Ontogeny of Adenylate Cyclase Activity in the Rat Lung: Guanine Nucleotides and Cytosolic Factors

Abstract

Summary: Catecholamines mediate surfactant release, smooth muscle tone, and lung fluid secretion presumably by their interactions with β-adrenergic receptors and adenylate cyclase, which results in the synthesis of 3′,5′ cyclic adenosine monophosphate (cAMP). Ontogenic changes in factors which regulate cAMP metabolism in the lung have been demonstrated previously and associated with developmental changes in lung structure and function in the perinatal period. In this study, developmental changes in characteristics of β-adrenergic receptors and the enzyme adenylate cyclase were characterized in rat lung from day 18 of gestation to adulthood. Although concentrations of pulmonary β-adrenergic receptors increased approximately 5-fold during the perinatal and suckling period, “basal” adenylate cyclase activity of lung homogenates decreased from 130 ± 8 pmole · mg⁻¹ · min⁻¹ at 20 days gestation to 25 ± 3 pmole · mg⁻¹ · min⁻¹ in the adult, P < 0.01. GTP and lung cytosol activated adenylate cyclase from fetal membranes to a significantly greater extent than they activated the enzyme from adult membrane. GTP enhanced adenylate cyclase activity 5–6-fold in fetal membranes, GTP being nearly as active as its non-hydrolyzable analogue Gpp(NH)p or NaF. Addition of GTP in the presence of PGE₁ or catecholamine resulted in nearly maximal activation of the enzyme from fetal but not from adult lung membranes. Cholera toxin enhanced GTP-dependent activity in adult samples but did not further increase GTP-dependent activity in fetal preparations.

Lung cytosolic factor(s), which activated adenylate cyclase, were separated into two components by gel filtration. One activating component was likely to be GTP, which was present in lung tissue in concentrations near the EC50 for activation of adenylate cyclase. A second (large molecular weight) cytosolic factor(s), which activated adenylate cyclase, was sensitive to heat or trypsin and was excluded by Sephadex G-50. An ATP-regenerating system was required for full activation of adenylate cyclase by lung cytosol. Effects of EGTA, Mg²⁺, guanine nucleotides, cell cytosol and Ca²⁺-calmodulin were partially characterized.

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