Prostacyclin release and receptor activation: differential control of human pulmonary venous and arterial tone

Abstract

1. In human pulmonary vascular preparations, precontracted arteries were more sensitive to the relaxant effect of acetylcholine (ACh) than veins (pD(2) values: 7.25+/-0.08 (n=23) and 5.92+/-0.09 (n=25), respectively). Therefore, the role of prostacyclin (PGI(2)) was explored to examine whether this mediator may be responsible for the difference in relaxation. 2. In the presence of the cyclooxygenase (COX) inhibitor, indomethacin (INDO), the ACh relaxations were reduced in arteries but not in veins. On the contrary, an inhibitor (l-NOARG) of the nitric oxide synthase blocked preferentially the relaxation in veins. 3. A greater release of 6-keto-PGF(1alpha), the stable metabolite of PGI(2), was observed in arterial preparations than in venous preparations when stimulated with either ACh or arachidonic acid (AA). 4. Exogenous PGI(2) produced a reduced relaxant effect in the precontracted vein when compared with the artery. In the presence of the EP(1)-receptor antagonist AH6809, the PGI(2) relaxation of veins was similar to arteries. 5. In veins, AA (0.1 mm) produced a biphasic response, namely, a contraction peak (0.4-0.5 g) followed by a relaxation. These contractions in venous preparations were abolished either in the absence of endothelium or in the presence of INDO or an EP(1)-receptor antagonist (AH6809, SC19220). In the arterial preparations AA induced only relaxations. 6. In both vascular preparations, COX-1 but not the COX-2 protein was detected in microsomal preparations derived from homogenized tissues or freshly isolated endothelial cells. 7. The differential vasorelaxations induced by ACh may be explained, in part, by a more pronounced production and release of PGI(2) in human pulmonary arteries than in the veins. In addition, while PGI(2) induced relaxation by activation of IP-receptors in both types of vessels, a PGI(2) constrictor effect was responsible for masking the relaxation in the veins by activation of the EP(1)-receptor.

Figure 1

Cumulative concentration–response curves induced by ACh in human pulmonary arteries and veins. The preparations were contracted with NA (10 μM) and cumulative concentrations of ACh were added into the baths. Responses are expressed in grams. Values are means±s.e.m. from 23 and 25 different lung samples for arteries and veins, respectively (see text for values significantly different).

Figure 2

Effect of INDO and L-NOARG on the relaxations induced by ACh in paired isolated human pulmonary vessels. After an incubation (30 min) with Tyrode's solution (Control) or Tyrode's solution containing indomethacin (INDO, 1.7 μM) and/or L-NOARG (0.1 mM), the preparations were contracted with NA (10 μM) and cumulative concentrations of ACh were added into the baths. Responses are expressed as the percent of the relaxation induced with papaverine (0.1 mM). Values are means±s.e.m. derived from five paired lung samples for the arteries and five other paired lung samples for the veins (see Table 1 for values significantly different).

Figure 3

Cumulative concentration–response curves induced by PGI₂ in paired isolated human pulmonary arteries (n=3) and veins (n=4). All the preparations were treated with BAY u3405 (1 μM). After an incubation (30 min) with Tyrode's solution (Control) or Tyrode's solution containing AH6809 (30 μM), the preparations were contracted with NA (10 μM) and cumulative concentrations of PGI₂ were added into the baths. Responses are expressed as the percent of the relaxation induced with papaverine (0.1 mM) and values are means±s.e.m.

Figure 4

Cumulative concentration–response curves induced by AA in human pulmonary arteries and veins precontracted with NA (10 μM). Responses are expressed in grams. Values are means±s.e.m. from 5–6 paired lung samples. ^*Indicates data significantly different (P<0.05) from values obtained 72 s after AA (100 μM) stimulation in arteries without treatment or in veins treated with INDO or from values measured 15 min after AA (10 μM) stimulation in veins (Student's paired t-test).

Figure 5

Physiological effects of AA (100 μM) on the basal tone of human pulmonary arteries and veins with or without endothelium. Responses are expressed in grams. Left panel, values are means±s.e.m. from seven paired lung samples for arteries and veins. ^*Indicates data significantly different (P<0.05) from appropriate control (Wilcoxon's signed rank test). Right panel, representative tracing of the effects induced by AA (100 μM) in two vascular preparations derived from the same lung sample.

Figure 6

Inhibition of the response induced by AA (100 μM) on the basal tone of human pulmonary veins with prostanoid receptor antagonists (SC19220 or AH6809; 30 min incubation). Responses are expressed in grams. Values are means±s.e.m. from four paired lung samples. ^*Indicates data significantly different (P⩽0.05) from appropriate control (Student's paired t-test).

Figure 7

Release of 6-keto-PGF_1α by isolated human pulmonary vessels stimulated with ACh. The preparations were contracted with NA (10 μM) and relaxed with ACh (1 or 10 μM). 6-Keto-PGF_1α was measured in aliquots collected (1) after a 5-min incubation of the preparations with the bath fluid prior to the NA stimulation (basal) and (2) 5 min after the ACh stimulation. Stimulated (NA) values were subtracted from stimulated values (ACh). The 6-keto-PGF_1α quantities were expressed as pg mg⁻¹ of tissue wet weight. Values are means±s.e.m. derived from 14–15 (basal), 9 (ACh 1 μM) or 10 (ACh 10 μM) lung samples. ^*Indicates data significantly different (P<0.05) from similar values obtained in arteries (Student's paired t-test).

Figure 8

Release of 6-keto-PGF_1α by isolated human pulmonary vessels stimulated with AA. The preparations were challenged with AA (10 or 100 μM). 6-Keto-PGF_1α was measured in aliquots collected (1) after a 15-min incubation of the preparations with the bath fluid just prior to the AA stimulation (basal) and (2) 15 min after the AA stimulation. Basal values were subtracted from stimulated values. The 6-keto-PGF_1α quantities were expressed as pg mg⁻¹ of tissue wet weight. Values are means±s.e.m. derived from 11 lung samples. ^*Indicates data significantly different (P<0.05) from similar values obtained in arteries (Student's paired t-test).

Figure 9

Western blot analysis of endothelial COX-1 and COX-2 proteins in human pulmonary arteries (PA) and veins (PV) obtained from three different patients. SDS–PAGE was performed on microsomal preparations derived from endothelial cells freshly isolated from human pulmonary vessels. Ovine COX-1 and COX-2 proteins were used as standard (St) for Western blots. Films were exposed 30 min on membranes. Arrows indicate the molecular weight of COX proteins in kDa.