Store-operated Ca2+ entry is mediated by plasma membrane store-operated Ca2+ channels (SOCCs) activated due to the depletion of endoplasmic reticulum (ER) Ca2+ stores. Activation of SOCCs is required for Ca2+signaling and ER refilling which both important for the protein synthesis and exocytosis. We measured free Ca2+ concentration in the cytosol ([Ca2+]cyt), inside the ER ([Ca2+]ER) and in mitochondria ([Ca2+]mit) using Fura-2/AM, Mag-fura-2/AM and Rhod-2/AM fluorescent dyes correspondingly. Experiments were performed on male Wistar rats (6-7 weeks old, 100-150 g). Gland was removed after intraperitoneal injection of pentobarbital. Submandibular salivary gland acinar cells were isolated by collagenase treatment. In the present study we first stimulated the acinar cells with acetylcholine (ACh) in Ca2+ free media (to deplete the ER and activate the store-operated Ca2+ channels (SOCCs)) and then readded Ca2+ to the media (to vizualize Ca2+entry). We showed that the amplitude and the initial rate of the ACh induced SOCC-mediated [Ca2+]i transients clearly depend on the potency and the duration of agonist stimulation. In particular, the initial slope and amplitude of [Ca2+]i transients were significantly decreased upon inhibition of mitochondria (Mit) Ca2+ uptake or Ca2+ release from the Mit. The latter indicate the decrease number of active SOCCs. In the whole cell patch-clamp experiments we directly showed that [Ca2+]mit transients are generated immediately after the SOCCs activation and their kinetics directly correlate with the potency of agonist stimulation. Notably, we found that under conditions of sustained ACh stimulation adding of BAPTA to the pipette solution caused dramatic decrease in the amplitude and the initial slope of the SOCE-induced [Ca2+]mit transients. In addition we showed that in the conditions of continious ACh stimulation the inhibition of Mit Ca2+ buffering caused almost complete inhibition of SOCCs Ca2+ influx the ER reffiling whereas upon short ACh action inhibition was much smaller. In summary, Mit are crucial for maintaining of SOC-mediated Ca2+ entry; ii) subplasmalemmal Mit effectively grade Ca2+ entry due to prevention of Ca2+-dependent inactivation of SOCCs; iii) in the condition of sustained ACh stimulation SOCE is dramatically increased in a direct proportion to potency of ER depletion; cell adapts its Ca2+ clearance system directing more Ca2+ into mitochondria via microdomains of high [Ca2+]cyt, providing strong positive feedback on SOCE. Furthermore, in the continuous presence of an agonist, Ca2+ transport inside mitochondria and its release through the Na+/Ca2+ exchanger supply ~90% of Ca2+ for ER refilling; whereas in the absence of an agonist ~30%. Robustly increased role of mitochondria Ca2+ dynamics in the maintenance of sustained SOCE and ER refilling represents a novel function of mitochondria as an intrinsic instrument enabling long lasting secretion.