Astrocyte Ca2+ induces ATP release to regulate myelin axon excitability and conduction velocity

Written by Zheng Yuanjia, edited by Zheng Yuanjia, Wang Sizhen In the central nervous system, astrocytes support neuronal functions
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In gray matter, they can: 1) regulate the number of synapses during development; 2) remove neurotransmitters released by synapses to reduce their overactivity and prevent excitotoxicity; 3) control the concentration of extracellular potassium to prevent excessive excitement ; 4) Regulate blood flow to ensure sufficient energy supply, and provide lactic acid for neurons to provide energy; 5) Respond to the increase of intracellular calcium ion (Ca2+) concentration, release adenosine triphosphate (ATP) and other glials Transmitters also act on neuronal receptors, regulate information processing, etc.
[1]
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However, unlike gray matter, in white matter, information transmission is mainly through axons wrapped in myelin sheath.
Myelin sheathing axons reduces axon capacitance, so the conduction speed of action potentials can be higher.
Produced in the initial segment of axon (AIS), sodium ions will flow through the nodes of Ranvier of the axon wrapped in myelin sheath to achieve rapid transmission of action potentials [2]
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However, the role of star gum in white matter is unclear
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 On October 15, 2021, the team of Jonathan Lezmy and David Attwell from the Department of Neurophysiology and Pharmacology, University of London, UK, published an article titled "Astrocyte Ca2+-evoked ATP release regulates myelinated axon excitability and conduction speed" on Science.
How the middle astrocytes are connected to the myelinated axons of neurons proves that changes in the level of adenosine (ATP) derived from astrocytes can control the white matter information flow and neural circuit function
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Specifically, first, the author observed that astrocytes (hereinafter referred to as astrocytes) are distributed throughout gray matter and white matter (Figure 1A); on cortical layer V pyramidal neurons or oligodendrocytes, astrocytes are associated with myeloid The axon (myelinated axon) and axonal internodal sheaths of the sheath are closely arranged (Figure 1 B, C)
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Figure 1 Astrocytes in AC gray matter and white matter (Source: J.
Lezmy, et al.
, Science, 2021) Use whole-cell patch clamp (technique) to act (load) on the astrocyte to make the V cone The neuron depolarizes, which triggers action potentials
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It was observed that during short-term excitation (1s), the intracellular Ca2+ concentration in astrocytes increased more near the dendrites of neurons than near the axons, but during long-term excitation (10s), the concentration of Ca2+ in both dendrites and axons The intracellular Ca2+ concentration increased (Figure 1 D, E)
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The Ca2+ released from the soma also increased the Ca2+ concentration of the entire astrocyte, and neuronal activity triggered a further Ca2+ concentration superposition (Figure 1F)
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Figure 1D-F Ca2+ concentration in astrocytes (source: J.
Lezmy, et al.
, Science, 2021) In gray matter, astrocyte regulates neuronal function by releasing ATP [2]
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Using the quinacrine labeling method, the author observed that ATP-containing vesicles in astrocyte are distributed around the myelinated axons of layer V pyramidal neurons, and the release of Ca2+ in the astrocytes will cause the vesicles Decrease in the number (43%) (Figure 1 GI).
When the intracellular Ca2+ concentration in the astrocyte is not high enough, this vesicle reduction phenomenon is not observed, and there is no ATP vesicle in the area outside the astrocyte (> 5 µm).
The decrease in the number of bubbles (Figure 1 J, K)
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These results (Figure 1) suggest that in response to the increase in the Ca2+ concentration in the star glue, the star glue will "stay" near the myelinated axons and release ATP to perform certain functions
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Figure 1G-K Ca2+ release causes the reduction of ATP vesicles (Source: J.
Lezmy, et al.
, Science, 2021) Figure 2A-C Ca2+ release causes extracellular ATP release (Source: J.
Lezmy, et al.
, Science, 2021) Therefore, next, the author used luciferin-luciferase to detect the amount of adenosine triphosphate (ATP) released outside the star gum
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1 mM ATP can cause a large increase in the luminescence of fluorescein (Figure 2A), and the Ca2+ released from the astrocytes also causes a similar reaction (Figure 2B, C)
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Immediately afterwards, the authors observed that the adenosine receptors on the myelin axons of layer V neurons—A2a receptors (A2aRs)—are present in 92% of AISs (Figure 2 DH) and 85% of Langfei’s knots (Figure 2).
2 FH), suggesting that A2aRs are expressed in myelinated axons of specific neuron types
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 A2aRs can increase the level of cyclic adenosine monophosphate (cAMP) to promote hyperpolarization activation in axons to activate the opening of hyperpolarization-activated cyclic nucleotide–gated (HCN) type channels [3 , 4], thereby affecting the excitability of cells [4-6]
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The author detected HCN2 subunits in 51% of AISs and 64% of Langfei’s junctions (Figure 2 IM)
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It is worth noting that neither A2aRs nor HCN channels have been reported to exist at the Langfei junction before
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Therefore, ATP derived from star gum may target the myelinated axons of layer V vertebral neurons (Figure 2)
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Figure 2D-MV layer neurons A2aRs and HCN channels on myelinated axons (Source: J.
Lezmy, et al.
, Science, 2021) Further, in order to explore the effect of these A2aRs activation, the author continues to use membrane With the patch clamp technique, adenosine or another A2aRs agonist (CGS 21680, 0.
5 μM) is applied to AIS for rat layer V pyramidal neurons
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It was found that A2aRs activation can depolarize these cells (Figure 3 A, B, E)
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It is also found that the response frequency of the action potential to small injection currents (100-300 pA) increases (Figure 3 C, F, G), but decreases at high injection currents (700-900 pA).
The author speculates that this may be due to Na+ Caused by enhanced channel inactivation (Figure 3 D, F, H)
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Figure 3A-H The activation effect of A2aRs (Source: J.
Lezmy, et al.
, Science, 2021) When the HCN channel is blocked, the inward current of hyperpolarized cells increases with time (Figure 3 I), apply A2aRs Agonist (CGS 21680) increases in amplitude at the end of the inward current, suggesting that the cell’s conductivity has increased (51%), moving up 50% of the point in the current activation curve in the depolarization direction, making the HCN channel more in the physiological range Internal activation (Figure 3 IK)
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A certain concentration of cAMP (50 μM) can also simulate this situation
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But CGS 21680 did not cause any further depolarization shift of these cells (Figure 3 JL)
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These results (Figure 3) indicate that the adenosine receptor-A2a receptor-regulates the axon initiation (AIS) excitation through cyclic adenosine monophosphate (cAMP) and cyclic nucleotide (HCN)-gated channels Sex
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Figure 3 I-L A2aRs regulates the excitability of AIS through cAMP and HCN channels (Source: J.
Lezmy, et al.
, Science, 2021) In order to further explore the function of A2aRs at the location of Langfei junction, the author applied a diaphragm The clamp technique records the part of the axon wrapped in myelin between the cell body of the mouse layer V pyramidal neuron and the bleb at the end of the axon (focus on the three Langfei knots between them), Combined with the method of immunohistochemistry, it was found that A2aRs were distributed at the AIS and Langfei junctions (Figure 4A)
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Injecting a certain current (500 pA, 5 ms) into the cell body of the layer V pyramidal neuron can induce the action potential of the cell body, but there is a delay in the induction of the action potential at the end of the axon (bubble), and it is given at the Langfei junction.
With a higher concentration of A2aRs agonist (CGS 21680), the action potential at the axon end is delayed longer (Figure 4B)
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However, the acceleration phase of the action potential (that is, the time at which the acceleration begins) is advanced (the dotted line in Figure 4B), and the response latency and peak width of the axon terminal vesicle (at the position) are also significantly increased (Figure 4D)
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Under normal circumstances, the forward (AIS middle or end to axon terminal vesicle) conduction velocity of the action potential is twice or three times the reverse (AIS middle or end to the cell body) conduction velocity [7], A2aRs agonist (CGS) 21680) After processing, the forward guide speed is reduced by 64% or 54% (Figure 4E)
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This series of results (Figure 4) shows that the A2aRs of the axon Langfei junction regulate the conduction velocity: the activation of A2aRs at this point will significantly reduce the conduction velocity
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Figure 4A-E The A2aRs in the Langfei junction regulate the conduction velocity (Source: J.
Lezmy, et al.
, Science, 2021) In order to specifically explore the effects of A2aRs and HCN channels on neuronal excitability and conduction velocity, the author uses MATLA [8] established a neuronal myelin axon model in the corpus callosum (Figure 5A)
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Adding the conductance of activated adenosine (ie activation of the HCN channel) to the distal end of the model AIS can significantly depolarize the cell body (Figure 5B), increase the discharge caused by the small injection current (20 pA), and reduce Discharge caused by high current (180 pA) (Figure 5 C, D)
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However, the magnitude of the action potentials simulated by the model has a larger drop than the experimental observations.
These changes are largely due to the influence of the HCN channel on the AIS, because the HCN channel is added to the Langfei junction to the cell body.
The resting potential has only a small effect (Figure 5B)
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Adding the HCN channel directly to the Langfei junction of the model will reduce the forward conduction velocity (Figure 5E), which is consistent with the experimental results observed by CGS 21680 acting on the Langfei junction (Figure 4E)
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Adding adenosine to the model axon to activate the HCN channel can depolarize the Langfei junction and reduce the conduction velocity by 48% (Figure 5F)
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These results (Figure 5) indicate that this computational model can effectively simulate (predict) phenomena and results similar to actual experiments, that is, it can effectively predict the reduction in axonal conduction velocity induced by adenosine (Figure 4)
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Figure 5A-F The calculation model predicts the decrease in axon conduction velocity caused by adenosine (Source: J.
Lezmy, et al.
, Science, 2021) The author's previous studies have shown that the increase in the intracellular Ca2+ concentration in astrocytes can induce ATP Release (Figure 1G-K, Figure 2A-C), and ATP can be converted into extracellular adenosine, and act on AIS and A2aRs of Langfei junction
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Based on this, the author also tested whether Ca2+ release in astrocytes regulates the excitability and conduction velocity of myelinated axons in cortical V pyramidal neurons
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Patch-clamp recordings showed that Ca2+ released from the star glue was distributed near the AIS of the V pyramidal neurons (Figure 6A).
Within 70 seconds after the release of Ca2+, the resting potential of the V pyramidal neurons was depolarized and an action potential occurred.
Change (Figure 6 B, C)
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Under the condition of low current injection, it shows a higher rate of action potential excitation, while under the condition of high current injection, it shows a lower excitation rate
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At the same time, the author also noticed that these changes are similar to the activation of AIS A2aRs observed in the previous experiment.
These changes are not regulated by astrocyte glutamate.
When A2aR is inhibited (blocker ZM 241385) or blocked After the HCN channel (blocker ZD7288), there will be no such changes, and if the Ca2+ concentration (released from the astrocytes) is not high enough, these changes will not be seen (Figure 6 BF)
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The author also observed that the Ca2+ released from the star gel can cause a decrease in axon conduction velocity (Figure 6 GI), which is similar to the situation when an A2aR agonist (CGS 21680) was administered at the Langfei junction in the previous experiment.
(Picture 4)
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At the same time, the author has calculated and simulated that the decrease in conduction velocity depends on the exact position of the action potential in the AIS and its forward and backward (forward and reverse) propagation velocity (Figure 6 I)
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These results (Figure 6) collectively indicate that the increase in the concentration of Ca2+ in the star gel can regulate the excitability and conduction velocity of the V pyramidal neurons
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Figure 6A-I Increased Ca2+ concentration in astrocytes regulates pyramidal cell excitability and axonal conduction velocity (Source: J.
Lezmy, et al.
, Science, 2021) Figure 7 Astrocytes regulate myelin The excitability and conduction velocity of sheath axons (source: J.
Lezmy, et al.
, Science, 2021).
Conclusion and discussion, inspiration and prospects Increased Ca2+ concentration in glial cells can regulate the excitability of pyramidal cells and axonal conduction velocity
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 Changes in AIS excitability lead to changes in the relationship between cell synaptic input and action potential output.
Changes in the conduction velocity of myelin sheath axons may change the function of neural circuits by changing the time for action potentials to reach the cell, thereby changing post-synaptic neuron signals Integration
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In gray matter, astrocyte Ca2+ regulates the release of ATP into the extracellular space of white matter.
After it is converted into adenosine, it regulates the excitability and conduction velocity of myelinated axons
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Specifically, when the level of adenosine rises, the conduction velocity of myelinated axons will decrease, causing the time delay for the action potential to reach the downstream synapse, which may affect the generation of oscillatory discharge (Figure 7)
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 This study reveals a new regulation of astrocytes on the function of neuronal circuits, which enables us to understand the interaction between astrocytes and neurons more deeply, and promotes the further in-depth study of the function of neural circuits
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 Original link: https://doi.
org/10.
1126/science.
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Recommended high-quality scientific research training courses [1] Discount countdown ︱ Near-infrared brain function data processing class (online: 11.
1~11.
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NJ Allen, Astrocyte regulation of synaptic behavior.
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