Background: In very preterm infants, clinical decision-making, such as closing a patent ductus arteriosus (PDA), may be aided by measuring circulating natriuretic and endothelial pro-peptides. Objectives: To investigate the association between perinatal characteristics, PDA echocardiography and plasma concentrations of stable pro-peptides of B-type natriuretic peptide (NT-proBNP), atrial natriuretic peptide (MR-proANP) and endothelin-1 (CT-proET-1). Methods: A prospective, cross-sectional, single-center study was performed in 66 infants who were less than 32 weeks of gestational age. Pro-peptide concentrations were determined at birth and at day 2–3 of life. Results: Plasma concentrations of all 3 pro-peptides increased on average 2- to 5-fold from birth to day 2–3 of life. NT-proBNP and MR-proANP were closely related at birth and at day 2–3 (Rs 0.902 and 0.897, respectively, p < 0.001), whereas CT-proET-1 was related to NT-proBNP and MR-proANP at birth (Rs 0.478 and 0.460, respectively, p < 0.001) but not at day 2–3. Birth weight was negatively related to all 3 pro-peptides at birth (p < 0.01); however, preeclampsia and compromised placental perfusion were associated with elevated NT-proBNP and MR-proANP concentrations at birth. At day 2–3, MR-proANP and NT-proBNP correlated significantly with the ductal diameter (Rs 0.416 and 0.415, respectively, both p = 0.011), whereas CT-proET-1 correlated with the left atrium/aorta ratio (Rs 0.506, p = 0.027). CT-proET-1 was elevated in infants with treated compared to untreated PDA [median (5–95% range) 388 (272–723) vs. 303 (152–422) pmol/l, p = 0.011], but not NT-proBNP or MR-proANP. Conclusion: CT-proET-1 is a promising predictor in determining the need for PDA intervention.