Abstracts

We recently described a novel pattern of interleaving excitatory-inhibitory (EI) activity (Ziburkus et al., J Neurophys., 2006) and calculated dynamic EI conductances during spontaneous in vitro seizures. The goal of the present study is to describe pharmacological effects, including gap junction blockade on inhibitory and excitatory conductances and EI network synchrony., Simultaneous single and dual voltage clamp whole-cell and extracellular recordings were performed in the CA1 region of rat transverse hippocampal slice preparations (n=P16-P30). To induce seizures, slices were bathed in 100-200[mu]M 4-Aminopyridine in decreased magnesium (0.6mM). Intracellular solution contained blockers for voltage-gated conductances. Excitatory conductances were recorded at -80 to -50mV and inhibitory at 0 to +40mV. Dynamic reversal potentials for interictal bursts and seizures were calculated using varying holding potential (-80mV to +40mV). Post-hoc, the cells were stained and identified using fluorescence conjugated antibody cocktails (neurobiotin, somatostatin, parvalbumin, and mGLUR1). Following immunocytochemical testing, the sections were processed for peroxidase-based neurobiotin visualization used for digital morphological reconstructions of the recorded cells., We performed measurements of dynamic synaptic inhibitory (Gi) and excitatory (Ge) conductances in E (n=12) and I (n=12) cells during seizures. During the initiation of the ictal events, inhibitory conductances were even or greater to the excitatory conductances. Inhibitory conductances peaked 1-2 seconds before the ictal discharge of pyramidal cells. During the persistent stage of the ictal event, Gi transiently diminished, and then increased with Ge during the post-ictal bursting.
Bursts and ictal events were observed in the presence of any one of NMDA (APV), AMPA (CNQX), or GABAa (picrotoxin or bicuculline) receptor blockers. A cocktail of excitatory blockers was necessary to completely abolish bursting and ictal events. Gap junction blockers (mefloquine, n=14 and carbenoxolone, n=12) had profound effects on E and I network synchronization, inhibitory currents, and seizure blockade. We measured effects of the individual blockers on burst dynamics and synchronization. Mefloquine preferentially affected inhibitory network synchronization (n=6), and both carbenoxolone and mefloquine had as strong of an effect on seizures as a mixture of excitatory conductance blockers., We report mechanisms of the first comparative interneuron-principal cell conductance study in a spontaneous in vitro seizure model. Gap junction blockers exhibit a potent effect on neuronal network synchronization and block 4-AP seizures., (Supported by Epilepsy Foundation Fellowship (JZ) and NIH grants: K02MH01493, and RO1MH50006.)