Alteration of A 3 adenosine receptors in human neutrophils and low frequency electromagnetic fields

Abstract

The present study was designed to evaluate the binding and functional characterization of A 3 adenosine receptors in human neutrophils exposed to low frequency, low energy, pulsing electromagnetic fields (PEMFs). Great interest has grown concerning the use of PEMF in the clinical practice for therapeutic purposes strictly correlated with inflammatory conditions. Saturation experiments performed using the high affinity and selective A 3 adenosine antagonist 5 N-(4-methoxyphenyl-carbamoyl)amino-8-propyl-2-(2-furyl)pyrazolo-[4,3- e]-1,2,4-triazolo[1,5- c]pyrimidine ([ 3 H ]-MRE 3008F20) revealed a single class of binding sites with similar affinity in control and in PEMF treated human neutrophils ( K D =2.36±0.16 and 2.45±0.15 nM, respectively). PEMFs treatment revealed that the receptor density was statistically increased ( P<0.01) ( B max=451±18 and 736±25 fmol mg −1 protein, respectively). Thermodynamic data indicated that [ 3 H ]-MRE 3008F20 binding in control and in PEMF-treated human neutrophils was entropy and enthalpy driven. Competition of radioligand binding by the high affinity A 3 receptor agonists, N 6-(3-iodo-benzyl)-2-chloro-adenosine-5′- N-methyluronamide (Cl-IB-MECA) and N 6-(3-iodo-benzyl)adenosine-5′- N-methyl-uronamide (IB-MECA), in the absence of PEMFs revealed high and low affinity values similar to those found in the presence of PEMFs. In both experimental conditions, the addition of GTP 100 μM shifted the competition binding curves of the agonists from a biphasic to a monophasic shape. In functional assays Cl-IB-MECA and IB-MECA were able to inhibit cyclic AMP accumulation and their potencies were statistically increased after exposure to PEMFs. These results indicate in human neutrophils treated with PEMFs the presence of significant alterations in the A 3 adenosine receptor density and functionality.