Tone-dependent responses to acetylcholine in the feline pulmonary vascular bed

The effects of an increase in base-line tone on pulmonary vascular responses to acetylcholine were investigated in the pulmonary vascular bed of the intact-chest cat. Under conditions of controlled blood flow and constant left atrial pressure, intralobar injections of acetylcholine under low-tone base-line conditions increased lobar arterial pressure in a dose-related manner. When tone was increased moderately by alveolar hypoxia, acetylcholine elicited dose-dependent decreases in lobar arterial pressure, and at the highest dose studied, acetylcholine produced a biphasic response. When tone was raised to a high steady level with the prostaglandin analogue, U46619, acetylcholine elicited marked dose-related decreases in lobar arterial pressure. Atropine blocked both vasoconstrictor responses at low tone and vasodilator responses at high tone, whereas meclofenamate and BW 755C had no effect on responses to acetylcholine at low or high tone. The vasoconstrictor response at low tone was blocked by pirenzepine (20 and 50 micrograms/kg iv) but not gallamine (10 mg/kg iv). The vasodilator response at high tone was not blocked by pirenzepine (50 micrograms/kg iv) or gallamine or pancuronium (10 mg/kg iv). The present data support the concept that pulmonary vascular responses to acetylcholine are tone dependent and suggest that the vasoconstrictor response under low-tone conditions is mediated by a high-affinity muscarinic (M1)-type receptor. These data also suggest that vasodilator responses under high-tone conditions are mediated by muscarinic receptors that are neither M1 nor M2 low-affinity muscarinic-type receptor and that responses to acetylcholine are not dependent on the release of cyclooxygenase or lipoxygenase products.     

Influence of SQ 30741 on thromboxane receptor-mediated responses in the feline pulmonary vascular bed.

The effects of SQ 30741, a thromboxane A2 (TxA2) receptor blocking agent, on responses to the TxA2 mimic, U-46619, were investigated in the pulmonary vascular bed of the intact-chest cat under constant-flow conditions. The administration of SQ 30741 in doses of 1-2 mg/kg iv markedly reduced vasoconstrictor responses to U-46619 without altering responses to prostaglandin (PG) F2 alpha or PGD2 and serotonin. SQ 30741 had no significant effect on mean vascular pressures in the cat, and the dose-response curve for U-46619 was shifted to the right in a parallel manner with a similar apparent maximal response. In addition to not altering responses to PGF2 alpha, PGD2 alpha, or serotonin, SQ 30741 (2 mg/kg iv) was without significant effect on pulmonary vasoconstrictor responses to the PGD2 metabolite 9 alpha, 11 beta-PGF2, norepinephrine, angiotensin II, BAY K 8644, endothelin 1, or endothelin 2. Although responses to vasoconstrictor agents, which act through a variety of mechanisms, were not altered, responses to the PG and TxA2 precursor, arachidonic acid, were reduced significantly. The duration of the TxA2 receptor blockade was approximately 30 and 75 min at the 1- and 2-mg/kg iv doses of the antagonist, respectively. The present data show that SQ 30741 selectively blocks TxA2 receptor-mediated responses in a competitive and reversible manner in the pulmonary vascular bed. These data suggest that responses to arachidonic acid are due in large part to the formation of TxA2 and that discrete TxA2 receptors unrelated to receptors activated by PGD2 or PGF2 alpha are most likely located in resistance vessel elements in the feline pulmonary vascular bed.     

Influence of cyclooxygenase blockade on responses to isoproterenol, bradykinin and nitroglycerin in the feline pulmonary vascular bed

We investigated the effect of indomethacin on responses to isoproterenol, bradykinin and nitroglycerin in the feline pulmonary vascular bed when pulmonary vascular resistance was actively increased by infusion of U46619 in order to determine if vasodilator responses to these agents were dependent on the integrity of the cyclooxygenase pathway. Since pulmonary blood flow and left atrial pressure were held constant, changes in lobar arterial pressure directly reflect changes in lobar vascular resistance. Intralobar injections of isoproterenol, bradykinin, and nitroglycerin decreased lobar arterial pressure in a dose-related manner. Pulmonary vasodilator responses to the lower and midrange doses of bradykinin and nitroglycerin were unchanged in the presence of indomethacin whereas pulmonary responses to the highest doses of nitroglycerin and bradykinin were increased by cyclooxygenase blockade. In contrast, pulmonary vasodilator responses to isoproterenol were significantly attenuated in the presence of propranolol, whereas pulmonary vasodilator responses to bradykinin and nitroglycerin were unchanged after beta blockade. The present data indicate that isoproterenol, bradykinin, and nitroglycerin have significant vasodilator activity in the cat when pulmonary vascular tone is actively increased. These data suggest that the formation of vasodilator cyclooxygenase products such as PGI2 do not mediate vasodilator responses to isoproterenol, bradykinin, and nitroglycerin in the feline pulmonary vascular bed.     

Analysis of responses to sympathetic nerve stimulation in the feline pulmonary vascular bed

The adrenergic receptor subtypes mediating the response to sympathetic nerve stimulation in the pulmonary vascular bed of the cat were investigated under conditions of controlled blood flow and constant left atrial pressure. The increase in lobar vascular resistance in response to sympathetic nerve stimulation was reduced by prazosin and to a lesser extent by yohimbine, the respective alpha 1- and alpha 2-adrenoceptor antagonists. Moreover, in animals pretreated with a beta-adrenoceptor antagonist to prevent an interaction between alpha- and beta 2-adrenoceptors, responses to nerve stimulation were reduced by prazosin, but yohimbine had no significant effect. On the other hand, in animals pretreated with a beta-adrenoceptor antagonist, yohimbine had an inhibitory effect on responses to tyramine and to norepinephrine. Propranolol had no significant effect on the response to nerve stimulation, whereas ICI 118551, a selective beta 2-adrenoceptor antagonist, enhanced responses to nerve stimulation and injected norepinephrine. The present data suggest that neuronally released norepinephrine increases pulmonary vascular resistance in the cat by acting mainly on alpha 1-adrenoceptors and to a lesser extent on postjunctional alpha 2-adrenoceptors but that this effect is counteracted by an action on presynaptic alpha 2-receptors. The present studies also suggest that neuronally released norepinephrine acts on beta 2-adrenoceptors and that the response to sympathetic nerve stimulation represents the net effect of the adrenergic transmitter on alpha 1-, alpha 2-, and beta 2-adrenoceptors in the pulmonary vascular bed.     

Influence of cyclooxygenase blockade on responses to isoproterenol, bradykinin and nitroglycerin in the feline pulmonary vascular bed

We investigated the effect of indomethacin on responses to isoproterenol, bradykinin and nitroglycerin in the feline pulmonary vascular bed when pulmonary vascular resistance was actively increased by infusion of U46619 in order to determine if vasodilator responses to these agents were dependent on the integrity of the cyclooxygenase pathway. Since pulmonary blood flow left atrial pressure were held constant, changes in lobar arterial pressure directly reflect changes in lobar vascular resistance. Intralobar injections of isoproterenol, bradykinin, and nitroglycerin decreased lobar arterial pressure in a dose-related manner. Pulmonary vasodilator responses to the lower and midrange doses of bradykinin and nitrogylcerin were unchanged in the presence of indomethacin whereas pulmonary responses to the highest doses of nitroglycerin and bradykinin were increased by cyclooxygenase blockade. In contrast, pulmonary vasodilator responses to isoproterenol were significantly attenuated in the presence of propranolol, whereas pulmonary vasodilator responses to bradykinin and nitroglycerin were unchanged after beta blockade. The present data indicate that isoproterenol, bladykinin, and nitroglycerin have significant vasodilator activity in the cat when pulmonary vascular tone is actively increased. These data suggest that the formation of vasodilator cyclooxygenase products such as PGI₂ do not mediate vasodilator responses to isoproterenol, bradykinin, and nitroglycerin in the feline pulmonary vascular bed.     

Effects of OKY 1581 on bronchoconstrictor responses to arachidonic acid and PGH2

The influence of OKY 1581, a thromboxane synthase inhibitor, on airway responses to arachidonic acid and endoperoxide, [prostaglandin (PG) H2], were investigated in anesthetized, paralyzed, mechanically ventilated cats. Intravenous injections of arachidonic acid and PGH2 caused dose-related increases in transpulmonary pressure and lung resistance and decreases in dynamic and static compliance. OKY 1581 significantly decreased airway responses to arachidonic acid but not to PGH2. Sodium meclofenamate, a cyclooxygenase inhibitor, abolished airway responses to arachidonic acid but had no effect on airway responses to PGH2. OKY 1581 or meclofenamate has no effect on airway responses to PGF2 alpha, PGD2, or U 46619, a thromboxane mimic. In microsomal fractions from the lung, OKY 1581 inhibited thromboxane formation without decreasing prostacyclin synthesis or cyclooxygenase activity. These studies show that OKY 1581 is a selective thromboxane synthesis inhibitor in the cat lung and suggest that a substantial part of the bronchoconstrictor response to arachidonic acid is due to thromboxane A2 formation. Moreover, the present data suggest that airway responses to endogenously released and exogenous PGH2 are mediated differently and that a significant part of the response to exogenous PGH2 may be due to activation of an endoperoxide/thromboxane receptor, since responses to PGH2 are blocked by the thromboxane receptor antagonist SQ 29548.     

Effects of PKC isozyme inhibitors on constrictor responses in the feline pulmonary vascular bed

The effects of G?-6976, a Ca(2+)-dependent protein kinase C (PKC) isozyme inhibitor, and rottlerin, a PKC-delta isozyme/calmodulin (CaM)-dependent kinase III inhibitor, on responses to vasopressor agents were investigated in the feline pulmonary vascular bed. Injections of angiotensin II, norepinephrine (NE), serotonin, BAY K 8644, and U-46619 into the lobar arterial constant blood flow perfusion circuit caused increases in pressure. G?-6976 reduced responses to angiotensin II; however, it did not alter responses to serotonin, NE, or U-46619, whereas G?-6976 enhanced BAY K 8644 responses. Rottlerin reduced responses to angiotensin II and NE, did not alter responses to serotonin or U-46619, and enhanced responses to BAY K 8644. Immunohistochemistry of feline pulmonary arterial smooth muscle cells demonstrated localization of PKC-alpha and -delta isozymes in response to phorbol 12-myristate 13-acetate and angiotensin II. Localization of PKC-alpha and -delta isozymes decreased with administration of G?-6976 and rottlerin, respectively. These data suggest that activation of Ca(2+)-dependent PKC isozymes and Ca(2+)-independent PKC-delta isozyme/CaM-dependent kinase III mediate angiotensin II responses. These data further suggest that Ca(2+)-independent PKC-delta isozyme/CaM-dependent kinase III mediate responses to NE. A rottlerin- or G?-6976-sensitive mechanism is not involved in mediating responses to serotonin and U-46619, but these PKC isozyme inhibitors enhanced BAY K 8644 responses in the feline pulmonary vascular bed.     

Influence of tone on responses to acetylcholine in the rabbit pulmonary vascular bed

Pulmonary vascular responses to acetylcholine were compared under resting and high tone conditions of the intact-chest rabbit. Under resting tone conditions, intralobar injections of acetylcholine increased lobar arterial pressure in a dose-related manner. The pressor responses to acetylcholine under resting conditions were blocked by meclofenamate, indomethacin, atropine, and pirenzepine. When lobar vascular resistance was raised to a high steady level, low doses of acetylcholine decreased lobar arterial pressure, whereas higher doses elicited a biphasic response with the pressor component predominating at the highest dose studied. Under high tone conditions, only the pressor component of the response was blocked by meclofenamate or indomethacin, whereas pressor and depressor responses were blocked by atropine or the 600-micrograms/kg iv dose of pirenzepine. Pressor responses to acetylcholine under resting and high tone conditions were blocked by pirenzepine (50 micrograms/kg iv), whereas gallamine had no effect on responses to acetylcholine. The 50-micrograms/kg iv dose of pirenzepine had no effect on depressor responses or the depressor component of the response to acetylcholine. The present data support the concept that acetylcholine has significant cyclooxygenase-dependent pressor activity in the rabbit pulmonary vascular bed and suggest that this response is mediated by a muscarinic M1-type receptor. These data also show that, under high tone conditions, a vasodilator response or a vasodilator component of a biphasic response is unmasked. This response is not dependent on the release of cyclooxygenase products and is mediated by a muscarinic receptor that is neither of the M1- nor the M2-type.     

Pulmonary vascular response to thrombin: effects of thromboxane synthetase inhibition with OKY-046 and OKY-1581

We examined the effects of thromboxane synthetase inhibition with OKY-1581 and OKY-046 on pulmonary hemodynamics and lung fluid balance after thrombin-induced intravascular coagulation. Studies were made in anesthetized sheep prepared with lung lymph fistulas. Pulmonary intravascular coagulation was induced by i.v. infusion of alpha-thrombin over a 15 min period. Thrombin infusion in control sheep resulted in immediate increases in pulmonary artery pressure (Ppa) and pulmonary vascular resistance (PVR), which were associated with rapid 3-fold increase in pulmonary lymph flow (Qlym) and a delayed increase in lymph-to-plasma protein concentration (L/P) ratio, indicating an increase in the pulmonary microvascular permeability to proteins. Thrombin-induced intravascular coagulation also increased arterial thromboxane B2 (a metabolite of thromboxane A2) and 6-keto-PGF1 alpha concentrations (a metabolite of prostacyclin). Both OKY-1581 and OKY-046 prevented thromboxane B2 and 6-keto-PGF1 alpha generation. The initial increments in Ppa and PVR were attenuated in both treated groups. The increases in Qlym were gradual in the treated groups but attained the same levels as in control group. However, the increases in Qlym were associated with decreases in L/P ratio. In both treated groups, the leukocyte count decreased after thrombin infusion but then increased steadily above the baseline value, whereas the leukocyte count remained depressed in the control group after thrombin. These studies indicate that a part of the initial pulmonary vasoconstrictor response to thrombin-induced intravascular coagulation is mediated by thromboxane generation. In addition, thromboxane may also contribute to the increase in lung vascular permeability to proteins that occurs after intravascular coagulation and this effect may be mediated by a thromboxane-neutrophil interaction.     

Pulmonary vascular response to thrombin: Effects of thromboxane synthetase inhibition with OKY-046 and OKY-1581

We examined the effects of thromboxane synthetase inhibition with OKY-1581 and OKY-046 on pulmonary hemodynamics and lung fluid balance after thrombin-induced intravascular coagulation. Studies were made in anesthetized sheep prepared with lyng lymph fistulas. Pulmonary intravascular coagulation was induced by i.v. infusion of α-thrombin over a 15 min period. Thrombin infusion in control sheep resulted in immediate increases in pulmonary artery pressure (P ) and pulmonary vascular resistance (PVR), which associated with rapid 3-fold increase in pulmonary lymph flow (Q̇lym) and a delayed increase in lymph-to-plasma protein concentration (L/P) ratio, indicating an increase in the pulmonary microvascular permeability to proteins. Thrombin-induced intravascular coagulation alos increased arterial thromboxane B₂ (a metabolite of thromboxane A₂) and 6-keto-PGF_1α concentrations (a metabolite of prostacyclin). Both OKY-1581 and OKY-046 prevented thromboxane B₂ and 6-keto-PGF_1α generation. The initial increments in P and PVR were attenuated in both treated groups. The increases in Q̇lym were gradual in the treated groups but attained the same levels as in control group. However, the increases in Q̇lym were associated with decreases in L/P ratio. In both treated groups, the leukocyte count decreased after thrombin infusion but then increased steadily above the baseline value, whereas the leukocyte count remained depressed in the control group after thrombin. These studies indicate that a part of the initial pulmonary vasoconstrictor response to thrombin-induced intravascular coagulation is mediated by thromboxane generation. In addition, thromboxane may also contribute to the increase in lung vascular permeability to proteins that occurs after intravascular coagulation and this effect may be mediated by a thromboxane-neutrophil interaction.     

Protective effect of a thromboxane synthetase inhibitor,OKY-1581, on increased lung vascular permeability in pulmonary microembolization in dogs

To evaluate the potential beneficial effect of a thromboxane synthetase inhibitor, OKY-1581, on increased pulmonary vascular permeability in pulmonary microembolization, we have measured the filtration coefficient in the nonembolized lung after unilateral microembolization in dogs. The unilateral microembolization caused marked elevations in pulmonary artery pressure and blood flow to the nonembolized lung, while pulmonary venous pressure in nonembolized lung did not change. The pulmonary vascular resistance in nonembolized lung did not increase significantly. The filtration coefficient (K_f) in nonembolized lung increased to 0.14 ± 0.02 from the baseline value of 0.07 ± 0.01 ml/min/mmHg/100g at 30 min after microembolization when the initial hemodynamic changes reduced toward the baseline value. In OKY-1581 treated dogs, similar hemodynamic changes did not result in the increase in the filtration coefficient in nonembolized lung. Platelet aggregation was also inhibited after microembolization in OKY-1581 treated dogs. Based on these results, we could conclude that OKY-1581 could prevent the increase in pulmonary vascular permeability following pulmonary microembolization by inhibiting platelet aggregation and possibly by preventing the release of thromboxane A₂.     

Nisoldipine inhibits vasoconstrictor responses in the cat pulmonary vascular bed

The influence of nisoldipine, a dihydropyridine calcium entry antagonist, on vascular resistance and vasoconstrictor responses was investigated in the feline pulmonary vascular bed under conditions of controlled blood flow. The calcium channel blocking agent caused a small reduction in lobar vascular resistance and blocked pulmonary vasoconstrictor responses to BAY K 8644, an agent which promotes calcium entry. The calcium entry blocking agent also reduced pulmonary vasoconstrictor responses to methoxamine and to BHT 933, alpha 1- and alpha 2-adrenoceptor agonists, and to U 46619, an agent which mimics the actions of thromboxane A2. Although there was a marked difference in vasoconstrictor potency in the pulmonary vascular bed, responses to the thromboxane mimic and to the alpha 1- and alpha 2-adrenoceptor agonists were reduced by approximately the same extent. The increases in systemic arterial pressure in response to BAY K 8644, methoxamine, and BHT 933 were also reduced by nisoldipine, and the calcium entry antagonist reduced systemic arterial pressure and systemic vascular resistance. The results of the present study suggest that an extracellular source of calcium is required for the maintenance of vascular tone and for the expression of vasoconstrictor responses, resulting from activation of alpha 1- and postjunctional alpha 2-adrenoceptors and thromboxane receptors in the feline pulmonary vascular bed.     

Prostanoid synthesis and vascular responses to exogenous arachidonic acid following cerebral ischemia in piglets

In newborn pigs, cerebral ischemia abolishes both increased cerebral prostanoid production and cerebral vasodilation in response to hypercapnia and hypotension. Attenuation of prostaglandin endoperoxide synthase activity could account for the failure to increase prostanoid systhesis and loss of responses to these stimuli. To test this possibility, arachidonic acid (3,6, or 30μg/ml) was placed under cranial windows in newborn pigs that been exposed to 20 min of cerebral ischemia. The conversion to prostanoids and pial arteriolar responses to the arachidonic acid were measured. At all three concentration, arachidonic acid caused similar increases in pial arteriolar diameter in sham control piglets and piglets 1 hr postischemia. Topical arachidonic acid caused dosedependent increases of PGE₂ in cortical periarachnoid cerebral spinal fluid. 6-keto-PGF_1α and TXB₂ only increased at the highest concentration of arachidonic acid (30 μg/ml). Cerebral ischemia did not decrease the conservation of any concentration of arachidonic acid to PGE₂, 6-keto-PGF_1α, or TXB₂. We conclude that ischemia and subsequent reperfusion do not result in inhibition of prostaglandin endoperoxide synthase in the newborn pig brain. Therefore, the mechanism for the impaired prostanoid production in response to hypercapnia and hypotension following cerebral ischemia appears to involve reduction in release of free arachidonic acid.     

Prostanoid synthesis and vascular responses to exogenous arachidonic acid following cerebral ischemia in piglets

In newborn pigs, cerebral ischemia abolishes both increased cerebral prostanoid production and cerebral vasodilation in response to hypercapnia and hypotension. Attenuation of prostaglandin endoperoxide synthase activity could account for the failure to increase prostanoid synthesis and loss of responses to these stimuli. To test this possibility, arachidonic acid (3, 6, or 30 micrograms/ml) was placed under cranial windows in newborn pigs that had been exposed to 20 min of cerebral ischemia. The conversion to prostanoids and pial arteriolar responses to the arachidonic acid were measured. At all three concentrations, arachidonic acid caused similar increases in pial arteriolar diameter in sham control piglets and piglets 1 hr postischemia. Topical arachidonic acid caused dose-dependent increases of PGE2 in cortical periarachnoid cerebral spinal fluid. 6-keto-PGF1 alpha and TXB2 only increased at the highest concentration of arachidonic acid (30 micrograms/ml). Cerebral ischemia did not decrease the conversion of any concentration of arachidonic acid to PGE2, 6-keto-PGF1 alpha, or TXB2. We conclude that ischemia and subsequent reperfusion do not result in inhibition of prostaglandin endoperoxide synthase in the newborn pig brain. Therefore, the mechanism for the impaired prostanoid production in response to hypercapnia and hypotension following cerebral ischemia appears to involve reduction in release of free arachidonic acid.