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Department of Pediatrics, Harbor-University of California at Los Angeles Medical Center, Torrance, California 90502
Platelet-activating factor (PAF) is a phospholipid with diverse biological functions mediated by a G protein-coupled receptor. We determined PAF receptor binding in lung membranes of four groups of perinatal lambs. Membrane protein (100 µg/ml) was incubated for 60 min at 30°C with 0.5-24 nM of acetyl-[3H]PAF in 30 mM Tris buffer, pH 7.2, containing 0.25% BSA, 10 mM MgCl2, and 125 mM choline chloride. PAF bound to membrane was isolated and quantified by scintillation spectrometry, followed with Scatchard analysis for receptor density (Bmax). The Bmax (means ± SE, fmol/mg protein) were 445.8 ± 12.3, 244.2 ± 3.3, 250.6 ± 3.6, and 419.9 ± 8.6 for the fetal, 90-min-old, <1-day-old, and 6- to 12-day-old lambs, respectively. The Bmax for the 90-min-old and <1-day-old lambs were not different but were significantly lower than those of either the term fetal or 6- to 12-day-old lambs. These data show a significant decrease in PAF binding to its receptor and in PAF Bmax in lung membranes of immediate newborn lambs. The dissociation constants (KD, nM) were 7.7 ± 0.52, 11.5 ± 0.34, 6.9 ± 0.48, and 5.0 ± 0.53 for fetal, 90-min-old, <1-day-old, and 6- to 12-day-old newborn lamb lungs, respectively. The KD of the 90-min-old lamb was the highest of all. PAF receptor gene measured by RT-PCR showed a significant downregulation of PAF receptor gene mRNA in lungs of lambs <1 day old, suggesting a transcriptional regulation of PAF receptor gene expression in the immediate newborn period. We speculate that decreased PAF receptor binding immediately after birth will facilitate the fall in pulmonary vascular resistance in the immediate newborn period.
reverse transcriptase-polymerase chain reaction; pulmonary circulation; receptor gene expression; perinatal
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