Campuses:

A Putative Channel that Opens to Drive Spreading Depolarization

Monday, February 12, 2018 - 9:00am - 10:00am
Lind 305
R. David Andrew (Queen's University)
Neurons of the higher brain quickly initiate spreading depolarization (SD) in response to ischemia from heart failure, traumatic brain injury or focal stroke. At SD onset, neurons cease firing, swell and can die within minutes. Accurately modeling SD requires characterizing the underlying current(s). 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, suggesting a minor role for voltage-dependent Na channels. To account for the massive SD current, the 'mystery' channel is either densely distributed in the plasma membrane or has a high unitary conductance. Like ischemic SD itself, this conductance resists blockers of standard voltage- and ligand-gated channels. Na channel blockers merely delay SD onset. We used neocortical slices from adult rat to record channel activity in patches under voltage clamp during oxygen-glucose deprivation (OGD) at 35oC. Using the cell-attached configuration (CAC), patches were recorded during bath superfusion with blockers of Na, K, Ca, 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 min 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). Cell-attached patch recording with 1 pM PLTX in the pipette (again 35oC, 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 patches at 35oC, 1 pM PLTX + blockers in the bath opened this channel (2.5 +/- 0.5 pA; n=11; h= -70 mV), as did aCSF after it has washed over an OGD-exposed slice (3.1 +/- 0.6 pA; n=21; h= -70 mV). These values appear to represent the same conductance.

We propose that OGD induces conversion of the Na/K pump (which is densely distributed in neuronal membrane) into an open, non-selective Na/K channel that then drives ischemic SD. This conductance may also underlie classic cortical spreading depression which generates migraine aura. R. David. Andrew and Peter. J. Gagolewicz. Queen`s University, Canada