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Refractory status epilepticus is a clinical emergency associated with high mortality and morbidity. Increasing evidence suggests neuroinflammation contributes to the development of drug-refractoriness during status epilepticus. Here, we have determined the contribution of the ATP-gated P2X7 receptor, previously linked to inflammation and increased hyperexcitability, to drug-refractory status epilepticus and its therapeutic potential.Status epilepticus was induced via a unilateral microinjection of kainic acid into the amygdala in adult mice. Severity of status epilepticus was compared in animals with overexpressing or knock-out of the P2X7 receptor, after inflammatory priming by pre-injection of bacterial lipopolysaccharide (LPS) and in mice treated with P2X7 receptor-targeting and anti-inflammatory drugs.Mice overexpressing P2X7 receptors were unresponsive to several anticonvulsants (lorazepam, midazolam, phenytoin and carbamazepine) during status epilepticus. P2X7 receptor expression increased in microglia during status epilepticus, at times when responses to anticonvulsants were reduced. Overexpression of P2X7 receptors induced a pro-inflammatory phenotype in microglia during status epilepticus and the anti-inflammatory drug minocycline restored normal responses to anticonvulsants in mice overexpressing P2X7 receptors. Pretreatment of wild-type mice with LPS increased P2X7 receptor levels in the brain and reduced responsiveness to anticonvulsants during status epilepticus, which was overcome by either genetic deletion of P2X7 receptors or treatment with the P2X7 receptor antagonists, AFC-5128 or ITH15004.Our results demonstrate that P2X7 receptor-induced pro-inflammatory effects contribute to resistance to pharmacotherapy during status epilepticus. Therapies targeting P2X7 receptors could be novel adjunctive treatments for drug-refractory status epilepticus.

Background and Purpose The synthetic compound efsevin was recently identified to suppress arrhythmogenesis in models of cardiac arrhythmia, making it a promising candidate for antiarrhythmic therapy. Its activity was shown to be dependent on the voltage‐dependent anion channel 2 (VDAC2) in the outer mitochondrial membrane. Here, we investigated the molecular mechanism of the efsevin–VDAC2 interaction. Experimental Approach To evaluate the functional interaction of efsevin and VDAC2, we measured currents through recombinant VDAC2 in planar lipid bilayers. Using molecular ligand‐protein docking and mutational analysis, we identified the efsevin binding site on VDAC2. Finally, physiological consequences of the efsevin‐induced modulation of VDAC2 were analysed in HL‐1 cardiomyocytes. Key Results In lipid bilayers, efsevin reduced VDAC2 conductance and shifted the channel's open probability towards less anion‐selective closed states. Efsevin binds to a binding pocket formed by the inner channel wall and the pore‐lining N‐terminal α‐helix. Exchange of amino acids N207, K236 and N238 within this pocket for alanines abolished the channel's efsevin‐responsiveness. Upon heterologous expression in HL‐1 cardiomyocytes, both channels, wild‐type VDAC2 and the efsevin‐insensitive VDAC2AAA restored mitochondrial Ca2+ uptake, but only wild‐type VDAC2 was sensitive to efsevin. Conclusion and Implications In summary, our data indicate a direct interaction of efsevin with VDAC2 inside the channel pore that leads to modified gating and results in enhanced SR‐mitochondria Ca2+ transfer. This study sheds new light on the function of VDAC2 and provides a basis for structure‐aided chemical optimization of efsevin.