In the CNS, action potentials are propagated through complex axonal arbours. It has been suggested that this process is subject to failure at axon branch points (Bielefeldt & Jackson, 1993; Debanne et al. 1997), although this has been disputed by others (Cox et al. 2000). Here we report some experiments showing that action potentials reliably invade axon collaterals of hippocampal CA1 neurons.

Eight-day-old rat pups were killed by cervical dislocation and hippocampal slices (250 µm thick) prepared and cultured as described by Stoppini et al. (1991) for 7-14 days before use. Whole-cell patch-clamp recordings were made from CA1 pyramidal cells. The pipette filling solution contained the calcium indicator 200 µM Oregon Green BAPTA-1. Once a cell had been successfully patched, it was imaged using two photon microscopy and the axon traced until a branch point was found. Thereafter, line scans were used to record the calcium transients elicited by action potentials triggered by depolarising the cell body. The presence of a calcium transient was taken as evidence of action potential invasion of the axon under investigation.

Calcium transients were recorded both from the main axon and axon collaterals at distances between 45 and 230 µm from the cell body. Following stimulation, the calcium concentration in the axon rose from baseline to peak within 2-4 ms. The fractional change in basal fluorescence (ΔF/F) was 0.41 ± 0.04 (n = 30; S.E.M.). We subsequently recorded evoked calcium transients from 15 main axons and their collaterals after clearly identified branch points. In no case did the action potential fail to invade the main axon or the collateral branch (n = 77 trials). Invasion of the axonal arbour was also reliable during short trains of action potentials (20-100 Hz). To estimate the safety factor for action potential invasion of collaterals, we recorded from eleven axon branches in the presence of lidocaine (120-250 µM) or procaine (500 µM) that reduced the inward current by 20-30 %. In all cases, the action potential successfully invaded the axon collaterals.

In conclusion, we find that action potential invasion of the axonal arbour is reliable, even when the peak inward sodium current is reduced by application of a local anaesthetic. Thus action potential invasion of axon collaterals has a significant safety factor.

This work was supported by the BBSRC Bioimaging initiative.