Tetrahydrobiopterin modulates the behavioral neuroinflammatory response to an LPS challenge in mice.

Tetrahydrobiopterin (BH4) is a necessary cofactor for the synthesis of monoamines from essential amino-acids, phenylalanine, tyrosine and tryptophan. The BH4 synthesis pathway is induced by inflammatory factors but highly regulated processes maintain levels in a physiological range. However, BH4 activity can be durably altered in inflammation-related pathologies, such as certain types of depression, potentially involving impairment of dopaminergic neurotransmission. The purpose of this study was to investigate the response of the brain BH4 pathway to the inflammatory stimulus induced by lipopolysaccharide (LPS) in mice. Brain expression of genes related to BH4 synthesis, levels of BH4, changes in L-aromatic amino acid precursors of monoamines and dopamine levels were determined. As secondary aim, the effect of acute BH4 supply under the inflammatory challenge was tested on these parameters and on the expression of inflammatory cytokines. Mice were also submitted to the sucrose preference test and to the open-field in order to asses hedonic and locomotor responses to LPS, in addition to their modulation by BH4 supply. The LPS challenge resulted in decreased striatal DA levels and increased Phenylalanine/Tyrosine ratio, suggesting reduced BH4 activity. BH4 supply was effective to increase striatal BH4 levels, to restore the LPS-induced decreased in DA levels in striatum and to dampen the LPS-induced expression of inflammatory cytokines. At the behavioral level, BH4 supply was able to restore the loss of locomotor response to amphetamine in the LPS treated mice, suggesting a modulation of the dopaminergic neurotransmission. These data suggest that BH4 can be considered as a potential add-on molecule, helping to maintain or restore dopaminergic neurotransmission in neuroinflammatory conditions..

Impact of perinatal exposure to high-fat diet and stress on responses to nutritional challenges, food-motivated behaviour and mesolimbic dopamine function.

BACKGROUND/OBJECTIVES: Energy-dense food exposure and stress during development have been suggested to contribute to obesity and metabolic disorders later in life. Although these factors are frequently associated, the effects of their combination have not yet been investigated. In this study, using an animal model, we examined the long-term impact of maternal high-fat diet (HFD) and early-life stress (ELS) on energy homoeostasis control and food motivation. METHODS: Body weight growth under HFD, adipose tissue, body weight control in response to fasting and refeeding, food-motivated behaviour and mesolimbic dopamine function were examined in adult male offspring exposed to maternal HFD (during gestation and lactation) and/or ELS (maternal separation 3 h per day from postnatal day 2 to 14). RESULTS: Maternal HFD or ELS alone had no significant effect on offspring body weight; however, the combination of these factors exacerbated body weight gain when animals were exposed to HFD after weaning. There are no other significant combinatory effects of these perinatal events. In contrast, independently of the maternal diet, ELS disrupted body weight control during a fasting-refeeding procedure, increased adipose tissue mass and altered lipid metabolism. Finally, maternal HFD and ELS both resulted in exacerbated food-motivated behaviour and blunted dopamine release in the nucleus accumbens during palatable food consumption. CONCLUSIONS: We report a synergistic effect of perinatal HFD exposure and stress on the susceptibility to gain weight under HFD. However, ELS has a stronger impact than maternal HFD exposure on energy homoeostasis and food motivation in adult offspring. Altogether, our results suggest a programming effect of stress and nutrition supporting the hypothesis of the developmental origin of health and disease.

Tetrahydrobioterin (BH4) Pathway: From Metabolism to Neuropsychiatry.

Tetrahydrobipterin (BH4) is a pivotal enzymatic cofactor required for the synthesis of serotonin, dopamine and nitric oxide. BH4 is essential for numerous physiological processes at periphery and central levels, such as vascularization, inflammation, glucose homeostasis, regulation of oxidative stress and neurotransmission. BH4 de novo synthesis involves the sequential activation of three enzymes, the major controlling point being GTP cyclohydrolase I (GCH1). Complementary salvage and recycling pathways ensure that BH4 levels are tightly kept within a physiological range in the body. Even if the way of transport of BH4 and its ability to enter the brain after peripheral administration is still controversial, data showed increased levels in the brain after BH4 treatment. Available evidence shows that GCH1 expression and BH4 synthesis are stimulated by immunological factors, notably pro-inflammatory cytokines. Once produced, BH4 can act as an anti- inflammatory molecule and scavenger of free radicals protecting against oxidative stress. At the same time, BH4 is prone to autoxidation, leading to the release of superoxide radicals contributing to inflammatory processes, and to the production of BH2, an inactive form of BH4, reducing its bioavailability. Alterations in BH4 levels have been documented in many pathological situations, including Alzheimer's disease, Parkinson's disease and depression, in which increased oxidative stress, inflammation and alterations in monoaminergic function are described. This review aims at providing an update of the knowledge about metabolism and the role of BH4 in brain function, from preclinical to clinical studies, addressing some therapeutic implications.

Tetrahydrobiopterin administration facilitates amphetamine-induced dopamine release and motivation in mice.

Dopamine (DA) is a critical neurotransmitter involved in motivational processes. Tetrahydrobiopterin (BH4) is an essential cofactor for tyrosine hydroxylase, the rate-limiting enzyme in DA synthesis. Decreases in BH4 levels are observed in several DA-related neuropsychiatric diseases involving impairment in motivation. Yet, whether BH4 could be used to treat motivational deficits has not been comprehensively investigated. To investigate the effects of exogenous BH4 administration on the dopaminergic system and related behaviors, we acutely injected mice with BH4 (50 mg/kg). Passage of BH4 through the blood brain barrier and accumulation in brain was measured using the in situ brain perfusion technique. DA release was then recorded using in-vivo micro-dialysis and motivation was evaluated through operant conditioning paradigms in basal condition and after an amphetamine (AMPH) injection. First, we showed that BH4 crosses the blood-brain barrier and that an acute peripheral injection of BH4 is sufficient to increase the concentrations of biopterins in the brain, without affecting BH4- and DA-related protein expression. Second, we report that this increase in BH4 enhanced AMPH-stimulated DA release in the nucleus accumbens. Finally, we found that BH4-induced DA release led to improved performance of a motivational task. Altogether, these findings suggest that BH4, through its action on the dopaminergic tone, could be used as a motivational enhancer.

Synergistic effects of stress and omega-3 fatty acid deprivation on emotional response and brain lipid composition in adult rats.

The aim was to determine the consequences of multi-generational n-3 polyunsaturated fatty acids (PUFA) deficiency on emotional response in rats subjected to maternal separation (MS) as chronic early life stress. Pups fed a control or an n-3 PUFA deficient diet were daily separated for 2 weeks before weaning. In adult rats, reward response was assessed by sucrose consumption and reactivity to novelty using openfield test. Both n-3 PUFA deficiency and MS increased reward response and impulsivity. Moreover, nutritional deficiency and stress acted in synergy to elevate sucrose intake by 80%, compared to control conditions. n-3 PUFA deprivation induced a depletion of docosahexanoeic acid of brain membranes by 70% compensated by increase in 22:5 n-6 and arachidonic acid (AA) levels. The diet-induced AA increase was, however, significantly higher in MS rats. This suggests that n-3 PUFA deficit could be an environmental risk increasing vulnerability to depressive-like response induced by chronic stress.

Hyperactivity in the rat is associated with spontaneous low level of n-3 polyunsaturated fatty acids in the frontal cortex.

Inattention, hyperactivity and impulsiveness are the main symptoms of the heterogeneous attention-deficit/hyperactivity disorder (ADHD). It has been suggested that ADHD is associated with an imbalance in polyunsaturated fatty acid (PUFA) composition, with abnormal low levels of the main n-3 PUFA, DHA (22: 6n-3). DHA is highly accumulated in nervous tissue membranes and is implicated in neural function. Animal studies have shown that diet-induced lack of DHA in the brain leads to alterations in cognitive processes, but the relationship between DHA and hyperactivity is unclear. We examined the membrane phospholipid fatty acid profile in frontal cortex of rats characterized for attention, impulsiveness and motricity in various environmental contexts to determine the relationship between brain PUFA composition and the symptoms of ADHD. The amounts of n-3 PUFA in the PE were significantly correlated with nocturnal locomotor activity and the locomotor response to novelty: hyperactive individuals had less n-3 PUFA than hypoactive ones. We conclude that spontaneous hyperactivity in rats is the symptom of ADHD that best predicts the n-3 PUFA content of the frontal cortex. This differential model in rats should help to better understand the role of PUFA in several psychopathologies in which PUFA composition is modified.

Impact of Early Consumption of High-Fat Diet on the Mesolimbic Dopaminergic System.

Increasing evidence suggest that consumption of high-fat diet (HFD) can impact the maturation of brain circuits, such as during adolescence, which could account for behavioral alterations associated with obesity. In the present study, we used behavioral sensitization to amphetamine to investigate the effect of periadolescent HFD exposure (pHFD) in rats on the functionality of the dopamine (DA) system, a central actor in food reward processing. pHFD does not affect responding to an acute injection, however, a single exposure to amphetamine is sufficient to induce locomotor sensitization in pHFD rats. This is paralleled by rapid neurobiological adaptations within the DA system. In pHFD-exposed animals, a single amphetamine exposure induces an increase in bursting activity of DA cells in the ventral tegmental area (VTA) as well as higher DA release and greater expression of (tyrosine hydroxylase, TH) in the nucleus accumbens (NAc). Post-synaptically, pHFD animals display an increase in NAc D2 receptors and c-Fos expression after amphetamine injection. These findings highlight the vulnerability of DA system to the consumption of HFD during adolescence that may support deficits in reward-related processes observed in obesity.

Plasma fatty acid levels in autistic children.

Phospholipid fatty acids are major structural components of neuronal cell membranes, which modulate membrane fluidity and hence function. Evidence from clinical and biochemical sources have indicated changes in the metabolism of fatty acids in several psychiatric disorders. We examined the phospholipid fatty acids in the plasma of a population of autistic subjects compared to mentally retarded controls. Our results showed a marked reduction in the levels of 22: 6n-3 (23%) in the autistic subjects, resulting in significantly lower levels of total (n-3) polyunsaturated fatty acids (PUFA) (20%), without significant reduction in the (n-6) PUFA series, and consequently a significant increase in the (n-6)/(n-3) ratio (25%). These variations are discussed in terms of potential differences in PUFA dietary intake, metabolism, or incorporation into cellular membranes between the two groups of subjects. These results open up interesting perspectives for the investigation of new biological indices in autism. Moreover, this might have new therapeutic implications in terms of child nutrition.

Analysis of biogenic amines by solid-phase microextraction and high-performance liquid chromatography with electrochemical detection.

We investigated the new solid-phase microextraction method by high-performance liquid chromatography with electrochemical detection for the analysis of biogenic amines. The Carbowax-Templated Resin 50 microm (purple) fibre coating offers good performances for dopamine and serotonin separation, i.e., good selectivity and high sensibility (0.1 microg l(-1)). We also tested this fibre for biogenic amines quantification of rat striatum. The coating seems to be selective towards the amines and has low affinity for the metabolites, allowing a good separation and preventing drawbacks from the biological matrix. These first results obtained using this original separation method offer large perspectives of application to many biological samples.

Polyunsaturated fatty acids and cerebral function: focus on monoaminergic neurotransmission.

More and more reports in recent years have shown that the intake of polyunsaturated fatty acids (PUFA) constitutes an environmental factor able to act on the central nervous system (CNS) function. We recently demonstrated that the effects of PUFA on behavior can be mediated through effects on the monoaminergic neurotransmission processes. Supporting this proposal, we showed that chronic dietary deficiency in alpha-linolenic acid in rats induces abnormalities in several parameters of the mesocortical and mesolimbic dopaminergic systems. In both systems, the pool of dopamine stored in presynaptic vesicles is strongly decreased. This may be due to a decrease in the number of vesicles. In addition, several other factors of dopaminergic neurotransmission are modified according to the system affected. The mesocortical system seems to be hypofunctional overall [e.g., decreased basal release of dopamine (DA) and reduced levels of dopamine D2 (DAD2) receptors]. In contrast, the mesolimbic system seems to be hyperfunctional overall (e.g., increased basal release of DA and increased levels of DAD2 receptors). These neurochemical changes are in agreement with modifications of behavior already described with this deficiency. The precise mechanisms explaining the effects of PUFA on neurotransmission remain to be clarified. For example, modifications of physical properties of the neuronal membrane, effects on proteins (receptors, transporters) enclosed in the membrane, and effects on gene expression and/or transcription might occur. Whatever the mechanism, it is therefore assumed that interactions exist among PUFA, neurotransmission, and behavior. This might be related to clinical findings. Indeed, deficits in the peripheral amounts of PUFA have been described in subjects suffering from neurological and psychiatric disorders. Involvement of the monoaminergic neurotransmission function has been demonstrated or hypothesized in several of these diseases. It can therefore be proposed that functional links exist among PUFA status, neurotransmission processes, and behavioral disorders in humans. Animal models are tools of choice for the understanding of such links. Improved prevention and complementary treatment of neurological and psychiatric diseases can be expected from these studies.

Omega-3 fatty acids and brain resistance to ageing and stress: body of evidence and possible mechanisms.

The increasing life expectancy in the populations of rich countries raises the pressing question of how the elderly can maintain their cognitive function. Cognitive decline is characterised by the loss of short-term memory due to a progressive impairment of the underlying brain cell processes. Age-related brain damage has many causes, some of which may be influenced by diet. An optimal diet may therefore be a practical way of delaying the onset of age-related cognitive decline. Nutritional investigations indicate that the ω-3 poyunsaturated fatty acid (PUFA) content of western diets is too low to provide the brain with an optimal supply of docosahexaenoic acid (DHA), the main ω-3 PUFA in cell membranes. Insufficient brain DHA has been associated with memory impairment, emotional disturbances and altered brain processes in rodents. Human studies suggest that an adequate dietary intake of ω-3 PUFA can slow the age-related cognitive decline and may also protect against the risk of senile dementia. However, despite the many studies in this domain, the beneficial impact of ω-3 PUFA on brain function has only recently been linked to specific mechanisms. This review examines the hypothesis that an optimal brain DHA status, conferred by an adequate ω-3 PUFA intake, limits age-related brain damage by optimizing endogenous brain repair mechanisms. Our analysis of the abundant literature indicates that an adequate amount of DHA in the brain may limit the impact of stress, an important age-aggravating factor, and influences the neuronal and astroglial functions that govern and protect synaptic transmission. This transmission, particularly glutamatergic neurotransmission in the hippocampus, underlies memory formation. The brain DHA status also influences neurogenesis, nested in the hippocampus, which helps maintain cognitive function throughout life. Although there are still gaps in our knowledge of the way ω-3 PUFA act, the mechanistic studies reviewed here indicate that ω-3 PUFA may be a promising tool for preventing age-related brain deterioration.

n-3 polyunsaturated fatty acid supplementation reverses stress-induced modifications on brain monoamine levels in mice.

The aim of this study was to examine the effects of supplementation with n-3 polyunsaturated fatty acids (PUFAs) on stress responses in mice subjected to an unpredictable chronic mild stress (UCMS) procedure. Stress-induced modifications in coat and aggressiveness were evaluated, and phospholipid PUFA profiles and monoamine levels were analyzed in the frontal cortex, hippocampus, and striatum. The results showed that repeated exposure to mild stressors induced degradation in the physical state of the coat, lowered body weight gain, and increased aggressiveness, without any effect of n-3 PUFA supplementation. The UCMS induced a significant decrease in the levels of norepinephrine in the frontal cortex and striatum, and a nonsignificant decrease in the hippocampus. The tissue levels of serotonin (5-HT) were 40% to 65% decreased in the three brain regions studied. Interestingly, the n-3 PUFA supplementation reversed this stress-induced reduction in 5-HT levels. These findings showed that supplementation in n-3 long-chain PUFAs might reverse certain effects of UCMS in cerebral structures involved in stress-related behaviors.

Dietary docosahexaenoic acid [22: 6(n-3)] as a phospholipid or a triglyceride enhances the potassium chloride-evoked release of acetylcholine in rat hippocampus.

We demonstrated previously that a dietary-induced depletion of docosahexaenoic acid (DHA) in cerebral phospholipids increases the spontaneous release of acetylcholine (Ach) in the rat hippocampus and reduces its potassium chloride evoked-release. In the present study, we investigated the effects in rats of DHA-enriched diets supplied by egg phospholipids (E-PL) or tuna oil (TO) on the PUFA in hippocampus membranes and on the synaptic release of Ach. Control rats were fed 3 g/kg of the DHA precursor, alpha-linolenic acid (LNA). Chronically (n-3) PUFA-deficient females were fed, starting 2 wk before mating, the deficient diet, a control diet, or a purified diet supplying 1, 2, or 3 g DHA/kg diet as E-PL or TO. Experiments were performed on the adult male progeny fed the same diet as their dams throughout life. The form of dietary DHA (TO or E-PL) did not influence its incorporation into the hippocampus. The 1 g DHA/kg diets allowed maximal incorporation into phosphatidylethanolamine (PE), but 2 g DHA/kg diet was needed for phosphatidylcholine (PC). A minimum of 2 g DHA/kg was needed to decrease the basal Ach release and to enhance the stimulated release to that of the control; the Ach release of the 1 g/kg DHA-groups did not differ from that of the deficient group. This suggests that >1 g DHA/kg diet is needed to ensure PUFA incorporation into PE and PC, and basal and stimulated Ach release in the rat hippocampus equivalent to the control group fed only LNA. PUFA incorporation into the hippocampus depends mainly on the PUFA concentration of the diet, not on the form of dietary DHA.

Alpha-linolenic acid deficiency modifies distractibility but not anxiety and locomotion in rats during aging.

In rodents, chronic dietary alpha-linolenic acid deficiency decreases learning and memory and alters dopaminergic and serotoninergic neurotransmission. However, these two neurotransmitter systems are related mainly to attention, emotion and locomotion. Therefore, we decided to investigate the effects of dietary alpha-linolenic acid deficiency in rats tested with animal models of distractibility (the distractometer procedure), anxiety (the elevated plus maze) and ambulatory activity (a circular corridor). Moreover, because these neurochemical modifications persist during aging, we decided to study the effects of aging on these behaviors by using rats aged 2, 6, 12 and 24 mo. An age-related decline in distractibility was observed that was accelerated by linolenic acid deficiency. Indeed, an age-related reduction in distractibility was found in so far as distraction time was reduced at the age of 12 mo in controls and at the age of 24 mo in deficient groups compared with 2-mo-old rats. Moreover, distraction time was significantly lower in 6- and 24-mo-old rats fed a deficient diet compared with age-matched controls. Anxiety was not modified by diet or age. Finally, a parallel decrease in locomotion was exhibited by rats fed both diets between 6 and 12 mo of age. Locomotion was not modified by diet. These results show that dietary alpha-linolenic deficiency alters behavior in a very specific way; distractibility is modified by diet, whereas anxiety and locomotion are not, suggesting that particular brain areas may be altered.

Dietary fish oil affects monoaminergic neurotransmission and behavior in rats.

We studied the effects of a fish oil enriched diet on fatty acid composition of cerebral membranes and on several neurochemical and behavioral variables of monoaminergic function in rats. The frontal cortex, striatum, hippocampus and cerebellum were studied in rats fed fish oil (FPO, 50% salmon oil + 50% palm oil), which provided an (n-6)/(n-3) polyunsaturated fatty acid (PUFA) ratio of 0.14 versus 6. 19 in controls fed a diet containing a mixture of African peanut oil and rapeseed oil. In the FPO group compared to the control group, the major modifications in fatty acid composition of cerebral membranes included the following: higher levels in 22:6(n-3), lower levels in 20:4(n-6) and a significantly greater proportion of phosphatidylserine. Dopamine levels were 40% greater in the frontal cortex of rats fed FPO than from those fed the control diet. In this cerebral region there was also a reduction in monoamine oxidase B (MAO-B) activity and greater binding to dopamine D2 receptors. By contrast, a lower binding to dopamine D2 receptors (-7%) was observed in the striatum. Ambulatory activity was also reduced in FPO-fed rats, possibly related to observed changes in striatal dopaminergic receptors. This suggested that the level of (n-6) PUFA, which was considerably lower in the FPO diet than in the control diet, could act on locomotion through an effect on striatal dopaminergic function, whereas the high level of (n-3) PUFA could act on cortical dopaminergic function.

Modification of dopamine neurotransmission in the nucleus accumbens of rats deficient in n-3 polyunsaturated fatty acids.

We studied the effects of a diet chronically deficient in alpha-linolenic acid, the precursor of long-chain n-3 polyunsaturated fatty acids, on dopaminergic neurotransmission in the shell region of the nucleus accumbens of rats. In vivo microdialysis experiments showed increased basal levels of dopamine and decreased basal levels of metabolites, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), in awake rats from the deficient group compared to controls. The release of dopamine under KCl stimulation was similar in both dietary groups. By contrast, the release of dopamine from the vesicular storage pool under tyramine stimulation was 90% lower in the deficient than in the control rats. Autoradiographic studies in the same cerebral region revealed a 60% reduction in the vesicular monoamine transporter sites in the deficient group. Dopamine D(2) receptors were 35% increased in these rats compared to controls, whereas no change occurred for D(1) receptors and membrane dopamine transporters. These results demonstrated that chronic n-3 polyunsaturated fatty acid deficiency modifies several factors of dopaminergic neurotransmission in the nucleus accumbens. These findings are in agreement with the changes in dopaminergic neurotransmission already observed in the frontal cortex, and with the behavioral disturbances described in these deficient rats.