Ex vivo characterization of neuroinflammatory and neuroreceptor changes during epileptogenesis using candidate...

Identification of patients at risk of developing epilepsy before the first spontaneous seizure may promote the development of preventive treatment providing opportunity to stop or slow down the disease.

As development of novel radiotracers and on-site setup of existing radiotracers is highly time-consuming and expensive, we used dual-centre in vitro autoradiography as an approach to characterize the potential of innovative radiotracers in the context of epilepsy development. Using brain slices from the same group of rats, we aimed to characterise the evolution of neuroinflammation and expression of inhibitory and excitatory neuroreceptors during epileptogenesis using translational positron emission tomography (PET) tracers; ¹⁸ F-flumazenil (¹⁸ F-FMZ; GABA_A receptor), ¹⁸ F-FPEB (metabotropic glutamate receptor 5; mGluR5), ¹⁸ F-flutriciclamide (translocator protein; TSPO, microglia activation) and ¹⁸ F-deprenyl (monoamine oxidase B, astroglia activation). Autoradiography images from selected time points after pilocarpine-induced status epilepticus (SE; baseline, 24 and 48 hours, 5, 10 and 15 days and 6 and 12-14 weeks after SE) were normalized to a calibration curve, co-registered to an MRI-based 2D region-of-interest atlas, and activity concentration (Bq/mm² ) was calculated.

In epileptogenesis-associated brain regions, ¹⁸ F-FMZ and ¹⁸ F-FPEB showed an early decrease after SE. ¹⁸ F-FMZ decrease was maintained in the latent phase and further reduced in the chronic epileptic animals, while ¹⁸ F-FPEB signal recovered from day 10, reaching baseline levels in chronic epilepsy. ¹⁸ F-flutriciclamide showed an increase of activated microglia at 24 hours after SE, peaking at 5-15 days and decreasing during the chronic phase. On the other hand, ¹⁸ F-deprenyl autoradiography showed late astrogliosis, peaking in the chronic phase.

Autoradiography revealed different evolution of the selected targets during epileptogenesis. Our results suggest an advantage of combined imaging of inter-related targets like glutamate and GABA_A receptors, or microglia and astrocyte activation, in order to identify important interactions, especially when using PET imaging for the evaluation of novel treatments.