Gianmaria Maccaferri, MD - US grants

DESCRIPTION (provided by applicant): The hippocampus is a brain region involved in higher cognitive functions, such as learning and memory, and is often affected by important neurological illnesses like epilepsy, schizophrenia and Alzheimer's disease. GABAergic interneurons of the hippocampus display a wide anatomical and functional diversity. Distinct types of interneurons innervate specific postsynaptic domains such as the axon initial segment, soma, as well as the proximal and distal dendrites of principal neurons. This anatomical specificity has been suggested to relate to different functional roles. Most interneurons have been studied "in vitro" during static network conditions, that is to say, from quiescent slices that lack the type of circuitry activity that is likely to occur "in vivo." As a consequence, the interactions between GABAergic interneurons and specific postsynaptic target domains during physiologically- or pathologically-relevant states of the network are not well understood. Therefore, the primary goal of this research is to re-evaluate the functional roles of interneuron diversity and GABAergic input domain specificity during activity similar to rhythms occurring "in vivo" in the brain. We will study the role of interneuron diversity and domain specific GABAergic input during hippocampal synchronous bursting, which is related to higher cognitive activity in the normal brain, but can progress to pathological levels in epileptic conditions. We will focus our project on three specific aims: (i) we will test the hypothesis that perisomatic and dendritic GABAergic inputs regulate different network functions, (ii) we will test the hypothesis that network-driven bursting in individual pyramidal cells can be controlled by specific sets of interneurons, and, finally, (iii) we will test the hypothesis that synaptic properties of specific sets of interneurons control the timing of GABAergic input during network activity. Relating different interneurons and GABAergic inputs to clear functions may lead to important insight into the organizing principles of cortical networks in the normal brain and during disease.

Project Summary The broad aim of this competitive renewal application is to shed new light on the structure and functions of the hippocampal network, with a specific emphasis on a rather mysterious and understudied neuronal cell type, the Cajal-Retzius cell (CR). The significance of studying these cells is highlighted by literature reports indicating increased densities of CRs in the hippocampus of a subpopulation of patients suffering from temporal lobe epilepsy, who also experienced febrile seizures at early ages. This observation has suggested that the physiological process controlling CR numbers and functions may be involved in the epileptogenic process. The scientific premise underlying this project relies on two main discoveries made by our laboratory. First, we have provided unequivocal evidence that hippocampal CRs are a third population of glutamatergic neurons (in addition to pyramidal and granule cells), which persist in the mature hippocampal network and are fully integrated in its microcircuits. Second, we have recently found that CRs express the polymodal, temperature-gated and Ca2+ permeable channel TRPV1. These discoveries provide unique opportunities to study the physiological and pathological functions of CRs and of the microcircuits they drive in genetically-altered animals with conditionally increased levels of TRPV1 expression or conditionally ablated vesicular glutamate transporters. In particular, we will test the hypotheses that the functional expression of TRPV1 by CRs determines their densities in the developing hippocampus and/or regulates their synaptic output. Lastly, we will test the hypothesis that temperatures in the febrile seizure range can impact hippocampal CR-dependent microcircuits via TRPV1 in vitro and in vivo.

Project Summary / Abstract A prominent theory regarding the development of epileptic hyper-synchronization in human and animal models of temporal lobe epilepsy proposes a key role for the specific down-regulation of the expression of the KCC2 transporter in subicular pyramidal cells. As KCC2 is essential to maintain the low intracellular chloride concentrations required for hyperpolarizing GABAergic signaling, loss of KCC2 expression would impair GABAergic inhibition and trigger a series of events leading to the emergence of subicular-initiated interictal activity. Furthermore, interictal discharges coupled to synaptic plasticity would result in interictal- ictal transitions and spread hyper-excitability to extra-hippocampal regions. In summary, if the essential aspects of this KCC2-based mechanistic theory of epileptogenesis were correct, the selective pharmacological reduction of KCC2 transporter activity in a naïve subiculum should be sufficient to generate epileptiform activity ranging from interictal-like to, possibly, full ictal-like events. Although this prediction was supported by computational modeling, direct experimental evidence has not yielded definitive results. Our preliminary data show that the application of highly selective KCC2 antagonists on isolated mini-slices of the mouse subiculum generate synchronous interictal-like bursting that depends on depolarizing GABAergic signaling, but are not, apparently, sufficient to trigger ictal-interictal transitions. We will take advantage of a variety of state of the art techniques (simultaneous patch-clamp recordings from synaptically coupled and uncoupled cells, optogenetic control of specific neuronal populations, and high resolution anatomical reconstructions) to investigate the impact of this type of pharmacologically- induced epileptiform activity in subicular mini-slices. We will explore its consequences on intrinsic and synaptic plasticity, reveal the underlying mechanisms played by different interneuron subtypes, and explore whether additional epileptogenic changes and/or synaptic input from extra-subicular regions are necessary to drive interictal-like to ictal-like transitions.