In the normal brain, the hippocampus receives input from the neocortex via the entorhinal cortex (EC). In the main canonical trisynaptic excitatory hippocampal network, signals propagate from layer II of the EC to the dentate gyrus (DG) through the perforant pathway, then from the granule cells of the DG to CA3 via the output axons of these neurons, the mossy fibers. Area CA3 pyramidal neurons interconnect reciprocally via recurrent axon collaterals and, in addition to sending output to the contralateral hippocampus, project to pyramidal neurons in area CA1 through an axon collateral termed the Schaffer collateral pathway. CA1 output to the layer 5 of EC directly or indirectly via the subiculum. This signal transduction loop is referred to as the trisynaptic pathway (TSP), although there are actually 4-5 synapses in a more comprehensive excitatory limbic loop. This type of excitatory interconnectivity is inherently unstable and prone to uncontrolled activation, and so intrinsic regulators of unchecked activity are also present in the hippocampus. As an example of one such regulator, in the normal hippocampus, DG -> CA3 excitatory relay is limited (gated or filtered) by complex inhibitory mechanisms as well as low intrinsic excitability of dentate granule cells. These inhibitory mechanisms include feed-forward GABA synaptic inhibition, feed-back inhibition, and tonic GABAergic currents.

How can the properties of this EC-> DG-> CA3 circuit can be visualized with functional imaging? Our laboratory use fluorescence VSD (RH795 and/or JPW 3031) to characterize hippocampal circuit function in brain slices. The VSD was bulk-loaded and imaged with a 4x objective lens using a fast CCD camera at a frame rate of 2 kHz. The VSD response to perforant path stimulation (100Hz, four pulses) was imaged in the DG/CA3 area. Upon stimulation, the inner molecular layer of DG showed a strong excitatory response. This excitatory response spread to the dentate granule neuron cell body layer and also to the proximal hilus, but only minimally activated the CA3 pyramidal cell layer. This indicates that dentate granule cells did not fire action potentials, and the response to perforant path activation was predominantly a sub-threshold postsynaptic response. If granule cells did fire synchronous action potentials, the excitatory response to perforant path activation should propagate through the mossy fibers and subsequently excite CA3 cells.

Breakdown of the dentate “gate” has been hypothesized to be a primary contributor to seizure generation in epilepsy. In fully developed epilepsy, we have investigated whether dentate ‘gating’ function was disrupted Surprisingly, the ‘gating’ function was intact in these chronic epilepsy models. This is consistent with several TLE studies that indicated that inhibition within the dentate remains operative, and suppresses seizure propagation from the entorhinal cortex. In contrast, recent studies from our laboratory, examining dentate gyrus function in the latent period preceding epilepsy onset, 1 week after pilocarpine-induced SE, showed markedly disrupted dentate ‘gating’ function, with 60% of cells activating in response to perforant path stimulation, comparable to control studies in the presence of a GABA antagonist. This suggests that ‘gating’ function is corrupted by SE-induced processes, and remains corrupted during the latent period (7 days after SE), only recovering at the point of developing chronic epilepsy (1 month or longer after SE).