Internal representation of hippocampal neuronal population spans a time-distance continuum.

The hippocampus plays a critical role in episodic memory: the sequential representation of visited places and experienced events. This function is mirrored by hippocampal activity that self organizes into sequences of neuronal activation that integrate spatiotemporal information. What are the underlying mechanisms of such integration is still unknown. Single cell activity was recently shown to combine time and distance information; however, it remains unknown whether a degree of tuning between space and time can be defined at the network level. Here, combining daily calcium imaging of CA1 sequence dynamics in running head-fixed mice and network modeling, we show that CA1 network activity tends to represent a specific combination of space and time at any given moment, and that the degree of tuning can shift within a continuum from 1 day to the next. Our computational model shows that this shift in tuning can happen under the control of the external drive power. We propose that extrinsic global inputs shape the nature of spatiotemporal integration in the hippocampus at the population level depending on the task at hand, a hypothesis which may guide future experimental studies.

GABAergic inhibition shapes interictal dynamics in awake epileptic mice.

Epilepsy is characterized by recurrent seizures and brief, synchronous bursts called interictal spikes that are present in-between seizures and observed as transient events in EEG signals. While GABAergic transmission is known to play an important role in shaping healthy brain activity, the role of inhibition in these pathological epileptic dynamics remains unclear. Examining the microcircuits that participate in interictal spikes is thus an important first step towards addressing this issue, as the function of these transient synchronizations in either promoting or prohibiting seizures is currently under debate. To identify the microcircuits recruited in spontaneous interictal spikes in the absence of any proconvulsive drug or anaesthetic agent, we combine a chronic model of epilepsy with in vivo two-photon calcium imaging and multiunit extracellular recordings to map cellular recruitment within large populations of CA1 neurons in mice free to run on a self-paced treadmill. We show that GABAergic neurons, as opposed to their glutamatergic counterparts, are preferentially recruited during spontaneous interictal activity in the CA1 region of the epileptic mouse hippocampus. Although the specific cellular dynamics of interictal spikes are found to be highly variable, they are consistently associated with the activation of GABAergic neurons, resulting in a perisomatic inhibitory restraint that reduces neuronal spiking in the principal cell layer. Given the role of GABAergic neurons in shaping brain activity during normal cognitive function, their aberrant unbalanced recruitment during these transient events could have important downstream effects with clinical implications.

Optimized temporally deconvolved Ca²âº imaging allows identification of spatiotemporal activity patterns of CA1 hippocampal ensembles.

Hippocampal activity is characterized by the coordinated firing of a subset of neurons. Such neuronal ensembles can either be driven by external stimuli to form new memory traces or be reactivated by intrinsic mechanisms to reactivate and consolidate old memories. Hippocampal network oscillations orchestrate this coherent activity. One key question is how the topology, i.e. the functional connectivity of neuronal networks supports their desired function. Recently, this has been addressed by characterizing the intrinsic properties for the highly recurrently connected CA3 region using organotypic slice cultures and Ca(2+) imaging. In the present study, we aimed to determine the properties of CA1 hippocampal ensembles at high temporal and multiple single cell resolution. Thus, we performed Ca(2+) imaging using the chemical fluorescent Ca(2+) indicator Oregon Green BAPTA 1-AM. To achieve most physiological conditions, we used acute hippocampal slices that were recorded in a so-called interface chamber. To faithfully reconstruct firing patterns of multiple neurons in the field of view, we optimized deconvolution-based detection of action potential associated Ca(2+) events. Our approach outperformed currently available detection algorithms by its sensitivity and robustness. In combination with advanced network analysis, we found that acute hippocampal slices contain a median of 11 CA1 neuronal ensembles with a median size of 4 neurons. This apparently low number of neurons is likely due to the confocal imaging acquisition and therefore yields a lower limit. The distribution of ensemble sizes was compatible with a scale-free topology, as far as can be judged from data with small cell numbers. Interestingly, cells were more tightly clustered in large ensembles than in smaller groups. Together, our data show that spatiotemporal activity patterns of hippocampal neuronal ensembles can be reliably detected with deconvolution-based imaging techniques in mouse hippocampal slices. The here presented techniques are fully applicable to similar studies of distributed optical measurements of neuronal activity (in vivo), where signal-to-noise ratio is critical.

Reliable optical detection of coherent neuronal activity in fast oscillating networks in vitro.

Cognitive and behavioral functions depend on the activation of stable neuronal assemblies, i.e. distributed groups of co-active neurons within neuronal networks. It is therefore crucial to monitor distributed patterns of activity in real time with single-neuron resolution. Microelectrode recordings allow detection of coincidence between discharges of identified units at high temporal resolution, but are not able to reveal the full spatial pattern of activity in multi-cellular assemblies. Therefore, observation of such distributed sets of neurons is a stronghold of optical techniques, but the required resolution, sensitivity, and speed are still challenging current technology. Here, we report a new approach for monitoring neuronal assemblies, using memory-related network oscillations in rodent hippocampal circuits as a model. The cytosolic calcium-sensitive fluorescent protein GCaMP3.NES was expressed using recombinant adeno-associated viral (rAAV)-mediated gene transfer in CA3 pyramidal neurons of cultured mouse hippocampal slices. After 14-21 days in culture, field potential recordings revealed spontaneous occurrence of sharp wave-ripple network events during which a fraction of local neurons is coherently activated. Using a custom-built epi-fluorescence microscope we could monitor a field of view of 410 µm × 410 µm with single-neuron optical resolution (20× objective, 0.4 NA). We developed a highly sensitive and specific wavelet-based method of cell identification allowing simultaneous observation of more than 150 neurons at frame rates of up to 60 Hz. Our recording configuration and image analysis provide a tool to investigate cognition-related activity patterns in the hippocampus and other circuits.

Field potential signature of distinct multicellular activity patterns in the mouse hippocampus.

Cognitive functions go along with complex patterns of distributed activity in neuronal networks, thereby forming assemblies of selected neurons. To support memory processes, such assemblies have to be stabilized and reactivated in a highly reproducible way. The rodent hippocampus provides a well studied model system for network mechanisms underlying spatial memory formation. Assemblies of place-encoding cells are repeatedly activated during sleep-associated network states called sharp wave-ripple complexes (SPW-Rs). Behavioral studies suggest that at any time the hippocampus harbors a limited number of different assemblies that are transiently stabilized for memory consolidation. We hypothesized that the corresponding field potentials (sharp wave-ripple complexes) contain a specific signature of the underlying neuronal activity patterns. Hence, they should fall into a limited number of different waveforms. Application of unbiased sorting algorithms to sharp wave-ripple complexes in mouse hippocampal slices did indeed reveal the reliable recurrence of defined waveforms that were robust over prolonged recording periods. Single-unit discharges tended to fire selectively with certain SPW-R classes and were coupled above chance level. Thus, field SPW-Rs of different waveforms are directly related to the underlying multicellular activity patterns that recur with high fidelity. This direct relationship between the coordinated activity of distinct groups of neurons and macroscopic electrographic signals may be important for cognition-related physiological studies in humans and behaving animals.

Awake hippocampal reactivations project onto orthogonal neuronal assemblies.

The chained activation of neuronal assemblies is thought to support major cognitive processes, including memory. In the hippocampus, this is observed during population bursts often associated with sharp-wave ripples, in the form of an ordered reactivation of neurons. However, the organization and lifetime of these assemblies remain unknown. We used calcium imaging to map patterns of synchronous neuronal activation in the CA1 region of awake mice during runs on a treadmill. The patterns were composed of the recurring activation of anatomically intermingled, but functionally orthogonal, assemblies. These assemblies reactivated discrete temporal segments of neuronal sequences observed during runs and could be stable across consecutive days. A binding of these assemblies into longer chains revealed temporally ordered replay. These modules may represent the default building blocks for encoding or retrieving experience.

Activity-dependent plasticity of mouse hippocampal assemblies in vitro.

Memory formation is associated with the generation of transiently stable neuronal assemblies. In hippocampal networks, such groups of functionally coupled neurons express highly ordered spatiotemporal activity patterns which are coordinated by local network oscillations. One of these patterns, sharp wave-ripple complexes (SPW-R), repetitively activates previously established groups of memory-encoding neurons, thereby supporting memory consolidation. This function implies that repetition of specific SPW-R induces plastic changes which render the underlying neuronal assemblies more stable. We modeled this repetitive activation in an in vitro model of SPW-R in mouse hippocampal slices. Weak electrical stimulation upstream of the CA3-CA1 networks reliably induced SPW-R of stereotypic waveform, thus representing re-activation of similar neuronal activity patterns. Frequent repetition of these patterns (100 times) reduced the variance of both, evoked and spontaneous SPW-R waveforms, indicating stabilization of pre-existing assemblies. These effects were most pronounced in the CA1 subfield and depended on the timing of stimulation relative to spontaneous SPW-R. Additionally, plasticity of SPW-R was blocked by application of a NMDA receptor antagonist, suggesting a role for associative synaptic plasticity in this process. Thus, repetitive activation of specific patterns of SPW-R causes stabilization of memory-related networks.

Coordinated network activity in the hippocampus.

The hippocampus expresses a variety of highly organized network states which bind its individual neurons into collective modes of activity. These patterns go along with characteristic oscillations of extracellular potential known as theta, gamma, and ripple oscillations. Such network oscillations share some important features throughout the entire central nervous system of higher animals: they are restricted to a defined behavioral state, they are mostly generated by subthreshold synaptic activity, and they entrain active neurons to fire action potentials at strictly defined phases of the oscillation cycle, thereby providing a unifying 'zeitgeber' for coordinated multineuronal activity. Recent work from the hippocampus of rodents and humans has revealed how the resulting spatiotemporal patterns support the formation of neuronal assemblies which, in our present understanding, form the neuronal correlate of spatial, declarative, or episodic memories. In this review, we introduce the major types of spatiotemporal activity patterns in the hippocampus, describe the underlying neuronal mechanisms, and illustrate the concept of memory formation within oscillating networks. Research on hippocampus-dependent memory has become a key model system at the interface between cellular and cognitive neurosciences. The next step will be to translate our increasing insight into the mechanisms and systemic functions of neuronal networks into urgently needed new therapeutic strategies.

Cholinergic plasticity of oscillating neuronal assemblies in mouse hippocampal slices.

The mammalian hippocampus expresses several types of network oscillations which entrain neurons into transiently stable assemblies. These groups of co-active neurons are believed to support the formation, consolidation and recall of context-dependent memories. Formation of new assemblies occurs during theta- and gamma-oscillations under conditions of high cholinergic activity. Memory consolidation is linked to sharp wave-ripple oscillations (SPW-R) during decreased cholinergic tone. We hypothesized that increased cholinergic tone supports plastic changes of assemblies while low cholinergic tone favors their stability. Coherent spatiotemporal network patterns were measured during SPW-R activity in mouse hippocampal slices. We compared neuronal activity within the oscillating assemblies before and after a transient phase of carbachol-induced gamma oscillations. Single units maintained their coupling to SPW-R throughout the experiment and could be re-identified after the transient phase of gamma oscillations. However, the frequency of SPW-R-related unit firing was enhanced after muscarinic stimulation. At the network level, these changes resulted in altered patterns of extracellularly recorded SPW-R waveforms. In contrast, recording of ongoing SPW-R activity without intermittent cholinergic stimulation revealed remarkably stable repetitive activation of assemblies. These results show that activation of cholinergic receptors induces plasticity at the level of oscillating hippocampal assemblies, in line with the different role of gamma- and SPW-R network activity for memory formation and -consolidation, respectively.