Методика оценки вклада различных типов ионотропных рецепторов в постсинаптический ответ в ходе эпилептоподобных разрядов in vitro

Abstract

Epileptiform activity in brain slices is the result of a complex integration of excitatory and inhibitory synaptic inputs. The temporal pattern of activation of various synaptic conductances during interictal discharges makes it possible to describe the neural network activity during seizures and to analyze the effect of different pharmacological agents on it. Existing methods of synaptic conductance estimation generally require an assumption of linearity of the current-voltage relationship (I-V relationship) of neuronal response and produce the estimates of only two types of conductances – excitatory and inhibitory, without isolating the contribution of specific types of postsynaptic receptors. This paper describes the theoretical basis for the assessment of postsynaptic conductances of three types of ionotropic receptors (AMPA, NMDA, and GABAa types) that are activated during the interictal discharges in the entorhinal cortex. The proposed algorithm utilizes the differences in the shape of the I-V relationships of these receptors to assess their contributions to the neuron response and requires the implementation of the whole-cell patch-clamp method in the voltage-clamp mode. The proposed method is demonstrated in three models of epileptiform activity in vitro. We compared the time course of synaptic conductances during interictal discharges generated with intact GABAa receptor-mediated transmission and when GABAa receptors are partially blocked by the antibiotic cefepime or by bicuculline. We demonstrated that even though cefepime is an antagonist of GABAa receptors, their total conductance during the late stages of interictal discharges is significantly increased when this antibiotic is applied, compared to the model in which these receptors were not blocked, or the one in which another competitive antagonist, bicuculline, was used. Thus, the proposed method allowed us to identify subtle differences in the temporal course of the three types of synaptic conductances and to demonstrate the differences in the proepileptic effects of GABAa receptor blockers.