J. David B. Roberts

Brain-state- and cell-type-specific firing of hippocampal interneurons in vivo

Neural-network oscillations at distinct frequencies have been implicated in the encoding, consolidation and retrieval of information in the hippocampus. Some GABA (γ-aminobutyric acid)-containing interneurons fire phase-locked to theta oscillations (4–8 Hz) or to sharp-wave-associated ripple oscillations (120–200 Hz), which represent different behavioural states. Interneurons also entrain pyramidal cells in vitro. The large diversity of interneurons poses the question of whether they have specific roles in shaping distinct network activities in vivo. Here we report that three distinct interneuron types—basket, axo-axonic and oriens–lacunosum-moleculare cells—visualized and defined by synaptic connectivity as well as by neurochemical markers, contribute differentially to theta and ripple oscillations in anaesthetized rats. The firing patterns of individual cells of the same class are remarkably stereotyped and provide unique signatures for each class. We conclude that the diversity of interneurons, innervating distinct domains of pyramidal cells, emerged to coordinate the activity of pyramidal cells in a temporally distinct and brain-state-dependent manner.

Spike timing of dendrite-targeting bistratified cells during hippocampal network oscillations in vivo.

Behavior-contingent network oscillations bring about transient, functionally coherent neuronal assemblies in the cerebral cortex, including the hippocampus. Inhibitory input on and close to the soma is believed to phase intrinsic oscillations and output of pyramidal cells, but the function of GABA release to pyramidal cell dendrites remains unknown. We recorded the oscillation-locked spike timing of identified bistratified interneurons in rats. These cells mainly innervated small dendritic shafts of pyramidal cells co-aligned with the glutamatergic Schaffer collateral/commissural input. During theta oscillations, bistratified cells fired at a phase when, on average, pyramidal cell dendrites are most hyperpolarized. Interneurons targeting the perisomatic domain discharge at an earlier phase. During sharp wave–associated ripples, bistratified cells fired with high frequency and in-phase with basket cells, on average 1–2 ms after the discharges in pyramidal cell somata and dendrites. Our results indicate that bistratified cells rhythmically modulate glutamatergic input to the dendrites of pyramidal cells to actively promote the precise input/output transformation during network oscillations.

Neuronal Diversity in GABAergic Long-Range Projections from the Hippocampus

The formation and recall of sensory, motor, and cognitive representations require coordinated fast communication among multiple cortical areas. Interareal projections are mainly mediated by glutamatergic pyramidal cell projections; only few long-range GABAergic connections have been reported. Using in vivo recording and labeling of single cells and retrograde axonal tracing, we demonstrate novel long-range GABAergic projection neurons in the rat hippocampus: (1) somatostatin- and predominantly mGluR1α-positive neurons in stratum oriens project to the subiculum, other cortical areas, and the medial septum; (2) neurons in stratum oriens, including somatostatin-negative ones; and (3) trilaminar cells project to the subiculum and/or other cortical areas but not the septum. These three populations strongly increase their firing during sharp wave-associated ripple oscillations, communicating this network state to the septotemporal system. Finally, a large population of somatostatin-negative GABAergic cells in stratum radiatum project to the molecular layers of the subiculum, presubiculum, retrosplenial cortex, and indusium griseum and fire rhythmically at high rates during theta oscillations but do not increase their firing during ripples. The GABAergic projection axons have a larger diameter and thicker myelin sheet than those of CA1 pyramidal cells. Therefore, rhythmic IPSCs are likely to precede the arrival of excitation in cortical areas (e.g., subiculum) that receive both glutamatergic and GABAergic projections from the CA1 area. Other areas, including the retrosplenial cortex, receive only rhythmic GABAergic CA1 input. We conclude that direct GABAergic projections from the hippocampus to other cortical areas and the septum contribute to coordinating oscillatory timing across structures.

Metabotropic glutamate receptor 8-expressing nerve terminals target subsets of GABAergic neurons in the hippocampus.

Presynaptic metabotropic glutamate receptors (mGluRs) show a highly selective expression and subcellular location in nerve terminals modulating neurotransmitter release. We have demonstrated that alternatively spliced variants of mGluR8, mGluR8a and mGluR8b, have an overlapping distribution in the hippocampus, and besides perforant path terminals, they are expressed in the presynaptic active zone of boutons making synapses selectively with several types of GABAergic interneurons, primarily in the stratum oriens. Boutons labeled for mGluR8 formed either type I or type II synapses, and the latter were GABAergic. Some mGluR8-positive boutons also expressed mGluR7 or vasoactive intestinal polypeptide. Interneurons strongly immunopositive for the muscarinic M2 or the mGlu1 receptors were the primary targets of mGluR8-containing terminals in the stratum oriens, but only neurochemically distinct subsets were innervated by mGluR8-enriched terminals. The majority of M2-positive neurons were mGluR8 innervated, but a minority, which expresses somatostatin, was not. Rare neurons coexpressing calretinin and M2 were consistently targeted by mGluR8-positive boutons. In vivo recording and labeling of an mGluR8-decorated and strongly M2-positive interneuron revealed a trilaminar cell with complex spike bursts during theta oscillations and strong discharge during sharp wave/ripple events. The trilaminar cell had a large projection from the CA1 area to the subiculum and a preferential innervation of interneurons in the CA1 area in addition to pyramidal cell somata and dendrites. The postsynaptic interneuron type-specific expression of the high-efficacy presynaptic mGluR8 in both putative glutamatergic and in identified GABAergic terminals predicts a role in adjusting the activity of interneurons depending on the level of network activity.

Synaptic and Nonsynaptic Localization of Benzodiazepine/GABAA Receptor/Cl− Channel Complex Using Monoclonal Antibodies in the Dorsal Lateral Geniculate Nucleus of the Cat

The two monoclonal antibodies, bd‐17 and bd‐24, are specific for β‐ and α‐subunits of the GABAA/benzodiazepine receptor/chloride channel complex respectively. An abundance of both subunits has been revealed in the visual thalamus of the cat by light microscopic immunocytochemistry using these antibodies. The α‐subunit specific antibody and electron microscopy were used to determine the subcellular distribution of immunoreactivity with respect to specific cell classes in the dorsal lateral geniculate nucleus. Immunoreactivity was always associated with membranes and the degree of immunoreactivity varied greatly between different types of cell as defined by: (i) immunoreactivity for GABA; (ii) soma area; (iii) presence or absence of cytoplasmic laminated bodies (CLB). GABA negative neurons with the smallest soma area showed the strongest immunoreactivity, mainly in the endoplasmic reticulum and also on the somatic plasma membrane. Cytoplasmic laminated bodies could be found in the majority of these neurons. Large GABA negative cells without CLBs were strongly immunoreactive on the plasma membrane of the soma and dendrites, but showed scant if any intracellular immunoreactivity. GABA‐positive cells showed weak intracellular immunoreactivity but negligible if any immunoreactivity at the somatic and proximal dendritic plasma membrane. A similar reaction pattern was found in GABA negative cells which contained no CLBs and which constituted a medium sized cell population. It is suggested that the degree of intracellular receptor immunoreactivity is positively correlated with receptor turnover. The dendrites of projection cells, particularly outside the glomeruli, showed strong immunoreactivity on the plasma membrane. The synaptic junctions formed by many boutons (F terminals) establishing symmetrical synapses with dendrites of relay cells were immunopositive, but no immunoreactivity could be detected at the synapses established by the presynaptic dendrites of the local interneurons. Many axo‐somatic F1 junctions were also immunoreactive. However, immunoreactivity for the receptor/channel complex was also widely distribution on nonsynaptic plasma membranes of somata and dendrites. Thus GABA may act at both synaptic and non‐synaptic sites. Furthermore, the correlation of immunoreactivity for the GABAA receptor complex with previously published properties of physiologically identified cells suggests that the strongly immunoreactive, small, GABA negative cells with CLBs might correspond to the ‘lagged’ X‐type cells, and the large GABA negative receptor outlined cells without CLBs might correspond to some of the Y‐type neurons.

High level of mGluR7 in the presynaptic active zones of select populations of GABAergic terminals innervating interneurons in the rat hippocampus

The release of neurotransmitters is modulated by presynaptic metabotropic glutamate receptors (mGluRs), which show a highly selective expression and subcellular location in glutamatergic terminals in the hippocampus. Using immunocytochemistry, we investigated whether one of the receptors, mGluR7, whose level of expression is governed by the postsynaptic target, was present in GABAergic terminals and whether such terminals targeted particular cells. A total of 165 interneuron dendritic profiles receiving 466 synapses (82% mGluR7a‐positive) were analysed. The presynaptic active zones of most GAD‐(77%) or GABA‐positive (94%) synaptic boutons on interneurons innervated by mGluR7a‐enriched glutamatergic terminals (mGluR7a‐decorated) were immunopositive for mGluR7a. GABAergic terminals on pyramidal cells and most other interneurons in str. oriens were mGluR7a‐immunonegative. The mGluR7a‐decorated cells were mostly somatostatin‐ and mGluR1α‐immunopositive neurons in str. oriens and the alveus. Their GABAergic input mainly originated from VIP‐positive terminals, 90% of which expressed high levels of mGluR7a in the presynaptic active zone. Parvalbumin‐positive synaptic terminals were rare on mGluR7a‐decorated cells, but on these neurons 73% of them were mGluR7a‐immunopositive. Some type II synapses innervating interneurons were immunopositive for mGluR7b, as were some type I synapses. Because not all target cells of VIP‐positive neurons are known it has not been possible to determine whether mGluR7 is expressed in a target‐cell‐specific manner in the terminals of single GABAergic cells. The activation of mGluR7 may decrease GABA release to mGluR7‐decorated cells at times of high pyramidal cell activity, which elevates extracellular glutamate levels. Alternatively, the presynaptic receptor may be activated by as yet unidentified endogenous ligands released by the GABAergic terminals or the postsynaptic dendrites.