Clearance of store-released Ca^sup 2+^ by the Na+-Ca^sup 2+^ exchanger is diminished in aortic smooth muscle from Na+-K+-ATPase [alpha]^sub 2^-isoform gene-ablated mice

Two α-isoforms of the Na...-K...-ATPase are expressed in vascular smooth muscle cells (VSMCs). The α...-isoform is proposed to serve a cytosolic housekeeping role, whereas the α...-isoform modulates Ca... storage via coupling to the Na...-Ca... exchanger (NCX) in a subsarcolemmal compartment. To evaluate the ramifications of this proposed interaction, Ca...-store load and the contributions of the primary Ca... transporters to Ca... clearance were studied in aortic VSMCs from embryonic wild-type (WT) and Na...-K...-ATPase α...-isoform gene-ablated, homozygous null knockout (α...-KO) mice. Ca... stores were unloaded by inhibiting the sarco(endo)plasmic reticulum Ca...-ATPase with cyclopiazonic acid (CPA) in Ca...-free media to limit Ca... influx. Ca... clearance by the plasma membrane Ca... ATPase (PMCA), NCX, or mitochondria was selectively inhibited. In WT VSMCs, NCX accounted for 90% of the Ca... efflux. In α...-KO VSMCs, preferential clearance of store-released Ca... by NCX was lost, whereas PMCA activity was increased. Selective inhibition of the α...-isoform (0.5 ...M ouabain for 20 min), before treatment with CPA enhanced the store load in VSMCs from WT, but not α...-KO mice. A subsequent analysis of capacitative Ca... entry (CCE) indicated that the magnitude of Ca... influx was significantly greater in α...-KO cells. Our findings support the concept of a subsarcolemmal space where the α...-isoform coupled with NCX modulates Ca...-store function and, thereby, CCE. (ProQuest: ... denotes formulae/symbols omitted.)