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Diffusion of Ca²⁺ from Small Boutons en Passant into the Axon Shapes AP-Evoked Ca²⁺ Transients.

ABSTRACT: Not only the amplitude but also the time course of a presynaptic Ca²⁺ transient determine multiple aspects of synaptic transmission. In small bouton-type synapses, the mechanisms underlying the Ca²⁺ decay kinetics have not been fully investigated. Here, factors that shape an action-potential-evoked Ca²⁺ transient were quantitatively studied in synaptic boutons of neocortical layer 5 pyramidal neurons. Ca²⁺ transients were measured with different concentrations of fluorescent Ca²⁺ indicators and analyzed based on a single-compartment model. We found a small endogenous Ca²⁺-binding ratio (7 ± 2) and a high activity of Ca²⁺ transporters (0.64 ± 0.03 ms^-1), both of which enable rapid clearance of Ca²⁺ from the boutons. However, contrary to predictions of the single-compartment model, the decay time course of the measured Ca²⁺ transients was biexponential and became prolonged during repetitive stimulation. Measurements of [Ca²⁺]_i along the adjoining axon, together with an experimentally constrained model, showed that the initial fast decay of the Ca²⁺ transients predominantly arose from the diffusion of Ca²⁺ from the boutons into the axon. Therefore, for small boutons en passant, factors like terminal volume, axon diameter, and the concentration of mobile Ca²⁺-binding molecules are critical determinants of Ca²⁺ dynamics and thus Ca²⁺-dependent processes, including short-term synaptic plasticity.

SUBMITTER: Tran V

PROVIDER: S-EPMC6170794 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Project description:BackgroundIntracellular Ca2+ modulates several microglial activities, such as proliferation, migration, phagocytosis, and inflammatory mediator secretion. Extracellular ATP, the levels of which significantly change during epileptic seizures, activates specific receptors leading to an increase of intracellular free Ca2+ concentration ([Ca2+]i). Here, we aimed to functionally characterize human microglia obtained from cortices of subjects with temporal lobe epilepsy, focusing on the Ca2+-mediated response triggered by purinergic signaling.MethodsFura-2 based fluorescence microscopy was used to measure [Ca2+]i in primary cultures of human microglial cells obtained from surgical specimens. The perforated patch-clamp technique, which preserves the cytoplasmic milieu, was used to measure ATP-evoked Ca2+-dependent whole-cell currents.ResultsIn human microglia extracellular ATP evoked [Ca2+]i increases depend on Ca2+ entry from the extracellular space and on Ca2+ mobilization from intracellular compartments. Extracellular ATP also induced a transient fivefold potentiation of the total transmembrane current, which was completely abolished when [Ca2+]i increases were prevented by removing external Ca2+ and using an intracellular Ca2+ chelator. TRAM-34, a selective KCa3.1 blocker, significantly reduced the ATP-induced current potentiation but did not abolish it. The removal of external Cl- in the presence of TRAM-34 further lowered the ATP-evoked effect. A direct comparison between the ATP-evoked mean current potentiation and mean Ca2+ transient amplitude revealed a linear correlation. Treatment of microglial cells with LPS for 48 h did not prevent the ATP-induced Ca2+ mobilization but completely abolished the ATP-mediated current potentiation. The absence of the Ca2+-evoked K+ current led to a less sustained ATP-evoked Ca2+ entry, as shown by the faster Ca2+ transient kinetics observed in LPS-treated microglia.ConclusionsOur study confirms a functional role for KCa3.1 channels in human microglia, linking ATP-evoked Ca2+ transients to changes in membrane conductance, with an inflammation-dependent mechanism, and suggests that during brain inflammation the KCa3.1-mediated microglial response to purinergic signaling may be reduced.

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Diffusion of Ca²⁺ from Small Boutons en Passant into the Axon Shapes AP-Evoked Ca²⁺ Transients.

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Dataset Information

Diffusion of Ca2+ from Small Boutons en Passant into the Axon Shapes AP-Evoked Ca2+ Transients.

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OmicsDI is part of the ELIXIR infrastructure

Tweets

Diffusion of Ca²⁺ from Small Boutons en Passant into the Axon Shapes AP-Evoked Ca²⁺ Transients.