Abstracts

Rationale: Prolonged status epilepticus (SE) generally has poor long term functional outcomes. Aims of this study were to a) assess neuroprotective and anticonvulsant effects of systemic cooling in a rat model of limbic SE; b) test a novel pharmacological method of cooling by blocking cold defense mechanism (shivering); c) validate MRI/MR spectroscopy as surrogate markers for evaluating neuroprotective efficacy. Methods: Adult male Sprague-Dawley rats (n = 96) were induced into SE for 4.5 hours using lithium-pilocarpine and were monitored continuously using EEG. Half of the rats were cooled (core temperature 31-33 degree C) following SE onset. Outcome was assessed at 24 hours using western blotting, bioluminescence imaging, histology and 7T MRI/MRS. Rats were tested in a Morris water maze at seventh week post SE. Anticonvulsant property of cooling was tested by injecting antiepileptic drugs (AEDs) at SE EEG Stage 3 and comparing the time to stop SE with/without AEDs. Pharmacological cooling was induced by blocking cold receptors transient receptor potential melastatin-8 (TRPM-8) using intravenous infusion of the drug M8-B (TRPM-8 antagonist) at the induction of SE. Results: 81% of rodents treated with cooling during SE survived compared to 32% in the non-treatment SE arm at day 1 (p <0.05; odds ratio of survival 9.53). At 2.5 months survival was 25% for the non-cooling and 50% for the cooling arm. Quantitative analysis of neuronal damage (H&E, cresyl violet stain) revealed less neuronal damage in CA1,CA 2 and dentate gyrus in the cooling arm (p<0.05). These neuroprotective effects were confirmed by western blot analysis using: NeuN (for intact neurons) and activated caspase-3 ( apoptosis). Bioluminescence imaging in vivo detected less inflammatory response in the SE rats treated with cooling. T2 weighted imaging, ADC maps showed robust gliotic changes which were attenuated by cooling. MR spectroscopy showed lower lactate peaks following cooling. Cooling did not stop SE, but adding AEDs at SE EEG Stage 3 decreased the total duration of SE by 38%, thereby suggesting a synergistic effect. Blocking TRPM-8 receptors by M8-B decreased core body temperature by 2.5 C° in comparison to vehicle at a fixed ambient temperature, whereas changing the ambient temperature modulated core body temperature during and after SE. At 7 weeks post SE there was no difference in learning between cooling and non-cooling arm following 2 days of cue testing (non hippocampal learning) in water maze. However in the spatial learning paradigm (hippocampal dependent) following 4 days training the cooling arm animals performed better. This was confirmed by probe trial. Conclusions: Hypothermia attenuates neuro-glial injury, improves survival and cognitive outcome following prolonged status epilepticus. Antiepileptic drugs together with cooling have synergistic anticonvulsant effects. Blocking TRPM-8 receptors lowers core body temperature in experimental status epilepticus, thereby demonstrating the feasibility of a novel method of cooling.