Abstract
It is now clearly established that microglial cells reciprocally interact with neurons. Increasing evidence indicates that these interactions are activity-dependent and influence the maturation, fate and activity of synapses. However, the mechanisms through which neuronal activity triggers microglial responses are not fully elucidated. We therefore used acute hippocampal and cortical slices of Cx3cr1-CreERT2::floxed-GCaMP5-Tomato mice to study the impact of neuronal activity on microglial Ca²+ signalling.In standard artificial cerebro-spinal fluid (aCSF ; 33°C), we observed Ca2+ transients occurring spontaneously at a low frequency (0.33 ± 0.08 events/min) in the soma or main processes of hippocampal microglia. Triggering epileptiform activity by bath applying low Mg²+ and 4-AP (200 µM)-containing aCSF induced an increased in the frequency of microglial Ca²+ transients (2.03 ± 0.22 events/min, p<0.05 paired t test ; n=11). Remarkably, these Ca²+ transients occurred simultaneously in all microglial cells of the field of view (200 µM in diameter) and in synchrony with neuronal bursts monitored by extracellular field recordings. In parallel experiments, we also tested whether the activity of a single neuron influenced Ca²+ signalling in microglia. Whole-cell recordings of cortical neurons were performed in standard aCSF and 100 ms depolarizing pulses (resulting in the emission of 15 ± 2 action potentials) were delivered at 1Hz while monitoring Ca²+ transients in microglia. We observed that after 1-2 minutes of stimulation, the frequency and the amplitude of Ca²+ transients were increased by 20% ± 6.94% (p=0.047 paired t test ; n=11) and by 59% ± 22.97% (p = 0.005 paired t test ; n=11) respectively. The frequency of Ca²+ transients induced in microglia by epileptiform activities or by single neuron stimulations was largely reduced upon bath application of the ATP/ADP-degrading enzyme apyrase (25 U/ml). These observations indicate that neuronal network activity enhances Ca2+ signalling in microglia through mechanisms involving purinergic signalling.