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Astroglial control of axonal excitability, adaptation and analogue signalling

Periodic Reporting for period 1 - AstroModulation (Astroglial control of axonal excitability, adaptation and analogue signalling)

Reporting period: 2017-09-01 to 2019-08-31

Neurons enjoy an elite status in biology as a diverse group of highly specialized cells responsible of high cognitive functions. Glial cells, by comparison, are still viewed by many as those sweeping the extracellular milieu free of excess potassium, neurotransmitter, and cellular debris, or wrapping axons with insulating lipid to facilitate conduction of nerve impulses. Only relatively recently, astrocytes in particular came into light as active contributors to signaling and information processing in the brain.
Action potentials (APs) are usually considered as an elementary unit of information conveyed by presynaptic neurons to their postsynaptic target. Thus, neuronal signals in brain circuits are traditionally thought to occur in an all-or none, or digital, fashion. However, an increasing amount of studies indicates that subthreshold analogue variation in presynaptic membrane potential modulates spike-evoked transmission. One of the main proposed mechanisms involved in this process is the modification of the AP shape via the regulation of axonal potassium and sodium conductances.
Our main hypothesis is that astrocytes engage their potassium channels and potassium-sodium exchange pumps, and probably release glutamate, to affect action potential shape and propagation speed in the nearby axons. Thus, local network activity sensed and integrated by individual astrocytes could modify presynaptic calcium entry and therefore synaptic efficacy. We aim to establish physiological mechanisms underlying this modification and the ensuing changes in short and long synaptic plasticity.
Overall objectives are as follow:
A) To establish through which molecular and cellular machinery the astrocyte activity controls axonal excitability and thus AP shape in the area they cover.
B) To test whether modification of axonal signalling by the astrocyte plays a role in adaptation of the synaptic transmission, including short-term use-dependent efficacy changes.
C) To establish whether modification of synaptic transmission by the astrocyte plays a role in long-term plasticity rules such as Spike-Timing Dependent Plasticity (STDP)
In order to achieve the defined objectives, we needed to establish two critical methods:
1) A reliable way to measure synaptic transmission and its modification by presynaptic potential.
2) A reliable way to record astrocytes and stimulate their fine processes.
Since the beginning of the project, the beneficiary set-up the first critical method for the measure of synaptic transmission in the mossy fibre of the dentate gyrus of the rat. This was done either by expressing the iGluSnFR construction in organotypic slices of hippocampus or by stereotaxic injections of iGluSnFR virus in live animals. This technical development led to some preliminary results, now published in a peer-reviewed journal (Rama, S., Jensen, T.P. and Rusakov, D.A. (2019). Glutamate Imaging Reveals Multiple Sites of Stochastic Release in the CA3 Giant Mossy Fiber Boutons. Front. Cell. Neurosci. 13.).
The second critical method was developed as well, which proved useful in a collaboration and is now published as a preprint (Henneberger, C., Bard, L., Panatier, A., Reynolds, J.P. Medvedev, N.I. Minge, D., Herde, M.K. Anders, S., Kraev, I., Heller, J.P. et al. (2019). LTP induction drives remodeling of astroglia to boost glutamate escape from synapses. BioRxiv 349233.).
In collaboration with other lab members, we have made a breakthrough showing that overexpression of astroglial Kir4.1 channels controls axonal excitability and thus impinge on short-term plasticity. Work is underway to demonstrate the effect on STDP.
Moreover, these first results were presented in an international conference (Rama, S., Jensen, T.P. and Rusakov, D.A. (2019). P3.011 - Glutamate imaging reveals co-operative release sites in the CA3 giant mossy fiber boutons. Poster at NeuroFrance 2019).
Obtained results were published in peer-reviewed journals and international conferences.
When working on the objectives, we showed new results on the model of the giant mossy fibre bouton (Rama, S., Jensen, T.P. and Rusakov, D.A. (2019). Glutamate Imaging Reveals Multiple Sites of Stochastic Release in the CA3 Giant Mossy Fiber Boutons. Front. Cell. Neurosci. 13.). They show that even if giant mossy fibre boutons are a single presynaptic structure, active zones can release glutamate independently from each other. This opens new insights into the precise function of giant mossy fibre boutons, work is underway to better characterize this new phenomenon.
Example of asynchronous release in the same bouton