Endoplasmic reticulum potassium-hydrogen exchanger and small conductance calcium-activated potassium channel activities are essential for ER calcium uptake in neurons and cardiomyocytes

Calcium pumping into the endoplasmic reticulum (ER) lumen is thought to be coupled to a countertransport of protons through sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) and the members of the ClC family of chloride channels. However, pH in the ER lumen remains neutral, which suggests a mechanism responsible for proton re-entry. We studied whether cation–proton exchangers could act as routes for such a re-entry. ER Ca 2+ uptake was measured in permeabilized immortalized hypothalamic neurons, primary rat cortical neurons and mouse cardiac fibers. Replacement of K + in the uptake solution with Na + or tetraethylammonium led to a strong inhibition of Ca 2+ uptake in neurons and cardiomyocytes. Furthermore, inhibitors of the potassium–proton exchanger (quinine or propranolol) but not of the sodium–proton exchanger reduced ER Ca 2+ uptake by 56–82%. Externally added nigericin, a potassium–proton exchanger, attenuated the inhibitory effect of propranolol. Inhibitors of small conductance calcium-sensitive K + (SK Ca ) channels (UCL 1684, dequalinium) blocked the uptake of Ca 2+ by the ER in all preparations by 48–94%, whereas inhibitors of other K + channels (IK Ca , BK Ca and K ATP ) had no effect. Fluorescence microscopy and western blot analysis revealed the presence of both SK Ca channels and the potassium–proton exchanger leucine zipper-EF-hand-containing transmembrane protein 1 (LETM1) in ER in situ and in the purified ER fraction. The data obtained demonstrate that SK Ca channels and LETM1 reside in the ER membrane and that their activity is essential for ER Ca 2+ uptake.