Neurounina-1, a Novel Compound That Increases Na⁺/Ca²⁺Exchanger Activity, Effectively Protects against Stroke Damage

Abstract

Previous studies have demonstrated that the knockdown or knockout of the three Na 1 /Ca 21 exchanger (NCX) isoforms, NCX1, NCX2, and NCX3, worsens ischemic brain damage. This suggests that the activation of these antiporters exerts a neuroprotective action against stroke damage. However, drugs able to increase the activity of NCXs are not yet available. We have here succeeded in synthesizing a new compound, named neurounina-1 (7-nitro-5-, provided with an high lipophilicity index and able to increase NCX activity. Ca 21 radiotracer, Fura-2 microfluorimetry, and patchclamp techniques revealed that neurounina-1 stimulated NCX1 and NCX2 activities with an EC 50 in the picomolar to low nanomolar range, whereas it did not affect NCX3 activity. Furthermore, by using chimera strategy and site-directed mutagenesis, three specific molecular determinants of NCX1 responsible for neurounina-1 activity were identified in the a-repeats. Interestingly, NCX3 became responsive to neurounina-1 when both a-repeats were replaced with the corresponding regions of NCX1. In vitro studies showed that 10 nM neurounina-1 reduced cell death of primary cortical neurons exposed to oxygen-glucose deprivation followed by reoxygenation. Moreover, in vitro, neurounina-1 also reduced gaminobutyric acid (GABA) release, enhanced GABA A currents, and inhibited both glutamate release and N-methyl-D-aspartate receptors. More important, neurounina-1 proved to have a wide therapeutic window in vivo. Indeed, when administered at doses of 0.003 to 30 mg/kg i.p., it was able to reduce the infarct volume of mice subjected to transient middle cerebral artery occlusion even up to 3 to 5 hours after stroke onset. Collectively, the present study shows that neurounina-1 exerts a remarkable neuroprotective effect during stroke and increases NCX1 and NCX2 activities.

Neuronal NCX1 overexpression induces stroke resistance while knockout induces vulnerability via Akt

“…Indeed, the compound developed by our research group that enhances both NCX1 and NCX2 activity, neurounina-1, exerts a neuroprotective effect in stroke with a good potential time-window of therapeutic intervention. 17 Lastly, the generation of these two genetically modified mice, besides having firmly established a role for NCX1 in ischemic stroke, will allow to explore more deeply the role of NCX1 in other physiological and pathophysiological conditions of the central nervous system.…”

“…Furthermore, a lack of highly selective compounds modulating NCX1 activity [14][15][16][17][18] has generated an array of conflicting results on the role of this antiporter in stroke. 6,[19][20][21] We hypothesized that NCX1 might play a relevant role in neuroprotection during brain ischemia since: (1) an increase in NCX1 expression/activity raises Akt phosphorylation, 22 a well-known signaling of neuronal survival [23][24][25] that increases neuronal resistance to brain ischemia; and (2) under anoxic conditions, NCX1 could work in the reverse mode, promoting Ca 2þ -influx and favoring Ca 2þ -refilling into the endoplasmic reticulum (ER), which is depleted under these conditions, thus allowing neurons to delay apoptosis.…”

Emerging Roles of microRNAs in Ischemic Stroke: As Possible Therapeutic Agents

“…Interestingly, the sodium-calcium exchanger-1 (NCX1) gene expression is influenced by cerebral ischemia, which is a plasma membrane transporter that regulates cellular calcium and sodium homeostasis in the brain [185-187]. NCX activation ameliorates the consequences of ischemic brain damage [188]. So, it has been showed that anti-miR-103-1 exerts a strong neuroprotective effect against ischemic damage through NCX1 activation and offers the opportunity to develop a new therapeutic strategy for ischemic stroke [189].…”

Ionic Transporter Activity in Astrocytes, Microglia, and Oligodendrocytes During Brain Ischemia

“…In particular, NCX1 and NCX3 downregulation or genetic ablation worsens ischemic damage in mice and rats, [56][57][58] whereas its pharmacological activation determines a reduction in the brain infarct volume. 59 Na þ /Ca 2 þ Exchanger in Astrocytes. Na þ /Ca 2 þ exchanger 1 represents the most expressed NCX isoform in astrocytes, 60 where when operating in the reverse mode, it can deliver Ca 2 þ to the cytosol.…”