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Science 331 (6017): 599-601

Copyright © 2011 by the American Association for the Advancement of Science

Action-Potential Modulation During Axonal Conduction

Takuya Sasaki1, Norio Matsuki1, and Yuji Ikegaya1,2,*

1 Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, University of Tokyo, Tokyo 113-0033, Japan.
2 Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency, 4-1-8 Honcho Kawaguchi, Saitama 332-0012, Japan.


Figure 1
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  		Fig. 1. Glutamate applied to axons induces AP broadening. (A) Confocal image of dual patch-clamping from the soma and axon of a CA3 PC. Drugs were locally puffed onto the path of the axon (red). Traces show somatic iAP and axonal eAP 300 μm from the axon hillock. (B) The half-maximal width of axonal eAPs (middle), but not of somatic iAPs (bottom), increased during local application of glutamate. Top: eAP traces at time points 1 and 2. (C) Effects of pharmacological reagents on eAP width. n = 4 to 7 slices, *P < 0.05, paired t test. (D) Glutamate-induced inward current in whole-cell recorded axon blebs. (E) Phase-space analysis of an eAP waveform. (Left) An eAP was plotted in the space of V versus dV/dt, where V represents the eAP voltage at a given time. The phase {theta} (blue arrow) was determined to maximize the difference between the orbits of eAPs before (black) and during drug application (red). Right: The {theta} values were similar for eAPs modulated by glutamate, high-K+ depolarization, or 4-AP, but not by TTX. n = 4 or 5 slices.

 

Figure 2
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  		Fig. 2. The AP-broadening effect reaches beyond the glutamate-activated region. (A) A glutamate-induced increase in axonal eAP width is plotted against the path distance between the glutamate puff and the cell-attached recording positions. Red indicates the puffed area (<100 μm {phi}). The line was best fit to a single exponential decay (R2 = 0.50, P = 0.01, n = 17 axon recordings). The decay constant {lambda} (223 μm) was smaller than the axon length constant reported previously (~500 μm) (2–4). Extracellular recording of APs may underestimate the effect of AP broadening. (B) Glutamate applied to axon branches in different locations than the recording site had no effect. n = 6 axons.

 

Figure 3
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  		Fig. 3. Glutamate applied to axons facilitates downstream synaptic efficacy. (A) Identification of the axon branch (no. 4) connecting a patched neuron pair by sequential application of TTX to spots 1 to 7. Red, axons; blue, somatodendrites. Only the main axon fibers are illustrated. (B) Individual uEPSC traces (gray traces) and their averages (thick traces) before and during glutamate application. (C) uEPSCs increased in amplitude during glutamate application. (D) Effects of local drug application on synaptic responses evoked by single-pulse (left) and paired-pulse (right) stimulation. n = 5 to 12 slices, *P < 0.05, paired t test.

 

Figure 4
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  		Fig. 4. Ca2+ uncaging in periaxonal astrocytes broadens APs and facilitates downstream synaptic transmission. (A) Astrocytes (magenta) bolus-loaded with NP-EGTA-AM near the axon whose soma and downstream axon (green) were patch-clamp–recorded. UV illuminated the circled area. (B) Photolysis of NP-EGTA induced axonal eAP broadening. Inset at top: eAPs at time points 1 and 2. (C) Summarized data, including slices that were not loaded with NP-EGTA-AM (UV alone) and slices in which the presynaptic neuron was intracellularly loaded with NP-EGTA (pre-neuron). (D) Astrocytes near the axon connecting two patch-clamped neurons were bolus-loaded with NP-EGTA-AM. (E) CNQX and AP5 were locally applied to the UV-illuminated area. UV was also applied to unloaded slices (UV alone) and slices in which NP-EGTA-AM was bolus-loaded into the astrocytes around irrelevant axon branches (other branch). n = 6 to 19 slices,*P < 0.05, paired t test.

 

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