SOCE Research Articles

Mechanism of 2-aminoethoxydiphenylboronic acid in acetylcholine-inducing Ca2+ signals in acute dissociated mouse pancreatic acinar cells

Objective To reveal the mechanism of 2-aminoethoxydiphenyl borate (2APB) modulates intracellular Ca2+ oscillations. Methods Intracellular Ca2+ signals was monitored directly using Ca2+ -dimensional dye (fluo-4) imaging technique in acute dissociated mouse pancreatic acinar cells. The effect and mechanism of 2APB on acetylcholine (Ach)-induced Ca2+ oscillations were examined by patch-clamp whole cell recording. Results (1) 2APB-induced modulations in acinar cell Ca2+ signals were dependent on 2APB concentrations, with increase of 2APB concentrations (1-100 μmol/L), the amplitude and width of Ca2+ oscillations were increased. (2) 2APB enhanced ACh-induced Ca2+ oscillations were also dependent on the concentrations of ACh. When ACh was less than 5 nmol/L, 2APB did not show effect, between ACh 10-30 nmol/L, 2APB potentiated ACh-induced oscillations, while ACh was higher than 100 nmol/L, 2APB inhibited Ca2+ responses. (3) 2APB enhanced ACh-induced Ca2+ oscillations through extracellular rather than intracellular targets because no effect was found by intracellular application of 100 μmol/L 2APB on 10 nmol/L ACh-induced Ca2+ oscillations (P=0.378, n=6); (4) 2APB significantly blocked SOCE, which can reduced the 3 μmol/L TG-induced Ca2+ oscillations current to (3.0±4.8)% (P=0.000, n=6), suggesting that SOCE may be the target of 2APB to enhance Ca2+ oscillations. (5) SOCE blocker GSK can mimic 2APB’s effect to enhance Ca2+ oscillations (P=0.000, n=7). (6) Using thapsigargin to empty Ca2+ pools, the Ca2+ oscillations current become (93.0±9.5)% (P=0.427, n=7) when 2APB added. In other words, 2APB’s effect was eliminated, suggesting that 2APB-induced Ca2+ oscillations is dependent on Ca2+ pool’s conditions. Conclusion At low ACh concentrations (10-30 nmol/L) (where Ca2+ pool is partially emptied and SOCE is open), 2APB blocks SOCC and promotes Ca2+ further release from Ca2+ pool, resulting in Ca2+ oscillation enhancement. Wherease at higher ACh concentrations (>100 nmol/L), 2APB blocks SOCC and prevents Ca2+ pool refilling from extracellular Ca2+ , thus reduces Ca2+ signal response. Key words: 2-aminoethoxydiphenyl borate; Calcium signals; Pancreatic acinar cells; Patch clam

Acetylcholine induces nitric oxide production by inducing intracellular Ca2+ oscillations in mouse brain endothelial cells

Basal forebrain neurons control intracortical arterioles by releasing acetylcholine (Ach), which stimulates endothelial cells (ECs) to produce the vasodilating gasotransmitter, nitric oxide (NO). Surprisingly, the mechanism by which Ach induces NO synthesis in brain ECs is still unknown. An increase in intracellular Ca2+ concentration ([Ca2+]i) recruits a multitude of endothelial Ca2+-dependent pathways, such as Ca2+/Calmodulin endothelial NO synthase (eNOS). The present investigation sought to investigate the role of intracellular Ca2+ signaling in Ach-induced NO production in bEnd5 cells, an established model of mouse brain microvascular ECs. Ach induced dose-dependent asynchronous Ca2+ oscillations in bEnd5 cells. Ach-evoked Ca2+ oscillations did not arise in the absence of external Ca2+ but rapidly resumed on Ca2+ restitution to the bath. However, nicotine, a selective agonist of the Ca2+-permeable nicotinic receptors, did not cause any detectable increase in [Ca2+]i. Pharmacological manipulation indeed revealed that Ach-induced Ca2+ spikes in bEnd5 cells are triggered by the interaction between intracellular Ca2+ release from InsP3 receptors (InsP3Rs) SOCE. Next, we found that Ach-induced NO production was hindered by L-NAME, a selective NOS inhibitor, and BAPTA, a membrane permeable intracellular Ca2+ buffer. Moreover, Ach-elicited NO synthesis was blocked by the pharmacological abrogation of the accompanying Ca2+ spikes. Ach stimulates bEnd5 cells by inducing a burst of intracellular Ca2+ spikes which is patterned by the interplay between ER-dependent Ca2+ mobilization and SOCE. Ach-elicited Ca2+ spikes result in NO production and are, therefore, predicted to control local CBF in mouse brain. Future experiments will assess whether this signaling pathway is altered in neurodegenerative disorders, such as Alzheimer’s Disease.

Attenuated store-operated Ca2+ entry in mesenteric artery endothelial cells of aged rat

Objective To observe attenuated store-operated Ca2+ entry in superior mesenteric artery endothelial cells of aged rat. Methods The pressure myograph was applied to measure the vasodilation of superior mesenteric artery induced by bradykinin (BK) in aged versus non-elder rats. Primarily cultured MAECs were treated by thapsigargin (TG) and BK to deplete intracellular Ca2+ stores for comparing SOCE. SOCE of MAECs was detected by calcium imaging technology and was compared between aged and non-elder rats. The expression levels of Orai 1 and stromal interaction factor 1 (STIM 1) were observed by immunofluorescence method. Results Compared with the non-elder rats (70.0±4.1%), BK-induced vasodilation in aged rats (27.0±4.9)% was declined by (60.7±4.3)%. SOCE induced by BK and TG in primarily cultured MAECs was attenuated by 37.6% and 39.2% in aged rats (1.71±0.18 and 1.06±0.03) respectively as compared with non-elder rats (2.72±0.39 and 1.75±0.06). The expressions of SOCE's components Orai 1 and STIM 1 in MAECs were decreased obviously in aged rats than in non-elder rats. Conclusions SOCE of MAECs and SOCE-induced vasodilation are significantly attenuated in aged rats. The decreased expression level of Orai 1 and STIM 1 may contribute to this alteration. Key words: Mesenteric artery superior; Endothelial cells; Calcium

Matrix Elements in Fermion Dynamical Symmetry Model**The project supported in part by National Natural Science Foundation of China under Grant No. 19 275 017

In a neutron–proton system, the matrix elements of the generators for SO(8) × SO(8) symmetry are constructed explicitly, and with these matrix elements the low-lying excitation spectra obtained by diagonalization are presented. The excitation spectra for SO(7) nuclei Pd and Ru isotopes and SO(6) r-soft rotational nuclei Xe, Ba, and Ce isotopes are calculated, and comparison with the experimental results is carried out.