Purinergic Signalling and Potassium Homeostasis Regulate Somatosensory Adaptation
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Adaptation is the inhibition of firing in response to background stimulation which functions to allow neurons to respond to novel stimuli (Graczyk et al., 2018; Lampl & Katz, 2017; Mathis et al., 2017). This process takes place in the somatosensory cortex and is constantly happening, but the pathway by which it occurs has not been fully elucidated. One area of interest in this process is the contribution of astrocytes. Astrocytes are part of the tripartite synapse and were originally thought to be passive elements in the brain, however it is now known that they have receptors that bind neurotransmitters released into the synapse, and in turn, can cause the release of gliotransmitters from the astrocyte (Araque, 2008; Araque et al., 1999; Parpura & Zorec, 2010; Perea et al., 2009). The most commonly released gliotransmitters include: ATP, glutamate (Glu), and D-serine (Bowser & Khakh, 2004; Jourdain et al., 2007; Quon et al., 2018). ATP has been shown to not only be released from astrocytes but also activate them causing the further release of gliotransmitters, specifically Glu (Gallagher & Salter, 2003). Subsequently, Glu released from astrocytes has been shown to act on metabotropic glutamate receptors (mGluRs), which can be coupled to inhibitory Gi proteins (Araque et al., 1999; Navarrete et al., 2012; Niswender & Conn, 2010). Aside from this, Glu can act on other receptors on astrocytes such as the N-methyl-D-aspartate (NMDA) receptor, and Gq linked mGluRs (Araque et al., 1999; Haydon, 2001). Activation of these receptors can cause a rise in Ca2+ and the release of gliotransmitters, primarily ATP (Araque et al., 1999; Quon et al., 2018; Yang et al., 2003). Additionally, ATP has been shown to activate GABAergic interneurons by binding the P2Y receptor (Bowser & Khakh, 2004; Quon et al., 2018). Aside from the release of gliotransmitters astrocytes also have an important role in regulating extracellular potassium (K+) concentrations. Modulation of extracellular K+ can lead to changes in the membrane potential resulting in variations in the amplitude and frequency of EPSPs (Hertz & Chen, 2016). Our knowledge of the many interactions between astrocytes, interneurons, and neurons that result in the modulation of firing led us to hypothesize that astrocytes regulate somatosensory adaptation through gliotransmitter release and modulation of extracellular K+ To test this hypothesis, we stimulated cortical slices using a ten-pulse adaptation paradigm in layer 4/5 of the somatosensory cortex and recorded in layer 2. To determine which gliotransmitters had a role in adaptation we used a variety of antagonists to block the receptors of common gliotransmitters including: the ATP P2Y receptor, adenosine A1/A2A receptors, the NMDA receptor (NMDAR), multiple mGluRs, and both the GABAA and GABAB receptors. We also tested the effects of endocannabinoids which are known to be released in a retrograde manner to act on CB1 receptors (CB1R) resulting in the inhibition of neurotransmitter release (Kreitzer & Regehr, 2002; Scheller & Kirchhoff, 2016). Additionally, we applied a solution with low K+ to prevent accumulation of extracellular K+ during sustained firing. Finally, to link these results back to astrocytes we inhibited astrocyte metabolism which is known to inhibit process within the astrocyte that require ATP such as the Na+/K+ ATPase (NKA). Our results revealed that GABA had the greatest role in adaptation. Further it appeared that the GABAA receptor had a larger role in the early phase of higher frequency adaptation and the GABAB receptor was the primary regulator of low frequency adaptation. We also found a large role for the P2Y receptor which supported research suggesting that ATP causes inhibition through activation of P2Y receptors found on interneurons. Finally, we observed a significant decrease in adaptation when applying our low K+ solution. This result suggested that accumulation of K+ in the extracellular space during a train of firing contributes to somatosensory adaptation. At this point we had indirect evidence that astrocytes were involved in adaptation. Furthermore, we were able to find direct evidence for an astrocyte role through the inhibiting astrocyte metabolism. Based on our results we performed occlusion experiments to determine how astrocytes were participating in adaptation and found significant interaction between astrocytes and ATP. These findings collectively suggested that astrocytes have a role in adaptation through the release of ATP. It is likely that this ATP activates interneurons via P2Y receptors to release GABA. Additionally, we cannot rule out that astrocytes also regulate adaptation through modulation of [K+]e, however, we did not find an interaction between K+ and astrocyte metabolism.
DegreeMaster of Science (M.Sc.)
CommitteeCampanucci, Veronica; Norton , Jonathan; Sawicki, Grzegorz