Components of neuronal chloride transport in rat and human neocortex

Considerable evidence indicates disturbances in the ionic gradient of GABAA receptor-mediated inhibition of neurones in human epileptogenic tissues. Two contending mechanisms have been proposed, reduced outward and increased inward Cl⁻ transporters. We investigated the properties of Cl⁻ transport in human and rat neocortical neurones (layer II/III) using intracellular recordings in slices of cortical tissue. We measured the alterations in reversal potential of the pharmacologically isolated inhibitory postsynaptic potential mediated by GABAA receptors (IPSPA) to estimate the ionic gradient and kinetics of Cl⁻ efflux after Cl⁻ injections before and during application of selected blockers of Cl⁻ routes (furosemide, bumetanide, 9-anthracene carboxylic acid and Cs+). Neurones from human epileptogenic cortex exhibited a fairly depolarized reversal potential of GABAA receptor-mediated inhibition (EIPSP-A) of -61.9 ± 8.3 mV. In about half of the neurones, the EIPSP-A averaged -55.2 ± 5.7 mV, in the other half, 68.6 ± 2.3 mV, similar to rat neurones (-68.9 ± 2.6 mV). After injections of Cl⁻, IPSPA recovered in human neurones with an average time constant (τ) of 19.0 ± 9.6 s (rat neurones: 7.2 ± 2.4 s). We calculated Cl⁻ extrusion rates (1/τ) via individual routes from the τ values obtained in different experimental conditions, revealing that, for example, the K+-coupled Cl⁻ transporter KCC2 comprises 45.3% of the total rate in rat neurones. In human neurones, the total rate of Cl⁻ extrusion was 63.9% smaller, and rates via KCC2, the Na+-K+-2Cl⁻ transporter NKCC1 and the voltage-gatedCl− channelClCwere smaller than in rat neurones by 80.0%, 61.7% and 79.9%, respectively. The rate via anion exchangers conversely was 14.4% larger in human than in rat neurones. We propose that (i) KCC2 is the major route of Cl⁻ extrusion in cortical neurones, (ii) reduced KCC2 is the initial step of disturbed Cl⁻ regulation and (iii) reductions in KCC2 contribute to depolarizing EIPSP-A of neurones in human epileptogenic neocortex.