Final Report Summary - NANOPHYS (Nanophysiology of fast-spiking, parvalbumin-expressing GABAergic interneurons)
Cortical neuronal networks are comprised of two types of neurons: Glutamatergic principal neurons (pyramidal cells and granule cells) and GABAergic interneurons. Although GABAergic interneurons numerically represent only 10–20% of the neuronal population, they serve key functions in the network. In contrast to the large amount of information available on the subcellular signaling properties of pyramidal neurons, little was known about the properties of GABAergic interneurons. The goal of the NANOPHYS project was to obtain a complete picture of the cellular and subcellular properties of a major interneuron subtype, the fast-spiking, parvalbumin-expressing GABAergic basket cell (PV+ interneuron), using cutting-edge subcellular patch-clamp methods in brain slices, imaging techniques, and computational approaches. We proposed to address four different aspects: (1) Analysis of interneuron dendrites and synaptic inputs, (2) analysis of interneuron axons, (3) analysis of presynaptic terminals, and (4) the relation between subcellular properties and activity of PV+ interneurons in neuronal microcircuits. The main goal was to reveal the molecular and cellular basis of the fast signaling properties of the cells.
To address these important questions, we developed techniques to record from the subcellular processes of PV+ interneurons. Combining confocal imaging and fluorescently targeted patch-clamp recording, we were able, for the first time, to record from dendrites and axons of PV+ interneurons. Using paired recordings from synaptically connected neurons, we were able to quantify transmitter release at the output synapses of PV+ interneurons with highest resolution. Finally, using whole-cell patch-clamp recordings in vivo, we were in a position to study the activity of PV+ interneurons and the output synapses in fully awake, behaving animals under natural conditions. Using this combined approach, we made several important discoveries. Using direct recordings from axons, we showed that action potentials were propagated with high speed and reliability. We revealed the mechanism underlying fast axonal action potential propagation, which is the expression of Na+ channels in supercritical density. We also found that the axonal action potential, despite its brief duration, is highly energy-efficient. Again, we were able to identify the underlying mechanism, which is a matching between the kinetics of Na+ channel inactivation and K+ channel activation. Using paired recordings between PV+ interneurons and their target cells, we demonstrated that PV is expressed at near millimolar concentrations in presynaptic terminals, and that PV therefore can act as an anti-facilitation factor. Furthermore, we identified synaptotagmin 2 as the main Ca2+ sensor that mediates transmitter release at these synapses. We demonstrated that synaptotagmin 2 conveys a kinetic advantage, providing the fastest possible transmitter release from the output synapses of PV+ interneurons. These results greatly widened our knowledge about the mechanisms underlying rapid signaling in PV+ interneurons. Finally, using whole-cell patch-clamp recordings in vivo, we demonstrated directly that PV+ interneurons provide a major contribution to the generation of high-frequency network oscillations, including gamma oscillations in the dentate gyrus and sharp wave-ripple oscillations in the hippocampal CA1 region. These results provided answers to the central question of how interneurons are activated in intact neuronal networks and how they contribute to high-frequency network oscillations.
In summary, the results that emerged from this project have substantially changed our way of thinking about GABAergic interneuron function. In particular, they shed new light on the mechanisms of rapid and energy-efficient signaling in these neurons. Accumulating evidence suggests that PV+ interneurons not only play an important role in the physiological activity of the hippocampal network, but also are involved in several brain diseases. The growing list includes schizophrenia, Rett syndrome, epilepsy, and neurodegenerative diseases. Thus, results obtained in the NANOPHYS project may, in the future, also help to develop new therapeutic strategies for brain disorders.
References:
Hu H, Gan J, Jonas P (2014) Fast-spiking, parvalbumin+ GABAergic interneurons: From cellular design to microcircuit function. Science 345, DOI: 10.1126/science.1255263.
To address these important questions, we developed techniques to record from the subcellular processes of PV+ interneurons. Combining confocal imaging and fluorescently targeted patch-clamp recording, we were able, for the first time, to record from dendrites and axons of PV+ interneurons. Using paired recordings from synaptically connected neurons, we were able to quantify transmitter release at the output synapses of PV+ interneurons with highest resolution. Finally, using whole-cell patch-clamp recordings in vivo, we were in a position to study the activity of PV+ interneurons and the output synapses in fully awake, behaving animals under natural conditions. Using this combined approach, we made several important discoveries. Using direct recordings from axons, we showed that action potentials were propagated with high speed and reliability. We revealed the mechanism underlying fast axonal action potential propagation, which is the expression of Na+ channels in supercritical density. We also found that the axonal action potential, despite its brief duration, is highly energy-efficient. Again, we were able to identify the underlying mechanism, which is a matching between the kinetics of Na+ channel inactivation and K+ channel activation. Using paired recordings between PV+ interneurons and their target cells, we demonstrated that PV is expressed at near millimolar concentrations in presynaptic terminals, and that PV therefore can act as an anti-facilitation factor. Furthermore, we identified synaptotagmin 2 as the main Ca2+ sensor that mediates transmitter release at these synapses. We demonstrated that synaptotagmin 2 conveys a kinetic advantage, providing the fastest possible transmitter release from the output synapses of PV+ interneurons. These results greatly widened our knowledge about the mechanisms underlying rapid signaling in PV+ interneurons. Finally, using whole-cell patch-clamp recordings in vivo, we demonstrated directly that PV+ interneurons provide a major contribution to the generation of high-frequency network oscillations, including gamma oscillations in the dentate gyrus and sharp wave-ripple oscillations in the hippocampal CA1 region. These results provided answers to the central question of how interneurons are activated in intact neuronal networks and how they contribute to high-frequency network oscillations.
In summary, the results that emerged from this project have substantially changed our way of thinking about GABAergic interneuron function. In particular, they shed new light on the mechanisms of rapid and energy-efficient signaling in these neurons. Accumulating evidence suggests that PV+ interneurons not only play an important role in the physiological activity of the hippocampal network, but also are involved in several brain diseases. The growing list includes schizophrenia, Rett syndrome, epilepsy, and neurodegenerative diseases. Thus, results obtained in the NANOPHYS project may, in the future, also help to develop new therapeutic strategies for brain disorders.
References:
Hu H, Gan J, Jonas P (2014) Fast-spiking, parvalbumin+ GABAergic interneurons: From cellular design to microcircuit function. Science 345, DOI: 10.1126/science.1255263.