Differential modulation of AMPA receptor mediated currents by evans blue in postnatal rat hippocampal neurones

Abstract

1. The modulation of non-N-methyl-D-aspartate (NMDA) receptor-mediated whole cell currents and of glutamatergic synaptic transmission by purified Evans Blue (EB) was investigated in rat cultured postnatal hippocampal neurones by use of patch clamp recordings and a fast drug application system. 2. Three different groups of neurones could be distinguished with respect to the type of modulation obtained with 10 microM EB: EB was either a predominant inhibitor of desensitization (13% of the neurones), a predominant inhibitor of current amplitudes (42%) or a mixed inhibitor of both properties (45%). Both effects were not use-dependent and reached maximal levels after 30 s of pre-equilibration with the diazo dye. 3. Dose-response curves obtained from glutamate activated whole cell currents yielded an IC50 value for EB of 13.3 microM (Hill coefficient: 1.3) for the inhibition of desensitization, and an IC50 value of 10.7 microM (Hill coefficient: 1.2) for the inhibition of current amplitudes. 4. Chicago acid SS (100 microM) which is one of the synthesis precursors of EB had no effect on current amplitudes of glutamate activated whole cell currents but was a weak inhibitor of desensitization in all hippocampal neurones investigated, irrespective of the type of modulation obtained with EB in the same neurone. 5. Oxidatively modified EB (the so-called VIMP (10 microM)) had no effect on the kinetics but was a partial inhibitor of glutamate-activated whole cell currents in all hippocampal neurones investigated. 6. EB (10 microM) inhibited the amplitudes of non-NMDA receptor mediated autaptic currents to the same extent (to 39 +/- 19% of control) as observed for glutamate activated whole cell currents (to 41 +/- 17% and 56 +/- 20%). However, the decay of the autaptic responses remained uninfluenced upon EB application, indicating that either receptor desensitization does not dominate the time course of the synaptic response or that the non-NMDA receptors sensitive to modulation of desensitization by EB are not present in the postsynaptic membrane. 7. In conclusion, EB differentially modulates alpha-amino-3-hydroxy-5-methyl -4-isoxazole propionic acid (AMPA) receptor gating in different subsets of neurones. Upon identification of the cellular determinants for the differential modulation (e.g. AMPA receptor subunit composition) EB could become a useful tool to investigate receptor subtypes during electrophysiological recordings.