1991, The British Journal of Radiology, 64, 768-770

Short communications

Effect of radiological iodinated contrast media on acetylcholinesterase activity By Akira Tamura, PhD, *Teruyasu Suzuki, MD, *Toshie Yamasaki, MD, *Rikushi Morita, MD, tTakashi Sato, PhD and tTatsuzo Fujii, PhD Department of Food and Nutrition, Faculty of Home Economics, Chukyo Women's University, Ohbu, Aichi 474, "Department of Radiology, Shiga University of Medical Science, Ohtsu, Shiga 520-21 and tDepartment of Biochemistry, Kyoto Pharmaceutical University, Yamashina, Kyoto 607, Japan {Received October 1990 and in revised form January 1991) Keywords: Contrast media, Acetylcholinesterase, Enzyme inhibition, Ioxaglate

The inhibitory effects of several contrast media on acetylcholinesterase (AchE) activity were studied in vitro. Their inhibiting power was in the order: iothalamate > ioxaglate > iopamidol > iohexol. Because both the Km and maximal velocity (Fmax) values of the enzyme were altered on adding ioxaglate, the inhibition mode seemed to be of a mixed type. Complete recovery from such inhibition was observed by removing the contrast media from the enzyme. Adding albumin (1%) to the reaction mixture led to a decrease in the inhibitory effect of ioxaglate. These results suggest that the contrast media, which have an inhibitory effect on AchE activity in vitro, may have no influence on its activity in blood which contains 4 - 5 % of albumin under physiological conditions. Radiological iodinated contrast media are relatively nontoxic, but they are employed in such large doses that mild toxic effects are common in clinical practice, and severe reactions sometimes occur (Labbe & Peyroux, 1984; Dawson, 1983, 1985). Since some symptoms of their toxicity such as peripheral vasodilation, bronchospasm and disturbances of cardiac rhythm are similar to those produced by acetylcholine, their inhibitory effects on AchE (acetylcholine acetylhydrolase, EC 3117) have been studied (Lasser & Lang, 1966; Dawson & Edgerton, 1983; Howell & Dawson, 1985; Guidollet et al, 1988). The published studies, however, have yielded conflicting results. In this short communication, we report more precisely the nature of the inhibition of AchE by some representative ionic and non-ionic contrast media.

colourimetric method (Ellman et al, 1961). Enzyme solution containing DTNB and 0 - 1 % albumin was incubated with varying concentrations of contrast medium (20-160 mg iodine/ ml) or with phosphate buffer (0.1M phosphate buffer, pH 8.0, as a control) at 25°C for 10-30 min. After the absorbance at 412 nm had reached a steady state, the substrate was added to the mixture and the absorbance increase for 10 min at 25°C was measured. The procedure in which the intact erythrocytes were used as an enzyme source was as follows: a washed cell suspension (haematocrit value of 0.04%) was mixed with ioxaglate (80 mg iodine/ml) and incubated at 25°C for 10 min. After incubation, an aliquot of the mixture was centrifuged at 1000 g for 5 min, the supernatant was discarded, and then the precipitated cells were washed twice with PBS. The AchE activities in the erythrocytes both before (in the presence of contrast medium) and after washing out the contrast medium were measured. Results and discussion The inhibitory effects of the contrast media on AchE activity are shown in Fig. 1. The enzyme activity is plotted against iodine concentration of the reaction mixture as a concentration

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Materials and methods Contrast agents The contrast media studied were iohexol (Nycomed Ltd) and iopamidol (Schering Health Care Ltd) as the non-ionic agents and sodium iothalamate (Dai-ichi Seiyaku Co.) and sodium meglumine ioxaglate (Guerbet Co.), a new low-osmolality ionic medium, as the ionic agents. Reagents AchE (partially purified from bovine erythrocytes) was obtained from Wako Pure Chemicals Co. (Japan). Intact human erythrocytes from healthy volunteers were also used as a source of the enzyme (Herz & Kaplan, 1973). Human serum albumin (fraction V, 96-99%) was obtained from Sigma Chemical Co. (USA). Acetylthiocholine as a substrate, dithiobis-2-nitrobenzoic acid (DTNB) as a chromogen and other reagents were of a pure grade. Method Erythrocytes were washed with phosphate buffered saline (PBS) (140.5 mM NaCl, 10 mM phosphate buffer, pH 7.4) and resuspended in the PBS. AchE activity was measured by a

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50 100 150 Concentration of the contrast media as expressed by iodine concentration (mg/ml) Figure 1. Inhibitory effects of contrast media on AchE activity. AchE (partially purified from bovine erythrocyte) activity determined in the presence of iodinated contrast media (Iohexol, • ; iopamidol, A ; sodium meglumine ioxaglate, # ; sodium iothalamate, A)-

The British Journal of Radiology, August 1991

Short communications

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of the contrast medium (mg iodine/ml). Their inhibiting power was in the order: iothalamate > ioxaglate > iopamidol > iohexol. The results obtained here are in good agreement with those reported previously by Dawson & Edgerton (1983). Figure 2 shows the Lineweaver-Burk plot of the data from the inhibition studies with ioxaglate. It can be seen that the mode of inhibition was of a mixed type, because both the apparent Km and Vmax values were affected on adding ioxaglate. Since ioxaglate and the other contrast media tested are structurally unrelated to the substrates of AchE they probably act by producing a sufficient conformational change of the enzyme protein to reduce, or eliminate, its activity rather than by directly binding to the active site. The inhibitory effects brought about by non-specific binding of contrast media to enzyme molecules have already been reported with several enzymes (Lasser & Lang, 1970a, b, 1971; Canillot et al, 1983; Guidollet et al, 1988). Binding of the contrast media to serum proteins, not only to some enzymes but also to albumin molecules, has been reported (Lasser et al, 1962). If such binding of the contrast media to both albumin and AchE is non-specific, addition of albumin to the reaction mixture containing the contrast media would suppress their inhibitory effect on AchE activity. Figure 3 shows the effect on the inhibition of AchE by ioxaglate when albumin was added. A marked depression in its inhibitory effect was observed in the presence of albumin at a concentration of 1% or less. The reduced inhibition may result from a decrease in the number of molecules of the contrast medium bound to the enzyme protein because of the presence of a large number of albumin molecules. It is possible that such easy binding of these contrast media to macromolecules in the body may cause toxicity. These results, however, also suggest that they may not actually suppress AchE activity in blood

Vol. 64, No. 764

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Effect of radiological iodinated contrast media on acetylcholinesterase activity.

1991, The British Journal of Radiology, 64, 768-770 Short communications Effect of radiological iodinated contrast media on acetylcholinesterase act...
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