BAPTA

BAPTA is a specific chelator of Ca2+, suppresses intracellular reactive oxygen species (ROS) levels.

In Vitro:Regarding ROS generation, a Ca2+ specific chelator, BAPTA, suppresses ROS generation of Sodium lauryl sulfate (SLS)-exposed HaCaT keratinocytes[1]. Depolarization does not increase the resting open probability of the mechanoelectrical transducer (MET) current of Tmc1Bth/Bth OHCs, whereas raising the intracellular concentration of the Ca2+ chelator BAPTA causes smaller increases in resting open probability in Bthmutant outer hair cells (OHCs) than in wild-type control cells. In the presence of 0.1 mM BAPTA, nonsaturating bundle displacements causes the MET current to adapt in both genotypes, exactly as seen when 1 mM EGTA is used in the intracellular solution. In the presence of 10 mM intracellular BAPTA, the time-dependent MET current decline is abolished and the resting Popen increased to near 50% of the maximal MET current in OHCs from both Tmc1+/+ and Tmc1Bth/Bth mice. The relation between the MET current and bundle displacement shows that increasing the intracellular BAPTA concentration from 0.1 to 10 mM significantly increased (p<0.0001) the resting Popen of the MET current in both Tmc1+/+ (0.1 mM, 8±1.6%, n=4; 10 mM, 39.6±2.7%, n=5) and Tmc1Bth/Bth (0.1 mM, 10.4±2.2%, n=3; 10 mM, 46.5±9.9%, n=6). No significant differences are seen between the two genotypes for both BAPTA concentrations. However, 3 and 5 mM BAPTA are less effective in shifting the MET current-bundle displacement curves in Tmc1Bth/Bth than in Tmc1+/+ OHCs. In Tmc1+/+, increasing the BAPTA concentration from 0.1 mM to either 3 or 5 mM produces a highly significant increase in Popen (post hoc test from one-way ANOVA, p<0.01 and p<0.001, respectively); in Tmc1Bth/Bth, the same comparison produced no or a much reduced increase in Popen (n.s. and p<0.05, respectively)[2].

References:
[1]. Mizutani T, et al. Sodium Lauryl Sulfate Stimulates the Generation of Reactive Oxygen Species through Interactions with Cell Membranes. J Oleo Sci. 2016 Dec 1;65(12):993-1001. [2]. Corns LF, et al. Tmc1 Point Mutation Affects Ca2+ Sensitivity and Block by Dihydrostreptomycin of the Mechanoelectrical Transducer Current of Mouse Outer Hair Cells. J Neurosci. 2016 Jan 13;36(2):336-49.

Aspirin-induced heat stress resistance in chicken myocardial cells can be suppressed by BAPTA-AM in vitro.[Pubmed:27262845]

Cell Stress Chaperones. 2016 Sep;21(5):817-27.

Our recent studies have displayed the protective functions of aspirin against heat stress (HS) in chicken myocardial cells, and it may be associated with heat shock proteins (HSPs). In this study, we further investigated the potential role of HSPs in the aspirin-induced heat stress resistance. Four of the most important HSPs including HspB1 (Hsp27), Hsp60, Hsp70, and Hsp90 were induced by aspirin pretreatment and were suppressed by BAPTA-AM. When HSPs were induced by aspirin, much slighter HS injury was detected. But more serious damages were observed when HSPs were suppressed by BAPTA-AM than those cells exposed to HS without BAPTA-AM, even the myocardial cells have been treated with aspirin in prior. Comparing to other HSPs, HspB1 presented the largest increase after aspirin treatments, 86-fold higher than the baseline (the level before HS). These findings suggested that multiple HSPs participated in aspirin's anti-heat stress function but HspB1 may contribute the most. Interestingly, during the experiments, we also found that apoptosis rate as well as the oxidative stress indicators (T-SOD and MDA) was not consistently responding to heat stress injury as expected. By selecting from a series of candidates, myocardial cell damage-related enzymes (CK-MB and LDH), cytopathological tests, and necrosis rate (measured by flow cytometry assays) are believed to be reliable indicators to evaluate heat stress injury in chicken's myocardial cells and they will be used in our further investigations.

Data on the concentrations of etoposide, PSC833, BAPTA-AM, and cycloheximide that do not compromise the vitality of mature mouse oocytes, parthenogencially activated and fertilized embryos.[Pubmed:27547800]

Data Brief. 2016 Jul 30;8:1215-20.

These data document the vitality of mature mouse oocytes (Metaphase II (MII)) and early stage embryos (zygotes) following exposure to the genotoxic chemotherapeutic agent, etoposide, in combination with PSC833, a selective inhibitor of permeability glycoprotein. They also illustrate the vitality of parthenogencially activated and fertilized embryos following incubation with the calcium chelator BAPTA-AM (1,2-Bis(2-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid tetrakis (acetoxymethyl ester)), cycloheximide (an antibiotic that is capable of inhibiting protein synthesis), and hydrogen peroxide (a potent reactive oxygen species). Finally, they present evidence that permeability glycoprotein is not represented in the proteome of mouse spermatozoa. Our interpretation and discussion of these data feature in the article "Identification of a key role for permeability glycoprotein in enhancing the cellular defense mechanisms of fertilized oocytes" (Martin et al., in press) [1].

BAPTA-AM decreases cellular pH, inhibits acidocalcisome acidification and autophagy in amino acid-starved T. brucei.[Pubmed:28274857]

Mol Biochem Parasitol. 2017 Apr;213:26-29.

To investigate the role of Ca(2+) signaling in starvation-induced autophagy in Trypanosoma brucei, the causative agent of human African trypanosomiasis, we used cell-permeant Ca(2+) chelator BAPTA-AM and cell impermeant chelator EGTA, and examined the potential involvement of several intracellular Ca(2+) signaling pathways in T. brucei autophagy. The results showed an unexpected effect of BAPTA-AM in decreasing cellular pH and inhibiting acidocalcisome acidification in starved cells. The implication of these results in the role of Ca(2+) signaling and cellular/organellar pH in T. brucei autophagy is discussed.

A novel combination treatment for breast cancer cells involving BAPTA-AM and proteasome inhibitor bortezomib.[Pubmed:27347145]

Oncol Lett. 2016 Jul;12(1):323-330.

Glucose-regulated protein 78 kDa/binding immunoglobulin protein (GRP78/BIP) is a well-known endoplasmic reticulum (ER) chaperone protein regulating ER stress by facilitating protein folding, assembly and Ca(2+) binding. GRP78 is also a member of the heat shock protein 70 gene family and induces tumor cell survival and resistance to chemotherapeutics. Bortezomib is a highly specific 26S proteasome inhibitor that has been approved as treatment for patients with multiple myeloma. The present study first examined the dose- and time-dependent effects of bortezomib on GRP78 expression levels in the highly metastatic mouse breast cancer 4T1 cell line using western blot analysis. The analysis results revealed that GRP78 levels were significantly increased by bortezomib at a dose as low as 10 nM. Time-dependent experiments indicated that the accumulation of GRP78 was initiated after a 24 h incubation period following the addition of 10 nM bortezomib. Subsequently, the present study determined the half maximal inhibitory concentration of intracellular calcium chelator BAPTA-AM (13.6 microM) on 4T1 cells. The combination effect of BAPTA-AM and bortezomib on the 4T1 cells was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and WST-1 assays and an iCELLigence system. The results revealed that the combination of 10 nM bortezomib + 5 microM BAPTA-AM is more cytotoxic compared with monotherapies, including 10 nM bortezomib, 1 microM BAPTA-AM and 5 microM BAPTA-AM. In addition, the present results revealed that bortezomib + BAPTA-AM combination causes cell death through the induction of apoptosis. The present results also revealed that bortezomib + BAPTA-AM combination-induced apoptosis is associated with a clear increase in the phosphorylation of stress-activated protein kinase/Jun amino-terminal kinase SAPK/JNK. Overall, the present results suggest that bortezomib and BAPTA-AM combination therapy may be a novel therapeutic strategy for breast cancer treatment.

Inhibition of phospholipase C activity in Drosophila photoreceptors by 1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA) and di-bromo BAPTA.[Pubmed:16140375]

Cell Calcium. 2005 Dec;38(6):547-56.

In vivo light-induced and basal hydrolysis of phosphatidyl inositol 4,5-bisphosphate (PIP2) by phospholipase C (PLC) were monitored in Drosophila photoreceptors using genetically targeted PIP2-sensitive ion channels (Kir2.1) as electrophysiological biosensors for PIP2. In cells loaded via patch pipettes with varying concentrations of Ca2+ buffered by 4 mM free BAPTA, light-induced PLC activity, showed an apparent bell-shaped dependence on free Ca2+ (maximum at "100 nM", approximately 10-fold inhibition at <10nM or approximately 1 microM). However, experiments where the total BAPTA concentration was varied whilst free [Ca2+] was maintained constant indicated that inhibition of PLC at higher (>100 nM) nominal Ca2+ concentrations was independent of Ca2+ and due to inhibition by BAPTA itself (IC50 approximately 8 mM). Di-bromo BAPTA (DBB) was yet more potent at inhibiting PLC activity (IC50 approximately 1mM). Both BAPTA and DBB also appeared to induce a modest, but less severe inhibition of basal PLC activity. By contrast, EGTA, failed to inhibit PLC activity when pre-loaded with Ca2+, but like BAPTA, inhibited both basal and light-induced PLC activity when introduced without Ca2+. The results indicate that both BAPTA and DBB inhibit PLC activity independently of their role as Ca2+ chelators, whilst non-physiologically low (<100 nM) levels of Ca2+ suppress both basal and light-induced PLC activity.