Perinatal undernourishment provokes long-lasting alterations of clusterin and fumarate hydratase expression in the rat nucleus accumbens.

BACKGROUND: Background: Perinatal malnutrition seems to provoke important neurochemical alterations in the brain that lead to higher vulnerability to develop neuropsychiatric disorders in the adulthood. OBJECTIVES: We have examined the persistence and reversibility of the changes induced by perinatal undernourishment on the expression of fumarate hydratase in the rat nucleus accumbens, bearing in mind that this expression has been previously linked with addictive disorders. Clusterin, a multifunctional protein known to be neuroprotective and possibly related to addiction in humans, was studied in parallel. METHODS: Female Wistar rats underwent a severe restriction of food during gestation and lactation. Upon weaning, a subgroup of undernourished animals was switched to normal chow and another one continued under food restriction. Control rats and their mothers were fed on chow along the experiment. Fumarate hydratase and clusterin were quantified by western blot after five months of postnatal life in the three experimental groups. RESULTS: Food restriction along the whole experimental period provoked a marked upregulation of both clusterin and fumarate hydratase in the mitochondrial fraction of the nucleus accumbens. In the case of clusterin, this upregulation was also observed in the cytosolic fraction of the nucleus accumbens. When undernourishment was limited to gestation and lactation the two proteins appeared downregulated with respect to controls. CONCLUSION: The results are consistent with the idea that perinatal malnutrition provokes marked changes in brain neurochemistry that are not fully corrected by the rehabilitation of normal feeding and could be linked to behavioural disturbances in the adulthood, that is, increased vulnerability to addiction.

Plasma midkine levels in patients with cocaine use disorder during abstinence. / Plasma midkine levels in patients with cocaine use disorder during abstinence.

Preclinical evidence suggests that endogenous midkine could play a key modulatory role on the neurotoxic and addictive effects of different kinds of drugs of abuse, including psychostimulants. However, this hypothesis has not yet been explored in humans. As a first approach to progress in this knowledge, we have comparatively studied plasma midkine levels in 75 patients with cocaine use disorder under abstinence and 26 control subjects matched for sex, age and body mass index. Patients were further segmented into early-abstinent (up to one month of abstinence, n = 30) and late-abstinent (more than one month of abstinence, n = 45). Midkine levels were quantified in plasma samples of all the participants by enzyme-linked immunosorbent assays. Early-abstinent patients exhibited a 60% increase of midkine plasma concentration in comparison with the controls. This elevation tended to normalize upon the progression of abstinence. The results obtained demonstrate that peripheral midkine levels are closely related to cocaine use and are consistent with the idea that this cytokine could play a protective role by limiting the biological activity of psychostimulants.

Diversos estudios preclínicos han sugerido que la midkina endógena podría jugar un papel modulador clave sobre los efectos neurotóxicos y adictivos de distintas drogas, incluidos los psicoestimulantes. Esta hipótesis no ha sido aún explorada en humanos. Como primer paso en esta dirección, en el presente trabajo hemos medido los niveles plasmáticos de midkina en 75 pacientes con trastorno por uso de cocaína en abstinencia y 26 controles apareados con los anteriores por sexo, edad e índice de masa corporal. Los pacientes fueron además divididos en un grupo de abstinencia temprana (menos de un mes, n = 30) y otro de abstinencia tardía (más de un mes, n = 45). Se cuantificaron los niveles plasmáticos de midkina de todos los participantes mediante un ensayo por inmunoabsorción ligado a enzimas. Los pacientes en abstinencia temprana mostraron un incremento del 60% en su concentración plasmática de midkina con respecto a los controles que tendió a desaparecer en los pacientes con periodos de abstinencia más prolongados. Los resultados demuestran que los niveles periféricos de midkina están estrechamente relacionados con el uso de cocaína y apoyan la idea de que dicha citoquina podría jugar un papel protector limitando la actividad biológica de los psicoestimulantes.

Niveles plasmáticos de midkina en pacientes con trastorno por uso de cocaína en abstinencia / Plasma midkine levels in patients with cocaine use disorder during abstinence

Diversos estudios preclínicos han sugerido que la midkina endógenapodría jugar un papel modulador clave sobre los efectos neurotóxicosy adictivos de distintas drogas, incluidos los psicoestimulantes. Estahipótesis no ha sido aún explorada en humanos. Como primer pasoen esta dirección, en el presente trabajo hemos medido los nivelesplasmáticos de midkina en 75 pacientes con trastorno por uso decocaína en abstinencia y 26 controles apareados con los anteriorespor sexo, edad e índice de masa corporal. Los pacientes fueronademás divididos en un grupo de abstinencia temprana (menos deun mes, n = 30) y otro de abstinencia tardía (más de un mes, n =45). Se cuantificaron los niveles plasmáticos de midkina de todoslos participantes mediante un ensayo por inmunoabsorción ligadoa enzimas. Los pacientes en abstinencia temprana mostraron unincremento del 60% en su concentración plasmática de midkina conrespecto a los controles que tendió a desaparecer en los pacientes conperiodos de abstinencia más prolongados. Los resultados demuestranque los niveles periféricos de midkina están estrechamenterelacionados con el uso de cocaína y apoyan la idea de que dichacitoquina podría jugar un papel protector limitando la actividadbiológica de los psicoestimulantes. (AU)

Preclinical evidence suggests that endogenous midkine couldplay a key modulatory role on the neurotoxic and addictive effectsof different kinds of drugs of abuse, including psychostimulants.However, this hypothesis has not yet been explored in humans. As afirst approach to progress in this knowledge, we have comparativelystudied plasma midkine levels in 75 patients with cocaine use disorderunder abstinence and 26 control subjects matched for sex, ageand body mass index. Patients were further segmented into earlyabstinent (up to one month of abstinence, n = 30) and late-abstinent(more than one month of abstinence, n = 45). Midkine levels werequantified in plasma samples of all the participants by enzyme-linkedimmunosorbent assays. Early-abstinent patients exhibited a 60%increase of midkine plasma concentration in comparison with thecontrols. This elevation tended to normalize upon the progressionof abstinence. The results obtained demonstrate that peripheralmidkine levels are closely related to cocaine use and are consistentwith the idea that this cytokine could play a protective role by limitingthe biological activity of psychostimulants. (AU)

Clusterin levels in undernourished SH-SY5Y cells.

Food-related disorders are increasingly common in developed societies, and the psychological component of these disorders has been gaining increasing attention. Both overnourishment with high-fat diets and perinatal undernourishment in mice have been linked to a higher motivation toward food, resulting in an alteration in food intake. Clusterin (CLU), a multifaced protein, is overexpressed in the nucleus accumbens (NAc) of over-fed rats, as well as in those that suffered chronic undernutrition. Moreover, an increase of this protein was observed in the plasma of obese patients with food addiction, suggesting the implication of CLU in this eating disorder. To characterize CLU's cellular mechanisms, in vitro experiments of undernutrition were performed using dopaminergic SH-SY5Y cells. To mimic in vivo dietary conditions, cells were treated with different fetal bovine serum (FBS) concentrations, resulting in control (C group) diet (10% FBS), undernourishment (U group) diet (0.5% FBS), and undernourishment diet followed by restoration of control diet (UC group) (0.5 + 10% FBS). Undernourishment compromised cell viability and proliferation, and concomitantly increased CLU secretion as well as the cytosolic pool of the protein, while decreasing the mitochondrial level. The restoration of normal conditions tended to recover cell physiology, and the normal levels and distribution of CLU. This research study is a step forward toward the characterization of clusterin as a potential marker for food addiction and nutritional status.

Clusterin overexpression as a potential neuroprotective response to the pathological effects of high fat dieting on the brain reward system.

High-fat diets (HFDs) can lead to pathological changes in the brain underlying several behavioral disturbances (e.g., reward deficiency). To further increase our knowledge of these associations, we studied the sucrose reward and the brain expression of clusterin, a protein that is overexpressed after several kind of brain damaging conditions. C57BL/6J male mice were differentially fed on an HFD or standard chow for 41 days and underwent 11 sucrose place conditioning sessions followed by 4 extinction sessions to monitor the effects of HFD on sucrose reward by means of free choice tests. We quantified clusterin expression by immunochemistry in the nucleus accumbens, dorsal striatum and cingulate cortex. HFD tended to provoke a transient potentiation in the acquisition of sucrose-conditioned place preference, but this effect was followed by a much more consistent reduction in sucrose preference, which spontaneously disappeared after 31 days of an HFD with no need for extinction learning. The HFD mice showed higher clusterin expression in the nucleus accumbens but not in the other brain areas studied. The results confirm that HFDs strongly influence the rewarding properties of palatable foods and suggest a direct connection with neurotoxic alterations in the brain reward system tagged by clusterin overexpression.

Undernutrition induces major alterations in the lipid droplets of white and brown adipose tissues in wistar rats.

Several studies have shown a relationship between the distribution of fat mass around the organism, metabolic disorders, and an increased risk of morbidity and mortality. It has been demonstrated that in obese animals there is a big rise in the white fat deposits due to hyperplasia and hypertrophy of the adipocytes. Studies related to weight and health have been more popular regarding obesity rather than extreme caquexia or calorico-proteic deficiencies, but these states are interesting from the point of view of the preferential atrophy of certain organs that may help us in the understanding of undernourishment. Moreover, the discovery of beige adipose tissue has instigated thoughts around the roles played by the different cells in the adipose tissue as well as its adaptability in pathological states. In our study we carried out morphometric, morphological, and quantitative measurements of the adipose tissue in an animal model based on a 40-50% diet restriction in comparison to control animals. We have found a decrease in the size of white adipocytes together with a variation in the lipid droplet size of brown adipocytes in undernourished animals, what may be considered as possible transformations between the types of adipose tissues, and that could be caused by an adaptive phenomenon to the undernourished state.

Clusterin: Always protecting. Synthesis, function and potential issues.

In the last years, clusterin, a challenging and paradoxical apolipoprotein, has been of growing interest amongst a rising number of scientists. This enigmatic protein is present in all fluids of the organism besides within the intracellular matrix, and it plays diverse, and at times contrary, roles in a growing number of pathologies. It seems to vary its location and function to assure cellular survival being cytoprotective hence its significance in neuroprotection and cancer along with chemotherapy resistance. However, it can also lead to cellular arrest and its modulation to apoptosis. Additionally, it has been described to modulate pain, as well as linked to inflammation, cardioprotection, satiety and hunger, and possibly to addictive behaviour development. Thus, it has been postulated to be used both as a biomarker and a very explorable new therapeutic target for several conditions.

Histamine H3 receptor gene variants associated with drug abuse in patients with cocaine use disorder.

BACKGROUND: Preclinical work revealed significant interactions between ligands of the histamine H3 receptor and different drugs of abuse. In the case of psychostimulants, the results reported are somewhat controversial and human data are still scarce, despite the fact that an inverse agonist of the H3 receptor (pitolisant) has reached the market after approval for the treatment of narcolepsy. AIMS: We have studied associations between histamine H3 receptor gene variants and cocaine use disorder to increase the knowledge of the possible involvement of histamine H3 receptor in drug abuse. METHODS: Seven single nucleotide polymorphisms of the histamine H3 receptor gene were genotyped by using a multiplexing assay in 248 samples of subjects with cocaine use disorder and 500 randomized samples of subjects representative of the Spanish population. RESULTS: The study of the epidemiological information associated to the samples revealed that subjects with cocaine use disorder broadly abused alcohol, tobacco and cannabinoids. Two single nucleotide polymorphisms (rs3787430 and rs74627870) were found significantly associated with the occurrence of addiction and one more (rs13042865) was specifically related to the severity of cocaine dependence within drug abusers. CONCLUSIONS: The associations found in this study further extend the hypothesis that histamine H3 receptor function could be relevant in drug abuse in general and cocaine addiction in particular.

Sciatic Nerve Ligation Downregulates Mitochondrial Clusterin in the Rat Prefrontal Cortex.

The concentration of the multifunctional protein clusterin is reduced in the plasma of subjects with degenerative scoliosis (DS) and carpal tunnel syndrome (CTS) but elevated in the cerebrospinal fluid of neuropathic pain patients successfully treated with spinal cord stimulation. The present work tries to increase the knowledge of pain-associated changes of plasma and brain clusterin by using an animal model of neuropathy. We studied the effects of sciatic nerve ligation on mechanical allodynia (von Frey test), anxiety (elevated plus maze test), plasma clusterin (enzyme-linked immunosorbent assay) and clusterin expression in the nucleus accumbens (NAC) and prefrontal cortex (PFC) of adult male Wistar rats (western blot). The possible modulatory role of high fat (HF) dieting was also studied, bearing in mind that obesity has been also reported to influence nociception, clusterin levels and prefrontal cortex activation. Animals with nerve ligation showed mechanical allodynia, anxiety and a marked downregulation of clusterin in the mitochondrial fraction of the prefrontal cortex. Animals fed on HF also exhibited a slight increase of the sensitivity to mechanical stimuli and anxiety; however, the diet did not potentiate the effects of nerve ligation. The results did not confirm a parallelism between neuropathy, obesity and alterations of plasma levels of clusterin, but strongly suggest that the protein could be involved in the functional reorganization of the prefrontal cortex which has been recently reported in chronic pain conditions.

The Neuroendocrine and Metabolic Outcomes of Bariatric Surgery Depend on Presurgical Control over Eating.

BACKGROUND: The outcomes of bariatric surgery are very irregular and mostly unpredictable. The search for variables of predictive value is encouraged to help preventing therapeutic failures. OBJECTIVE: We aimed to confirm the hypothesis that preexisting eating behaviors could predict neuroendocrine and metabolic outcomes of gastric bypass surgery in morbidly obese subjects. METHODS: Twenty-one morbidly obese patients from the Bariatric Surgery Program of our hospital were selected according to the specific inclusion and exclusion criteria for this study. The subjects filled out a validated questionnaire to quantify the "loss-of-control" (LC) dimension of food craving and provided serum samples at the onset of the study and 1 year after gastric bypass surgery. Hematological, metabolic, and hormonal variables were studied by conventional clinical tests and enzyme immunoassays and checked for correlations with LC both before and after surgery. RESULTS: Those patients that had exhibited worse eating control at the beginning of the study experienced a better metabolic response 1 year after surgery in terms of reduction of serum insulin, HOMA1-IR, HOMA2-IR, and vitamin D1; all these variables were inversely correlated with presurgical LC. Serum brain-derived neurotrophic factor (BDNF) levels showed the same tendency; in fact, BDNF significantly decreased only in those patients with worse eating control. CONCLUSIONS: Problematic eating behaviors may predict a better response of insulin resistance and a specific reduction of serum BDNF in morbidly obese patients after gastric bypass surgery.

Antiproliferative and cytotoxic effects of grape pomace and grape seed extracts on colorectal cancer cell lines.

Grape pomace is the source of bioactive compounds (anthocyanins, flavonols, flavan-3-ols, and stilbenes) which exhibit antiproliferative actions on cell cultures. We have investigated the antitumoral effects of grape pomace and grape seed extracts on colon cancer cells (Caco-2, HT-29) and fibroblasts. Crude extracts prepared from white and red pomace, and grape seeds, reduced the viability and proliferation of Caco-2. HT-29 cells were resistant to these actions. Purified extracts were then prepared from the same sources and compared with the LDH test; again, all three extracts were active and purified extract from grape seed was the most potent and specific on Caco-2 cells. HT-29 cells were more sensitive to these purified extracts. The biological activity resided almost exclusively in the flavonol and flavan-3-ols subfractions, rather than the anthocyanin subfraction. Preliminary results on the mechanisms involved in these effects revealed downregulation of Myc gene expression in HT-29 and upregulation of Ptg2 in Caco-2 cells.

Inflammatory stress and altered angiogenesis evoked by very high-fat diets in mouse liver / Las dietas ricas en grasa estimulan procesos inflamatorios y angiogénicos en hígado de ratón

Background: Non-alcoholic fatty liver disease (NAFLD), a condition that leads to fibrosis, is caused by intake of very high-fat diets (HFDs). However, while the negative impact on the liver of these diets has been an issue of interest, systematic research on the effect of HFDs are lacking. Objective: To characterize the overall impact of HFDs on both molecular and morphological signs of liver remodeling. Methods: A study was conducted on male C57BL/6J mice to assess the effect of 4- and 8-week HFDs (60% kcal from fat) on (I) liver steatosis and fibrosis, and (II) expression of factors involved in inflammation and angiogenesis. Results: After an 8-week HFD, vascular endothelial growth factor type-2 receptor (VEGF-R2) and fatty acid translocase/trombospondin-1 receptor (CD36) were overexpressed in liver tissue of mice given HFDs. These changes suggest impaired liver angiogenesis and occurred together with (I) increased GPR78-BiP and EIF2alpha phosphorylation, suggesting endoplasmic reticulum stress, (II) induction of Col1a1 gene expression, a marker of fibrosis, and (III) increased CD31 immunolabeling, consistent with active angiogenesis and fibrosis. Conclusion: Our data show that very HFDs promote a rapid inflammatory response, as well as deregulation of angiogenesis, both consistent with development of liver fibrosis

Antecedentes: El hígado graso no alcohólico (HGNA) es una enfermedad hepática que ocasiona fibrosis y se genera por la ingesta de dietas ricas en grasa. Aunque los efectos nocivos de este tipo de dietas son de gran interés, no son muy abundantes los estudios sistemáticos sobre las consecuencias que su consumo puede tener en el hígado. Objetivo: Evaluar los efectos de una dieta rica en grasa sobre el remodelado hepático, tanto a nivel morfológico como molecular. Métodos: Se utilizaron ratones macho C57BL/6J tratados durante 4/8 semanas con una dieta que contenía un 60% de las kilocalorías procedentes de grasa sobre: 1) la aparición de esteatosis y/o fibrosis hepática y 2) la expresión de factores implicados en procesos de inflamación y angiogénesis. Resultados: Tras 8 semanas de dieta se observó un incremento en el receptor del factor de crecimiento vascular endotelial tipo 2 (R2-FCVE) y en la translocasa de ácidos grasos (CD36). Estos cambios, que sugieren que los procesos angiogénicos del hígado están alterados, fueron simultáneos a: 1) un aumento del GPR78-BiP y de la fosforilación de EIF2alpha, marcadores de estrés del retículo endoplásmico, 2) la inducción en la expresión del gen de colágeno Col1a1, indicador de fibrosis y 3) un incremento de células inmunofluorescentes para CD31 compatible con procesos de angiogénesis y de fibrosis. Conclusiones: Nuestros resultados muestran que las dietas con alto contenido en grasa inducen la rápida activación de respuestas inflamatorias, así como alteraciones en la angiogénesis, siendo ambos procesos compatibles con el desarrollo de fibrosis hepática

Inflammatory stress and altered angiogenesis evoked by very high-fat diets in mouse liver.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD), a condition that leads to fibrosis, is caused by intake of very high-fat diets (HFDs). However, while the negative impact on the liver of these diets has been an issue of interest, systematic research on the effect of HFDs are lacking. OBJECTIVE: To characterize the overall impact of HFDs on both molecular and morphological signs of liver remodeling. METHODS: A study was conducted on male C57BL/6J mice to assess the effect of 4- and 8-week HFDs (60% kcal from fat) on (i) liver steatosis and fibrosis, and (ii) expression of factors involved in inflammation and angiogenesis. RESULTS: After an 8-week HFD, vascular endothelial growth factor type-2 receptor (VEGF-R2) and fatty acid translocase/trombospondin-1 receptor (CD36) were overexpressed in liver tissue of mice given HFDs. These changes suggest impaired liver angiogenesis and occurred together with (i) increased GPR78-BiP and EIF2α phosphorylation, suggesting endoplasmic reticulum stress, (ii) induction of Col1a1 gene expression, a marker of fibrosis, and (iii) increased CD31 immunolabeling, consistent with active angiogenesis and fibrosis. CONCLUSION: Our data show that very HFDs promote a rapid inflammatory response, as well as deregulation of angiogenesis, both consistent with development of liver fibrosis.

Proteomic Identification of Biomarkers Associated with Eating Control and Bariatric Surgery Outcomes in Patients with Morbid Obesity.

BACKGROUND: The current therapeutics of morbid obesity could be significantly improved after the identification of novel biomarkers associated with the food addiction endophenotype of obesity and with bariatric surgery outcomes. METHODS: We applied differential expression proteomics and enzyme-linked immunosorbent confirmatory assays to identify (a) proteins that varied according to loss of control over eating in morbidly obese patients and (b) proteins that varied between normoweight controls and patients before and 1 year after bariatric surgery. RESULTS: Clusterin was the only protein that consistently varied according to eating control in patients. Patients showed increased levels of serum amyloid P protein, apolipoprotein A4, serotransferrin, complement factors B and C3 and haptoglobin with respect to controls; the levels of all these proteins tended to return to control values 1 year after surgery. In contrast, apolipoprotein A1 and transthyretin were initially downregulated in patients and were scarcely changed by surgery. Leucine-rich alpha-2-glycoprotein was markedly increased in patients only after surgery. CONCLUSIONS: Clusterin could be of interest as a putative biomarker for food addiction diagnosis in people with morbid obesity. In addition, postsurgical normalization of the proteins initially dysregulated in obese subjects might help monitor clinical improvements after surgery, while lasting or newly detected alterations (i.e., those affecting transthyretin and leucine-rich alpha-2-glycoprotein) could reflect partial refractoriness and/or contribute to the early prediction of clinical problems.

Cocaine and amphetamine regulated transcript and brain-derived neurotrophic factor in morbid obesity. One-year follow-up after gastric bypass.

BACKGROUND: The identification of biomarkers associated with obesity and response to treatment could represent an important advance to design more effective and personalized therapeutic strategies. The complexity of morbid obesity could be explained as the combination of genetic, biochemical, cultural, and behavioral factors, among others. The study of biomarkers should be considered a determinant factor taken into account in this equation. OBJECTIVES: The aim of this study was to define better biomarker profiles potentially associated to the short-term outcome of bariatric surgery by paying attention to cocaine and amphetamine regulated transcript and brain-derived neurotrophic factor, 2 neuropeptides related to eating behavior. SETTING: University General Hospital of Ciudad Real, Spain. METHODS: Twenty-seven morbidly obese patients and 30 healthy weight individuals matched by age and sex were selected for the study. RESULTS: Patients underwent bariatric surgery by Roux-en-Y gastric bypass and responded adequately in terms of weight loss and normalization of many biochemical parameters 1 year postsurgery. A multivariate analysis showed that the hormonal/neuropeptidic profile explained 82% of the variability of the weight loss response. The evolution of cocaine and amphetamine regulated transcript paralleled that of insulin and leptin, serum levels of this peptide were initially elevated in patients (4.24 ± .47 ng/mL) with respect to controls (2.94 ± .2 ng/mL), but this difference disappeared 1 year after Roux-en-Y gastric bypass (3.14 ± .26 ng/mL). Brain-derived neurotrophic factor levels were also decreased by Roux-en-Y gastric bypass (11.93 ± .96 ng/mL postsurgery versus 15.3 ± 1.02 ng/mL presurgery), even when this peptide was not elevated in patients before surgery (14.23 ± .86 ng/mL in controls). CONCLUSIONS: The results suggest that cocaine and amphetamine regulated transcript and brain-derived neurotrophic factor could be envisaged as new candidate biomarkers of short-term outcome after surgery.

Lewis and Fischer 344 rats as a model for genetic differences in spatial learning and memory: Cocaine effects.

Lewis (LEW) and Fischer 344 (F344) rats are considered a model of genetic vulnerability to drug addiction. We previously showed important differences in spatial learning and memory between them, but in contrast with previous experiments demonstrating cocaine-induced enhanced learning in Morris water maze (MWM) highly demanding tasks, the eight-arm radial maze (RAM) performance was not modified either in LEW or F344 rats after chronic cocaine treatment. In the present work, chronically cocaine-treated LEW and F344 adult rats have been evaluated in learning and memory performance using the Y-maze, two RAM protocols that differ in difficulty, and a reversal protocol that tests cognitive flexibility. After one of the RAM protocols, we quantified dendritic spine density in hippocampal CA1 neurons and compared it to animals treated with cocaine but not submitted to RAM. LEW cocaine treated rats showed a better performance in the Y maze than their saline counterparts, an effect that was not evident in the F344 strain. F344 rats significantly took more time to learn the RAM task and made a greater number of errors than LEW animals in both protocols tested, whereas cocaine treatment induced deleterious effects in learning and memory in the highly difficult protocol. Moreover, hippocampal spine density was cocaine-modulated in LEW animals whereas no effects were found in F344 rats. We propose that differences in addictive-like behavior between LEW and F344 rats could be related to differences in hippocampal learning and memory processes that could be on the basis of individual vulnerability to cocaine addiction.

A high-fat diet combined with food deprivation increases food seeking and the expression of candidate biomarkers of addiction.

A mouse model has been developed to study the effect of dietary fat combined with food deprivation periods on palatable food seeking and on the expression of three potential addiction biomarkers in the nucleus accumbens: fumarate hydratase (FH), ATP synthase subunit alpha (ATP5a1) and transketolase (TKT). Forty C57BL/6 J male mice, four-week old, were fed either with a high-fat (HF) diet or standard diet along the experiment. After 3 weeks of differential feeding, animals underwent a two-week training period of two daily sessions where visual cues were paired either to palatable food (chocolate cereals) or no food at all. This training was prolonged one more week with similar, one daily sessions preceded by 12 hours of food deprivation. A behavioural test was finally conducted where mice were confined for 30 minutes either in food unpaired compartments or in compartments previously paired with cereals, but now with empty food trays. Total activity during this behavioural test and serum corticosterone levels right after it were similar in all experimental groups. Mice tested in food-paired compartments showed a marked preference for the empty food tray that gradually disappeared in standard diet-fed individuals but persisted in HF-fed mice. HF-fed mice also overexpressed FH, ATP5a1 and TKT, which positively correlated with the persistence of preference for the empty food tray. It is suggested that HF diets combined with food deprivation may enhance food seeking behaviours while upregulating FH/ATP5a1/TKT, which are further envisaged as biomarkers of addiction.

Target identification and validation in brain reward dysfunction.

Addictive disorders (substance-use disorder and gambling disorder) are collected together in the fifth edition of The Diagnostic and Statistical Manual of Mental Disorders (DSM-5) partially because of a common brain origin, which seems to involve dysfunction of the reward system. Beyond these disorders, other neuropsychiatric diseases also share abnormal reward sensitivity, maladaptive impulsivity or compulsive behaviours, and have been reunited under the 'reward deficiency syndrome' (RDS) umbrella. Research in this field could then provide novel drugs with positive actions in all these diseases, but many animal models used for this purpose lack enough translational value to enable the identification of novel targets and should be then avoided. As we discuss here, only selected protocols could provide reliable targets that would be common to the whole family of diseases, thus qualifying for further validation in patients.

Pro-apoptotic properties of morphine in neuroblastoma × glioma NG108-15 hybrid cells: modulation by yohimbine.

Short-term incubation with pharmacologically relevant concentrations of morphine has been shown to transiently affect the metabolism and redox status of NG108-15 cells through δ-opioid receptor stimulation, but apparently did not provoke cell death. The present work tries to determine if incubation with morphine at longer time intervals (24 h) provokes apoptosis and/or necrosis, as it has been described in other cell lines. We have also checked the potential modulatory role of yohimbine on these effects, on the basis of the previously described interactions between this drug and opioid receptor ligands. Incubation with morphine 0.1 and 10 µM provoked the appearance of images compatible with apoptosis (bebbling, pyknotic cells with cytoplasmic and nuclear condensation) and necrosis (cells swollen with vacuolated cytoplasm lacking cell processes) that could be observed directly and/or after staining with methylene blue, crystal violet and propidium iodide/4',6-diamidino-2-phenylindole (IP/DAPI). Quantification of apoptosis by activation of caspases 3 and 7 and DNA fragmentation with the Tunel assay revealed a modest but significant increase after incubation with the two concentrations of morphine used. Co-incubation with 10 µM yohimbine prevented all these effects of the opioid. The results extend previous findings of a yohimbine-sensitive, neurotoxic effect of morphine on NG108-15 cells.

Remodeling of energy metabolism and absence of electrophysiological changes in the heart of obese hyperleptinemic mice. New insights into the pleiotropic role of leptin.

Dietary treatment with high-fat diets (HFD) triggers diabetes and hyperleptinemia, concomitantly with a partial state of leptin resistance that affects hepatic and adipose tissue but not the heart. In this context, characterized by widespread steatosis, cardiac lipid content remains unchanged. As previously reported, HFD-evoked hyperleptinemia could be a pivotal element contributing to increase fatty-acid (FA) metabolism in the heart and to prevent cardiac steatosis. This metabolic adaptation might theoretically reduce energy efficiency in cardiomyocytes and lead to cardiac electrophysiological remodeling. Therefore the aim of the current study has been to investigate the impact of long-term HFD on cardiac metabolism and electrophysiological properties of the principal ionic currents responsible of the action potential duration in mouse cardiomyocytes. Male C57BL/6J mice were fed a control (10 kcal% from fat) or HFD (45 kcal% from fat) during 32 weeks. Quantification of enzymatic activities regulating mitochondrial uptake of pyruvate and FA showed an increase of both carnitine-palmitoyltransferase and citrate synthase activities together with a decrease of lactate dehydrogenase and pyruvate dehydrogenase activities. Increased expression of uncoupling protein-3, Mn-, and Cu/Zn-superoxide dismutases and catalase were also detected. Total glutathione/oxidized glutathione ratios were unaffected by HFD. These data suggest that HFD triggers adaptive mechanisms aimed at (i) facilitating FA catabolism, and (ii) preventing oxidative stress. All these changes did not affect the duration of action potentials in cardiomyocytes and only slightly modified electrocardiographic parameters.