Non-Convulsive Status Epilepticus in a periadolescent rat model: evidence for alterations in hippocampal synaptic plasticity and later life disturbances in cognitive and emotional behaviors

Abstract

Background: Non-convulsive status epilepticus (NCSE) refer to prolonged seizures with minimal motor involvement. In clinical practice, these seizures are often underdiagnosed due to subtle manifestations such as transient behavioral changes. While convulsive status epilepticus (tonic-clonic motor seizures) is universally aggressively treated within minutes to prevent its indisputable harmful consequences and ensuing brain damage, NCSE management remains controversial and often not aggressive over hours to days as its consequences on the brain remain elusive. Pilot experiments in our laboratory, in line with few recent reports, have revealed possible early learning deficits days following NCSE in our recently established periadolescent rodent model. Here, we aim at confirming these early deficits in emotionally-relevant learning following one or two NCSE episodes given that this condition often recurs prior to coming to medical attention. Furthermore, we examine possible electrophysiological, structural, and molecular underlying mechanisms for such deficits. Additionally, we investigate whether emotional and cognitive behavioral deficits are present one month following NCSE.

Methods: Male postnatal day 43 (P43) peri-adolescent rats received one (SKA group), or two 24 hours apart (RKA group), intra-hippocampal 0.00625μg injections of kainic acid to induce prolonged non-convulsive seizures (NCSE) under continuous EEG monitoring. Controls were sham treated with normal saline. In the short-term experimental paradigm, animals were subjected to the modified active-avoidance (MAAV) test at P45, then sacrificed to perform immunohistochemistry for neuronal nuclear antigen (NeuN), glial fibrillary acidic protein (GFAP), and the synaptic plasticity marker, synaptophysin (Syp). In the long-term paradigm: animals were subjected to continuous EEG monitoring followed by a battery of behavioral tests one month post-NCSE. The behavioral panel included tests for anxiety-like behaviors (lightdark box (LD) and open field (OF) tests), depressive-like behaviors (forced swim test (FST)), and hippocampal-dependent visuospatial navigation (Morris water maze (MWM)), followed by the MAAV test.

Results: All rats developed one (SKA) or two (RKA) episodes of hippocampal NCSE following KA injections, with electrographic patterns of evolving rhythmic fast spikes and polyspikes, accompanied by behavioral arrest, staring, and oromotor automatisms. Average post-KA latencies to NCSE were comparable between SKA and RKA on the first day of induction, as were the average seizure durations. On the second day of induction, RKA rats had comparable latencies and durations to the first day of induction. In the short-term paradigm, daily acquisition of tone-signaled electrical footshock avoiding behaviors in the MAAV revealed a statistically significant (p < 0.05) deficit in avoidance rates of both the SKA and RKA groups when compared to controls. In the auditory retention subset, while SKA rats had avoidance rates comparable to ontrols, RKA rats exhibited statistically significant (p < 0.05) lower retention rates. SKA and control groups were significantly (p < 0.05) faster in acquiring adaptive context-cued shock avoidance (p < 0.05) when compared to the RKA group that also exhibited deficits in contextual retention (p < 0.05). No overt cell loss was observed in hippocampal NeuN stained sections. However, compared to controls, both the SKA and RKA groups had a statistically significant increase in reactive astrogliosis in both the left and right hippocampal hilar regions, and CA2-CA3 regions (p < 0.05). RKA rats had significantly decreased hippocampal Syp levels in the hilar and CA2-CA3 regions (p < 0.05) when compared to both SKA and control groups which were comparable. In the long-term paradigm, continuous EEG recordings revealed no seizure recurrence but an increase in spike and polyspike frequencies in both the SKA and RKA groups compared to controls. One month post-NCSE, the RKA rats had deficits in visuospatial navigation in the MWM and in contextual learning in the MAAV test compared to both SKA and control rats. The LD and OF tests pointed to potential hyperactivity and anxiety-like behaviors with decreased exploratory tendencies in the RKA group. Both the SKA and RKA groups showed increased immobility in the FST, suggestive of possible depressive-like behaviors.

Conclusions: Here we confirmed the presence of early behavioral deficits following NCSE, and our pilot data points to the persistence of such deficits along with depressive and anxiety-like behaviors one month following two, but not one, episodes of NCSE. These deficits were accompanied by hippocampal injury evidenced by reactive astrocytosis and alteration in synaptic homeostasis. Ongoing studies in our laboratory aim at confirming the persistence of these molecular changes and detrimental behavioral deficits. These harmful consequences, and the impact of seizure burden, if confirmed, call for a more urgent diagnosis and treatment of NCSE.