Abstracts

Rationale: Therapeutic hypothermia (TH), the artificial lowering of body temperature, is used as a neuroprotective strategy in patients with brain damage, e.g., during status epilepticus (SE). It has been shown that TH can delay seizure onset, reduce seizure activity, and prevent neuronal loss. However, TH carries the risk of severe, sometimes life-threatening side effects. We have uncovered the mechanism that controls RNA-binding motif- 3 (RBM3) expression (Neumann et al., EMBO Rep 2020), a cold-shock protein that largely mediates the neuroprotective effect of hypothermia. Next, we have developed antisense oligonucleotides (ASOs) that lead to a 3-4-fold increase in RBM3 mRNA and protein and have validated its long-term neuroprotective effect in a model of prion disease (Preußner et al., EMBO Mol Med 2023). We are now studying the effects of hypothermia-induced changes on epileptogenesis without resorting to whole-body cooling ('hypothermia in a syringe').


Methods: We tested the effects of RBM3-inducing ASOs on epileptogenesis in the intrahippocampal kainate (KA) mouse model. Neuronal damage within the hippocampus can be detected already hours after KA injection and progressively increases over the course of the disease. PBS-injected animals served as non-epileptic controls. RBM3-inducing ASO or control ASO were injected intracerebroventricularly in the same surgery. Mice were monitored for the occurrence of SE and sacrificed at 7, 28, or 90 days post-SE, when neurodegeneration was assessed histologically. Intrahippocampal depth electrodes were implanted in a subset of mice, and the effect of RBM3-inducing ASO treatment on chronic seizure frequency and severity is currently being analyzed.


Results: A total of 11 PBS-controls, 14 KA-only, 11 KA-ASO control, and 18 KA-ASO were enrolled in the study. KA-induced SE led to a 5-10% reduction in body weight and KA-only and KA-ASO-control animals slowly recovered their baseline weight by 7 days, while PBS and KA ASO-treated mice returned to their initial weight faster, and showed a significant weight gain after one week.

Pathohistological analysis of hippocampal neurodegeneration revealed a significant neuronal loss in the KA-injected hippocampus in KA-only and KA-ASO control mice at 7 days post-SE compared to their own non-injected contralateral hippocampus. Neuron counts in KA-ASO-treated mice were not statistically different from non-epileptic PBS controls in both hemispheres. Results from later time points will be presented at the meeting, along with chronic seizure data. While following unilateral injection of 20 ul ASO or control ASO 8/29 mice showed transient circling during SE; a modified Irwin Screen did not show any behavioral alterations in ASO-treated mice compared to controls four weeks after SE.


Conclusions: RBM3-inducing ASOs are well tolerated and show promising disease-modifying effects on animal recovery after SE, as well as strong neuroprotective effects in a mouse model of temporal lobe epilepsy.


Funding: This study is supported by grants to TH and SB (VAL120/2023) of the Investitionsbank Berlin (IBB).