세포내 칼슘 조절 기전에 대한 연구

Abstract

The present study was designed to investigate the role in intracellular calcium mobilization of two representative endoplasmic reticulum (ER) proteins, STIM1 (stromal interaction molecule 1) and SARAF (store-operated calcium entry associated regulatory factor), in pacemaker activity of cultured interstitial cells of Cajal (ICCs) isolated from mouse small intestine, and to clarify the mechanism of actions. The whole-cell patch clamp technique was applied for analysis of intracellular calcium mobilization in cultured ICCs, which overexpressed STIM1 or SARAF. In current-clamping mode, the ICCs generated spontaneous pacemaker potentials. External carbachol, one of cholinceptor agonists, continuously produced membrane depolarization, which was recovered within a few seconds into the level of control resting membrane potentials. In STIM1-overexpressed ICCs, frequency of pacemaker potential was markedly elevated, while in SARAF-overexpressed ICCs, significantly inhibited. To check connection between external calcium and pacemaker potential frequency, when treated with gadolinium (a non-selective cation channel blocker) or a calcium-free solution into culture media, the generation of pacemaker potentials was completely abolished. Also, in order to establish relationship between intracellular calcium concentration and STIM1 or SARAF, when given with Fluo-4AM (a calcium indicator), frequency of spontaneous intracellular calcium oscillations was significantly elevated, but rather distinctly inhibited in SARAF-overexpressed ICCs. Taken together, these results suggest that the frequency of pacemaker activity in cultured ICCs isolated from mouse small intestine is regulated through ER proteins (STIM1 and SARAF) due to calcium mobilization. Moreover, based on these findings, it is thought that expression levels of STIM1 and SARAF may affect slow wave patterns in the gastrointestinal tract.