Proteins related to ictogenesis and seizure clustering in chronic epilepsy.

Seizure clustering is a common phenomenon in epilepsy. Protein expression profiles during a seizure cluster might reflect the pathomechanism underlying ictogenesis. We performed proteomic analyses to identify proteins with a specific temporal expression pattern in cluster phases and to demonstrate their potential pathomechanistic role. Pilocarpine epilepsy model mice with confirmed cluster pattern of spontaneous recurrent seizures by long-term video-electroencpehalography were sacrificed at the onset, peak, or end of a seizure cluster or in the seizure-free period. Proteomic analysis was performed in the hippocampus and the cortex. Differentially expressed proteins (DEPs) were identified and classified according to their temporal expression pattern. Among the five hippocampal (HC)-DEP classes, HC-class 1 (66 DEPs) represented disrupted cell homeostasis due to clustered seizures, HC-class 2 (63 DEPs) cluster-onset downregulated processes, HC-class 3 (42 DEPs) cluster-onset upregulated processes, and HC-class 4 (103 DEPs) consequences of clustered seizures. Especially, DEPs in HC-class 3 were hippocampus-specific and involved in axonogenesis, synaptic vesicle assembly, and neuronal projection, indicating their pathomechanistic roles in ictogenesis. Key proteins in HC-class 3 were highly interconnected and abundantly involved in those biological processes. This study described the seizure cluster-associated spatiotemporal regulation of protein expression. HC-class 3 provides insights regarding ictogenesis-related processes.

Possible epigenetic regulatory effect of dysregulated circular RNAs in Alzheimer's disease model.

As circular RNAs (circRNAs) regulates the effect of micro RNAs (miRNAs), circRNA-miRNA-mRNA network might be implicated in various disease pathogenesis. Therefore, we evaluated the dysregulated circRNAs in the Tg2576 mouse Alzheimer's disease (AD) model, their possible regulatory effects on downstream target mRNAs, and their pathomechanistic role during the disease progression. The microarray-based circRNA expression analysis at seven- and twelve-months of ages (7 M and 12 M) returned 101 dysregulated circRNAs at 7 M (55 up-regulated and 46 down-regulated) and twelve dysregulated circRNAs at 12 M (five up-regulated and seven down-regulated). For each dysregulated circRNA, potential target miRNAs and their downstream target mRNAs were searched. Dysregulation of circRNAs was associated with increased frequency of relevant dysregulation of their downstream target mRNAs. Those differentially expressed circRNA-miRNA-mRNA regulatory network included 2,275 networks (876 for up-regulated circRNAs and 1,399 for down-regulated circRNAs) at 7 M and 38 networks (25 for up-regulated circRNAs and 13 for down-regulated circRNAs) at 12 M. Gene ontology (GO) and pathway analyses demonstrated that the dysregulated mRNAs in those networks represent the AD pathomechanism at each disease stage. We concluded that the dysregulated circRNAs might involve in the AD pathogenesis by modulating disease relevant mRNAs via circRNA-miRNA-mRNA regulatory networks.

Possible epigenetic regulatory effect of dysregulated circular RNAs in epilepsy.

Circular RNAs (circRNAs) involve in the epigenetic regulation and its major mechanism is the sequestration of the target micro RNAs (miRNAs). We hypothesized that circRNAs might be related with the pathophysiology of chronic epilepsy and evaluated the altered circRNA expressions and their possible regulatory effects on their target miRNAs and mRNAs in a mouse epilepsy model. The circRNA expression profile in the hippocampus of the pilocarpine mice was analyzed and compared with control. The correlation between the expression of miRNA binding sites (miRNA response elements, MRE) in the dysregulated circRNAs and the expression of their target miRNAs was evaluated. As miRNAs also inhibit their target mRNAs, circRNA-miRNA-mRNA regulatory network, comprised of dysregulated RNAs that targets one another were searched. For the identified networks, bioinformatics analyses were performed. As the result, Forty-three circRNAs were dysregulated in the hippocampus (up-regulated, 26; down-regulated, 17). The change in the expression of MRE in those circRNAs negatively correlated with the change in the relevant target miRNA expression (r = -0.461, P<0.001), supporting that circRNAs inhibit their target miRNA. 333 dysregulated circRNA-miRNA-mRNA networks were identified. Gene ontology and pathway analyses demonstrated that the up-regulated mRNAs in those networks were closely related to the major processes in epilepsy. Among them, STRING analysis identified 37 key mRNAs with abundant (≥4) interactions with other dysregulated target mRNAs. The dysregulation of the circRNAs which had multiple interactions with key mRNAs were validated by PCR. We concluded that dysregulated circRNAs might have a pathophysiologic role in chronic epilepsy by regulating multiple disease relevant mRNAs via circRNA-miRNA-mRNA interactions.

Experimental Induction of Cerebral Aneurysms by Developmental Low Copper Diet.

Optimal models are needed to understand the pathophysiology of human cerebral aneurysms (CA). We investigated the development of experimental CA by decreasing the activity of lysyl oxidases by dietary copper deficiency from the time of gestation and then augmenting vascular stress by angiotensin II infusion in adulthood. Rats were fed copper-free, low-copper, or normal diets at different time periods from gestation to adulthood. The incidences of CAs were evaluated and autopsies performed to determine the coexistence of cardiovascular diseases. A copper-free diet from gestation was associated with high mortality rates (79.1%) resulting from rupture of ascending aorta aneurysms; a low-copper diet led to acceptable mortality rates (13.6%) and produced CAs and subarachnoid hemorrhage in 46.4% and 3.6% of animals, respectively. Higher proportions of CAs (up to 33.3%) in the rats primed for copper deficiency from gestation ruptured following angiotensin II infusion from adulthood. Gene expression array analyses of the CAs indicated that genes involving extracellular matrix and vascular remodeling were altered in this model. This model enables future research to understand the entire pathogenetic basis of CA development and rupture in association with systemic vasculopathies.

Determination of electron transfer kinetic parameters by fourier transform electrochemical impedance spectroscopic analysis.

A new attempt to obtain electron transfer kinetic parameters at an electrified electrode/electrolyte interface using Fourier transform electrochemical impedance spectroscopic (FTEIS) analyses of small potential step chronoamperometric currents is presented. The kinetic parameters thus obtained allowed mass transport free voltammograms to be constructed in an overpotential region, where the diffusion limits the electron transfer reaction, using the Butler-Volmer (B-V) relation. The B-V voltammograms clearly distinguish electrode reactions that are not much different in their electron transfer kinetic parameters, thus showing very similar normal linear sweep voltammetric (SCV) behaviors. Electrochemical reduction of p-benzoquinone, which displays nearly the same SCV responses at a gold electrode regardless whether the electrode is covered by a thiolated beta-cyclodextrin self-assembled monolayer, was taken as an example for the demonstration. The results show that the two voltametrically similar systems display very different electron transfer characteristics.

Programmed potential sweep voltammetry for lower detection limits.

We report a novel programmed potential sweep voltammetry for a much lower detection limit than those achieved by any other known electroanalyitcal techniques. In this technique, an input waveform is programmed such that the background current would become flat or any other predefined form in the potential region of interest where the peak current arising from the analyte is observed, followed by the amplification of the background subtracted peak current. The current thus obtained showed a much better signal integrity at very low analyte concentrations than those obtained by the traditional linear sweep voltammetric and other related voltammetric techniques. The technique was applied to the analysis of dopamine at a carbon ultramicroelectrode (10-microm diameter). The background-compensated currents showed excellent dynamic linearity for dopamine concentrations of more than 3 orders of magnitudes between 500 pM and 100 nM with an estimated detection limit of 127 pM. This method can provide a convenient way for determining biogenic amines in real time with a much higher sensitivity.

Real-time impedance measurements during electrochemical experiments and their application to aniline oxidation.

Development of an in situ technique for measuring electrochemical impedance spectra in real time during an electrochemical experiment is described. The technique is based on staircase voltammetry with relatively large step heights, in which a series of increasing potential steps are applied to an electrochemical system, and the resulting currents are sampled. The first derivatives of the currents thus obtained are then converted to ac current signals in frequency domain, and impedances are computed from them. To demonstrate the technique as a tool for studying the electrode/electrolyte interface during the electrochemical reaction, we chose an electrochemical oxidation reaction of aniline, whose reaction products have been known to continuously change the electrode surface due to the polymer film growth on its surface, and report a number of observations that would not have been obtained without such in situ experiments. A suggestion is also made on the use of staircase voltammetry for mechanistic studies on complex electrochemical reactions by simply varying the sampling time.

Solvolysis-decomposition of 2-adamantyl chloroformate: evidence for two reaction pathways.

Reaction of 2-adamantyl chloroformate under a variety of solvolytic conditions leads to 2-adamantyl chloride accompanied by solvolysis products, some with and some without retention of the CO(2) unit. For example, in 100% ethanol, only 4.8% 2-adamantyl chloride is formed with the mixed carbonate (88%) being the dominant product, and in 100% 2,2,2-trifluoroethanol, the products are both formed with loss of CO(2), 59% of the chloride and 41% of the ether. With exclusion of the specific rates in 100% and 90% ethanol and methanol, a good Grunwald-Winstein plot against Y(Cl) values (solvent ionizing power) is obtained, with a slope of 0.47 +/- 0.03. The results are compared with those reported earlier for 1-adamantyl chloroformate and isopropyl chloroformate and mechanistic conclusions are drawn.