Aim: To evaluate the role of swelling-induced activation of volume-regulated anion channels (VRACs) in a neonatal hypoxic-ischemic injury model using the selective VRAC blocker 4-(2-butyl-6,7-dichloro-2-cyclopentyl-indan-1-on5-yl) oxobutyric acid (DCPIB). Methods: Cerebral hypoxic-ischemic injury was induced in 7-day-old mouse pups with Rice-Vannucci method. Prior to the onset of ischemia, the animals were ip administered DCPIB (10 mg/kg). The animals were sacrificed 24 h afterwards, coronal sections of the brains were cut and the areas of infarct were examined using TTC staining and an image-analysis system. Cultured PC12 cells were subjected to oxygen-glucose deprivation (OGD) for 4 h. The cellular viability was assessed using Cell Counting Kit 8. Intracellular chloride concentration [Cl -] i was measured using 6-methoxy-N-ethylquinolinium iodide. Results: DCPIB-treated mice showed a significant reduction in hemispheric corrected infarct volume (26.65%±2.23%) compared to that in vehicle-treated mice (45.52%±1.45%, P<0.001). DCPIB-treated mice also showed better functional recovery as they were more active than vehicle-treated mice at 4 and 24 h post injury. In cultured PC12 cells, DCPIB (10 μmol/L) significantly reduced OGDinduced cell death. Moreover, DCPIB (20 μmol/L) blocked hypotonic-induced decrease in [Cl -] i in PC12 cells of both control and OGD groups. Conclusion:The results further support the pathophysiological role of VRACs in ischemic brain injury, and suggest DCPIB as a potential, easily administrable agent targeting VRACs in the context of perinatal and neonatal hypoxic-ischemic brain injury.

The origin of spheroidal dolomitized burrow from Al-Subiya sabkha in Kuwait was previously described as enigmatic as no evidence of precursor calcium carbonate was found in the siliciclastic sediment. An assumption for the genesis of spheroidal dolomite from the same area was attributed to hydrocarbon seepage but no evidence was provided. Here, we investigated a recently discovered early-middle Miocene coastal mud volcano outcrop in Al-Subiya sabkha where dolomitized burrows and spheroidal dolomite are found in bioturbated marine zones, and associated with traces of salt. Conversely, the continental zone lacks bioturbation features, dolomite and traces of salt, which together contrast with bioturbated rich marine zones. Geochemical signatures of Rare Earth Elements + Yttrium show a true positive Ce anomaly (Ce/Ce* > 1.2) and positive Eu/Eu* anomaly of spheroidal dolomite indicating strictly anoxic conditions, and sulphate reduction to sulphide, respectively. Our results are suggestive of a relationship between dolomite formation and interdependent events of hydrocarbon seepage, flux of hypersaline seawater, bioturbation, and fluid flow in the marine zones of the mud volcano. The bioturbation activity of crustaceans introduced channels/burrows in the sediment–water interface allowing for the mixing of seeped pressurized hydrocarbon-charged fluids, and evaporitic seawater. In the irrigated channels/burrows, the seeped pressurized hydrocarbon-charged fluids were oxidized via microbial consortia of methanotrophic archaea and sulphate-reducing bacteria resulting in elevated alkalinity and saturation index with respect to dolomite, thus providing the preferential geochemical microenvironment for dolomite precipitation in the bioturbated sediment.

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