Time-dependent phenotypical changes of microglia drive alterations in hippocampal synaptic transmission in acute
Laura Ferrucci1, Bernadette Basilico1,2, Ingrid Reverte1,3
1Department of Physiology and Pharmacology, Sapienza University, Laboratory Affiliated to Institute Pasteur Italia-Fondazione Cenci Bolognetti, Rome, Italy.
Abstract:
It is widely acknowledged that microglia actively regulate synaptic function in the brain. Remarkably, much of our understanding regarding the role of microglia in synaptic regulation is derived from studies in acute brain slices. However, it is still uncertain to what extent the preparation and maintenance of acute slices can influence microglial function and whether microglial changes may affect synaptic transmission. In this study, we examined the impact of acute slice resting time on hippocampal CA1 microglia, by assessing morphological and functional parameters at two distinct time intervals. We report that after 4 h from slicing microglia undergo morphological, functional, and transcriptional changes, including a decrease in the number of branches and in their movement speed. Furthermore, microglia acquire a reactive phenotype, characterized by increased amplitude of outward rectifying K+ currents, increased expression of the pro-inflammatory cytokine Tnfα and altered expression of the microglial receptors Cx3cr1 and P2y12r. We also examined time-dependent changes of excitatory synaptic transmission in CA1 pyramidal neurons from acute hippocampal slices, reporting time-dependent decrease in both amplitude and frequency of postsynaptic currents (sEPSCs), along with a decrease in spine density. Noticeably, sEPSCs amplitude decrease was absent in slices prepared from PLX5622 microglia-depleted mice, suggesting that this time-dependent effect on synaptic transmission is microglia-dependent. Our findings highlight possible causal relation between microglia phenotypic changes in the hours following slice preparation and concomitant synaptic changes, pointing to the mechanisms of acute synaptic modulation, whose understanding is crucial for unraveling microglia-neurons interplay in nature. Furthermore, they emphasize the potential issues associated with experimental time windows in ex vivo samples.
Insights
Acute brain slice preparation alters microglia function and brain synapses. These changes, observed within hours, highlight the need to consider experimental time windows for accurate microglia-neuron interplay studies.
Area of Science:
- Neuroscience
- Cell Biology
- Neuroimmunology
Background:
- Microglia are crucial for regulating brain synaptic function.
- Current understanding relies heavily on acute brain slice studies.
- The influence of slice preparation on microglial function remains unclear.
Purpose of the Study:
- To investigate the impact of acute slice resting time on hippocampal CA1 microglia.
- To assess morphological, functional, and transcriptional changes in microglia over time.
- To determine if microglial alterations affect synaptic transmission in acute slices.
Main Methods:
- Assessed microglial morphology and function at two time points post-slicing.
- Analyzed microglial transcriptomic and electrophysiological properties.
- Measured excitatory synaptic transmission (sEPSCs) and spine density in CA1 neurons.
- Utilized microglia-depleted mice (PLX5622) for comparative analysis.
Main Results:
- Microglia showed morphological (reduced branching, slower movement) and functional (increased K+ currents, altered receptor expression) changes after 4h.
- Transcriptional changes included increased Tnfα expression, indicating a reactive phenotype.
- Synaptic transmission decreased over time (reduced sEPSC amplitude/frequency, lower spine density).
- The decrease in sEPSC amplitude was absent in microglia-depleted slices, confirming microglia's role.
Conclusions:
- Acute slice preparation induces time-dependent microglial activation and synaptic alterations.
- Microglial phenotypic changes are causally linked to observed synaptic modifications.
- Experimental time windows are critical considerations for ex vivo brain slice research.
- Findings underscore the importance of understanding microglia-neuron interactions in physiological contexts.
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