Simultaneous multicompartment intracellular Ca²⁺ measurements in the perfused heart using ¹⁹F NMR spectroscopy

Abstract

Although Ca2+ transport regulation at subcellular organelles is of great interest, only limited methodology has been available for measuring organellar [Ca2+] levels. The present study employs the 19F NMR resonance frequency of 4F-BAPTA to measure free [Ca2+]. In 4F-BAPTA loaded perfused rabbit hearts, two 19F NMR resonances were clearly observed. The frequency of one was consistent with cytosolic [Ca2+] levels. Responses to agents that after sarcoplasmic reticulum function identified the other resonance as originating from that organelle. The experiments demonstrate the utility of NMR shift indicator methodology in obtaining simultaneous multi-compartment intracellular [Ca2+] measurements and in enabling organellar [Ca2+] measurements to be made from within intact living tissue.

A method for multiparametric statistical quantification of the heterogeneity of free Na+concentration by19F MR spectroscopy: Proof of principlein silicoandin vitro

“…The molecular weights of these BAPTA derivatives are of the same order of magnitude as that of FCrown‐1 (≈ 500 Da), thus suggesting similar line widths. Indeed, 19 F MR spectra from FCrown‐1 solutions exhibited line widths no larger than those from BAPTA‐FF or 4F‐BAPTA solutions at comparable field strengths. Consequently, useful 5‐min 19 F MR acquisitions should also be feasible for in vivo FCrown‐1 experiments.…”

“…The sensitivity of future in vivo experiments with fluorinated Na + reporter molecules can readily be estimated based on similar experiments using fluorinated Ca 2+ reporter molecules. For instance, it has been demonstrated that useful 19 F MR spectra of BAPTA‐FF can be obtained within five minutes on a 9.4 T magnet from perfused rabbit or rat heart, as can 4‐FBAPTA spectra on a 8.5 T magnet, and 5F‐BAPTA spectra from intact ferret heart on the same magnet. The molecular weights of these BAPTA derivatives are of the same order of magnitude as that of FCrown‐1 (≈ 500 Da), thus suggesting similar line widths.…”

Aggregation-Induced Emission Probe for Light-Up and in Situ Detection of Calcium Ions at High Concentration

“…For example, the concentration of free Ca 2+ ions in cytoplasm and extracellular fluids varied from nanomolar (nM) to millimolar (mM) range. 3 Although various methods have been developed for the detection of Ca 2+ ions, such as atomic absorption spectrometry, 4 colorimetric titration, 5 ionselective electrode, 6 and 19 F NMR spectroscopy, 7,8 they suffer from expensive equipments and complicated procedures for sample pretreatment. Compared with these traditional methods, fluorescent sensing has advantages in terms of high sensitivity, low cost, and easy operation.…”

Multiparametric quantification of the heterogeneity of free Ca2+ concentration by 19F MR spectroscopy

“…Nonetheless, we assess the potential of 4F-BAPTA by numerically modeling 19 F MR spectra as a function of Ca 2+ concentration. These are not ab-initio, but empirical simulations; both chemical shift and linewidth values are taken from actual 19 F MR spectroscopy experiments previously conducted on 4F-BAPTA solutions in the presence of varying Ca 2+ concentrations [5,21], at a time when 4F-BAPTA was still available commercially. This bestows high practical relevance upon our calculated models.…”

“…Our phantom [Ca 2+ ] concentrations were chosen as to be within the range characteristic of realworld tissue exhibiting elevated [Ca 2+ ] i values. For instance, Ca 2+ concentration attributed to heterogeneous sarcoplasmic reticulum populations in perfused rabbit heart has been shown to decrease from about 1.5 to 0.3 mM[11], and even further down to < 0.1 mM[21] upon perfusion with a Ca 2+ -ATPase inhibitor. Thus, our BAPTA-FF solutions prepared to model unimodal and bimodal [Ca 2+ ] distributions (0.18 and 0.31 mM Ca2+) were within the [Ca 2+ ] range given in these references[11,21].Moreover, the [Ca 2+ ]-sensitive 5F/5'F resonance of this compound is also more broadened by spurious effects (essentially due to fast-to-intermediate exchange with [Ca 2+ ]).…”