Axon transport defects are a common theme in neurodegenerative diseases,35
and may have a role in the pathogenesis of RGC death.8,38
Little is known on the relationship between reduced axon transport and RGC function.39
Here, we addressed this problem by recording the PERG—a signal that depends on the physiologic integrity of RGC—before and after manipulations at postretinal level that impair retrograde signaling in the retinocollicular pathway. Lidocaine is a well established method to block axon transport40–42
without damaging optic nerve structures,43
and acts at very low concentrations.44
At sufficient concentrations, lidocaine also is known to alter signal conduction by blocking voltage-gated sodium channels in the neuronal cell membrane,45
thereby suppressing postsynaptic activity. As FVEPs were not altered significantly after retrobulbar lidocaine, our results indicated that the action of lidocaine on sodium channels was insufficient to impair signal conduction along the optic nerve. Insufficient blockage of sodium channels also suggests that a direct action of lidocaine on action potentials of RGCs is unlikely. It also is unlikely that lidocaine entered the retina via the cardiovascular system and impaired directly RGC spiking activity, as in this case the concentration of lidocaine at the retina would have been minimal. Finally, it is unlikely that optic nerve crush caused ischemic damage to the retina, as shown in previous studies.21,22
For lidocaine injection and optic nerve crush, the FERG—a signal originating in the outer retina—was unaltered. This suggested that the effects of all manipulations did not cause generalized retinal dysfunction.
Altogether, as the effects of all postretinal manipulations on PERG were qualitatively similar and major unspecific effects on RGC could be ruled out, the results strongly suggested that PERG changes were linked to altered supply of retrogradely-delivered material via axon transport. Reversible blockade of axon transport was not expected to cause damage to RGC,43,46
whereas for optic nerve crush RGC loss was expected to start approximately 5 days after surgery.21,47
However, the effects of the optic nerve crush could include an acute physiologic effect on RGC signaling48
that added to the effect mediated by impairment of retrograde signaling.
The main result of our study is that lidocaine injections caused rapid, reversible reduction of PERG signal to at least 50% of its baseline value. PERG effects after intracollicular lidocaine injection could be measured within 1 hour after treatment, and recovered progressively over 4 days. After retrobulbar lidocaine injection, PERG effects could be measured as early as 10 to 20 minutes posttreatment, reached a maximum at 30 minutes, and recovered within 5 days. The effects of retrobulbar optic nerve crush on PERG also could be measured as early 10 to 20 minutes after surgery, and remained stable at reduced level over 1-month follow-up, after which histology demonstrated a drastic (88%) decrease of TUJ1-positive RGC counts in the retina. That the earliest effects on PERG of retrobulbar lidocaine were rather similar in magnitude and time course to those obtained with retrobulbar optic nerve crush suggested that the latter also were mediated largely by impairment of retrograde axon transport rather that direct injury to RGC.48
Dramatic losses of PERG signal after either optic nerve section49
in the mouse have been reported before, but these studies were not designed to monitor early postsurgical events.
Our study provided only a reasonable approximation of the time course of the PERG effects, as there were constraints due the time needed for the experimental procedures. For retrobulbar injection and optic nerve crush, the earliest opportunity to record a postprocedural PERG was approximately 10 minutes. For intracollicular injections, the earliest opportunity was 1 hour later. Thus, we cannot exclude that the PERG effects could have manifested somewhat earlier compared to the values we were able to measure. If we assume 10 to 15 minutes postprocedural delay for proximal interventions (retrobulbar lidocaine, optic nerve crush) and 60 minutes delay for distal intervention (intracollicular lidocaine); if we also assume that the delay was due to the speed of retrograde axon transport, and the retinocollicular distance would be approximately 11 mm,50
then the corresponding velocities of axon transport would be approximately 0.12 mm/min (1.5 mm distance/12.5 min) and 0.18 mm/min (11 mm distance/60 min). These calculated velocities are well in the range of those reported for fast retrograde transport in a number of studies.1,51
It should be taken into account that the mouse optic nerve has an unmyelinated portion of 0.6 to 0.8 mm immediately adjacent to the sclera.52
This might have represented a vulnerable location that drove the earliest effect of retrobulbar lidocaine.
What is the retrograde signal(s) whose reduced supply caused reduced electric responsiveness of RGC? Our study does not provide direct answers to this important question, which will be addressed in a subsequent study. A reasonable hypothesis is that target-derived brain derived neurotrophic factor (BDNF) may represent a likely molecular candidate. Available evidence shows that intraocular pressure elevation in an experimental glaucoma model causes obstructed axon transport of BDNF and its receptor, TrkB, eventually leading to RGC death.38
BDNF has been shown to depolarize neurons just as rapidly as the neurotransmitter glutamate, even at very low concentrations.53–55
Rapid actions of neurotrophins include changes in neuronal excitability, synaptic transmission, and neural plasticity. Of interest, intracollicular injection of saline also temporarily reduced the PERG signal, although to a smaller/shorter extent compared to intracollicular lidocaine injection. It is possible that intracollicular injection of 1 μL saline mechanically caused temporary impairment of the target-derived supply of retrogradely-transported material that sustains RGC responsiveness. RGC responsiveness also may be sustained in part by other postsynaptic targets, such as the dorsal geniculate nucleus and the suprachiasmatic nucleus.
In conclusion, our results showed that impairment of retrograde signaling causes rapid, substantial decrease of PERG amplitude and increase in PERG latency that may be reversible. Results implied that intact retrograde signaling is necessary for the normal electrical responsiveness of retinal ganglion cells. Numerous reports of early PERG impairment in glaucoma,56,57
optic nerve diseases,58
at least in part may be due to altered axon transport. Also, recovery of PERG amplitude losses after either IOP lowering15,17,18,61,62
or removal of pituitary tumors63
may be related to restoration of axon transport. Thus, the PERG may represent a promising marker of early, reversible axonal dysfunction preceding RGC death in glaucoma and optic nerve diseases.64