, we identified the NF-κB signaling pathway participated in MAGL-mediated epithelial-mesenchymal transition (EMT) using HCC cell lines with different metastatic potentials.

Results

The expression of MAGL was significantly higher in HCC tumors than in matched peritumor tissues. Specifically, high MAGL expression was found in tumors with larger tumor size, microvascular invasion, poor differentiation, or advanced TNM stage. In addition, the clinical prognosis for the MAGL^high group was markedly poorer than that for the MAGL^low group in the 1-, 3-, and 5-year overall survival times and recurrence rates of HCC patients. MAGL expression was an independent prognostic factor for both survival and recurrence after curative resection. Furthermore, the upregulation of MAGL in HCC cells promoted cell growth and invasiveness abilities, and accompanied by EMT. In contrast, downregulation of MAGL obviously inhibited these characteristics. Moreover, further investigations verified that MAGL facilitates HCC progression via NF-κB-mediated EMT process.

Conclusions

Our findings demonstrate MAGL could promote HCC progression by the induction of EMT and suggest a potential therapeutic target, as well as a biomarker for prognosis, in patients with HCC.

     

Therapeutic potential of monoacylglycerol lipase inhibitors

Marijuana and aspirin have been used for millennia to treat a wide range of maladies including pain and inflammation. Both cannabinoids, like marijuana, that exert anti-inflammatory action through stimulating cannabinoid receptors, and cyclooxygenase (COX) inhibitors, like aspirin, that suppress pro-inflammatory eicosanoid production have shown beneficial outcomes in mouse models of neurodegenerative diseases and cancer. Both cannabinoids and COX inhibitors, however, have untoward effects that discourage their chronic usage, including cognitive deficits and gastrointestinal toxicity, respectively. Recent studies have uncovered that the serine hydrolase monoacylglycerol lipase (MAGL) links the endocannabinoid and eicosanoid systems together through hydrolysis of the endocannabinoid 2-arachidonoylglycerol (2-AG) to provide the major arachidonic acid (AA) precursor pools for pro-inflammatory eicosanoid synthesis in specific tissues. Studies in recent years have shown that MAGL inhibitors elicit anti-nociceptive, anxiolytic, and anti-emetic responses and attenuate precipitated withdrawal symptoms in addiction paradigms through enhancing endocannabinoid signaling. MAGL inhibitors have also been shown to exert anti-inflammatory action in the brain and protect against neurodegeneration through lowering eicosanoid production. In cancer, MAGL inhibitors have been shown to have anti-cancer properties not only through modulating the endocannabinoid–eicosanoid network, but also by controlling fatty acid release for the synthesis of protumorigenic signaling lipids. Thus, MAGL serves as a critical node in simultaneously coordinating multiple lipid signaling pathways in both physiological and disease contexts. This review will discuss the diverse (patho)physiological roles of MAGL and the therapeutic potential of MAGL inhibitors in treating a vast array of complex human diseases.     

Effects of prenatal administration of testosterone and cortisone on the reproductive system of the female rat.

Testosterone propionate, cortisone, or sesame oil vehicle were given to rats during the last week of pregnancy so that effects of the hormones on anogenital distance, breeding capacity and vaginal opening of the female progeny could be contrasted. Testosterone significantly increased anogenital distance and delayed vaginal opening of progeny. When females that had been exposed to testosterone in utero were tested for breeding capacity, a significantly smaller number mated than in the control group. Female rats that had been exposed to cortisone in utero exhibited premature vaginal opening but did not differ from controls in anogenital distance, and, unlike the testosterone-exposed rats, mated. Cortisone-exposed rats carried litters to term and the litters did not differ from those of controls in numbers of pups or numbers of living pups at birth. The pups born to cortisone-exposed rats had greater birth weights and a higher survival rate to 20 days of age than pups of controls. Results indicate that testosterone administration to rats during pregnancy is far more detrimental to the development and subsequent function of the reproductive system of female progeny than cortisone and suggest that similar changes which occur in response to maternal stress or to administration of ACTH during pregnancy are more likely to result from increases in testosterone than from increases in glucocorticoid secretion.     

Female preference for large males in sailfin mollies,<Emphasis Type="Italic"> Poecilia latipinna</Emphasis>: the importance of predation pressure and reproductive status

We examined the effect of predation risk on female association patterns in the live-bearing sailfin molly (Poecilia latipinna). We tested two classes of females, with and without the risk of predation by a green sunfish (Lepomis cyanellus): (1) postpartum females (maintained with males until visibly gravid, then isolated and tested within 24–48 h of dropping a brood); and (2) non-postpartum females (different females, isolated from males for >50 days). When there was no apparent risk of predation, postpartum females showed a significant preference for large over small males, whereas non-postpartum females showed no size preference at all. When there was an apparent risk of predation, postpartum females maintained their preference for larger males and non-postpartum females continued to show no preference for large or small males. These results suggest that reproductive status (receptivity) plays a greater role in mate preference than predation risk. For postpartum females, the cost of not choosing a preferred mate may outweigh the potential cost of predation. Non-postpartum females either do not benefit from mating or are being indecisive about mating and thus are less likely to be choosy whether or not a predator is present.Communicated by I. Schlupp     

Olanzapine effects on body composition, food preference, glucose metabolism and insulin sensitivity in the rat

The atypical antipsychotic drug olanzapine induces weight gain and defects in glucose metabolism in patients. Using a rat model we investigated the effects of acute and long term olanzapine treatment on weight gain, food preference and glucose metabolism. Olanzapine treated rats fed a chow diet grew more slowly than vehicle controls but olanzapine treated animals fed a high fat/sugar diet grew faster than control animals on the same diet. These changes in weight were paralleled by changes in fat mass. Olanzapine also induced a strong preference for a high fat/high sugar diet. Acute exposure to olanzapine rapidly induced severe impairments of glucose tolerance and increased insulin secretion but did not impair insulin tolerance. These results indicate the defect in glucose metabolism induced by acute olanzapine treatment was most likely due to increased hepatic glucose output associated with a reduction in active GLP-1 levels and correspondingly high glucagon levels.     

The design, synthesis and biological evaluation of novel URB602 analogues as potential monoacylglycerol lipase inhibitors

We have synthesised an extensive series of URB602 analogues as inhibitors of monoacylglycerol lipase (MAGL), which is the major enzyme responsible for metabolising the endocannabinoid 2-arachidonylglycerol. The recently identified crystal structure of MAGL was used in the design strategy and revealed three possible binding sites for URB602 and the proposed analogues. A test series of carbamate analogues were docked into the identified sites to predict the most favourable binding location. The synthesised analogues of URB602 explored the biological effects of isosteric replacement, ring size and substitution, para substitution of the biphenyl moiety and the incorporation of a bicyclic element. The compounds were tested for their ability to inhibit human MAGL. The carbamate analogue 16 displayed the most significant inhibitory activity, reducing MAGL activity to 26% of controls at 100 μM compared to 73% for the parent compound URB602.     

Design,synthesis, molecular modelling and in vitro cytotoxicity analysis of novel carbamate derivatives as inhibitors of Monoacylglycerol lipase

Monoacylglycerol lipase (MAGL) has an essential role in the catabolic pathway of the endocannabinoid 2-arachidonoylglycerol, which makes it a potential target for highly specific inhibitors for the treatment of a number of diseases. We designed and synthesized a series of carbamate analogues of URB602. We evaluated their inhibitory activity toward human MAGL in vitro both in cell culture and lysates. The target compounds exhibited moderate to excellent inhibitory activity against MAGL. The most promising compound 2b showed good inhibitory activity with IC₅₀ value of 4.5?±?0.70?μM reducing MAGL activity to 82% of controls at 10?μM compared to 66% for the parent compound URB602. Interestingly, compounds 2b and 2c induce cell death through the inhibition of MAGL. Molecular modelling approaches and docking studies, used to investigate inhibitory profiles, indicated that trifluoromethyl substitutions of the aryl group and the benzene ring present at the oxygen side of the carbamate molecule had a significant impact on the activity.     

Maternal high-fat diet induces sex-specific endocannabinoid system changes in newborn rats and programs adiposity,energy expenditure and food preference in adulthood

Early life inadequate nutrition triggers developmental adaptations and adult chronic disease. Maternal high-fat (HF) diet promotes visceral obesity and hypothalamic leptin resistance in male rat offspring at weaning and adulthood. Obesity is related to over active endocannabinoid system (ECS). The ECS consists mainly of endogenous ligands, cannabinoid receptors (CB1 and CB2), and the enzymes fatty acid anandamide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). We hypothesized that perinatal maternal HF diet would regulate offspring ECS in hypothalamus and brown adipose tissue (BAT) at birth, prior to visceral obesity development, and program food preference and energy expenditure of adult offspring. Female rats received control diet (C, 9% fat) or isocaloric high-fat diet (HF, 28% fat) for 8 weeks before mating, and throughout gestation and lactation. We evaluated C and HF offspring at birth and adulthood. At birth, maternal HF diet decreased leptinemia and increased hypothalamic CB1, orexin-A, and proopiomelanocortin while it decreased thyrotropin-releasing hormone (Trh) in male pups. Differentially, maternal HF diet increased hypothalamic CB2 in female pups. In BAT, maternal HF diet decreased CB1 and increased CB2 in male and female pups, respectively. Besides presenting different molecular ECS profile at birth, HF adult offspring developed overweight, higher adiposity and high-fat diet preference, independently of the sex, but only males presented hyperleptinemia and higher energy expenditure. In conclusion, maternal HF diet alters ECS components and energy metabolism targets in hypothalamus and BAT of offspring at birth, in a sex-specific manner, which may contribute for hyperphagia, food preference and higher adiposity later in life.