Na+/Ca2+ exchanger isoform 1 takes part to the Ca2+-related prosurvival pathway of SOD1 in primary motor neurons exposed to beta-methylamino-l-alanine

Background: The cycad neurotoxin beta-methylamino-l-alanine (L-BMAA), one of the environmental trigger factor for amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar Ca2+ homeostasis. Through the activation of Akt/ERK1/2 pathway, the Cu,Zn-superoxide dismutase (SOD1) and its non-metallated form, ApoSOD1, prevent endoplasmic reticulum (ER) stress-induced cell death in motor neurons exposed to L-BMAA. This occurs through the rapid increase of intracellular Ca2+ concentration ([Ca2+]i) in part flowing from the extracellular compartment and in part released from ER. However, the molecular components of this mechanism remain uncharacterized. Methods: By an integrated approach consisting on the use of siRNA strategy, Western blotting, confocal double- labeling immunofluorescence, patch-clamp electrophysiology, and Fura 2-/SBFI-single-cell imaging, we explored in rat motor neuron-enriched cultures the involvement of the plasma membrane proteins Na+/Ca2+ exchanger (NCX) and purinergic P2X7 receptor as well as that of the intracellular cADP-ribose (cADPR) pathway, in the neuroprotective mechanism of SOD1. Results: We showed that SOD1-induced [Ca2+]i rise was prevented neither by A430879, a P2X7 receptor specific antagonist or 8-bromo-cADPR, a cell permeant antagonist of cADP-ribose, but only by the pan inhibitor of NCX, CB-DMB. The same occurred for the ApoSOD1. Confocal double labeling immunofluorescence showed a huge expression of plasmalemmal NCX1 and intracellular NCX3 isoforms. Furthermore, we identified NCX1 reverse mode as the main mechanism responsible for the neuroprotective ER Ca2+ refilling elicited by SOD1 and ApoSOD1 through which they promoted translocation of active Akt in the nuclei of a subset of primary motor neurons. Finally, the activation of NCX1 by the specific agonist CN-PYB2 protected motor neurons from L-BMAA-induced cell death, mimicking the effect of SOD1. Conclusion: Collectively, our data indicate that SOD1 and ApoSOD1 exert their neuroprotective effect by modulating ER Ca2+ content through the activation of NCX1 reverse mode and Akt nuclear translocation in a subset of primary motor neurons. [MediaObject not available: see fulltext.

The Na+/Ca2+ Exchanger 3 Is Functionally Coupled With the NaV1.6 Voltage-Gated Channel and Promotes an Endoplasmic Reticulum Ca2+ Refilling in a Transgenic Model of Alzheimer’s Disease

The remodelling of neuronal ionic homeostasis by altered channels and transporters is a critical feature of the Alzheimer’s disease (AD) pathogenesis. Different reports converge on the concept that the Na+/Ca2+ exchanger (NCX), as one of the main regulators of Na+ and Ca2+ concentrations and signalling, could exert a neuroprotective role in AD. The activity of NCX has been found to be increased in AD brains, where it seemed to correlate with an increased neuronal survival. Moreover, the enhancement of the NCX3 currents (INCX) in primary neurons treated with the neurotoxic amyloid β 1–42 (Aβ1–42) oligomers prevented the endoplasmic reticulum (ER) stress and neuronal death. The present study has been designed to investigate any possible modulation of the INCX, the functional interaction between NCX and the NaV1.6 channel, and their impact on the Ca2+ homeostasis in a transgenic in vitro model of AD, the primary hippocampal neurons from the Tg2576 mouse, which overproduce the Aβ1–42 peptide. Electrophysiological studies, carried in the presence of siRNA and the isoform-selective NCX inhibitor KB-R7943, showed that the activity of a specific NCX isoform, NCX3, was upregulated in its reverse, Ca2+ influx mode of operation in the Tg2576 neurons. The enhanced NCX activity contributed, in turn, to increase the ER Ca2+ content, without affecting the cytosolic Ca2+ concentrations of the Tg2576 neurons. Interestingly, our experiments have also uncovered a functional coupling between NCX3 and the voltage-gated NaV1.6 channels. In particular, the increased NaV1.6 currents appeared to be responsible for the upregulation of the reverse mode of NCX3, since both TTX and the Streptomyces griseolus antibiotic anisomycin, by reducing the NaV1.6 currents, counteracted the increase of the INCX in the Tg2576 neurons. In agreement, our immunofluorescence analyses revealed that the NCX3/NaV1.6 co-expression was increased in the Tg2576 hippocampal neurons in comparison with the WT neurons. Collectively, these findings indicate that NCX3 might intervene in the Ca2+ remodelling occurring in the Tg2576 primary neurons thus emerging as a molecular target with a neuroprotective potential, and provide a new outcome of the NaV1.6 upregulation related to the modulation of the intracellular Ca2+ concentrations in AD neurons

Rebound effects of NCX3 pharmacological inhibition: A novel strategy to accelerate myelin formation in oligodendrocytes

The Na+/Ca2+ exchanger NCX3 is an important regulator of sodium and calcium homeostasis in oligodendrocyte lineage. To date, no information is available on the effects resulting from prolonged exposure to NCX3 blockers and subsequent drug washout in oligodendroglia. Here, we investigated, by means of biochemical, morphological and functional analyses, the pharmacological effects of the NCX3 inhibitor, the 5–amino-N-butyl-2–(4–ethoxyphenoxy)-benzamide hydrochloride (BED), on NCXs expression and activity, as well as intracellular [Na+]i and [Ca2+]i levels, during treatment and following drug washout both in human MO3.13 oligodendrocytes and rat primary oligodendrocyte precursor cells (OPCs). BED exposure antagonized NCX activity, induced OPCs proliferation and [Na+]i accumulation. By contrast, 2 days of BED washout after 4 days of treatment significantly upregulated low molecular weight NCX3 proteins, reversed NCX activity, and increased intracellular [Ca2+]i. This BED-free effect was accompanied by an upregulation of NCX3 expression in oligodendrocyte processes and accelerated expression of myelin markers in rat primary oligodendrocytes. Collectively, our findings show that the pharmacological inhibition of the NCX3 exchanger with BED blocker maybe followed by a rebound increase in NCX3 expression and reversal activity that accelerate myelin sheet formation in oligodendrocytes. In addition, they indicate that a particular attention should be paid to the use of NCX inhibitors for possible rebound effects, and suggest that further studies will be necessary to investigate whether selective pharmacological modulation of NCX3 exchanger may be exploited to benefit demyelination and remyelination in demyelinating diseases

New Insights into the Structure-Activity Relationship and Neuroprotective Profile of Benzodiazepinone Derivatives of Neurounina-1 as Modulators of the Na+/Ca2+Exchanger Isoforms

Due to the neuroprotective role of the Na+/Ca2+ exchanger (NCX) isoforms NCX1 and NCX3, we synthesized novel benzodiazepinone derivatives of the unique NCX activator Neurounina-1, named compounds 1-19. The derivatives are characterized by a benzodiazepinonic nucleus linked to five- or six-membered cyclic amines via a methylene, ethylene, or acetyl spacer. The compounds have been screened on NCX1/NCX3 isoform activities by a high-throughput screening approach, and the most promising were characterized by patch-clamp electrophysiology and Fura-2AM video imaging. We identified two novel modulators of NCX: compound 4, inhibiting NCX1 reverse mode, and compound 14, enhancing NCX1 and NCX3 activity. Compound 1 displayed neuroprotection in two preclinical models of brain ischemia. The analysis of the conformational and steric features led to the identification of the molecular volume required for selective NCX1 activation for mixed NCX1/NCX3 activation or for NCX1 inhibition, providing the first prototypal model for the design of optimized isoform modulators

The MASSIMO system for the safeguarding of historic buildings in a seismic area: operationally-oriented platforms

In this paper, the non-invasive system MASSIMO is presented for the monitoring and the seismic vulnerability mitigation of the cultural heritage. It integrates ground-based, airborne and space-borne remote sensing tools with geophysical and in situ surveys to provide the multi-spatial (regional, urban and building scales) and multi-temporal (long-term, short-term, near-real-time and real-time scales) monitoring of test areas and buildings. The measurements are integrated through web-based GIS and 3D visual platforms to support decision-making stakeholders involved in urban planning and structural requalification. An application of this system is presented over the Calabria region for the town of Cosenza and a test historical complex

Una metodologia di indagine basata su tecniche NDT per la mitigazione del rischio sismico di beni storici soggetti a tutela

Sommario In questo lavoro viene presentata una metodologia di indagine NDT (acronimo inglese di Non Destructing Testing), basata su tecniche classiche di misura, telerilevamento prossimale e satellitare ad alta risoluzione spaziale, analisi sismogenetiche ed indagini geofisiche, per la mitigazione del rischio sismico di edifici storici monumentali soggetti a tutela. I risultati sperimentali, ottenuti per il caso di studio del complesso monumentale di Sant’Agostino ubicato nella città di Cosenza, dimostrano l’efficacia dell’approccio proposto sia per applicazioni correnti (il monitoraggio materiale e strutturale di un manufatto), sia per applicazioni emergenti (mitigazione del rischio sismico di beni storici)

Principi di farmacologia clinica, materna e fetale

Lo scopo principale degli autori del presente trattato è stato quello di correlare le conoscenze sempre più approfondite sull’azione molecolare dei farmaci con gli effetti farmacologici esercitati nell’uomo e che costituiscono i presupposti razionali della terapia medica. Uno spazio adeguato è stato inoltre riservato alla trattazione degli aspetti farmacocinetici clinici, delle reazioni avverse e delle interazioni tra i farmaci, che rappresentano un aspetto in continuo divenire e che sono spesso alla base di eventi avversi clinicamente rilevanti.[...] L’ intento del testo è stato quello di fornire un’informazione la più completa possibile con l’inserimento nella trattazione di argomenti che hanno assunto negli ultimi anni particolare rilevanza nel campo delle possibili prospettive terapeutiche più moderne. Nel trattato sono presenti infatti capitoli innovativi quali quello sui Farmaci Biotecnologici a bersaglio definito, sulla Farmacogenetica e sulla Terapia Genica. Completano la trattazione argomenti inerenti gli aspetti socio-sanitari , economici e normativi della Farmacologia contenuti nei capitoli sulla Sperimentazione Clinica dei Farmaci, sulla Farmacoeconomia e sulla Farmacologia del Doping. Infine, in considerazione del largo uso che si fa dei prodotti di origine vegetale , si è ritenuto utile affrontare tale problematica nel capitolo Principi di fitoterapia nella pratica clinic