Molecular Neurobiology

GAP-43 is a presynaptic membrane phosphoprotein that has been implicated in both the development and the modulation of neural connections. The availability of cDNA clones for GAP-43 makes it possible to examine with greater precision its role in neuronal outgrowth and physiology. We used Northern blots andin situ hybridization with GAP-43 antisense RNA probes to show that GAP-43 is expressed selectively in associative regions of the adult brain. Immunocytochemical analyses showed alterations in the pattern of GAP-43 expression in the hippocampus during reactive synaptogenesis following lesions of the perforant pathway. Genetic intervention methodology was used to analyze the molecular nature of GAP-43 involvement in synaptic plasticity. GAP-43-transfected PC12 cells displayed an enhanced response to nerve growth factor, suggesting that GAP-43 may be directly involved in neurite extension and in the modulation of the neuronal response to extrinsic trophic factors. Studies of PC12 cell transfectants, in which the synthesis of GAP-43 was blocked by expression of GAP-43 antisense RNA, showed that evoked dopamine release was significantly attenuated in these cells. The use of gene transfer into neurons with the HSV-1 vector is presented as a method of analyzing the interaction of GAP-43 with signal transduction systems during neurotransmitter release.

The effects of the consumption of high-fat diets (HFD) have been studied to unravel the molecular pathways they are altering in order to understand the link between increased caloric intake, metabolic diseases, and the risk of cognitive dysfunction. The saturated fatty acid, palmitic acid (PA), is the main component of HFD and it has been found increased in the circulation of obese and diabetic people. In the central nervous system, PA has been associated with inflammatory responses in astrocytes, but the effects on neurons exposed to it have not been largely investigated. Given that PA affects a variety of metabolic pathways, we aimed to analyze the transcriptomic profile activated by this fatty acid to shed light on the mechanisms of neuronal dysfunction. In the current study, we profiled the transcriptome response after PA exposition at non-toxic doses in primary hippocampal neurons. Gene ontology and Reactome pathway analysis revealed a pattern of gene expression which is associated with inflammatory pathways, and importantly, with the activation of lipid metabolism that is considered not very active in neurons. Validation by quantitative RT-PCR (qRT-PCR) of Hmgcs2, Angptl4, Ugt8, and Rnf145 support the results obtained by RNAseq. Overall, these findings suggest that neurons are able to respond to saturated fatty acids changing the expression pattern of genes associated with inflammatory response and lipid utilization that may be involved in the neuronal damage associated with metabolic diseases.

Acute inflammation is important for tissue repair; however, chronic inflammation contributes to neurodegeneration in Alzheimer's disease (AD) and occurs when glial cells undergo prolonged activation. In the brain, stress or damage causes the release of nucleotides and activation of the Gq protein-coupled P2Y2 nucleotide receptor subtype (P2Y2R) leading to pro-inflammatory responses that can protect neurons from injury, including the stimulation and recruitment of glial cells. P2Y2R activation induces the phosphorylation of the epidermal growth factor receptor (EGFR), a response dependent upon the presence of a SH3 binding domain in the intracellular C terminus of the P2Y2R that promotes Src binding and transactivation of EGFR, a pathway that regulates the proliferation of cortical astrocytes. Other studies indicate that P2Y2R activation increases astrocyte migration. P2Y2R activation by UTP increases the expression in astrocytes of αVβ3/5 integrins that bind directly to the P2Y2R via an Arg-Gly-Asp (RGD) motif in the first extracellular loop of the P2Y2R, an interaction required for Go and G12 protein-dependent astrocyte migration. In rat primary cortical neurons (rPCNs) P2Y2R expression is increased by stimulation with interleukin-1β (IL-1β), a pro-inflammatory cytokine whose levels are elevated in AD, in part due to nucleotide-stimulated release from glial cells. Other results indicate that oligomeric β-amyloid peptide (Aβ1-42), a contributor to AD, increases nucleotide release from astrocytes, which would serve to activate upregulated P2Y2Rs in neurons. Data with rPCNs suggest that P2Y2R upregulation by IL-1β and subsequent activation by UTP are neuroprotective, since this increases the non-amyloidogenic cleavage of amyloid precursor protein. Furthermore, activation of IL-1β-upregulated P2Y2Rs in rPCNs increases the phosphorylation of cofilin, a cytoskeletal protein that stabilizes neurite outgrowths. Thus, activation of pro-inflammatory P2Y2Rs in glial cells can promote neuroprotective responses, suggesting that P2Y2Rs represent a novel pharmacological target in neurodegenerative and other pro-inflammatory diseases.

Circadian clocks synchronize the physiology and behavior of most animals with the day to night cycle. A fundamental property of the molecular pacemakers generating circadian rhythms is their self-sustained nature: they keep oscillating even under constant conditions, with a period close to, but not exactly, 24 h. However, circadian pacemakers have to be sensitive to environmental cues to be beneficial. They need to be reset every day to keep a proper phase relationship with the day to night cycle, and they have to be able to adjust to seasonal changes in day length and temperature. Here, we review our current knowledge of the molecular and neural mechanisms contributing to the plasticity of Drosophila circadian rhythms, which are proving to be remarkably sophisticated and complex.

Pigment-dispèrsing factor (PDF) neuropeptide is an important neurochemical that carries circadian timing information originating from the central oscillator in Drosophila. Several core-clock factors function as upstream pdf regulators; the dClock and cycle genes control pdf transcription, whereas the period and timeless genes regulate post-translational processes of PDF via unknown mechanisms. For a downstream neural path, PDF most likely acts as a local modulator, which binds to its receptors that are possibly linked to Ras/MAPK signaling pathways. PDF receptor-containing cells seem to localize in the vicinity of nerve terminals from pace-making neurons. Although PDF is likely to be a principal clock-output factor, our recent evidence predicts the presence of other neuropeptides with rhythm-relevant functions. Furthermore, recent microarray screens have identified numerous potential clock-controlled genes, suggesting that diverse physiological processes might be affected by the biological clock system.

Oxidative stress increases serum thioredoxin (TRX), a redox-regulating protein with antioxidant activity recognized as an oxidative stress marker. The aim of this study was to assess the clinical significance of serum TRX levels in Chinese patients with acute ischemic stroke (AIS). From January 1, 2012, to December 31, 2013, all patients with first-ever acute ischemic stroke were recruited to participate in the study. Serum levels of TRX were assayed with solid-phase sandwich enzyme-linked immunosorbent assay (ELISA), and the severity of stroke was evaluated with the National Institutes of Health Stroke Scale (NIHSS) score on admission. The results indicated that the median serum TRX levels were significantly (P < 0.0001) higher in stroke patients as compared to normal cases [15.03 ng/mL (interquartile range (IQR), 10.21–32.42) and 8.95 ng/mL (6.79–11.05), respectively]. We found the serum TRX reflected the disease severity of AIS. There was a significant positive association between serum TRX levels and NIHSS scores (r = 0.476, P < 0.0001). After adjusting for all other possible covariates, TRX remained as an independent marker of AIS with an adjusted OR of 1.245 (95 % confidence interval (CI), 1.164–1.352; P < 0.0001). Based on the receiver operating characteristic (ROC) curve, the optimal cutoff value of serum TRX levels as an indicator for auxiliary diagnosis of AIS was projected to be 11.0 ng/mL, which yielded a sensitivity of 80.3 % and a specificity of 73.7 %, with the area under the curve at 0.807 (95 % CI, 0.766–0.847). Further, in our study, we found that an increased risk of AIS was associated with serum TRX levels ≥11.0 ng/mL (adjusted OR 6.99; 95 % CI, 2.87–12.87) after adjusting for possible confounders. Our study demonstrated that serum TRX levels at admission were associated with stroke severity and lesion volumes. Elevated levels could be considered as a novel, independent diagnosis marker of AIS in a Chinese sample.

Different factors may contribute to the development of neurodegenerative diseases. Among them, metabolic syndrome (MS), which has reached epidemic proportions, has emerged as a potential element that may be involved in neurodegeneration. Furthermore, studies have shown the importance of the sirtuin family in neuronal survival and MS, which opens the possibility of new pharmacological targets. This study investigates the influence of sirtuin metabolic pathways by examining the functional capacities of glucose-induced obesity in an excitotoxic state induced by a quinolinic acid (QA) animal model. Mice were divided into two groups that received different diets for 8 weeks: one group received a regular diet, and the other group received a high-fat diet (HF) to induce MS. The animals were submitted to a stereotaxic surgery and subdivided into four groups: Standard (ST), Standard-QA (ST-QA), HF and HF-QA. The QA groups were given a 250 nL quinolinic acid injection in the right striatum and PBS was injected in the other groups. Obese mice presented with a weight gain of 40 % more than the ST group beyond acquiring an insulin resistance. QA induced motor impairment and neurodegeneration in both ST-QA and HF-QA, although no difference was observed between these groups. The HF-QA group showed a reduction in adiposity when compared with the groups that received PBS. Therefore, the HF-QA group demonstrated a commitment-dependent metabolic pathway. The results suggest that an obesogenic diet does not aggravate the neurodegeneration induced by QA. However, the excitotoxicity induced by QA promotes a sirtuin pathway impairment that contributes to metabolic changes.

The major genetic risk factor for sporadic Alzheimer’s disease (AD) is the lipid binding and transporting carrier protein apolipoprotein E, epsilon 4 allele (ApoE4). One of the unsolved mysteries of AD is how the presence of ApoE4 elicits this age-associated, currently incurable neurodegenerative disease. Recently, we showed that ApoE4 acts as a transcription factor and binds to the promoters of genes involved in a range of processes linked to aging and AD disease pathogenesis. These findings point to novel therapeutic strategies for AD and aging, resulting in an extension of human healthspan, the disease-free and functional period of life. Here, we review the effects and implications of the putative transcriptional role of ApoE4 and propose a model of Alzheimer’s disease that focuses on the transcriptional nature of ApoE4 and its downstream effects, with the aim that this knowledge will help to define the role ApoE4 plays as a risk factor for AD, aging, and other processes such as inflammation and cardiovascular disease.