Abstracts

Rationale: The neuronal network linking the hippocampus proper and the medial entorhinal cortex (mEC) is implicated in physiological and pathophysiological functions. We have earlier shown that a Pentylenetetrazol (PTZ)-induced Status Epilepticus (SE) at postnatal day 20 impacts on the adult hippocampal interictal high-frequency oscillations (HFOs) modifying their cholinergic control (Mikroulis A et al. Neuroscience. 2018;369:386-398). Here, we investigated post-immature SE communication modalities in the adult hippocampal CA3-mEC loop by disrupting their functional connections and by measuring interictal epileptiform discharge (IED) frequency and concurrent HFOs. Methods: Combined temporal hippocampal-mEC slices were prepared from adult Sprague-Dawley rats, >40 days after a PTZ-induced-SE (SE) or from their normal (saline injected) littermates (N). Spontaneous IEDs were recorded simultaneously from the CA3 pyramidal and mEC deep layers of intact slices during 50 µM 4-aminopyridine perfusion, following which specific network connections were abolished by 2 successive incisions (figure 1): 1^st disrupting the CA3-CA1 path (Schaffer collateral cut)2^nd the mEC-CA3 path (Parasubiculum cut)Recording electrodes were withdrawn and repositioned again as close as possible to their former position. Stored waveforms (25kHz/8KHz anti-aliasing filter) were band-pass filtered (FIR) at 80-200 Hz (Ripples, R) and 200-600 Hz (Fast Ripples, FR). HFOs were visually identified as local power maxima in the time-frequency domain (figure 2) and their power (µV²/Hz) was measured in 5min-long traces. All datasets were tested for normality (K-S test) and statistical comparisons were performed by paired or unpaired Student’s t-test as required.  Results: Paired recordings from N and SE slices showed that mEC IED frequency was significantly lower than in CA3 (n=16N, p<0.0001; n=9SE, p=0.002); isolation of the 2 areas did not alter IED frequency or this relationship. CA3 IED frequency was similar in N & SE slices, however, mEC IED frequency was higher in SE vs N slices (n=16N, 9SE, p=0.014). Temporal raster plots indicated no specific pattern of coincidence between CA3 and mEC IEDs either in intact slices or after the successive cuts (N, SE). CA3 HFOs had 100-times higher power than those of mEC. Interestingly, the direction of post-SE power changes of Rs and FRs was a mirror image between CA3 & mEC and also between N & SE slices. In intact slices, FR/R ratio was lower in CA3 vs mEC (n=14N, p=0.032; n=8SE, p=0.009), a difference that disappeared post-isolation, because of CA3’s FR/R ratio increase (N, p=0.0007; SE, p=0.005). Furthermore, CA3 FR/R ratio was increased in SE vs N sices both in the presence and also in the absence of mEC in the network (p= 0.0008).  Conclusions: Findings from this in vitro 4-AP model suggest that (i) functional network connections between CA3-mEC exert an “inhibitory” role on hippocampal HFO output; moreover, their disruption affects HFOs not IED frequency supporting the notion that these represent surrogate indices of excitability. Also, (ii) an early-life SE may affect in the long-term neuronal synchronization properties (HFOs) in a region-dependent way (cortex vs hippocampus). Ref: Mikroulis, A.*, Lisgaras, C. P.*, & Psarropoulou, C. Immature Status Epilepticus: In Vitro Models Reveal Differences In Cholinergic Control and HFO Properties of Adult CA3 Interictal Discharges In Temporal Vs Septal Hippocampus. Neuroscience. doi:https://doi.org/10.1016/j.neuroscience.2017.11.029 *equal contribution Funding: Funded by the University of Ioannina.