Carbon-Fiber Microbiosensor for Monitoring Rapid Lactate Fluctuations in Brain Tissue Using Fast-Scan Cyclic Voltammetry.

Recent studies have described a role for lactate in brain energy metabolism and energy formation, challenging the conventional view that glucose is the principle energy source for brain function. To date, lactate dynamics in the brain are largely unknown, limiting insight into function. We addressed this by developing and characterizing a lactate oxidase-modified carbon-fiber microelectrode coupled with fast-scan cyclic voltammetry. This new tool boasts a sensitivity for lactate of 22 ± 1 nA·mM-1 and LOD of 7.0 ± 0.7 µM. The approach has enabled detection of rapid lactate fluctuations with unprecedented spatiotemporal resolution as well as excellent stability, selectivity, and sensitivity. The technology was characterized both in vitro and in vivo at discrete recording sites in rat striatum. We provide evidence that striatal lactate availability increases biphasically in response to electrical stimulation of the dopaminergic midbrain in the anesthetized rat. This new tool for real-time detection of lactate dynamics promises to improve understanding of how lactate availability underscores neuronal function and dysfunction.

Quantitative Comparison of Enzyme Immobilization Strategies for Glucose Biosensing in Real-Time Using Fast-Scan Cyclic Voltammetry Coupled with Carbon-Fiber Microelectrodes.

Electrochemical monitoring of non-electroactive species requires a biosensor that is stable and selective, with sensitivity to physiological concentrations of targeted analytes. We have combined glucose oxidase-modified carbon-fiber microelectrodes with fast-scan cyclic voltammetry for real-time measurements of glucose fluctuations in brain tissue. Work presented herein quantitatively compares three approaches to enzyme immobilization on the microelectrode surface-physical adsorption, hydrogel entrapment, and entrapment in electrospun nanofibers. The data suggest that each of these methods can be used to create functional microbiosensors. Immobilization of glucose oxidase by physical adsorption generates a biosensor with poor sensitivity to glucose and unstable performance. Entrapment of glucose oxidase in poly(vinyl alcohol) nanofibers generates microbiosensors that are effective for glucose measurements over a large linear range, and that may be particularly useful when targeting glucose concentrations in excess of 3 mm, such as in blood. Hydrogel entrapment is the most effective in terms of sensitivity and stability. These microbiosensors can be used for simultaneous monitoring of glucose and dopamine in real time. The findings outlined herein should be applicable to other oxidase enzymes, and thus they are broadly important for the development of new tools for real-time measurements of fluctuating molecules that are not inherently electroactive.

Characterization of Early Alzheimer's Disease-Like Pathological Alterations in Non-Human Primates with Aging: A Pilot Study.

BACKGROUND: Sporadic or late onset Alzheimer's disease (LOAD) is a multifactorial neurodegenerative disease with aging the most known risk factor. Non-human primates (NHPs) may serve as an excellent model to study LOAD because of their close similarity to humans in many aspects including neuroanatomy and neurodevelopment. Recent studies reveal AD-like pathology in old NHPs. OBJECTIVE: In this pilot study, we took advantage of brain samples from 6 Cynomolgus macaques that were divided into two groups: middle aged (average age 14.81 years) and older (average age 19.33 years). We investigated whether AD-like brain pathologies are present in the NHPs. METHODS: We used immunohistochemical method to examine brain Aß pathology and neuron density. We applied biochemical assays to measure tau phosphorylation and multiple signaling pathways indicated in AD. We performed electron microscopy experiments to study alterations of postsynaptic density and mitochondrial morphology in the brain of NHPs. RESULTS: We found multiple AD-like pathological alteration in the prefrontal cortex (but not in the hippocampus) of the older NHPs including tau hyperphosphorylation, increased activity of AMP-activated protein kinase (AMPK), decreased expression of protein phosphatase 2A (PP2A), impairments in mitochondrial morphology, and postsynaptic densities formation. CONCLUSION: These findings may provide insights into the factors contributing to the development of LOAD, particularly during the early stage transitioning from middle to old age. Future endeavors are warranted to elucidate mechanisms underlying the regional (and perhaps cellular) vulnerability with aging and the functional correlation of such pathological changes in NHPs.

Repression of eEF2 kinase improves deficits in novel object recognition memory in aged mice.

The normal aging process is commonly associated with mild cognitive deficits including memory decline. Previous studies indicate a role of dysregulated messenger ribonucleic acid translation capacity in cognitive defects associated with aging and aging-related diseases, including hyperphosphorylation of eukaryotic elongation factor 2 (eEF2). Phosphorylation of eEF2 by the kinase eEF2K inhibits its activity, hindering general protein synthesis. Here, we sought to determine whether cognitive deficits in aged mice can be improved by genetically deleting eEF2K (eEF2K KO) and consequently reduction of eEF2 phosphorylation. We found that suppression of eEF2K prevented aging-related deficits in novel object recognition memory. Interestingly, deletion of eEF2K did not alter overall protein synthesis in the hippocampus. Ultrastructural analysis revealed increase size and larger active zone lengths of postsynaptic densities in the hippocampus of aged eEF2K KO mice. Biochemical assays showed hippocampal eIF2α hyperphosphorylation in aged eEF2K KO mice, indicating inhibition of translation initiation. Our findings may provide insight into mechanistic understanding and thus development of novel therapeutic strategies for aging-related cognitive decline.