Adenosine released by astrocytes contributes to hypoxia‐induced modulation of synaptic transmission

Abstract

Astrocytes play a critical role in brain homeostasis controlling the local environment in normal as well as in pathological conditions, such as during hypoxic/ischemic insult. Since astrocytes have recently been identified as a source for a wide variety of gliotransmitters that modulate synaptic activity, we investigated whether the hypoxia-induced excitatory synaptic depression might be mediated by adenosine release from astrocytes. We used electrophysiological and Ca 21 imaging techniques in hippocampal slices and transgenic mice, in which ATP released from astrocytes is specifically impaired, as well as chemiluminescent and fluorescence photometric Ca 21 techniques in purified cultured astrocytes. In hippocampal slices, hypoxia induced a transient depression of excitatory synaptic transmission mediated by activation of presynaptic A1 adenosine receptors. The glia-specific metabolic inhibitor fluorocitrate (FC) was as effective as the A1 adenosine receptor antagonist CPT in preventing the hypoxia-induced excitatory synaptic transmission reduction. Furthermore, FC abolished the extracellular adenosine concentration increase during hypoxia in astrocyte cultures. Several lines of evidence suggest that the increase of extracellular adenosine levels during hypoxia does not result from extracellular ATP or cAMP catabolism, and that astrocytes directly release adenosine in response to hypoxia. Adenosine release is negatively modulated by external or internal Ca 21 concentrations. Moreover, adenosine transport inhibitors did not modify the hypoxia-induced effects, suggesting that adenosine was not released by facilitated transport. We conclude that during hypoxia, astrocytes contribute to regulate the excitatory synaptic transmission through the release of adenosine, which acting on A1 adenosine receptors reduces presynaptic transmitter release. Therefore, adenosine release from astrocytes serves as a protective mechanism by down regulating the synaptic activity level during demanding conditions such as transient hypoxia.

Cultured Astrocytes do not Release Adenosine during Hypoxic Conditions

“…Adenosine deaminase is added in a second step after photon production in the first step has stabilized (Kather et al, 1987). Our study was unable to reproduce the mildly hypoxic conditioninduced adenosine release (Martin et al, 2007). It is possible that the difference in results reflect the use of serial two-step instead of single-step procedure.…”

“…However, if astrocytes release adenosine during mildly hypoxic condition, this mechanism could have an important neuroprotective role by suppressing the activity of nearby neurons. We have here compared adenosine release from cultured astrocytes using a chemiluminescence assay (Kather et al, 1987;Martin et al, 2007) with the more traditional HPLC detection of purines (Cui et al, 2009;Goldman et al, 2010). Our analysis show that cultured astrocytes do not release adenosine during mildly hypoxic conditions and that the chemiluminescence assay is unreliable.…”

“…We further evaluated whether astrocytes could release adenosine in response to mildly hypoxic conditions. Figure 3A illustrates the procedure for a mildly hypoxic condition replicating the method described by Martin et al (2007). Cells were washed with artificial cerebrospinal fluid, pH 7.4, and 24-well plates were placed into special chambers equipped with thermostat housing.…”

“…However, more recent observations suggest that cultured astrocytes are also sensitive to hypoxia and release adenosine in response to mild hypoxia. These observations are of potential importance, because astrocytic adenosine release may control excitatory transmission (Martin et al, 2007). Nevertheless, it is a concern that data on adenosine release from astrocytes have been collected using a chemiluminescent assay that heavily depends on serial enzymatic reactions and nonlinear generation of photons (Kather et al, 1987;Martin et al, 2007).…”

Measurement of purine release with microelectrode biosensors

“…Under pathological conditions, it seems that adenosine is indeed released directly [22,23], although how this occurs and the molecules and transporters involved still have not been settled.…”