Advanced age protects microvascular endothelium from aberrant Ca²⁺ influx and cell death induced by hydrogen peroxide

Endothelial cell Ca²⁺ signalling is integral to blood flow control in the resistance vasculature yet little is known of how its regulation may be affected by advancing age. We tested the hypothesis that advanced age protects microvascular endothelium by attenuating aberrant Ca²⁺ signalling during oxidative stress. Intact endothelial tubes (width, ~60 μm; length, ∼1000 μm) were isolated from superior epigastric arteries of Young (3-4 months) and Old (24-26 months) male C57BL/6 mice and loaded with Fura-2 dye to monitor [Ca²⁺]_i. At rest there was no difference in [Ca²⁺]_i between age groups. Compared to Young, the [Ca²⁺]_i response to maximal stimulation with acetylcholine (3 μm, 2 min) was ∼25% greater in Old, confirming signalling integrity with advanced age. Basal H₂O₂ availability was ∼33% greater in Old while vascular catalase activity was reduced by half. Transient exposure to elevated H₂O₂ (200 μm, 20 min) progressively increased [Ca²⁺]_i to ∼4-fold greater levels in endothelium of Young versus Old. With no difference between age groups at rest, Mn²⁺ quench of Fura-2 fluorescence revealed 2-fold greater Ca²⁺ influx in Young during elevated H₂O₂; this effect was attenuated by ∼75% using ruthenium red (5 μm) as a broad-spectrum inhibitor of transient receptor potential channels. Prolonged exposure to H₂O₂ (200 μm, 60 min) induced ∼7-fold greater cell death in endothelium of Young versus Old. Thus, microvascular endothelium can adapt to advanced age by reducing Ca²⁺ influx during elevated oxidative stress. Protection from cell death during oxidative stress will sustain endothelial integrity during ageing. copy; 2015 The Physiological Society.