Masahiro Mori

The hippocampal CA3 area, an associational network implicated in memory function, receives monosynaptic excitatory as well as disynaptic inhibitory input through the mossy-fiber axons of the dentate granule cells. Synapses made by mossy fibers exhibit low release probability, resulting in high failure rates at resting discharge frequencies of 0.1 Hz. In recordings from functionally connected pairs of neurons, burst firing of a granule cell increased the probability of glutamate release onto both CA3 pyramidal cells and inhibitory interneurons, such that subsequent low-frequency stimulation evoked biphasic excitatory/inhibitory responses in a CA3 pyramidal cell, an effect lasting for minutes. Analysis of the unitary connections in the circuit revealed that granule cell bursting caused powerful activation of an inhibitory network, thereby transiently suppressing excitatory input to CA3 pyramidal cells. This phenomenon reflects the high incidence of spike-to-spike transmission at granule cell to interneuron synapses, the numerically much greater targeting by mossy fibers of inhibitory interneurons versus principal cells, and the extensively divergent output of interneurons targeting CA3 pyramidal cells. Thus, mossy-fiber input to CA3 pyramidal cells appears to function in three distinct modes: a resting mode, in which synaptic transmission is ineffectual because of high failure rates; a bursting mode, in which excitation predominates; and a postbursting mode, in which inhibitory input to the CA3 pyramidal cells is greatly enhanced. A mechanism allowing the transient recruitment of inhibitory input may be important for controlling network activity in the highly interconnected CA3 pyramidal cell region.

The hippocampus, a brain structure essential for memory and cognition, is classically represented as a trisynaptic excitatory circuit. Recent findings challenge this view, particularly with regard to the mossy fibre input to CA3, the second synapse in the trisynaptic pathway. Thus, the powerful mossy fibre input to CA3 pyramidal cells might mediate both synaptic excitation and inhibition. Here we show, by recording from connected cell pairs in rat entorhinal-hippocampal slice cultures, that single action potentials in a dentate granule cell evoke a net inhibitory signal in a pyramidal cell. The hyperpolarization is due to disynaptic feedforward inhibition, which overwhelms monosynaptic excitation. Interestingly, this net inhibitory synaptic response changes to an excitatory signal when the frequency of presynaptic action potentials increases. The process responsible for this switch involves the facilitation of monosynaptic excitatory transmission coupled with rapid depression of inhibitory circuits. This ability to immediately switch the polarity of synaptic responses constitutes a novel synaptic mechanism, which might be crucial to the state-dependent processing of information in associative hippocampal networks.

Interneurons are critical in regulating the excitability of principal cells in neuronal circuits, thereby modulating the output of neuronal networks. We investigated synaptically evoked inhibitory responses in CA3 pyramidal cells mediated by metabotropic glutamate receptors (mGluRs) expressed somatodendritically by interneurons. Although pharmacological activation of mGluRs in interneurons has been shown to enhance their excitability, the inability to record mGluR-mediated synaptic responses has precluded detailed characterization of mGluR function in hippocampal interneurons. We found that a single extracellular pulse in CA3 stratum pyramidale was sufficient to induce disynaptic inhibitory responses mediated by postsynaptic mGluRs of the interneurons in CA3 pyramidal cells of hippocampal slice cultures. The disynaptic inhibitory response followed a short-latency monosynaptic inhibitory response, and was observed at stimulus intensities evoking half-maximal monosynaptic IPSCs. Synergistic activation of mGluR1 and mGluR5 was required to induce the full inhibitory response. When recordings were obtained from interneurons in CA3 stratum radiatum or stratum oriens, a single extracellular stimulus induced a slow inward cationic current with a time course corresponding to the slow inhibitory response measured in pyramidal cells. DCG IV, a group II mGluR agonist, which specifically blocks synaptic transmission through mossy fibres, had no effect on mGluR-mediated synaptic responses in interneurons, suggesting that feed-forward inhibition via mossy fibres is not involved. Thus, postsynaptic mGluR1 and mGluR5 in hippocampal interneurons cooperatively mediate slow feedback inhibition of CA3 pyramidal cells. This mechanism may allow interneurons to monitor activity levels from populations of neighbouring principal cells to adapt inhibitory tone to the state of the network.

Electrophysiological and pharmacological properties of glycine receptors were characterized in hippocampal organotypic slice cultures. In the presence of ionotropic glutamate and GABA_B receptor antagonists, pressure‐application of glycine onto CA3 pyramidal cells induced a current associated with increased chloride conductance, which was inhibited by strychnine. Similar chloride currents could also be induced with β‐alanine or taurine. Whole‐cell glycine responses were significantly greater in CA3 pyramidal cells than in CA1 pyramidal cells and dentate granule cells, while responses to GABA were similar among these three cell types. Although these results demonstrate the presence of functional glycine receptors in the hippocampus, no evidence for their activation during synaptic stimulation was found. Gabazine, a selective GABA_A receptor antagonist, totally blocked evoked IPSCs in CA3 pyramidal cells. Glycine receptor activation is not dependent on transporter‐controlled levels of extracellular glycine, as no chloride current was observed in response to sarcosine, an inhibitor of glycine transporters. In contrast, application of guanidinoethanesulfonic acid, an uptake inhibitor of β‐alanine and taurine, induced strychnine‐sensitive chloride current in the presence of gabazine. These data indicate that modulation of transporters for the endogenous amino acids, β‐alanine and taurine, can regulate tonic activation of glycine receptors, which may function in maintenance of inhibitory tone in the hippocampus.

1. A fast ATP-mediated synaptic current was identified in CA3 pyramidal cells in organotypic hippocampal slice cultures. In the presence of inhibitors for ionotropic glutamate and GABA receptors, extracellular stimulation in the pyramidal cell layer evoked fast synaptic currents that reversed near 0 mV, reflecting an increase in a non-selective cationic conductance. This response was mimicked by focal application of ATP. Antagonists of ionotropic P2X receptors reduced both synaptic and ATP-induced currents. 2. Using a pharmacological approach, the source of synaptically released ATP was determined. Synaptic ATP responses were insensitive to presynaptic blockade of GABAergic transmission between interneurons and CA3 pyramidal cells with the mu-opioid receptor agonist D-Ala(2),MePhe(4),Met(O)(5)-ol-enkephalin (FK33-824), but were blocked by adenosine, which inhibits glutamate release from synaptic terminals in the hippocampus. However, selective inhibition of mossy fibre glutamatergic transmission with the metabotropic glutamate receptor group II agonist (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG IV) did not affect the response. This result points to the associational fibres as the source of the ATP-mediated synaptic response. 3. These results suggest that ATP, coreleased with glutamate, induces a synaptic response in CA3 pyramidal cells that is observed mainly under conditions of synchronous discharge from multiple presynaptic inputs.