Richard Miles

Differences between Somatic and Dendritic Inhibition in the Hippocampus

Hippocampal synaptic inhibition is mediated by distinct groups of inhibitory cells. Some contact pyramidal cells perisomatically, while others terminate exclusively on their dendrites. We examined perisomatic and dendritic inhibition by recording from CA3 inhibitory and pyramidal cells and injecting biocytin to visualize both cells in light and electron microscopy. Single perisomatic inhibitory cells made 2-6 terminals clustered around the soma and proximal pyramidal cell processes. Dendritic cells established 5-17 terminals, usually on different dendrites of a pyramidal cells. Perisomatic terminals were larger than those facing dendritic membrane. Perisomatic inhibitory cells initiated the majority of simultaneous IPSPs seen in nearby pyramidal cells. Single IPSPs initiated by perisomatic sodium-dependent action potentials. Activation of inhibitory fibers terminating on dendrites could suppress calcium-dependent spikes. Thus, distinct inhibitory cells may differentially control dendritic electrogenesis and axonal output of hippocampal pyramidal cells.

Perturbed Chloride Homeostasis and GABAergic Signaling in Human Temporal Lobe Epilepsy

Changes in chloride (Cl−) homeostasis may be involved in the generation of some epileptic activities. In this study, we asked whether Cl− homeostasis, and thus GABAergic signaling, is altered in tissue from patients with mesial temporal lobe epilepsy associated with hippocampal sclerosis. Slices prepared from this human tissue generated a spontaneous interictal-like activity that was initiated in the subiculum. Records from a minority of subicular pyramidal cells revealed depolarizing GABAA receptor-mediated postsynaptic events, indicating a perturbed Cl− homeostasis. We assessed possible contributions of changes in expression of the potassium–chloride cotransporter KCC2. Double in situ hybridization showed that mRNA for KCC2 was absent from ∼30% of CaMKIIα (calcium/calmodulin-dependent protein kinase IIα)-positive subicular pyramidal cells. Combining intracellular recordings with biocytin-filled electrodes and KCC2 immunochemistry, we observed that all cells that were hyperpolarized during interictal events were immunopositive for KCC2, whereas the majority of depolarized cells were immunonegative. Bumetanide, at doses that selectively block the chloride-importing potassium–sodium–chloride cotransporter NKCC1, produced a hyperpolarizing shift in GABAA reversal potentials and suppressed interictal activity. Changes in Cl− transporter expression thus contribute to human epileptiform activity, and molecules acting on these transporters may be useful antiepileptic drugs.

Disinhibition of rat hippocampal pyramidal cells by GABAergic afferents from the septum.

1. Slices were prepared from rat forebrain to include both the septum and the hippocampus in order to examine the effects of septal stimulation on hippocampal inhibitory circuits. 2. Repetitive stimulation of septo‐hippocampal fibres caused a maintained decrease in the frequency of spontaneous IPSPs recorded from CA3 pyramidal cells in the presence of antagonists of excitatory amino acid receptors and of muscarine receptors. 3. In records made from pyramidal cells with CsCl‐filled electrodes, IPSPs were examined at potentials both more positive and more negative than their reversal potential. Single septal stimuli hyperpolarized pyramidal cells when IPSPs were depolarizing events and depolarized them when IPSPs were hyperpolarizing. The GABAA receptor antagonist picrotoxin abolished the effects of septal stimulation. 4. Activation of septal afferents initiated an IPSP in hippocampal inhibitory cells but not in pyramidal cells. Septal IPSPs had similar kinetics to those initiated by local hippocampal stimulation and could suppress inhibitory cell discharge. 5. In pyramidal cells recorded with potassium acetate‐filled electrodes, septal stimuli initiated a depolarization that increased with the driving force for Cl‐ and that could cause firing. 6. Rhythmic stimulation of septo‐hippocampal fibres at 5 Hz initiated, in the hippocampus, a maintained out‐of‐phase oscillation of pyramidal cell discharge and inhibitory cell firing, as detected by the occurrence of spontaneous IPSPs. 7. These results suggest that GABAergic septo‐hippocampal afferents selectively inhibit hippocampal inhibitory cells and so disinhibit pyramidal cells. This disinhibition could contribute to the transmission of the theta rhythm from the septum to the hippocampus.

How many subtypes of inhibitory cells in the hippocampus

Hippocampal inhibitory cells are diverse. It is supposed that they fall into functionally distinct subsets defined by a similar morphology and physiology. Switching between functions could be accomplished by activating receptors for modulating transmitters expressed selectively by different subsets of interneurons. We tested this hypothesis by comparing morphology, physiology, and neurotransmitter receptor expression for CA1 hippocampal interneurons. We distinguished 16 distinct morphological phenotypes and 3 different modes of discharge. Subsets of inhibitory cells were excited or inhibited by agonists at receptors for noradrenaline, muscarine, serotonin, and mGluRs. Most cells responded to 2 or 3 agonists, and 25 different response combinations were detected. Subsets defined by morphology, physiology, and receptor expression did not coincide, suggesting that hippocampal interneurons cannot easily be segregated into a few well-defined groups.

Interneuron Diversity series: Fast in, fast out – temporal and spatial signal processing in hippocampal interneurons

Abstract The operation of neuronal networks crucially depends on a fast time course of signaling in inhibitory interneurons. Synapses that excite interneurons generate fast currents, owing to the expression of glutamate receptors of specific subunit composition. Interneurons generate brief action potentials in response to transient synaptic activation and discharge repetitively at very high frequencies during sustained stimulation. The ability to generate short-duration action potentials at high frequencies depends on the expression of specific voltage-gated K + channels. Factors facilitating fast action potential initiation following synaptic excitation include depolarized interneuron resting potential, subthreshold conductances and active dendrites. Finally, GABA release at interneuron output synapses is rapid and highly synchronized, leading to a faster inhibition in postsynaptic interneurons than in principal cells. Thus, the expression of distinct transmitter receptors and voltage-gated ion channels ensures that interneurons operate with high speed and temporal precision.

EPSP Amplification and the Precision of Spike Timing in Hippocampal Neurons

The temporal precision with which EPSPs initiate action potentials in postsynaptic cells determines how activity spreads in neuronal networks. We found that small EPSPs evoked from just subthreshold potentials initiated firing with short latencies in most CA1 hippocampal inhibitory cells, while action potential timing in pyramidal cells was more variable due to plateau potentials that amplified and prolonged EPSPs. Action potential timing apparently depends on the balance of subthreshold intrinsic currents. In interneurons, outward currents dominate responses to somatically injected EPSP waveforms, while inward currents are larger than outward currents close to threshold in pyramidal cells. Suppressing outward potassium currents increases the variability in latency of synaptically induced firing in interneurons. These differences in precision of EPSP-spike coupling in inhibitory and pyramidal cells will enhance inhibitory control of the spread of excitation in the hippocampus.

Glutamatergic pre-ictal discharges emerge at the transition to seizure in human epilepsy

The mechanisms involved in the transition to an epileptic seizure remain unclear. To examine them, we used tissue slices from human subjects with mesial temporal lobe epilepsies. Ictal-like discharges were induced in the subiculum by increasing excitability along with alkalinization or low Mg2+. During the transition, distinct pre-ictal discharges emerged concurrently with interictal events. Intracranial recordings from the mesial temporal cortex of subjects with epilepsy revealed that similar discharges before seizures were restricted to seizure onset sites. In vitro, pre-ictal events spread faster and had larger amplitudes than interictal discharges and had a distinct initiation site. These events depended on glutamatergic mechanisms and were preceded by pyramidal cell firing, whereas interneuron firing preceded interictal events that depended on both glutamatergic and depolarizing GABAergic transmission. Once established, recurrence of these pre-ictal discharges triggered seizures. Thus, the subiculum supports seizure generation, and the transition to seizure involves an emergent glutamatergic population activity.

Cell‐attached measurements of the firing threshold of rat hippocampal neurones

1 The cell‐attached configuration of the patch‐clamp technique was used to assess resting membrane potential and firing threshold of CA1 pyramidal cells and interneurones of rat hippocampal slices. 2 Resting potential was inferred from the reversal potential of voltage‐gated K+ currents with symmetrical intracellular and pipette K+ concentrations. Its mean value was −74 ± 9 mV for silent interneurones (mean ± s.d.; n= 17) and −84 ± 7 mV for silent pyramidal cells (n= 8). Spontaneous action currents occurred in thirteen out of thirty‐two interneurones and two out of ten pyramidal cells. In active cells, membrane potential values fluctuated by up to 20 mV, due in part to the large hyperpolarizations that followed an action current. 3 Membrane potential values determined from K+ current reversal were 13 ± 6 mV more hyperpolarized than those measured in whole‐cell recordings from the same neurones (n= 8), probably due to a Donnan equilibrium potential between pipette and cytoplasm. 4 Firing threshold of silent cells was determined by elevating external K+ until action currents were generated, while membrane potential was monitored from the cell‐attached K+ current reversal. Spike threshold was attained at −49 ± 8 mV for interneurones (n= 17) and at −60 ± 8 mV for pyramidal cells (n= 8). Increasing external Ca2+ from 2 to 4 mM shifted the neuronal voltage threshold by +5 mV, without affecting resting potential. 5 For comparison with these values, we examined how the rate of membrane polarization influenced firing threshold in whole‐cell records. Ramp current injections, of duration 15–1500 ms, revealed that current threshold followed a classical strength‐duration relationship. In contrast voltage threshold, determined from current injection or by elevating extracellular K+, varied little with the rate of membrane polarization. 6 The state of activation and inactivation of Na+ and K+ currents might contribute to the stability of the voltage threshold. Cell‐attached records showed that 79 ± 10 % of Na+ channels and 64 ± 10 % of K+ channels were available for activation at resting potential in silent cells (n= 8). As cells were depolarized to threshold, Na+ current availability was reduced to 23 ± 10 %, and K+ current availability to 31 ± 12 %. 7 The speed of transition into the inactivated states also appears to contribute to the invariance of threshold for all but the fastest depolarizations. At potentials close to threshold, the rate of inactivation of Na+ and K+ followed a double exponential time course, such that Na+ currents were 62 % inactivated and K+ currents were 63 % inactivated within 15 ms.

cAMP oscillations and retinal activity are permissive for ephrin signaling during the establishment of the retinotopic map

Spontaneous activity generated in the retina is necessary to establish a precise retinotopic map, but the underlying mechanisms are poorly understood. We demonstrate here that neural activity controls ephrin-A–mediated responses. In the mouse retinotectal system, we show that spontaneous activity of the retinal ganglion cells (RGCs) is needed, independently of synaptic transmission, for the ordering of the retinotopic map and the elimination of exuberant retinal axons. Activity blockade suppressed the repellent action of ephrin-A on RGC growth cones by cyclic AMP (cAMP)-dependent pathways. Unexpectedly, the ephrin-A5–induced retraction required cAMP oscillations rather than sustained increases in intracellular cAMP concentrations. Periodic photo-induced release of caged cAMP in growth cones rescued the response to ephrin-A5 when activity was blocked. These results provide a direct molecular link between spontaneous neural activity and axon guidance mechanisms during the refinement of neural maps.

Contributions of intrinsic and synaptic activities to the generation of neuronal discharges in in vitro hippocampus

1 Extracellular and intracellular records were made from guinea‐pig hippocampal slices to examine the contributions of intrinsic cellular properties and synaptic events to the generation of neuronal activity. Extracellular signals were filtered to pass action potentials, which could be detected within a distance of about 80 μm from a discharging cell. 2 Spontaneous action potentials were invariably detected in records from the stratum pyramidale of CA3 region. Blocking excitatory synaptic transmission with NBQX and APV reduced their frequency by 23 ± 35 %. Suppressing synaptic inhibition, while excitation was already blocked, increased the rate of spike discharge to 177 ± 71 % of its control value. 3 Most action potentials recorded intracellularly from CA3 pyramidal cells were initiated in the absence of a detectable synaptic event. In contrast, most action potentials generated by inhibitory cells located close to stratum pyramidale were preceded by an EPSP. 4 In 31 simultaneous recordings, intracellular pyramidal cell action potentials appeared consistently to initiate extracellular spikes with a mean latency of 2·2 ± 1·0 ms. Single inhibitory cell action potentials could initiate a reduction in the frequency of extracellular spikes of duration 10–30 ms. 5 Some identified extracellular spikes (n= 9) consistently preceded intracellularly recorded IPSPs. IPSPs were initiated monosynaptically with latencies of 0·9‐1·5 ms. In reciprocal interactions, single pyramidal cell action potentials could trigger the discharge of an identified unit that in turn appeared to initiate an IPSP in the same pyramidal cell. 6 These data suggest that intrinsic cellular mechanisms underly much of the spontaneous activity of pyramidal cells of the CA3 region of the hippocampus in vitro. Both synaptic inhibition and a strong excitation of inhibitory cells by pyramidal cells act to reduce population activity.