Cristina L. Sánchez

Effects of acute tryptophan depletion on brain serotonin function and concentrations of dopamine and norepinephrine in C57BL/6J and BALB/cJ mice.

Acute tryptophan depletion (ATD) is a method of lowering brain serotonin (5-HT). Administration of large neutral amino acids (LNAA) limits the transport of endogenous tryptophan (TRP) across the blood brain barrier by competition with other LNAAs and subsequently decreases serotonergic neurotransmission. A recent discussion on the specificity and efficacy of the ATD paradigm for inhibition of central nervous 5-HT has arisen. Moreover, side effects such as vomiting and nausea after intake of amino acids (AA) still limit its use. ATD Moja-De is a revised mixture of AAs which is less nauseating than conventional protocols. It has been used in preliminary clinical studies but its effects on central 5-HT mechanisms and other neurotransmitter systems have not been validated in an animal model. We tested ATD Moja-De (TRP−) in two strains of mice: C57BL/6J, and BALB/cJ, which are reported to have impaired 5-HT synthesis and a more anxious phenotype relative to other strains of mice. ATD Moja-De lowered brain TRP, significantly decreased 5-HT synthesis as indexed by 5-HTP levels after decarboxlyase inhibition, and lowered 5-HT and 5-HIAA in both strains of mice, however more so in C57BL/6J than in BALB/cJ. Dopamine and its metabolites as well as norepinephrine were not affected. A balanced (TRP+) control mixture did not raise 5-HT or 5-HIAA. The present findings suggest that ATD Moja-De effectively and specifically suppresses central serotonergic function. These results also demonstrate a strain- specific effect of ATD Moja-De on anxiety-like behavior.

Pharmacometabolomics reveals that serotonin is implicated in aspirin response variability.

While aspirin is generally effective for prevention of cardiovascular disease, considerable variation in drug response exists, resulting in some individuals displaying high on‐treatment platelet reactivity. We used pharmacometabolomics to define pathways implicated in variation of response to treatment. We profiled serum samples from healthy subjects pre‐ and postaspirin (14 days, 81 mg/day) using mass spectrometry. We established a strong signature of aspirin exposure independent of response (15/34 metabolites changed). In our discovery (N = 80) and replication (N = 125) cohorts, higher serotonin levels pre‐ and postaspirin correlated with high, postaspirin, collagen‐induced platelet aggregation. In a third cohort, platelets from subjects with the highest levels of serotonin preaspirin retained higher reactivity after incubation with aspirin than platelets from subjects with the lowest serotonin levels preaspirin (72 ± 8 vs. 61 ± 11%, P = 0.02, N = 20). Finally, ex vivo, serotonin strongly increased platelet reactivity after platelet incubation with aspirin (+20%, P = 4.9 × 10−4, N = 12). These results suggest that serotonin is implicated in aspirin response variability.

Effects of serotonin depletion on punishment processing in the orbitofrontal and anterior cingulate cortices of healthy women

Diminished synthesis of the neurotransmitter serotonin (5-HT) has been linked to disrupted impulse control in aversive contexts. However, the neural correlates underlying a serotonergic modulation of female impulsivity remain unclear. The present study investigated punishment-induced inhibition in healthy young women. Eighteen healthy female subjects (aged 20-31) participated in a double-blinded, counterbalanced, placebo-controlled, within subjects, repeated measures study. They were assessed on two randomly assigned occasions that were controlled for menstrual cycle phase. In a randomized order, one day, acute tryptophan depletion (ATD) was used to reduce 5-HT synthesis in the brain. On the other day, participants received a tryptophan-balanced amino acid load (BAL) as a control condition. Three hours after administration of ATD/BAL, neural activity was recorded during a modified Go/No-Go task implementing reward or punishment processes using functional magnetic resonance imaging (fMRI). Neural activation during No-Go trials in punishment conditions after BAL versus ATD administration correlated positively with the magnitude of central 5-HT depletion in the ventral and subgenual anterior cingulate cortices (ACC). Furthermore, neural activation in the medial orbitofrontal cortex (mOFC) and the dorsal ACC correlated positively with trait impulsivity. The results indicate reduced neural sensitivity to punishment after short-term depletion of 5-HT in brain areas related to emotion regulation (subgenual ACC) increasing with depletion magnitude and in brain areas related to appraisal and expression of emotions (mOFC and dorsal ACC), increasing with trait impulsivity. This suggests a serotonergic modulation of neural circuits related to emotion regulation, impulsive behavior, and punishment processing in females.

Estradiol replacement enhances cocaine-stimulated locomotion in female C57BL/6 mice through estrogen receptor alpha.

Psychostimulant effects are enhanced by ovarian hormones in women and female rodents. Estradiol increases behavioral responses to psychostimulants in women and female rats, although the underlying mechanism is unknown. This study utilized mice to investigate the time frame and receptor mediation of estradiols enhancement of cocaine-induced behavior as mice enable parallel use of genetic, surgical and pharmacological methods. The spontaneous behavior of Sham and Ovariectomized (Ovx) female wildtype (WT) mice was determined during habituation to a novel environment and after cocaine administration. Ovx mice were replaced with vehicle (sesame oil) or 17β-estradiol (E2) for 2 days or 30 min prior to a cocaine challenge to investigate the time course of E2s effects. To examine receptor mediation of estradiol effects, Ovx mice replaced for 2 days with either the ERα-selective agonist PPT or the ERβ-selective agonist DPN were compared to Sham mice, and mice lacking either ERα (αERKO) or ERβ (βERKO) were compared to WT littermates. Ovx mice exhibited fewer ambulations during habituation than Sham females. Cocaine-induced increases in behavioral ratings were greater in Sham than in Ovx mice. Two days but not 30 min of E2 replacement in Ovx mice increased cocaine responses to Sham levels. PPT replacement also increased the cocaine response relative to vehicle- or DPN- treated Ovx mice. αERKO mice displayed modestly attenuated behavioral responses to novelty and cocaine compared to αWT littermates, but no behavioral differences were found between βERKO and βWT mice. These results suggest that E2 enhances cocaine-stimulated locomotion in mice predominantly through ERα.

Dietary manipulation of serotonergic and dopaminergic function in C57BL/6J mice with amino acid depletion mixtures

Amino acid (AA) depletion techniques have been used to decrease serotonin (5-HT) and/or dopamine (DA) synthesis after administration of a tryptophan (acute tryptophan depletion, ATD) or phenylalanine/tyrosine-free (phenylalanine–tyrosine depletion, PTD) AA formula and are useful as neurochemical challenge procedures to study the impact of DA and 5-HT in patients with neuropsychiatric disorders. We recently demonstrated that the refined Moja-De ATD paradigm decreases brain 5-HT synthesis in humans and mice and lowers brain 5-HT turnover. In the present study we validated the neurochemical effects of three developed AA formulas on brain 5-HT and DA function in mice. To distinguish the direct and indirect effects of such mixtures on 5-HT and DA and to determine whether additive depletion of both could be obtained simultaneously, we compared the effects of ATD for 5-HT, PTD for DA, and a combined monoamine depletion mixture (CMD) compared to a control condition consisting of a balanced amino acid mixture. Food-deprived male C57BL/6J mice were gavaged with AA mixtures. Serum and brain samples were collected and analyzed for determination of tryptophan (Trp), tyrosine (Tyr), 5-HT, 5-HIAA, DA, DOPAC and HVA levels. ATD was the most effective at decreasing Trp, 5-HT and 5-HIAA. In contrast, PTD reduced Tyr globally but HVA only in certain brain regions. Although CMD affected both 5-HT and DA synthesis, it was less effective when compared with ATD or PTD alone. The present results demonstrate that two newly developed PTD and CMD formulas differentially impact brain 5-HT and DA synthesis relative to 5-HT-specific ATD Moja-De. Different effects on 5-HT and DA function by these mixtures suggest that the exact composition may be a critical determinant for effectiveness with respect to the administered challenge procedure.

The Role of Serotonin (5-HT) in Behavioral Control: Findings from Animal Research and Clinical Implications

The neurotransmitters serotonin and dopamine both have a critical role in the underlying neurobiology of different behaviors. With focus on the interplay between dopamine and serotonin, it has been proposed that dopamine biases behavior towards habitual responding, and with serotonin offsetting this phenomenon and directing the balance toward more flexible, goal-directed responding. The present focus paper stands in close relationship to the publication by Worbe et al. (2015), which deals with the effects of acute tryptophan depletion, a neurodietary physiological method to decrease central nervous serotonin synthesis in humans for a short period of time, on the balance between hypothetical goal-directed and habitual systems. In that research, acute tryptophan depletion challenge administration and a following short-term reduction in central nervous serotonin synthesis were associated with a shift of behavioral performance towards habitual responding, providing further evidence that central nervous serotonin function modulates the balance between goal-directed and stimulus-response habitual systems of behavioral control. In the present focus paper, we discuss the findings by Worbe and colleagues in light of animal experiments as well as clinical implications and discuss potential future avenues for related research.

Simplified dietary acute tryptophan depletion: effects of a novel amino acid mixture on the neurochemistry of C57BL/6J mice

Background Diet and nutrition can impact on the biological processes underpinning neuropsychiatric disorders. Amino acid (AA) mixtures lacking a specific neurotransmitter precursor can change the levels of brain serotonin (5-HT) or dopamine (DA) in the central nervous system. The availability of these substances within the brain is determined by the blood–brain barrier (BBB) that restricts the access of peripheral AA into the brain. AA mixtures lacking tryptophan (TRP) compete with endogenous TRP for uptake into the brain across the BBB, which in turn leads to a decrease in central nervous 5-HT synthesis. Objective The present study compared the effects of a simplified acute tryptophan depletion (SATD) mixture in mice on blood and brain serotonergic and dopaminergic metabolites to those of a commonly used acute tryptophan depletion mixture (ATD Moja-De) and its TRP-balanced control (BAL). Design The SATD formula is composed of only three large neutral AAs: phenylalanine (PHE), leucine (LEU), and isoleucine (ILE). BAL, ATD Moja-De, or SATD formulas were delivered to adult male C57BL/6J mice by gavage. TRP, monoamines, and their metabolites were quantified in blood and brain regions (hippocampus, frontal cortex, amygdala, caudate putamen, and nucleus accumbens). Results Both ATD Moja-De and SATD significantly decreased levels of serum and brain TRP, as well as brain 5-HIAA and 5-HT compared with BAL. SATD reduced HVA levels in caudate but did not alter total DA levels or DOPAC. SATD decreased TRP and serotonergic metabolites comparably to ATD Moja-De administration. Conclusion A simplified and more palatable combination of AAs can manipulate serotonergic function and might be useful to reveal underlying monoamine-related mechanisms contributing to different neuropsychiatric disorders.

Studying the effects of dietary body weight-adjusted acute tryptophan depletion on punishment-related behavioral inhibition

Background Alterations in serotonergic (5-HT) neurotransmission are thought to play a decisive role in affective disorders and impulse control. Objective This study aims to reproduce and extend previous findings on the effects of acute tryptophan depletion (ATD) and subsequently diminished central 5-HT synthesis in a reinforced categorization task using a refined body weight–adjusted depletion protocol. Design Twenty-four young healthy adults (12 females, mean age [SD]=25.3 [2.1] years) were subjected to a double-blind within-subject crossover design. Each subject was administered both an ATD challenge and a balanced amino acid load (BAL) in two separate sessions in randomized order. Punishment-related behavioral inhibition was assessed using a forced choice go/no-go task that incorporated a variable payoff schedule. Results Administration of ATD resulted in significant reductions in TRP measured in peripheral blood samples, indicating reductions of TRP influx across the blood–brain barrier and related brain 5-HT synthesis. Overall accuracy and response time performance were improved after ATD administration. The ability to adjust behavioral responses to aversive outcome magnitudes and behavioral adjustments following error contingent punishment remained intact after decreased brain 5-HT synthesis. A previously observed dissociation effect of ATD on punishment-induced inhibition was not observed. Conclusions Our results suggest that neurodietary challenges with ATD Moja–De have no detrimental effects on task performance and punishment-related inhibition in healthy adults.

Acute tryptophan depletion – converging evidence for decreasing central nervous serotonin synthesis in rodents and humans

We read the comment provided by Simon N. Young (1) on the articles (2–5) in the special issue of Acta Psychiatrica Scandinavica (6) dealing with the acute tryptophan depletion (ATD) methodology with great interest. ATD is a pharmacological method designed to lower central nervous system (CNS) synthesis of the neurotransmitter serotonin (5-HT) for a brief period that can also be used in both adults and young people (7). As 5-HT plays an important role in behavioral inhibition (8– 10) and other important processes in the brain (11–14), ATD is a translational method to study the effects of changes in CNS 5-HT function that has particular value, as discussed at a recent symposium dedicated to the role of 5-HT in psychopathology (7–11, 15). The author of this particular comment expressed concerns that ATD might not always decrease CNS 5-HT synthesis and that the lack of the amino acid histidine (HIS) in the depletion mixtures used might influence the results due to the potential role of 5-HT–histamine interactions in any observed outcome. We appreciate the comments made and would like to address the issues raised, point by point. Young argues that ‘there is no evidence that ATD does always decrease serotonin release (in humans)’. This is contradictory by decades of work in rodents and in humans demonstrating that ATD can decrease 5-HT synthesis and release in rodents and lower 5-HIAA in human CSF (16–19). In one of our laboratories, the acute tryptophan depletion (ATD) protocol termed ‘Moja-De’ has been shown to decrease 5-HT release in rodents (20, 21) and to lower tryptophan (TRP) comparably in humans (22), suggesting that this mixture successfully decreases 5-HT synthesis as postulated. While some experiments (23) fail to detect changes in central 5-HT function after ATD, this is the exception rather than the rule in published studies. The author of this comment was also concerned that there would be regional variations in the inhibition of serotonin function. This is logical and consistent with published data on the effects of Moja-De ATD in mice. Mouse studies indicated that depletion of TRP was comparable across different brain areas but that the extent of decrease in 5-HIAA varied by region (20, 21). Regional release of 5-HT is controlled by a combination of cell firing including regionally selective input, the concentration of 5-HT1b receptors on terminals, the amount of tryptophan hydroxylase, and many other factors (24). However, there is no evidence that 5-HT release happens only in selective regions, but we agree the magnitude of ATD effects on release is likely to vary between regions despite comparable depletion of TRP. As regards potential interactions between 5-HT and histamine, we agree that measurement of histidine after depletion of TRP or any other formula lacking HIS is of interest. Young has questioned the results of ATD experiments in which HIS was not included, stating that ‘histidine is an essential amino acid’. However, the essentiality of this amino acid is not clearly established (25). It has been reported that HIS was not necessary for the maintenance of nitrogen balances in short-term (26, 27). Kriengsunyos et al. (28) observed after a long-term histidine depletion administered to healthy adults that there were no effects on the protein metabolism (urinary nitrogen excretion and nitrogen balance). They suggested that the essentiality of this amino acid in healthy adults is still unclear as there are some components that may serve as sources of HIS, although the data they reported indicate that this amount may not be enough for maintain the HIS pool. The other concern expressed by Young was that effects of ATD could reflect disruption of a histamine–serotonin interaction, as ATD would cause a dramatic decrease in histamine synthesis. This is possible, as it is well established that the neurotransmitter histamine is formed from HIS (29), and histamine turnover seems to occur faster than other biogenic amines, such as norepinephrine or 5-HT (30). Therefore, in the absence of HIS, competition from the amino acid mixtures could indeed lower histamine production. However, neither the control nor the ATD mixture in most studies contains histidine, and so histamine would not be differentially affected by the ATD mixture, but should be comparably depleted in both control and ATD mixtures. Nevertheless, it is possible that some interaction between histamine depletion and 5-HT depletion could have behavioral effects. Unfortunately, no behavioral effects of histamine depletion have been clearly established in the literature. A study by Young and his collaborators of HIS depletion effects on sensory and motor behavior in healthy adults (31) showed that HIS in plasma decreased 20% and the ratio HIS/ΣLNAA decreased 59%, but there were no behavioral effects of this depletion. Finally, we disagree with the statement that ‘the relevance of such animal studies to the far more complex human brain is uncertain’. It is well known that validation of translational methods has allowed modeling many aspects of the neuropsychopathology with the use of appropriate animal models, the majority of them throughout the use of rodents (32, 33). Translation of behavioral findings is challenging, due to limits in extrapolating simple behavioral tasks in rodents to sophisticated behaviors in humans. However, biochemical studies of ATD effects in humans and rodents have shown considerable concordance. For example, our studies in humans (5, 22) have been validated in mice (20, 21), consistent with the field as described above (16–19). As Dr. Young points out, detailed anatomic studies of 5-HT synthesis in the human brain are technologically demanding and rarely conducted. However, the concordance between the dependent measures that can be collected in humans (CSF 5-HIAA for example) and comparable measures in rodents (tissue 5-HIAA content, 5-HT and 5-HIAA content in microdialysate) supports the concordance of findings after ATD in humans. In summary, there is convincing and converging evidence that ATD decreases 5-HT synthesis in the brain in both rodents and humans. Interactions between 5-HT and

Perinatal western-type diet and associated gestational weight gain alter postpartum maternal mood

The role of perinatal diet in postpartum maternal mood disorders, including depression and anxiety, remains unclear. We investigated whether perinatal consumption of a Western‐type diet (high in fat and branched‐chain amino acids [BCAA]) and associated gestational weight gain (GWG) cause serotonin dysregulation in the central nervous system (CNS), resulting in postpartum depression and anxiety (PPD/A).