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P. W. Yates, S. Husain, M. Haracznak, J. Fant, C. E. Crosson; Effects of the Adenosine A3 Agonist (IB-MECA) on ERK1/2 Activation in Human Ciliary Muscle Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3926.
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© ARVO (1962-2015); The Authors (2016-present)
Previous studies have shown that adenosine agonists can modulate IOP; however, the adenosine receptor subtypes and cellular actions responsible for these chages are not fully understood. The objective of this study was to investigate the effects of adenosine A3 agonist, IB-MECA, on extracellular regulated kinase 1/2 (ERK1/2) activation in human ciliary muscle cells.
Primary cultures of human ciliary muscle cells were established from human ciliary muscle explants and grown in DMEM media with 10% serum. Prior to experiments, cells were maintained in serum-free medium for 16 hours. To activate ERK1/2, cells were incubated in the presence of the adenosine A3 receptor agonist, N6-(3-iodobenzyl) adenosine-5'-N-methyluronamide (IB-MECA), for 10 minutes. In experiments evaluating the response of adenosine A3 receptor antagonist, 3-ethyl-5-benzyl-2-methyl-4-phenylethynyl-6-phenyl-1, 4-(±)-dihydropyridine-3, 5-dicarboxylate (MRS-1191; 1 µmol/L), and A1 receptor antagonist, 8-cyclopentyl-1, 3-dimethylxanthine (CPT; 1 µmol/L), cells were pretreated for 30 minutes with the antagonist prior to the addition of the agonist. The activation of ERK1/2 was determined by Western blot analysis.
In cultured human ciliary muscle cells, the addition ofIB-MECA (10-7 mol/L) produced a significant 246 ± 22 % increase in ERK1/2 activation. This increase in ERK1/2 activation was concentration-dependent with an EC50 value of 1.2 x 10-8 mol/L. In cells pretreated with adenosine A3 receptor antagonist, MRS-1191 (1 µmol/L), ERK1/2 activation in response to 100 nmol/L IB-MECA was significantly reduced by 64%. In contrast, pretreatment with adenosine A1 receptor antagonist, CPT (1 µmol/L) did not inhibit ERK1/2 activation induced by IB-MECA.
These results provide evidence for the existence of functional adenosine A3 receptors in the human ciliary smooth muscle cells. These receptors are coupled to ERK1/2 activation. As adenosine receptors have been linked to the regulation of intraocular pressure, these studies support the idea that part of this regulatory process may involve the activation of adenosine A3 receptors in the ciliary muscle.
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