Reduced Efficacy of the KCC2 Cotransporter Promotes Epileptic Oscillations in a Subiculum Network Model

Abstract

Pharmacoresistant epilepsy is a chronic neurological condition in which a basal brain hyperexcitability results in paroxysmal hypersynchronous neuronal discharges. Human temporal lobe epilepsy has been associated with dysfunction or loss of the potassium-chloride cotransporter KCC2 in a subset of pyramidal cells in the subiculum, a key structure generating epileptic activities. KCC2 regulates intraneuronal chloride and extracellular potassium levels by extruding both ions. Absence of effective KCC2 may alter the dynamics of chloride and potassium levels during repeated activation of GABAergic synapses due to interneuron activity. In turn, such GABAergic stress may itself affect Cl^- regulation. Such changes in ionic homeostasis may switch GABAergic signaling from inhibitory to excitatory in affected pyramidal cells and also increase neuronal excitability. Possibly these changes contribute to periodic bursting in pyramidal cells, an essential component in the onset of ictal epileptic events. We tested this hypothesis with a computational model of a subicular network with realistic connectivity. The pyramidal cell model explicitly incorporated the cotransporter KCC2 and its effects on the internal/external chloride and potassium levels. Our network model suggested the loss of KCC2 in a critical number of pyramidal cells increased external potassium and intracellular chloride concentrations leading to seizure-like field potential oscillations. These oscillations included transient discharges leading to ictal-like field events with frequency spectra as in vitro Restoration of KCC2 function suppressed seizure activity and thus may present a useful therapeutic option. These simulations therefore suggest that reduced KCC2 cotransporter activity alone may underlie the generation of ictal discharges.

Significance statement: Ion regulation in the brain is a major determinant of neural excitability. Intracellular chloride in neurons, a partial determinant of the resting potential and the inhibitory reversal potentials, is regulated together with extracellular potassium via kation chloride cotransporters. During temporal lobe epilepsy, the homeostatic regulation of intracellular chloride is impaired in pyramidal cells, yet how this dysregulation may lead to seizures has not been explored. Using a realistic neural network model describing ion mechanisms, we show that chloride homeostasis pathology provokes seizure activity analogous to recordings from epileptogenic brain tissue. We show that there is a critical percentage of pathological cells required for seizure initiation. Our model predicts that restoration of the chloride homeostasis in pyramidal cells could be a viable antiepileptic strategy.

Keywords: GABA; KCC2; epilepsy; extracellular potassium; intracellular chloride; subiculum.

Figure 1.

Absence of KCC2 leads to a depolarizing GABA reversal potential. A, Scheme of the PY model with intrinsic currents. B, Ionic pathways in the single-cell model. C, Current via KCC2 cotransporter I_KCC2 as function of extracellular potassium K_OUT⁺ when Cl_IN⁻ = 4 mm and I_KCC2^max = 2 μA/cm². D, Experimental and model voltage traces during GABAergic stimulation. E, Amplitude of the PSP during experimental stimulation (red and blue dots) and in the model (red and blue lines). Experimental traces of D and E are taken from Cohen et al. (2002).

Figure 2.

Consequences of the KCC2(−) pathology in single PYs. A, B, voltage trajectories of the KCC2(−) and KCC2(+) PY neurons after stimulation with periodic AMPA, GABA, and NMDA synaptic input for 5 s. Bottom, Changes in K⁺ and Cl⁻ levels. C, State diagram of the model and changes in ionic concentrations. Red and blue trajectories correspond to the KCC2(−) and KCC2(+) cells, respectively. D, Relations between stimulus intensity and firing after stimulation for KCC2(−) (red) and KCC2(+) (blue) cells. E, Voltage trajectories of the KCC2(+) with and without KCC2 contribution to the extracellular potassium after the same synaptic stimulation as in A [KCC2(+) I_KCC2(+) and KCC2(+) I_KCC2(−)]. F, The corresponding extracellular potassium concentration changes.

Figure 3.

Mechanisms of extracellular potassium regulation. A, Capacity of the glial buffer depending on K_OUT⁺. B, Current via I_Na−K^pump depending on K_OUT⁺. C, Current via KCC2 when different K_OUT⁺ regulation mechanisms are present in the KCC2(+) cell. D, Corresponding voltage traces (same stimulus as in Fig. 2A,B).

Figure 4.

Ion concentrations during gamma oscillations in the PY–IN network in the presence of 8 mm potassium in the bath solution. A, Raster plot showing activity for a representative part of the PY and IN populations. B, Voltage traces from representative PYs and IN cells. C, K_OUT⁺ and Cl_IN⁻ concentrations during network activity. D, LFP computed from PY activity and, below, power spectrum of LFP signal.

Figure 5.

KCC2(−) pathology in the subiculum circuit. A, Raster plot of firing in PYs and INs during seizure initiation. B, LFP computed from the network and experimental LFP recordings. C, Extracellular potassium, K_OUT⁺, and intracellular chloride, Cl_IN⁻, changes during seizure initiation. D, Power spectrum of the LFP from the model and experimental recordings.

Figure 6.

Analysis of epileptic oscillations. A, Relations between the number of KCC2(−) cells in the network, and time until seizure initiation and peak frequency of LFP spectrum. The insets show characteristic LFP traces. B, LFP peak frequency as a function of the mean strength of AMPA and GABA conductances in the network. C, Population bursts generated after blocking inhibition in the model and experimental records (block with bicuculline). Synaptic conductances were varied from 0 to 150%: PY–PY, 0–2.25 nS/cm²; PY–IN, 0–1.5 nS/cm²; IN–PY, 0–1.05 nS/cm².

Figure 7.

Adding KCC2(−) cells to the network with endogenous ion concentrations leads to the development of pathological oscillations A, Activity of PYs and INs during seizure initiation. B, LFP trace computed from neuronal activity and corresponding power spectrum. Inset, PY activity during the seizure. C, Changes in the extracellular potassium, K_OUT⁺, and intracellular chloride, Cl_IN⁻, concentrations during seizure initiation. D, Seizure frequency plotted against the proportion of KCC2(−) cells. The inset shows a typical LFP signal.

Figure 8.

Restoring KCC2(−) function with realistic ionic levels prevents seizure. A, B, raster plots of PY and IN activities during seizure oscillations are shown on the left. Those during the transition to normal activity after restoring KCC2 function (black line at 5 s) are shown on the right. C, Changes in the extracellular potassium, K_OUT⁺, and intracellular chloride, Cl_IN⁻, concentrations after restoring KCC2 function, at the black line. D, LFP power spectra during epileptic oscillations and after KKC2 function was restored in KCC2(−) cells.