Denis Guilloteau

α‐Linolenic Acid Dietary Deficiency Alters Age‐Related Changes of Dopaminergic and Serotoninergic Neurotransmission in the Rat Frontal Cortex

Abstract: The effects of α‐linolenic acid diet deficiency on rat dopaminergic and serotoninergic neurotransmission systems were investigated in the frontal cortex, striatum, and cerebellum of male rats 2, 6, 12, and 24 months of age. The diet deficiency induced a severe decrease in the 22:6n‐3 fatty acid levels in all regions and a compensatory increase in n‐6 fatty acid levels. A recovery in the levels of 22:6n‐3 was observed in deficient rats between 2 and 12 months of age; however, this recovery was lower in frontal cortex than in striatum and cerebellum. In the striatum and the cerebellum, dopaminergic and serotoninergic receptor densities and endogenous dopamine and serotonin levels were affected by aging regardless of the diet. In contrast, a 40–75% lower level of endogenous dopamine in the frontal cortex occurred in deficient rats according to age. The deficiency also induced an 18–46% increase in serotonin 5‐HT2 receptor density in the frontal cortex during aging, without variation in endogenous serotonin level, and a 10% reduction in density of dopaminergic D2 receptors. Monoamine oxidase‐A and ‐B activities showed specific age‐related variations but regardless of the diet. Our results suggest that a chronically α‐linolenic‐deficient diet specifically affects the monoaminergic systems in the frontal cortex.

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 α-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 behavioural 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.

Serotoninergic neurotransmission is affected by n-3 polyunsaturated fatty acids in the rat

We explored the effects of chronic α‐linolenic acid dietary deficiency on serotoninergic neurotransmission. In vivo synaptic serotonin (5‐HT) levels were studied in basal and pharmacologically stimulated conditions using intracerebral microdialysis in the hippocampus of awake 2‐month‐old rats. We also studied the effects of reversion of the deficient diet on fatty acid composition and serotoninergic neurotransmission. A balanced (control) diet was supplied to deficient rats at different stages of development, i.e. from birth, 7, 14 or 21 days of age. We demonstrated that chronic n‐3 polyunsaturated fatty acid dietary deficiency induced changes in the synaptic levels of 5‐HT both in basal conditions and after pharmacological stimulation with fenfluramine. Higher levels of basal 5‐HT release and lower levels of 5‐HT‐stimulated release were found in deficient than in control rats. These neurochemical modifications were reversed by supply of the balanced diet provided at birth or during the first 2 weeks of life through the maternal milk, whereas they persisted if the balanced diet was given from weaning (at 3 weeks of age). This suggests that provision of essential fatty acids is durably able to affect brain function and that this is related to the developmental stage during which the deficiency occurs.

DPA-714, a New Translocator Protein–Specific Ligand: Synthesis, Radiofluorination, and Pharmacologic Characterization

The translocator protein (18 kDa) (TSPO), formerly known as the peripheral benzodiazepine receptor, is dramatically upregulated under pathologic conditions. Activated microglia are the main cell type expressing the TSPO at sites of central nervous system pathology. Radioligands for the TSPO can therefore measure active disease in the brain. This article details the synthesis, radiofluorination, and pharmacologic evaluation of a new TSPO-specific pyrazolopyrimidine, DPA-714. Methods: The affinity of DPA-714 for the TSPO was measured in rat kidney membranes with 3H-PK11195. The in vitro functional activity of DPA-714 was measured in a steroidogenic assay in which the ability of DPA-714 to increase pregnenolone synthesis was measured with rat C6 glioma cells. The radiofluorination of DPA-714 was achieved by nucleophilic 18F-fluoride displacement of the tosylate precursor. 18F-DPA-714 was assessed in rats harboring unilateral quinolinic acid (QA) lesions. In addition, pretreatment experiments were performed with PK11195 (5 mg/kg), DPA-714 (1 mg/kg), and DPA-713 (1 mg/kg). The in vivo binding and biodistribution of 18F-DPA-714 were determined in a baboon with PET. Experiments involving presaturation with PK11195 (1.5 mg/kg) and displacement with DPA-714 (1 mg/kg) were conducted to evaluate the specificity of radioligand binding. Results: In vitro binding studies revealed that DPA-714 displayed a high affinity for the TSPO (dissociation constant, 7.0 nM). DPA-714 stimulated pregnenolone synthesis at levels 80% above the baseline. 18F-DPA-714 was prepared at a 16% radiochemical yield and a specific activity of 270 GBq/μmol. In rats harboring unilateral QA lesions, an 8-fold-higher level of uptake of 18F-DPA-714 was observed in the ipsilateral striatum than in the contralateral striatum. Uptake in the ipsilateral striatum was shown to be selective because it was inhibited to the level in the contralateral striatum in the presence of PK11195, nonlabeled DPA-714, or DPA-713. PET studies demonstrated rapid penetration and good retention of 18F-DPA-714 in the baboon brain. Pretreatment with PK11195 effectively inhibited the uptake of 18F-DPA-714 in the whole brain, indicating its selective binding to the TSPO. The injection of nonlabeled DPA-714 20 min after the injection of 18F-DPA-714 resulted in radioligand washout, demonstrating the reversibility of 18F-DPA-714 binding. Conclusion: 18F-DPA-714 is a specific radioligand for the TSPO, displaying promising in vivo properties and thus warranting further investigation.

Chronic n-3 polyunsaturated fatty acid deficiency alters dopamine vesicle density in the rat frontal cortex

We studied the effects of a chronic deficiency in n-3 polyunsaturated fatty acids (n-3 PUFA) on the vesicle dopaminergic compartment in the frontal cortex of rats. Electronic micrographic analysis showed that the synaptic density and the clear vesicle density were similar in deficient and control rats. However, dopaminergic immunolabeling revealed a significantly decreased number of gold-labeled vesicles in the dopaminergic presynaptic terminals of the deficient rats. These findings demonstrate that dopamine cortical vesicles are specifically decreased in n-3 PUFA deficiency. The mechanism leading to this modification could involve several abnormalities (vesicle turn-over, membrane fluidity, vesicular monoamine transporter). This reduction in the dopaminergic vesicle pool constitutes the first structural support for the previously described modifications of dopamine metabolism in the frontal cortex. Such changes in dopamine neurotransmission could be involved in behavioral abnormalities occurring in n-3 PUFA deficient rats.

Region-Specific Hierarchy between Atrophy, Hypometabolism, and β-Amyloid (Aβ) Load in Alzheimer's Disease Dementia

Gray matter atrophy, glucose hypometabolism, and β-amyloid Aβ deposition are well-described hallmarks of Alzheimers disease, but their relationships are poorly understood. The present study aims to compare the local levels of these three alterations in humans with Alzheimers disease. Structural magnetic resonance imaging, 18F-fluorodeoxyglucose positron emission tomography (PET), and 18F-florbetapir PET data from 34 amyloid-negative healthy controls and 20 demented patients with a high probability of Alzheimers disease etiology (attested using neuroimaging biomarkers as recently recommended) were analyzed. For each patient and imaging modality, age-adjusted Z-score maps were computed, and direct between-modality voxelwise comparison and correlation analyses were performed. Significant differences in the levels of atrophy, hypometabolism, and Aβ deposition were found in most brain areas, but the hierarchy differed across regions. A cluster analysis revealed distinct subsets of regions: (1) in the hippocampus, atrophy exceeded hypometabolism, whereas Aβ load was minimal; (2) in posterior association areas, Aβ deposition was predominant, together with high hypometabolism and lower but still significant atrophy; and (3) in frontal regions, Aβ deposition was maximal, whereas structural and metabolic alterations were low. Atrophy and hypometabolism significantly correlated in the hippocampus and temporo-parietal cortex, whereas Aβ load was not significantly related to either atrophy or hypometabolism. These findings provide direct evidence for regional variations in the hierarchy and relationships between Aβ load, hypometabolism, and atrophy. Altogether, these variations probably reflect the differential involvement of region-specific pathological or protective mechanisms, such as the presence of neurofibrillary tangles, disconnection, as well as compensation processes.

Partial recovery of dopaminergic pathway after graft of adult mesenchymal stem cells in a rat model of Parkinson's disease

Cellular therapy with adult stem cells appears as an opportunity for treatment of Parkinsons disease. To validate this approach, we studied the effects of transplantation of rat adult bone-marrow mesenchymal stem cells in a rat model of Parkinsons disease. Animals were unilaterally lesioned in the striatum with 6-hydroxydopamine. Two weeks later, group I did not undergo grafting, group II underwent sham grafting, group III was intra-striatal grafted with cells cultured in an enriched medium and group IV was intra-striatal grafted with cells cultured in a standard medium. Rotational amphetamine-induced behavior was measured weekly until animals were killed 6 weeks later. One week after graft, the number of rotations/min was stably decreased by 50% in groups III and IV as compared with groups I and II. At 8 weeks post-lesion, the density of dopaminergic markers in the nerve terminals and cell bodies, i.e. immunoreactive tyrosine hydroxylase, membrane dopamine transporter and vesicular monoamine transporter-2 was significantly higher in group III as compared with group I. Moreover, using microdialysis studies, we observed that while the rate of pharmacologically induced release of dopamine was significantly reduced in lesioned versus intact striatum in no grafted rats, it was similar in both sides in animals transplanted with mesemchymal stem cells. These data demonstrate that graft of adult mesemchymal stem cells reduces behavioral effects induced by 6-hydroxydopamine lesion and partially restores the dopaminergic markers and vesicular striatal pool of dopamine. This cellular approach might be a restorative therapy in Parkinsons disease.

Fetal exposure to teratogens: evidence of genes involved in autism.

Environmental challenges during the prenatal period can result in behavioral abnormalities and cognitive deficits that appear later in life such as autism. Prenatal exposure to valproic acid, ethanol, thalidomide and misoprostol has been shown to be associated with an increased incidence of autism. In addition, rodents exposed in utero to some of these drugs show autism-like abnormalities, including brain changes and lifelong behavior dysfunction. Our aim is to summarize current understanding of the relationship between in utero exposure to these drugs and autism in humans and in autism-like animal model phenotypes. It also highlights the importance of these models to understanding the neurobiology of autism, particularly in the identification of susceptibility genes. These drugs are able to modulate the expression of many genes involved in processes such as proliferation, apoptosis, neuronal differentiation and migration, synaptogenesis and synaptic activity. It seems essential to focus research on genes expressed during early neurodevelopment which may be the target of mutations or affected by drugs such as those included in this review.

Molecular Imaging of Microglial Activation in Amyotrophic Lateral Sclerosis

There is growing evidence of activated microglia and inflammatory processes in the cerebral cortex in amyotrophic lateral sclerosis (ALS). Activated microglia is characterized by increased expression of the 18 kDa translocator protein (TSPO) in the brain and may be a useful biomarker of inflammation. In this study, we evaluated neuroinflammation in ALS patients using a radioligand of TSPO, 18F-DPA-714. Ten patients with probable or definite ALS (all right-handed, without dementia, and untreated by riluzole or other medication that might bias the binding on the TSPO), were enrolled prospectively and eight healthy controls matched for age underwent a PET study. Comparison of the distribution volume ratios between both groups were performed using a Mann-Whitney’s test. Significant increase of distribution of volume ratios values corresponding to microglial activation was found in the ALS sample in primary motor, supplementary motor and temporal cortex (p = 0.009, p = 0.001 and p = 0.004, respectively). These results suggested that the cortical uptake of 18F-DPA-714 was increased in ALS patients during the “time of diagnosis” phase of the disease. This finding might improve our understanding of the pathophysiology of ALS and might be a surrogate marker of efficacy of treatment on microglial activation.

Chronic n-3 polyunsaturated fatty acid diet-deficiency acts on dopamine metabolism in the rat frontal cortex: a microdialysis study

The effects of alpha-linolenic acid diet deficiency on rat dopaminergic metabolism were investigated in the frontal cortex of male 2-3 month-old rats using the microdialysis method. Increased basal levels of dopamine metabolites were observed in the frontal cortex of awake deficient rats, without modification of dopamine levels. Moreover, using KCl perfusion which releases newly synthesized dopamine, no difference was observed in anaesthetized deficient rats versus control rats. In addition, a decrease in dopamine release was observed in anaesthetized deficient rats versus control rats after tyramine stimulation, which is known to induce release of dopamine from vesicular stores. A working model is proposed which suggests that a chronic n-3 polyunsaturated fatty acids (PUFA) deficiency may lead to modifications in the internalization of dopamine in the storage pool in the frontal cortex.