The effect of choline availability from gestation to early development on brain and retina functions and phospholipid composition in a male mouse model.

OBJECTIVES: Although choline is essential for brain development and neural function, the effect of choline on retina function is not well understood. This study examined the effects of choline on neural tissues of brain and retina, and membrane phospholipid (PL) composition during fetal development. METHODS: Pregnant C57BL/6 mice were fed one of 4 choline modified diets: i) control (Cont, 2.5g/kg), ii) choline deficient (Def, 0g/kg), iii) supplemented with choline chloride (Cho, 10g/kg) and iv) supplemented with egg phosphatidylcholine (PC, 10g/kg). At postnatal day (PD) 21, pups were weaned onto their mothers' respective diets until PD 45. Spatial memory was measured using the Morris Water Maze; retina function by electroretinogram (ERG); and PL composition with nuclear magnetic resonance spectroscopy. RESULTS: Cho and PC supplementation enhanced cued learning and spatial memory abilities, respectively (p Def > PC > Cho, with no statistically significant alterations in cone-driven responses. There were no differences in the composition of major PLs in the brain and retina. In the brain, subclasses of ether PL, alkyl acyl- phosphatidylethanolamine (PEaa) and phosphatidylcholine (PCaa) were significantly greater among the PC supplemented group in comparison to the Def group. DISCUSSION: These results indicate that while choline supplementation during gestation to an early developmental period is beneficial for spatial memory, contributions to retina function are minor. Assessment with a larger sample size of retinas could warrant the essentiality of choline for retina development.

Sex-Specific Effects of Chronic Creatine Supplementation on Hippocampal-Mediated Spatial Cognition in the 3xTg Mouse Model of Alzheimer's Disease.

The creatine (Cr) energy system has been implicated in Alzheimer's disease (AD), including reductions in brain phosphoCr and Cr kinase, yet no studies have examined the neurobehavioral effects of Cr supplementation in AD, including the 3xTg mouse model. This studied investigated the effects of Cr supplementation on spatial cognition, plasticity- and disease-related protein levels, and mitochondrial function in the 3xTg hippocampus. Here, 3xTg mice were fed a control or Cr-supplemented (3% Cr (w/w)) diet for 8-9 weeks and tested in the Morris water maze. Mitochondrial oxygen consumption (Seahorse) and protein levels (Western blots) were measured in the hippocampus in subsets of mice. Overall, 3xTg females exhibited impaired memory as compared to males. In females, Cr supplementation decreased escape latency and was associated with increased spatial search strategy use. In males, Cr supplementation decreased the use of spatial search strategies. Pilot data indicated mitochondrial enhancements with Cr supplementation in both sexes. In females, Cr supplementation increased CREB phosphorylation and levels of IκB (NF-κB suppressor), CaMKII, PSD-95, and high-molecular-weight amyloid ß (Aß) species, whereas Aß trimers were reduced. These data suggest a beneficial preventative effect of Cr supplementation in females and warrant caution against Cr supplementation in males in the AD-like brain.

Early Onset of Sex-Dependent Mitochondrial Deficits in the Cortex of 3xTg Alzheimer's Mice.

Alzheimer's disease (AD) is a major public health concern worldwide. Advanced age and female sex are two of the most prominent risk factors for AD. AD is characterized by progressive neuronal loss, especially in the cortex and hippocampus, and mitochondrial dysfunction has been proposed to be an early event in the onset and progression of the disease. Our results showed early perturbations in mitochondrial function in 3xTg mouse brain, with the cortex being more susceptible to mitochondrial changes than the hippocampus. In the cortex of 3xTg females, decreased coupled and uncoupled respiration were evident early (at 2 months of age), while in males it appeared later at 6 months of age. We observed increased coupled respiration in the hippocampus of 2-month-old 3xTg females, but no changes were detected later in life. Changes in mitochondrial dynamics were indicated by decreased mitofusin (Mfn2) and increased dynamin related protein 1 (Drp1) (only in females) in the hippocampus and cortex of 3xTg mice. Our findings highlight the importance of controlling and accounting for sex, brain region, and age in studies examining brain bioenergetics using this common AD model in order to more accurately evaluate potential therapies and improve the sex-specific translatability of preclinical findings.

Chronic dietary creatine enhances hippocampal-dependent spatial memory, bioenergetics, and levels of plasticity-related proteins associated with NF-κB.

The brain has a high demand for energy, of which creatine (Cr) is an important regulator. Studies document neurocognitive benefits of oral Cr in mammals, yet little is known regarding their physiological basis. This study investigated the effects of Cr supplementation (3%, w/w) on hippocampal function in male C57BL/6 mice, including spatial learning and memory in the Morris water maze and oxygen consumption rates from isolated mitochondria in real time. Levels of transcription factors and related proteins (CREB, Egr1, and IκB to indicate NF-κB activity), proteins implicated in cognition (CaMKII, PSD-95, and Egr2), and mitochondrial proteins (electron transport chain Complex I, mitochondrial fission protein Drp1) were probed with Western blotting. Dietary Cr decreased escape latency/time to locate the platform (P < 0.05) and increased the time spent in the target quadrant (P < 0.01) in the Morris water maze. This was accompanied by increased coupled respiration (P < 0.05) in isolated hippocampal mitochondria. Protein levels of CaMKII, PSD-95, and Complex 1 were increased in Cr-fed mice, whereas IκB was decreased. These data demonstrate that dietary supplementation with Cr can improve learning, memory, and mitochondrial function and have important implications for the treatment of diseases affecting memory and energy homeostasis.

Mechanisms of Mitochondrial Dysfunction in Alzheimer's Disease.

Mitochondria are the primary source for energy generation in the cell, which manifests itself in the form of the adenosine triphosphate (ATP). Nicotinamide dinucleotide (NADH) molecules are the first to enter the so-called electron transport chain or ETC of the mitochondria. The ETC represents a chain of reducing agents organized into four major protein-metal complexes (I-IV) that utilize the flow of electrons to drive the production of ATP. An additional integral protein that is related to oxidative phosphorylation is ATP synthase, referred to as complex V. Complex V carries out ATP synthesis as a result of the electron flow through the ETC. The coupling of electron flow from NADH to molecular oxygen to the production of ATP represents a process known as oxidative phosphorylation. In this review, we describe mainly the bioenergetic properties of mitochondria, such as those found in the ETC that may be altered in Alzheimer's disease (AD). Increasing evidence points to several mitochondrial functions that are affected in AD. Furthermore, it is becoming apparent that mitochondria are a potential target for treatment in early-stage AD. With growing interest in the mitochondria as a target for AD, it has been hypothesized that deficit in this organelle may be at the heart of the progression of AD itself. The role of mitochondria in AD may be significant and is emerging as a main area of AD research.

Early growth response 2 (Egr-2) expression is triggered by NF-κB activation.

Transcription factors are known to play multiple roles in cellular function. Investigators report that factors such as early growth response (Egr) protein and nuclear factor kappa B (NF-κB) are activated in the brain during cancer, brain injury, inflammation, and/or memory. To explore NF-κB activity further, we investigated the transcriptomes of hippocampal slices following electrical stimulation of NF-κB p50 subunit knockout mice (p50-/-) versus their controls (p50+/+). We found that the early growth response gene Egr-2 was upregulated by NF-κB activation, but only in p50+/+ hippocampal slices. We then stimulated HeLa cells and primary cortical neurons with tumor necrosis factor alpha (TNFα) to activate NF-κB and increase the expression of Egr-2. The Egr-2 promoter sequence was analyzed for NF-κB binding sites and chromatin immunoprecipitation (ChIP) assays were performed to confirm promoter occupancy in vivo. We discovered that NF-κB specifically binds to an NF-κB consensus binding site within the proximal promoter region of Egr-2. Luciferase assay demonstrated that p50 was able to transactivate the Egr-2 promoter in vitro. Small interfering RNA (siRNA)-mediated p50 knockdown corroborated other Egr-2 expression studies. We show for the first time a novel link between NF-κB activation and Egr-2 expression with Egr-2 expression directly controlled by the transcriptional activity of NF-κB.