Trans-synaptic increase of hypoxic tolerance in hippocampus upon physical challenge with two-photon microscopy

Neuronal hypoxic tolerance is modulated by preceding challenges. We investigated hypoxic tolerance in CA1 pyramidal cells of murine hippocampal slices upon preceding physical challenge with two-photon illumination in close spatial proximity to the recorded area, at distant presynaptic neurons, or preceding chemical treatment with acetylsalicylic acid while zinc fluorescence was assessed with fluorescence measurement upon staining with N-(6-methoxy-8-quinolyl)-para-toluenesulfonamide (TSQ). Posthypoxic recovery (15 min hypoxia, 45 min recovery) of CA1 population spike amplitude (PSAP) upon stimulation of Schaffer collaterals in hippocampal region CA3 was 20 +/- 38% (mean +/- SD; n = 15) in control slices. At the end of hypoxia, zinc fluorescence increased to 120 +/- 16% (P < 0.05 to control) in slices that later recovered and 141 +/- 20% in slices that did not recover (P < 0.01 to control; P < 0.05 compared with returns). Multi-photon illumination alone was an appropriate physical challenge to improve hypoxic tolerance, even trans-synaptically. Depending on the number of illuminations posthypoxic PSAP increased up to 84 +/- 25% (P < 0.01 to control) upon illumination of hippocampal region CA1 and 85 +/- 28% (P < 0.01 to control) upon illumination of CA3. With the latter treatment, zinc fluorescence in CA1 increased to 126 +/- 20% before hypoxia (P < 0.05 to control), and no further zinc increase was observed upon subsequent hypoxia. Similar results were obtained upon chemical preconditioning with acetylsalicylate. We conclude that observation of live specimen with multi-photon imaging alters the physiology of neuronal cell ensembles, including hypoxic tolerance, even trans-synaptically at long distances from the imaged area. This is mediated in part through endogenous modulation by zinc. Mild zinc increase improves hypoxic tolerance while pronounced increase predicts neuronal cell death.