Generation of physiological and pathological high frequency oscillations: the role of perisomatic inhibition in sharp-wave ripple and interictal spike generation

Sharp-wave-ripple complexes (SWRs) and interictal-spikes are physiological and pathological forms of irregularly occurring transient high activity events in the hippocampal EEG. They share similar features and carry high-frequency oscillations with different spectral features. Recent results reveal similarities and differences in the generation of the two types of transients, and argue that parvalbumin containing basket cells (PVBCs) are crucial in synchronizing neuronal activity in both cases. SWRs are generated in the reciprocally connected network of inhibitory PVBCs, while in the pathological case, synchronous failure of perisomatic inhibition triggers massive pyramidal cell burst firing. While physiological ripple oscillation is primarily the result of phasic perisomatic inhibitory currents, pathological high-frequency ripples are population spikes of partially synchronous, massively bursting, uninhibited pyramidal cells

Mechanisms of sharp wave initiation and ripple generation

Replay of neuronal activity during hippocampal sharp wave-ripples (SWRs) is essential in memory formation. To understand the mechanisms underlying the initiation of irregularly occurring SWRs and the generation of periodic ripples, we selectively manipulated different components of the CA3 network in mouse hippocampal slices. We recorded EPSCs and IPSCs to examine the buildup of neuronal activity preceding SWRs and analyzed the distribution of time intervals between subsequent SWR events. Our results suggest that SWRs are initiated through a combined refractory and stochastic mechanism. SWRs initiate when firing in a set of spontaneously active pyramidal cells triggers a gradual, exponential buildup of activity in the recurrent CA3 network. We showed that this tonic excitatory envelope drives reciprocally connected parvalbumin-positive basket cells, which start ripple-frequency spiking that is phase-locked through reciprocal inhibition. The synchronized GABAA receptor-mediated currents give rise to a major component of the ripple-frequency oscillation in the local field potential and organize the phase-locked spiking of pyramidal cells. Optogenetic stimulation of parvalbumin-positive cells evoked full SWRs and EPSC sequences in pyramidal cells. Even with excitation blocked, tonic driving of parvalbumin-positive cells evoked ripple oscillations. Conversely, optogenetic silencing of parvalbumin-positive cells interrupted the SWRs or inhibited their occurrence. Local drug applications and modeling experiments confirmed that the activity of parvalbumin-positive perisomatic inhibitory neurons is both necessary and sufficient for ripple-frequency current and rhythm generation. These interneurons are thus essential in organizing pyramidal cell activity not only during gamma oscillation, but, in a different configuration, during SWRs

Dendritic spikes induce ripples in parvalbumin interneurons during hippocampal sharp waves.

Sharp-wave ripples are transient oscillatory events in the hippocampus that are associated with the reactivation of neuronal ensembles within specific circuits during memory formation. Fast-spiking, parvalbumin-expressing interneurons (FS-PV INs) are thought to provide fast integration in these oscillatory circuits by suppressing regenerative activity in their dendrites. Here, using fast 3D two-photon imaging and a caged glutamate, we challenge this classical view by demonstrating that FS-PV IN dendrites can generate propagating Ca(2+) spikes during sharp-wave ripples. The spikes originate from dendritic hot spots and are mediated dominantly by L-type Ca(2+) channels. Notably, Ca(2+) spikes were associated with intrinsically generated membrane potential oscillations. These oscillations required the activation of voltage-gated Na(+) channels, had the same frequency as the field potential oscillations associated with sharp-wave ripples, and controlled the phase of action potentials. Furthermore, our results demonstrate that the smallest functional unit that can generate ripple-frequency oscillations is a segment of a dendrite

자유롭게 움직이는 생쥐의 해마에서 sharp wave-ripple의 전기생리학적 신호와 칼슘 신호를 동시에 기록하는 방법과 활용

학위논문(박사) -- 서울대학교대학원 : 의과대학 의과학과, 2023. 2. 김상정.Various experiences occur in our daily life in diverse context, and among them, some experiences become a life-long memory while some are easily forgotten. Not only encoding of experiences, this phenomenon of memory selection is heavily dependent on the consolidation of memory. In the investigation of long-term memory formation, hippocampal SWR signal and its neuronal contents have been extensively studied using neuronal decoding analysis of electrophysiological signal to recognize SWRs in the neuronal signals. However, as this signal has low spatial resolution and hard to track neurons across time, it has been difficult to analyze the individual contribution of neurons to task-specific SWRs. In this work, I focused on the investigation of the hippocampal SWRs in spatial aspect not only in temporal aspect, and identification of cellular ensembles consisting of the activity to improve contents of consolidated memory by SWRs. To understand the composition of SWRs and its change by the environment in detail, I divided the research process into two parts. In the first part, to investigate individual hippocampal neuronal activity participating in SWRs activity, I developed a Microdrive array with tetrodes, that combines with UCLA miniscope, a 1-p calcium imaging device. This method enables us to observe SWRs activity not only populational electrophysiological manner, as well as individual cellular activity using calcium indicators from freely behaving animals. The acquired data show that a group of hippocampal neurons was identified to have increased activity on the onset of SWRs, while activities were found to be decreased when SWRs are disrupted. This result implies the potential contribution of individual neuronal activity in the memory consolidation process. In the second part, the calcium transient signals acquired from hippocampal neurons was compared by the environment of the animal. While animals are exploring two different environments, SWRs were detected in real-time and hippocampal neuronal activities were observed simultaneously. From the result, we found that different subsets of neurons are firing during SWRs depending on the environment of the animals, suggesting that SWR signals are collective signals of multiple neurons but their compositions are different by the contents of experience. This result has a potential to improve decoding accuracy when investigating replay contents and neuronal composition. Overall, this thesis covers comprehensive strides from the development of tools to analysis of scientific findings in search of neuronal constitution of memory engraved in the hippocampus.우리의 일상 생활에서는 다양한 경험이 다양한 환경에서 일어난다. 그 중 일부는 평생 지속되는 강렬한 경험이 되기도 하고, 다른 경험들은 기억도 되지 못하고 쉽게 잊혀진다. 이러한 기억의 선택적 저장에는, 경험의 입력 과정(encoding) 뿐 만 아니라 경험의 강화 (consolidation) 과정도 큰 영향을 끼친다. 해마의 sharp wave-ripples (SWRs) 와 그 구성 뉴런들은 장기 기억의 형성 과정을 연구하는 데에 큰 비중을 차지해왔다. 특히 이 뉴런의 신호들은 전기생리학적 방식으로 기록되고 연구되었다. 그러나, 이러한 신호들은 뉴런의 위치에 대한 정보가 부족하고 장기간에 걸쳐 같은 뉴런을 인식할 수 없기 때문에, 특정 환경에서 일어나는 SWRs이 어떠한 뉴런으로 구성되어있는지 연구하는 데에는 어려움이 있었다. 이 논문에서는, 해마의 SWRs을 구성하는 뉴런들을 시간적 측면 뿐 만 아니라 공간적 측면에서도 관찰하여, SWRs로 인해 강화되는 기억을 구성하는 뉴런들을 식별하고자 하였다. SWRs의 구성과 환경에 따른 구성의 변화를 알아보기 위해, 본 연구는 아래의 두 부분으로 나누어 진행되었다. 첫번째 부분에서는 전기생리학적 방법과 칼슘 이미징 방법을 통해, 자유롭게 움직이는 생쥐의 해마에서 발생하는 SWRs을 두 가지 방법으로 기록하고, 그 신호에 따라 변화하는 뇌세포의 활동을 알아보고자 한다. 이를 위하여 살아있는 동물에서 실시간으로 전기신호를 측정할 수 있는 초소형 기구를 만들고, 이를 단광자 칼슘 이미징 장치와 결합시켰다. 이러한 방식을 통해 해마의 SWRs을 전기생리학적 방식과 칼슘 신호의 두 가지 방식으로 기록하였다. 기록 결과, 해마의 뇌세포들의 활동성이 SWRs이 시작됨에 따라 증가하는 것이 확인되었다. 반면, SWRs이 방해된 경우에는 해마 뇌세포의 활동성 증가가 관찰되지 않았다. 이러한 결과는, SWRs을 구성하는 세포들이 기억 강화 과정에 영향을 미칠 수 있음을 암시한다. 두 번째 부분에서는, 해마 뉴런에서 얻어진 칼슘 신호들을 동물들의 실험 환경에 따라 비교하여 보았다. 동물들이 두 개의 다른 환경을 경험하고 있는 동안, 해마에서 발생하는 SWRs 신호가 실시간으로 기록되었다. 그 결과, 동물들이 처한 환경에 따라서, 서로 다른 해마 뉴런으로 구성된 그룹들이 SWRs을 구성하고 있다는 것을 발견하였다. 즉, 해마의 SWRs들은 여러 뉴런의 신호들로 이루어져 있으나 그 각각을 구성하는 뉴런은 경험의 환경에 따라 달라질 수 있음을 보여준다. 또한 이러한 결과는, 기억의 재생 (replay)과 그 구성 뉴런을 식별하기 위한 신호의 해독(decoding) 과정에서의 정확성을 높이는 데에 기여할 수 있음을 보여준다. 정리하면, 이 논문을 통하여, 해마에 저장되어 있는 기억을 구성하는 뉴런들을 식별하기 위한 실험 도구의 개발부터 그 결과 발견한 과학적인 내용의 분석에 이르기까지의 단계들을 포괄적으로 기술하고 소개하고자 한다.Abstract 4 Table of contents 7 List of figures and tables 8 Chapter 1. Introduction 10 Chapter 2. Simultaneous cellular imaging, electrical recording and stimulation of hippocampal activity in freely behaving mice 39 Summary 39 Introduction 41 Materials and Methods 43 Results 53 Discussion 58 Chapter 3. Simultaneous cellular imaging and electrical recording for dissecting neuronal ensemble activity of SWRs in multiple environments 76 Summary 76 Introduction 77 Materials and Methods 78 Results 79 Discussion 81 Chapter 4. Conclusion and future perspective 88 References 92 국문요약 113박

Synaptic GABA release prevents GABA transporter type-1 reversal during excessive network activity.

GABA transporters control extracellular GABA, which regulates the key aspects of neuronal and network behaviour. A prevailing view is that modest neuronal depolarization results in GABA transporter type-1 (GAT-1) reversal causing non-vesicular GABA release into the extracellular space during intense network activity. This has important implications for GABA uptake-targeting therapies. Here we combined a realistic kinetic model of GAT-1 with experimental measurements of tonic GABAA receptor currents in ex vivo hippocampal slices to examine GAT-1 operation under varying network conditions. Our simulations predict that synaptic GABA release during network activity robustly prevents GAT-1 reversal. We test this in the 0 Mg(2+) model of epileptiform discharges using slices from healthy and chronically epileptic rats and find that epileptiform activity is associated with increased synaptic GABA release and is not accompanied by GAT-1 reversal. We conclude that sustained efflux of GABA through GAT-1 is unlikely to occur during physiological or pathological network activity

Fast-spiking parvalbumin^+ GABAergic interneurons: From cellular design to microcircuit function

The success story of fast-spiking, parvalbumin-positive (PV+) GABAergic interneurons (GABA, γ-aminobutyric acid) in the mammalian central nervous system is noteworthy. In 1995, the properties of these interneurons were completely unknown. Twenty years later, thanks to the massive use of subcellular patch-clamp techniques, simultaneous multiple-cell recording, optogenetics, in vivo measurements, and computational approaches, our knowledge about PV+ interneurons became more extensive than for several types of pyramidal neurons. These findings have implications beyond the “small world” of basic research on GABAergic cells. For example, the results provide a first proof of principle that neuroscientists might be able to close the gaps between the molecular, cellular, network, and behavioral levels, representing one of the main challenges at the present time. Furthermore, the results may form the basis for PV+ interneurons as therapeutic targets for brain disease in the future. However, much needs to be learned about the basic function of these interneurons before clinical neuroscientists will be able to use PV+ interneurons for therapeutic purposes

Volume-transmitted GABA waves pace epileptiform rhythms in the hippocampal network

Mechanisms that entrain and pace rhythmic epileptiform discharges remain debated. Traditionally, the quest to understand them has focused on interneuronal networks driven by synaptic GABAergic connections. However, synchronized interneuronal discharges could also trigger the transient elevations of extracellular GABA across the tissue volume, thus raising tonic conductance (Gtonic) of synaptic and extrasynaptic GABA receptors in multiple cells. Here, we monitor extracellular GABA in hippocampal slices using patch-clamp GABA "sniffer" and a novel optical GABA sensor, showing that periodic epileptiform discharges are preceded by transient, region-wide waves of extracellular GABA. Neural network simulations that incorporate volume-transmitted GABA signals point to a cycle of GABA-driven network inhibition and disinhibition underpinning this relationship. We test and validate this hypothesis using simultaneous patch-clamp recordings from multiple neurons and selective optogenetic stimulation of fast-spiking interneurons. Critically, reducing GABA uptake in order to decelerate extracellular GABA fluctuations-without affecting synaptic GABAergic transmission or resting GABA levels-slows down rhythmic activity. Our findings thus unveil a key role of extrasynaptic, volume-transmitted GABA in pacing regenerative rhythmic activity in brain networks

An Approach for Reliably Investigating Hippocampal Sharp Wave-Ripples In Vitro

Among the various hippocampal network patterns, sharp wave-ripples (SPW-R) are currently the mechanistically least understood. Although accurate information on synaptic interactions between the participating neurons is essential for comprehensive understanding of the network function during complex activities like SPW-R, such knowledge is currently notably scarce. counterpart. We show that slice storage in the interface chamber close to physiological temperature is the required condition to preserve network integrity that is necessary for the generation of SPW-R. Moreover, we demonstrate the utility of our method for studying synaptic and network properties of SPW-R, using electrophysiological and imaging methods that can only be applied in the submerged system.The approach presented here demonstrates a reliable and experimentally simple strategy for studying hippocampal sharp wave-ripples. Given its utility and easy application we expect our model to foster the generation of new insight into the network physiology underlying SPW-R