Age attenuates the T-type Ca 3.2-RyR axis in vascular smooth muscle.

Caveolae position Ca 3.2 (T-type Ca channel encoded by the α-3.2 subunit) sufficiently close to RyR (ryanodine receptors) for extracellular Ca influx to trigger Ca sparks and large-conductance Ca -activated K channel feedback in vascular smooth muscle. We hypothesize that this mechanism of Ca spark generation is affected by age. Using smooth muscle cells (VSMCs) from mouse mesenteric arteries, we found that both Ca 3.2 channel inhibition by Ni (50 µM) and caveolae disruption by methyl-ß-cyclodextrin or genetic abolition of Eps15 homology domain-containing protein (EHD2) inhibited Ca sparks in cells from young (4 months) but not old (12 months) mice. In accordance, expression of Ca 3.2 channel was higher in mesenteric arteries from young than old mice. Similar effects were observed for caveolae density. Using SMAKO Ca 1.2 mice, caffeine (RyR activator) and thapsigargin (Ca transport ATPase inhibitor), we found that sufficient SR Ca load is a prerequisite for the Ca 3.2-RyR axis to generate Ca sparks. We identified a fraction of Ca sparks in aged VSMCs, which is sensitive to the TRP channel blocker Gd (100 µM), but insensitive to Ca 1.2 and Ca 3.2 channel blockade. Our data demonstrate that the VSMC Ca 3.2-RyR axis is down-regulated by aging. This defective Ca 3.2-RyR coupling is counterbalanced by a Gd sensitive Ca pathway providing compensatory Ca influx for triggering Ca sparks in aged VSMCs.