Abstracts

Rationale: The piriform cortex (PC) is a three-layer cortex consisting of the plexiform layer I and the cellular layers II and III. Layer I contains unmyelinated fibers coming from the lateral olfactory tract (LOT) and has been identified as the site of a large and fast increase of extracellular potassium concentration ([K⁺]_o) recorded at the onset of seizure-like events induced by the K⁺ channel blocker 4-aminopyridine (4AP) in the isolated guinea-pig brain in vitro (Uva et al., 2017). We aim to prove that in a condition of excessive discharge the release of K⁺ is larger in the extracellular space in a region with unmyelinated fibers compared to regions where cells bodies and myelinated axons are present, because of the lack of the oligodendrocytes’ ensheathment around the axons and their buffering action. In this case the uptake of  K⁺ relies on astrocytes and when they are not able to buffer this K⁺ excess, this can contribute to the epileptiform activity.

Methods: In the isolated guinea-pig brains maintained in vitro by arterial perfusion, we activated the fibers of LOT with trains of electrical stimuli (30 Hz trains every 2-5 sec) to mimic the preictal activity observed when 4AP is arterially applied. During LOT high frequency stimulation (HFS), we recorded field potentials in both PC and within LOT and measured the corresponding [K⁺]_o shifts with ion-sensitive electrodes inserted in both PC superficial and deeper positions and in the LOT. We monitored [K⁺]_o changes induced by HFS before and after the arterial application of the K_ir4.1 channel blocker BaCl₂ (100µM; 30 min) to prevent the K⁺ buffering carried out by the glia. Differences between [K⁺]_o augmentations in LOT and PC were assessed also after arterial application of the proconvulsant 4AP (50µM; 5 min).

Results: Series of HFS delivered to the LOT at increasing intensities induced increasing shifts of [K⁺]_o that were larger and faster in the superficial (unmyelinated) layer I than in the deeper cellular layers II/III. Trains could result in spike afterdischarges that outlasted HFS; these were associated to a prolongation of [K⁺]_o increase in the deeper PC sites compared to layer I. When BaCl₂ was arterially applied, spike afterdischarges could be elicited after trains at lower stimulus amplitude, indicating a decrease of the threshold for the generation of  hyperexcitability; [K⁺]_o rises in presence of BaCl₂ were larger in both superficial and deeper positions. In the LOT, which is formed by myelinated axons, the [K⁺]_o increment was delayed compared to the PC layer I shifts after both HFS and arterial 4AP perfusion, confirming a role of myelin in the [K⁺]_o increase buffering.

Conclusions: In an unmyelinated fiber layer an excessive [K⁺]_o increase is favored during activity. The [K⁺]_o rise represents a proconvulsant condition that is exacerbated by impairments in K⁺ buffering. In a condition of myelin loss and gliosis, as described in models of epilepsy and in patients suffering from epilepsy, these pathological K⁺ changes could represent a fertile soil for hyperexcitability.

Funding: This work was supported by Italian Health Ministry (Finanziamento di Ricerca Corrente and Progetto 5X1000).