Rubabe S Saharan1, Kwaku D Nantwi. 1. Department of Anatomy and Cell Biology, Wayne State University, Detroit, Michigan 48202, USA.
Abstract
BACKGROUND/ OBJECTIVE: Adenosine A1 receptors localized in the phrenic motoneurons (PMNs), where the axons of the descending bulbospinal respiratory make synaptic contacts, may be involved in theophylline-induced respiratory-related activity in rats. The objective of this study was to characterize the biochemical profiles of adenosine A1 receptors in 2 groups of rats: (a) naïve and (b) theophylline-treated (3-day oral administration). METHODS: Biochemical binding characteristics of adenosine A1 receptors in the C3 to C5 (PMN) of adult rats were assessed in naïve (n = 6) and theophylline-treated animals (n = 6) using [3H]-DPCPX (10 pmol/L to 30 nmol/L), the specific adenosine A1 receptor antagonist in saturation-binding assays. Competition assays used theophylline as the competing ligand (20 mmol/L to 20 pmol/L), and protein concentration was determined with the Bradford assay using a range of standards (0.016-1.0 mg/mL). RESULTS: In saturation-binding assays in naïve animals, the A1 receptor was characterized by a single binding site with Bmax and Kd values of 256.00 +/- 32.13 fmol/mg protein and 2.89 +/- 0.45 nmol/L, respectively. Analysis of the isotherm in theophylline-treated animals showed 1 site with Bmax and Kd values of 219.00 +/- 26.3 fmol/mg protein and 0.60 +/- 0.21 nmol/L, respectively, and a second site characterized by Bmax and Kd values of 492.6 +/- 3.15 fmol/mg protein and 14.09 +/- 2.06 nmol/L, respectively. CONCLUSIONS: Theophylline administration revealed 2 binding sites on receptors (characterized by the specific adenosine A1 antagonist, [3H]-DPCPX) located in the vicinity of phrenic motoneurons (C3-C5). Alteration of the receptor profiles after theophylline may underlie the respiratory-related actions of the drug.
BACKGROUND/ OBJECTIVE: Adenosine A1 receptors localized in the phrenic motoneurons (PMNs), where the axons of the descending bulbospinal respiratory make synaptic contacts, may be involved in theophylline-induced respiratory-related activity in rats. The objective of this study was to characterize the biochemical profiles of adenosine A1 receptors in 2 groups of rats: (a) naïve and (b) theophylline-treated (3-day oral administration). METHODS: Biochemical binding characteristics of adenosine A1 receptors in the C3 to C5 (PMN) of adult rats were assessed in naïve (n = 6) and theophylline-treated animals (n = 6) using [3H]-DPCPX (10 pmol/L to 30 nmol/L), the specific adenosine A1 receptor antagonist in saturation-binding assays. Competition assays used theophylline as the competing ligand (20 mmol/L to 20 pmol/L), and protein concentration was determined with the Bradford assay using a range of standards (0.016-1.0 mg/mL). RESULTS: In saturation-binding assays in naïve animals, the A1 receptor was characterized by a single binding site with Bmax and Kd values of 256.00 +/- 32.13 fmol/mg protein and 2.89 +/- 0.45 nmol/L, respectively. Analysis of the isotherm in theophylline-treated animals showed 1 site with Bmax and Kd values of 219.00 +/- 26.3 fmol/mg protein and 0.60 +/- 0.21 nmol/L, respectively, and a second site characterized by Bmax and Kd values of 492.6 +/- 3.15 fmol/mg protein and 14.09 +/- 2.06 nmol/L, respectively. CONCLUSIONS:Theophylline administration revealed 2 binding sites on receptors (characterized by the specific adenosine A1 antagonist, [3H]-DPCPX) located in the vicinity of phrenic motoneurons (C3-C5). Alteration of the receptor profiles after theophylline may underlie the respiratory-related actions of the drug.