MicroRNAs miR-629-3p, miR-1202 and miR-1225-5p as potential diagnostic and surgery outcome biomarkers for mesial temporal lobe epilepsy with hippocampal sclerosis.

BACKGROUND: Mesial temporal lobe epilepsy (MTLE) is a symptomatic epilepsy syndrome clinically characterized by high prevalence, pharmacoresistance, good surgical prognosis and hippocampal sclerosis (HS); however, no singular criteria can be considered sufficient for the MTLE-HS diagnosis. MicroRNAs (miRNAs) are small non-coding molecules that act as important gene-expression regulators at post-transcriptional level. Evidences on the involvement of miRNAs in epilepsy pathogenesis as well as their potential to be employed as biomarkers claim for investigations on miRNAs' applicability as epilepsy diagnosis and prognosis biomarkers. Consequently, the present study aimed to evaluate the applicability of three specific miRNAs as biomarkers of diagnosis and surgical outcomes in adult patients with MTLE-HS. METHOD: Hippocampus, amygdala and blood samples from 20 patients with MTLE-HS were analyzed, 10 with favorable surgical prognosis (Engel I) and 10 with unfavorable surgical prognosis (Engel III-IV). For the control groups, hippocampus and amygdala from necropsy and blood samples from healthy individuals were adopted. The miRNAs expression analysis was performed using Real-Time Quantitative Polymerase Chain Reaction for miRNAs highlighted from microarray as being involved in GABAergic neurotransmission. RESULTS: The miRNAs miR-629-3p, miR-1202 and miR-1225-5p were found to be hyper-expressed in MTLE-HS patients' blood. CONCLUSIONS: Our data suggest the existence of three circulating miRNAs (miR-629-3p, miR-1202 and miR-1225-5p) that could possibly act as additional tools in the set of factors that contribute to MTLE-HS diagnose.

Carotid jugular fistula after Le Fort I osteotomy.

Le Fort I osteotomy is the technique most applied worldwide in the treatment of dentoskeletal deformity involving the maxilla. Even though it is considered a very safe technique with good intra- and postoperative results, many complications have been described. This paper presents a case of carotid jugular fistula developed in a 22-year-old white male submitted to Le Fort I osteotomy for the treatment of anteroposterior maxillary deficiency, and discusses the possible aetiology and management of this serious complication.

Modelling zinc changes at the hippocampal mossy fiber synaptic cleft.

Zinc, a transition metal existing in very high concentrations in the hippocampal mossy fibers from CA3 area, is assumed to be co-released with glutamate and to have a neuromodulatory role at the corresponding synapses. The synaptic action of zinc is determined both by the spatiotemporal characteristics of the zinc release process and by the kinetics of zinc binding to sites located in the cleft area, as well as by their concentrations. This work addresses total, free and complexed zinc concentration changes, in an individual synaptic cleft, following single, short and long periods of evoked zinc release. The results estimate the magnitude and time course of the concentrations of zinc complexes, assuming that the dynamics of the release processes are similar to those of glutamate. It is also considered that, for the cleft zinc concentrations used in the model (≤ 1 µM), there is no postsynaptic zinc entry. For this reason, all released zinc ends up being reuptaken in a process that is several orders of magnitude slower than that of release and has thus a much smaller amplitude. The time derivative of the total zinc concentration in the cleft is represented by the difference between two alpha functions, corresponding to the released and uptaken components. These include specific parameters that were chosen assuming zinc and glutamate co-release, with similar time courses. The peak amplitudes of free zinc in the cleft were selected based on previously reported experimental cleft zinc concentration changes evoked by single and multiple stimulation protocols. The results suggest that following a low amount of zinc release, similar to that associated with one or a few stimuli, zinc clearance is mainly mediated by zinc binding to the high-affinity sites on the NMDA receptors and to the low-affinity sites on the highly abundant GLAST glutamate transporters. In the case of higher zinc release brought about by a larger group of stimuli, most zinc binding occurs essentially to the GLAST transporters, having the corresponding zinc complex a maximum concentration that is more than one order of magnitude larger than that for the high and low affinity NMDA sites. The other zinc complexes considered in the model, namely those formed with sites on the AMPA receptors, calcium and KATP channels and with ATP molecules, have much smaller contributions to the synaptic zinc clearance.

Blockade of presynaptic K ATP channels reduces the zinc-mediated posttetanic depression at hippocampal mossy fiber synapses.

Zinc is one of the most abundant transition metals in the brain, being present in a variety of synaptic processes. The mossy fiber terminals in area CA3 of the hippocampus contain large amounts of vesicular zinc and have an extremely high density of ATP-sensitive potassium (KATP) channels. The activation of these channels by zinc leads to rapid hyperpolarization of these presynaptic terminals and inhibition of transmitter release. It has been previously shown that intense stimulation of the synapses between mossy fibers and CA3 pyramidal cells evokes a posttetanic depression of synaptic activity, accompanied by a decrease in presynaptic calcium and vesicular zinc signals. These results suggest a neuromodulatory role for zinc at these synapses, which could be mediated by inhibition of presynaptic voltage-dependent calcium channels (VDCCs) and/or activation of presynaptic KATP channels. In order to evaluate the contribution of the second mechanism we have applied multiple tetanic stimulations in the absence and presence of the KATP channel blocker tolbutamide. Under control conditions, it was observed that the delivery of six tetani caused a posttetanic depression of synaptic activity. In the presence of tolbutamide, the depression was smaller and had a shorter time course. A similar depression was also observed in the presynaptic zinc and calcium signals. These findings are in agreement with the hypothesis that the activation of KATP channels by tetanically released zinc leads to cell hyperpolarization and subsequent reduction of presynaptic calcium entry, followed by the inhibition of both zinc and glutamate release. Thus, these results suggest that the inhibition of mossy fiber synaptic transmission by intensely released zinc is partially mediated by the activation of KATP channels.

Tetanically released zinc inhibits hippocampal mossy fiber calcium, zinc and synaptic responses.

At the zinc-enriched mossy fiber synapses from hippocampal CA3 area, electrical or chemical stimulation evokes zinc release from glutamatergic synaptic vesicles that may cause different pre- or postsynaptic actions. Besides zinc that can be co-localized with glutamate and GABA, the mossy fibers contain a very high density of ATP-sensitive potassium channels that are activated by zinc. We have investigated the possibility that intensely released zinc inhibits presynaptic calcium changes and consequently zinc and glutamate release. The studies were made combining optical recording of fast presynaptic calcium and zinc signals, using the fluorescent indicators Fura-2 and N-(6-methoxy-8-quinolyl)-para-toluenesulfonamide, respectively, with measurements of field potentials. We have observed that strong tetanic stimulation caused posttetanic depressions of electrically induced presynaptic calcium and zinc signals and of synaptic responses, the depressions being blocked by zinc chelators. These results suggest that endogenously released zinc has an inhibitory role, mediated by presynaptic ATP-sensitive potassium channels and/or presynaptic calcium channels, that leads to the depression of zinc and glutamate release.

Measurement of presynaptic zinc changes in hippocampal mossy fibers.

The hippocampal mossy fiber terminals of CA3 area contain high levels of vesicular zinc that is released in a calcium-dependent way, following high-frequency stimulation. However the properties of zinc release during normal synaptic transmission, paired-pulse facilitation and mossy fiber long-term potentiation are still unknown. Using the fluorescent zinc probe N-(6-methoxy-8-quinolyl)-para-toluenesulfonamide, we measured fast mossy fiber zinc changes indicating that zinc is released following single and low levels of electrical stimulation. The observed presynaptic zinc signals are maintained during the expression of mossy fiber long-term potentiation, assumed to be mediated by an increase in transmitter release, and are enhanced during paired-pulse facilitation. This zinc enhancement is, like paired-pulse facilitation, reduced during established long-term potentiation. The correlation between the paired-pulse evoked zinc and field potential responses supports the idea that zinc is co-released with glutamate.

Hippocampal mossy fiber calcium transients are maintained during long-term potentiation and are inhibited by endogenous zinc.

The hippocampal mossy fiber long-term potentiation (LTP) is an N-methyl-d-aspartate (NMDA) receptor-independent form of long-lasting synaptic plasticity characteristic of the zinc-enriched mossy fiber synapses. Its expression is generally considered to have a presynaptic locus and to be mediated by a persistent increase of evoked transmitter release. Because the release process is calcium-dependent, the observed increase in synaptic efficacy could be due to a persistent modification of presynaptic calcium mechanisms, triggered by the large calcium influx associated with long-term potentiation induction. Alternatively, it might be caused by an enhancement in the sensitivity to calcium of some components of the synaptic vesicle release system, following the large intraterminal calcium accumulation. We investigated the first hypothesis by measuring presynaptic Fura-2 calcium signals associated with electrically induced mossy fiber long-term potentiation. We have observed that like residual calcium, single presynaptic calcium changes are not enhanced during the maintenance phase of mossy fiber long-term potentiation. This result supports the idea that this form of long-term potentiation may be mediated by persistent changes of some process occurring after calcium entry. It has been established that voltage-dependent calcium channels are inhibited by zinc and that endogenous zinc is released in a calcium-dependent way following intense mossy fiber activation. Because there is evidence that at these synapses zinc is also released following single electrical stimulation, we investigated the effect of endogenous zinc on single presynaptic calcium signals and on field potentials associated with mossy fiber LTP. We have observed that this form of LTP could be induced in the presence of the permeant heavy metal chelator N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) and that application of this chelator, during LTP, caused an enhancement of the presynaptic calcium signals without affecting synaptic transmission. This enhancement is consistent with the idea that mossy fiber zinc, released following individual stimuli, inhibits presynaptic calcium mechanisms.