Related Experiment Video
Updated: Jul 5, 2026
Culturing In Vivo-like Murine Astrocytes Using the Fast, Simple, and Inexpensive AWESAM Protocol
Published on: January 10, 2018
Maxi-anion channel as a candidate pathway for osmosensitive ATP release from mouse astrocytes in primary culture
Hong-Tao Liu1, Abduqodir H Toychiev, Nobuyuki Takahashi
1Department of Cell Physiology, National Institute for Physiological Sciences, Myodaiji-cho, Okazaki 444-8585, Japan.
Abstract:
In the present study, we aimed to evaluate the pathways contributing to ATP release from mouse astrocytes during hypoosmotic stress. We first examined the expression of mRNAs for proteins constituting possible ATP-releasing pathways that have been suggested over the past several years. In RT-PCR analysis using both control and osmotically swollen astrocytes, amplification of cDNA fragments of expected size was seen for connexins (Cx32, Cx37, Cx43), pannexin 1 (Px1), the P2X7 receptor, MRP1 and MDR1, but not CFTR. Inhibitors of exocytotic vesicular release, gap junction hemi-channels, CFTR, MRP1, MDR1, the P2X7 receptor, and volume-sensitive outwardly rectifying chloride channels had no significant effects on the massive ATP release from astrocytes. In contrast, the hypotonicity-induced ATP release from astrocytes was most effectively inhibited by gadolinium (50 muM), an inhibitor of the maxi-anion channel, which has recently been shown to serve as a pathway for ATP release from several other cell types. Thus, we propose that the maxi-anion channel constitutes a major pathway for swelling-induced ATP release from cultured mouse astrocytes as well.
Insights
Mouse astrocytes release adenosine triphosphate (ATP) during hypoosmotic stress. The maxi-anion channel is identified as a primary pathway for this swelling-induced ATP release, as shown by gadolinium inhibition.
Area of Science:
- Neuroscience
- Cell Biology
- Biochemistry
Background:
- Astrocytes play crucial roles in brain function.
- Adenosine triphosphate (ATP) release from astrocytes is implicated in cell signaling.
- Understanding ATP release mechanisms under stress is vital.
Purpose of the Study:
- To investigate the specific pathways responsible for ATP release from mouse astrocytes during hypoosmotic stress.
- To identify the primary channel mediating ATP release under hypotonic conditions.
Main Methods:
- Reverse transcription-polymerase chain reaction (RT-PCR) to analyze mRNA expression of potential ATP release channels.
- Pharmacological inhibition of various ATP release pathways, including connexins, pannexin 1, P2X7 receptor, MRP1, MDR1, CFTR, and maxi-anion channels.
- Assessment of ATP release levels using biochemical assays.
Main Results:
- mRNA for connexins (Cx32, Cx37, Cx43), pannexin 1, P2X7 receptor, MRP1, and MDR1 were detected in astrocytes.
- Inhibitors targeting exocytotic release, gap junction hemichannels, CFTR, MRP1, MDR1, P2X7 receptor, and volume-sensitive outwardly rectifying chloride channels did not significantly affect ATP release.
- Gadolinium, an inhibitor of the maxi-anion channel, significantly inhibited hypotonicity-induced ATP release.
Conclusions:
- The maxi-anion channel is proposed as a major pathway for swelling-induced ATP release from cultured mouse astrocytes.
- Other previously suggested ATP release pathways were not found to be significant under these experimental conditions.
