Campuses:

A Putative Channel Driving Spreading Depolarization

Monday, February 12, 2018 - 9:00am - 10:00am
Lind 305
R. David Andrew (Queen's University)
Neurons of the higher brain immediately undergo spreading depolarization (SD) in response to ischemia resulting from heart failure, traumatic brain injury or focal stroke. The current(s) driving SD remain unidentified. At SD onset, neurons cease firing, swell and can die within minutes. To account for the massive SD current, the channel has to be either densely distributed in the plasma membrane, display a high unitary conductance, or both. A study of pyramidal neurons using whole-cell voltage clamp (Czeh et al. 1993) showed that the macro-conductance driving SD is inward, cationic, non-selective and reverses near zero millivolts. Like ischemic SD itself, this conductance resists blockers of standard voltage- and ligand-gated channels. Na + channel blockers merely delay SD onset and glutamate receptor antagonists are without effect. We used neocortical slices from adult rat to record channel activity in membrane patches from pyramidal neurons under voltage clamp during oxygen-glucose deprivation (OGD) at 35 o C. Using the cell-attached configuration (CAC), patches were recorded during bath superfusion with blockers of Na + , K + , Ca 2+ , pannexin and glutamate-related channels. The blockers were also included in the recording pipette solution. This silenced all spontaneous channel activity within 2 minutes of commencing a recording. Nonetheless within 6 to 8 minutes of OGD, novel channel opening commenced. The mean unitary current (+/- st. dev.) was 1.7 +/- 0.17 pA at holding potential (h) = -70 mV (n=5 patches). Unitary event frequency increased, as did multiple channel openings, until the patch was lost during full-onset SD. More positive h values reversed the unitary current near 0 mV, implicating a Na + /K + conductance. In support, the channel properties appeared unaltered by substituting K + for Na + in the patch pipette. In the CAC, the channel slope conductance was ~28 pS based on unitary pA values from 23 neurons spanning h= -90 to +50 mV.

The marine poison palytoxin (PLTX) specifically binds externally to the Na + /K + pump, converting it into an open Na + /K + channel. Bath PLTX also induces SD in neocortical slices (10 to 100 nM). CAC patch recording with 1 pM PLTX in the pipette (again, blockers in the pipette and extracellularly) opened a unitary channel of 1.7 +/- 0.3 pA (n=7; h= -70 mV), similar to the OGD-evoked channel described above. In outside-out (o-o) patches, 1 pM PLTX + blockers in the bath appeared to open the same channel (2.5 +/- 0.5 pA; n=11; h= -70 mV). When artificial CSF was washed over an OGD-exposed slice, a similar channel opened (3.1 +/- 0.6 pA; n=21; h= -70 mV). The IV relation plotted from 28 o-o patches under OGD reversed near h= -10 mV, again implicating a Na + /K + conductance.

We propose that OGD induces conversion of the Na + /K + pump into an open Na + /K + channel that then drives ischemic SD. If correct, this conductance likely also underlies classic cortical spreading depressionthat generates migraine aura.