Investigating GluCEST and its specificity for pH mapping at low temperatures

Abstract

Chemical exchange saturation transfer (CEST) from glutamate to water (GluCEST) is a powerful tool for mapping glutamate concentration and intracellular pH. GluCEST could also be helpful to understand the physiology of lower aquatic vertebrates and invertebrates. Therefore, this study aimed to investigate the GluCEST effect and the exchange rate ksw from amine protons of glutamate to water in a broad range of temperatures (1-37°C) and pH (5.5-8.0). Z-spectra were measured from glutamate solutions at different pH values and temperatures and analysed by numerically solving the Bloch-McConnell equation. As expected, a strong dependence of the GluCEST effect and the determined ksw values on pH and temperature was observed. In addition, a strong dependence of the GluCEST effect on phosphate buffer concentration was confirmed. The in vitro data show that GluCEST is detectable in the whole temperature range, even at 1°C. An interpolation function for the exchange rate ksw was determined for the considered range of temperatures and pH values, showing a bijective relation between the exchange rate and pH at a given temperature. To investigate the specificity of GluCEST imaging at low temperatures, the CEST effect was investigated for several metabolites relevant for CEST imaging of the brain. As an example, the contribution of GluCEST to the total CEST effect at 3 ppm was estimated for zebrafish (Danio rerio). It is shown that also at lower temperatures glutamate is the major contributor to the total CEST effect, particularly if the experimental parameters are optimized.

CEST imaging of fast exchanging amine pools with corrections for competing effects at 9.4 T

“…CEST effects can be roughly divided into three categories based on how fast the chemical exchange rates ( k sw ) of the protons are relative to their resonance frequency offsets ( Δ r ): amide protons on the backbone of proteins/peptides ( k sw ≈ 30 s −1 and Δ r = 3.5 ppm (Reference )) are in a slow exchange regime; guanidinium amine protons ( k sw = 500–1000 s −1 and Δ r = 2 ppm (References )) are in an intermediate exchange regime; most other amine protons (e.g. glutamate amine and protein lysine amine protons) and hydroxyl protons ( k sw is several thousand s −1 and Δ r = 1–3 ppm) are in a fast exchange regime. Imaging of fast exchanging amine pools at 3 ppm at high field strength is of special interest, because these protons are part of important molecules such as glutamate and proteins .…”

“…glutamate amine and protein lysine amine protons) and hydroxyl protons ( k sw is several thousand s −1 and Δ r = 1–3 ppm) are in a fast exchange regime. Imaging of fast exchanging amine pools at 3 ppm at high field strength is of special interest, because these protons are part of important molecules such as glutamate and proteins . However, these pools are not well suited to produce CEST signals, because they break the CEST condition that exchange should be slow‐intermediate on the NMR time scale (i.e.…”

“…To remove the DS and semi‐solid MT effects, a reference signal that ideally has the same contributions from DS and non‐specific semi‐solid MT effects but without chemical exchange is required to compare to the exchange‐labeled signal. The magnetization transfer ratio (MTR) may then be calculated as the difference between the labeled and reference signals normalized to a control signal with no saturating pulses or a reference signal, which assumes that CEST, DS, and semi‐solid MT effects add linearly. MTR asym describes this ratio when the reference signal is obtained from the offset frequency symmetric about the water resonance.…”

“…MTR asym describes this ratio when the reference signal is obtained from the offset frequency symmetric about the water resonance. MTR asym has been previously used to quantify fast exchanging amine CEST signals at high irradiation powers . However, recent studies have shown that the CEST, DS, and non‐specific semi‐solid MT effects have mutual interactions, and do not add linearly .…”

Temperature dependence of 1H NMR chemical shifts and its influence on estimated metabolite concentrations

“…This study focused on the upfield signals of important brain metabolites exhibiting also downfield signals of amine protons, because an accurate quantification based on the upfield signals of these metabolites is essential for evaluating CEST effects [20,48,49]. In the future, the temperature dependence of chemical shifts of other brain metabolites has to be studied to build up a complete database for MR quantification of data measured at a certain temperature.…”