Hypotension induced by growth-hormone-releasing peptide is mediated by mast cell serotonin release in the rat.

Growth-hormone-releasing peptide (GH-RP-6) is a synthetic hexapeptide that selectively releases growth hormone (GH) when administered to a number of animals species. In the rat, maximal GH release occurs after intravenous administration of 100 micrograms/kg GH-RP-6. Intravenous administration of 5 mg/kg GH-RP-6 produced 100% lethality within 2-5 min of drug administration. Further investigative studies demonstrated that the lethal effect of GH-RP-6 was preceded by an initial hypertensive episode, followed by a rapid, profound hypotension and bradycardia. The rise and fall in blood pressure also were observed in pithed rats treated with GH-RP-6, suggesting that the central nervous system was not responsible for the changes in blood pressure. However, the GH-RP-6-induced bradycardia was not observed in pithed rats, indicating the fall in heart rate was mediated through a central reflex mechanism. No direct effects of GH-RP-6 were seen in the isolated rat aorta or canine saphenous vein. Pretreatment of conscious rats with naloxone (10 mg/kg, iv), an opiate receptor antagonist, did not prevent the hypertensive response to GH-RP-6, but the hypotension and lethality were attenuated. Pretreatment with cyproheptadine (2.5 mg/kg, iv), a dual serotonin/histamine antagonist, or ketanserin (3 mg/kg, iv), a selective serotonin antagonist, prevented the GH-RP-6-induced hypotension and lethality. Cyproheptadine unmasked a 40 mm Hg rise in mean arterial pressure which persisted for over 10 min. In addition, degranulation of mast cells with compound 48/80 inhibited the toxicity of GH-RP-6, suggesting that mast cell degranulation and the subsequent release of autocoids is responsible for the cardiovascular effects of GH-RP-6. In vitro, GH-RP-6 (10(-5) - 10(-3) M) produced a concentration-related release of histamine from rat peritoneal mast cells. However, the histamine release by GH-RP-6 (10(-4) M) was not inhibited by naloxone (10(-4) M) in isolated mast cells, suggesting either that peritoneal mast cells are not responsible or that the mast cell degranulation in vitro is not opiate mediated. In conclusion, it appears that GH-RP-6 degranulates mast cells releasing serotonin, which produces hypotension, bradycardia, and death. This degranulation of mast cells is apparently inhibited by naloxone in vivo, suggesting that opiate receptors are involved in the hypotension and lethality associated with the administration of GH-RP-6.

In vivo and in vitro cardiotoxicity of a gold-containing antineoplastic drug candidate in the rabbit.

Bis[1,2-bis(diphenylphosphino)ethane] gold(I) chloride (Au(DPPE)+2), a cytotoxic antineoplastic drug candidate, was cardiotoxic in rabbits. Intravenous administration of Au(DPPE)+2 (15 mg/kg) as a single dose produced multiple, 2- to 5-mm subendocardial and myocardial lesions, macroscopically appearing as pale tan foci. Histologically, these lesions consisted of widely scattered zones of myocardial cell necrosis and mineralization. The myocardium also contained multifocal areas of contraction band necrosis in which aggregated clumps of disorganized myofilaments were contiguous with areas of sarcoplasm which were relatively devoid of myofilaments. In a series of in vitro studies, electron microscopic examination of isolated rabbit myocytes treated with 30 microM Au(DPPE)+2 for 15 min showed evidence of mitochondrial swelling and electron translucent mitochondrial matrices. After 60 min of incubation, myocytes had mitochondria that were condensed and disrupted but the cristae had retained their tubular profiles. Isolated rabbit myocytes exposed to 30 microM Au(DPPE)+2 had significant increases in the leakage of lactate dehydrogenase, an index of cell death. Cellular ATP content in myocytes exposed to 30 microM Au(DPPE)+2 was significantly reduced by 30 min. State 4 respiration in isolated rabbit mitochondria was significantly increased by Au(DPPE)+2 (30 microM) while state 3 respiration was unaffected. Au(DPPE)+2 also caused a rapid dissipation of the mitochondrial inner membrane electrochemical potential in a concentration-dependent manner and was accompanied by a ruthenium red-sensitive calcium efflux. These data suggest that disruption of mitochondrial function, leading to uncoupling of oxidative phosphorylation, decreased ATP synthesis, and altered mitochondrial calcium homeostasis, may be a contributing factor leading to cardiac myofibril necrosis produced by Au(DPPE)+2.

An investigation of the pathological and physiological effects of intraosseous sodium bicarbonate in pigs.

Recent interest in the intraosseous (IO) route as an alternative venous access for drug and fluid administration has increased. This study examined the physiological and skeletal pathological effects of IO NaHCO3 in pigs. In the pathological studies, swine (8-10 kg) received NaHCO3 (1 mEq/kg) in one tibia and saline (1 ml/kg) in the other tibia via an 18-gauge spinal needle inserted into the anteromedial surface of the bone. The animals were then observed for one month, sacrificed, and the tibias were isolated, sectioned, and stained for pathological examinations. The physiological effects of IO NaHCO3 infusion were studied and compared with that of intravenous (IV) administration using a cardiac arrest model as previously described. The results demonstrated that NaHCO3 had no effect on the mean arterial blood pressure and plasma catecholamine levels, but increased arterial pH values within two minutes of administration. Similar effects were found with IV NaHCO3. Pathological data indicated signs of minimal local increase in skeletal turnover associated with IO NaHCO3 infusion. It is concluded that the IO route is a safe alternative venous access for NaHCO3 administration in swine.

Pathogenesis of arterial lesions induced by dopaminergic compounds in the rat.

Fenoldopam mesylate (FM), a selective post-junctional dopaminergic (DA1) vasodilator, causes lesions of large caliber splanchnic arteries (100-800 microns) in the rat characterized by necrosis of medial smooth muscle cells and hemorrhage. FM does not induce lesions in other vascular beds of the rat, or in dogs or monkeys. Dopamine, like FM, causes hemorrhagic lesions of large caliber splanchnic arteries in the rat, as well as fibrinoid necrosis of small caliber arteries (less than 100 microns) of the splanchnic, cerebral, coronary and renal vascular beds. Dopamine is an alpha- and beta-adrenoceptor and a dopaminergic receptor agonist. Because these arterial lesions are thought to result from the pharmacologic activity of these 2 compounds, we sought to ascertain the presence of DA1 receptors in mesenteric arteries of the rat and to determine the role of these or other vascular receptor subtypes in lesion induction. We also studied the process of repair after arterial injury caused by FM or dopamine. The presence of DA1 receptors was confirmed in isolated perfused mesenteric arteries by standard pharmacologic techniques; stimulation by FM resulted in vasodilation which was inhibited by the DA1 receptor antagonist SK&F 83566-C. Likewise, SK&F 83566-C prevented the induction of hemorrhagic lesions of large caliber arteries in rats upon infusion of FM or dopamine. In rats co-exposed to the alpha-adrenoreceptor antagonist phenoxybenzamine (PBZ) and either FM or dopamine, the incidence and severity of hemorrhagic lesions of large caliber arteries were increased, but PBZ prevented the formation of dopamine-induced fibrinoid lesions in arteries of small caliber. Rats exposed concurrently to dopamine, phenoxybenzamine, and SK&F 83566-C were free of all arterial lesions. Thus, the induction of splanchnic arterial lesions in the rat by dopamine and FM is caused by stimulation of, and interaction between, alpha-adrenoceptors and dopaminergic DA1 receptors. Fibrinoid lesions of small arteries (alpha-adrenoceptor-mediated) were repaired, as observed morphologically by 14 d after exposure to dopamine. Hemorrhagic lesions of large caliber arteries (DA1 receptor-mediated) had undergone significant repair by 28 d after exposure to FM but these arteries possessed a thicker media surrounded by adventitial fibrosis. Thus, morphologically distinct receptor-mediated splanchnic arterial lesions induced by dopaminergic and alpha-adrenoceptor agonists follow a markedly different course of repair. Arterial lesions induced by FM or dopamine by activation of post-junctional dopaminergic DA1 receptors may represent a model of polyarteritis nodosa.

An investigation of the cardiotoxic action of vancomycin in the isolated working rat heart.

The mechanism of cardiotoxic action of vancomycin was examined in preparations of isolated working rat hearts and spontaneously beating right atria. Vancomycin produced a concentration-dependent decrease in aortic flow, coronary flow and heart rate in the isolated working rat heart, with no significant effect on other haemodynamic parameters. At 5 mm, vancomycin produced a statistically significant decrease (compared with control values) in aortic flow (24.1 +/- 7.5%) and in the basal heart rate (34.3 +/- 3.5%) after 15 min incubation. Coronary flow was also reduced by 23.5 +/- 9.2%. Prolonged exposure of the preparation to 5 mm-vancomycin produced a marked time-dependent bradycardia accompanied by a time-dependent increase in the leakage of lactic dehydrogenase (LDH) into the perfusion medium. Moreover, a correlation (r = -0.94) was found between the time-dependent bradycardia and LDH leakage induced by Vancomycin. In the isolated spontaneously beating right atria pretreated with atropine, vancomycin (5 mm) also produced a time-dependent bradycardia similar to that found in the isolated working rat heart. Moreover, (45)Ca(2+)-flux studies indicated that vancomycin had no significant effect on the (45)Ca(2+) uptake into the right atrial muscle. These data suggest that: (1) vancomycin has a direct and acute cardiotoxic action at high concentrations (> 1 mm); (2) time-dependent bradycardia is a sensitive functional index for the cardiotoxicity induced by vancomycin; (3) the bradycardia elicited by vancomycin is due neither to the release of acetylcholine from the parasympathetic nerves innervating the heart nor to blockade of Ca(2+) entry.

Suppression of pentylenetetrazol-elicited seizure activity by intraosseous propranolol in pigs.

The intraosseous (IO) route provides a rapid and effective alternative venous access in the pediatric population when the conventional intravenous (IV) route cannot be easily obtained. DL-propranolol, a beta-adrenoceptor antagonist, exhibits antiepileptic activity in various animal seizure models. This study assessed the efficacy of IO propranolol in suppressing pentylenetetrazol (PTZ)-induced seizure activity in pigs. Domestic swine (13-20 kg) were prepared for recordings of arterial blood pressure, ECG and electrocortical activity. Seizure activity was induced by pentylenetetrazol (PTZ; 100 mg/kg; IV). Sixty seconds after the onset of seizure activity, the animals either received no drug (control) or propranolol (IV or IO via an 18-gauge spinal needle placed in the right proximal tibia). A transient increase (16.3-50.0%) in the mean arterial blood pressure (MAP) was observed following PTZ administration. Both IO and IV propranolol significantly suppressed the seizure duration (SD) (sec/min interval) at 1 min following drug administration; SD control, 36.3 +/- 4.8; IV propranolol, 12.3 +/- 5.1; IO propranolol, 18.3 +/- 6.0. In addition, both IV and IO propranolol produced a maximal decrease of 32-38% in the basal heart rate; and reduced the transient increase in MAP elicited by PTZ, with no significant effect on the basal MAP. The data demonstrate that 1) propranolol possesses anticonvulsant activity against PTZ-induced seizure in the pig, and 2) the intraosseous route is a rapid and effective alternative venous access for propranolol administration in swine.

Role of calcium in phorbol-ester-induced contractions of the canine saphenous vein.

The role of calcium in phorbol-12,13-dibutyrate (PDB)-induced contractions of the canine saphenous vein (CSV) was examined. Phorbol-12,13-dibutyrate elicited concentration-dependent contractions in CSV (EC50 = 1.6 +/- 0.2 X 10(-7) M) which were not affected by atropine (10(-6) M), pyrilamine (10(-6) M), and phentolamine (10(-5) M). The maximum contraction induced by PDB (10(-6) M) was slightly greater than that elicited by phenylephrine (PE; 10(-4) M). Phorbol-12,13-dibutyrate produced maximal 45Ca2+ uptake (0.80 +/- 0.07 mmol/kg wet wt) comparable with that induced by PE (0.90 +/- 0.04 mmol/kg wet wt) which was approximately fourfold above basal 45Ca2+ uptake (0.21 +/- 0.02 mmol/kg wet wt). The increase in 45Ca2+ uptake stimulated by PDB and PE was completely abolished by La3+ (5 mM). In the absence of Ca2+ entry, the contractions to PDB were reduced by only 29 +/- 3.4%. Substantial responses to PDB (51.3 +/- 4.8% of control) remained after reduction of intracellular Ca2+ store by repeated challenges with PE (10(-4) M) in the presence of La3+. Similar results were obtained when the contractions of CSV to PDB were determined in zero external Ca2+ medium. The data suggest that PDB utilizes both extracellular and intracellular Ca2+ for contractions of CSV.

Role of receptor reserve in the inhibition of alpha-1 adrenoceptor-mediated pressor responses by calcium antagonists in the pithed rat.

The effect of the calcium channel antagonists nifedipine and FR 34235 on the vasopressor response to alpha-1 adrenoceptor stimulation in the pithed normotensive rat was investigated. The maximal pressor response elicited by the full alpha-1 adrenoceptor agonist SK&F l-89748 was slightly but significantly reduced by 1-mg/kg doses of nifedipine (21 +/- 2%) and FR 34235 (34 +/- 4%). In comparison, the maximal pressor response to alpha-1 adrenoceptor stimulation by the partial alpha-1 agonist SK&F 88444 was markedly inhibited by nifedipine (51 +/- 1%) and FR 34235 (65 +/- 3%). Partial inactivation of the postsynaptic alpha-1 adrenoceptors with phenoxybenzamine (0.1 mg/kg) resulted in a maximal increase in diastolic pressure to alpha-1 adrenoceptor activation by SK&F l-89748 less than that induced by SK&F 88444. After phenoxybenzamine treatment, nifedipine and FR 34235 produced even greater reductions in the maximal vasopressor response to alpha-1 adrenoceptor stimulation by SK&F l-89748 (77 +/- 8 and 85 +/- 1%, respectively). Moreover, an inverse linear correlation (r = 1.00) was observed between the sensitivity of the maximal vasopressor response to nifedipine and FR 34235 and the magnitude of the maximal pressor response. The data suggest that the sensitivity of the alpha-1 adrenoceptor-mediated pressor response to inhibition by calcium antagonists in the pithed rat is inversely related to the magnitude of the pressor response, and they are consistent with the notion that the presence of "spare" alpha-1 adrenoceptors may determine the sensitivity of the pressor response to calcium antagonists.

Evidence for and against heterogeneity of alpha 1-adrenoceptors.

Recent experimental evidence has suggested that the alpha 1 adrenoceptor may need to be further subdivided. It can no longer be stated categorically that alpha 1-adrenoceptors are present only at postjunctional sites, in view of several reports of alpha 1-mediated modulation of adrenergic and cholinergic neurotransmission. Furthermore, comparison of the pharmacologic characteristics of the alpha 1-adrenoceptor in different species and/or tissues can show clear differences in sensitivity to selective agonists and antagonists, and differences in the degree of dependence on extracellular calcium. However, in other cases, alpha 1-adrenoceptors at diverse sites have been found to have identical characteristics. Furthermore, the subcategories identified by the various selective agents do not fall into the same discrete groups, in contrast to division of alpha-adrenoceptors into alpha 1 and alpha 2-adrenoceptors. Therefore, at this time it seems premature to subdivide the alpha 1-adrenoceptor further.

Characterization of the antihypertensive activity of SK&F 86466, a selective alpha-2 antagonist, in the rat.

The effects of SK&F 86466 (6-chloro-N-methyl-2,3,4,5-tetrahydro-1-H-3-benzazepine), a potent and selective alpha-2 adrenoceptor antagonist, on blood pressure and heart rate were examined in normotensive and hypertensive rats. SK&F 86466 was approximately 10-fold more potent in lowering blood pressure in deoxycorticosterone acetate (DOCA)-salt hypertensive than in normotensive rats, when administered by i.v. infusion. A dose-related antihypertensive effect was observed in conscious DOCA-salt rats after p.o. doses of SK&F 86466 (2-15 mg/kg), with a duration of over 6 hr after the highest dose. Oral administration of SK&F 86466 was associated with tachycardia, which reached maximal levels within 15 min after dosing, followed by an interval of bradycardia. The duration of antihypertensive activity observed with SK&F 86466 was at least 6 hr, compared to less than 90 min with phentolamine, at doses which produced equal peak reductions in blood pressure in conscious DOCA-salt rats. In contrast, heart rate was still significantly elevated by phentolamine 90 min postdosing, whereas the transient tachycardia induced by SK&F 86466 had dissipated within 30 min. SK&F 86466 was also an effective antihypertensive agent in the spontaneously hypertensive rat, although lower antihypertensive efficacy was observed than in the DOCA-salt model. SK&F 86466 was even less effective in the two-kidney one-clip Goldblatt model in which hypertension is maintained by the renin-angiotensin system rather than by the sympathetic nervous system.(ABSTRACT TRUNCATED AT 250 WORDS)

Selective alpha-2 adrenoceptor blockade by SK&F 86466: in vitro characterization of receptor selectivity.

SK&F 86466 (6-chloro-N-methyl-2,3,4,5-tetrahydro-1-H-3-benzazepine) is a potent and selective antagonist at alpha-2 adrenoceptors. Prejunctional alpha-2 adrenoceptor antagonism can be demonstrated either by blockade of the alpha-2 adrenoceptor-mediated neuroinhibitory effect of clonidine or B-HT 920 in the guinea-pig atrium [receptor dissociation constant (KB) = 13-17 nM] or by potentiation of nerve-evoked release of [3H]norepinephrine from prelabeled guinea-pig atria, dog splenic artery or rabbit ear artery. Blockade of the postjunctional alpha-2 adrenoceptor was also seen, as demonstrated by a parallel shift to the right of the concentration-response curve for B-HT 920 as a constrictor agent in the dog saphenous vein. The KB for SK&F 86466 in this test system was 42 nM. The affinity of SK&F 86466 for the alpha-1 adrenoceptor is much lower, with a KB of 900 nM against norepinephrine-mediated constriction in the rabbit ear artery, or 1100 nM vs. SK&F 89748-induced constriction in the dog saphenous vein. The alpha-2/alpha-1 adrenoceptor selectivity ratio of SK&F 86466 is comparable to that obtained with agents such as yohimbine, making SK&F 86466 a useful tool for characterization of alpha-2 adrenoceptors and for investigation of their physiological role.

An evaluation of the ability of a series of full alpha-1 adrenoceptor agonists to release internal calcium in venous smooth muscle.

Our previous studies have shown that activation of postsynaptic alpha-2 adrenoceptors mainly utilizes extracellular calcium whereas activation of postsynaptic alpha-1 adrenoceptors mobilizes both extracellular and intracellular calcium to produce contractions in the canine saphenous vein. In the present study, the abilities of several full alpha-1 adrenoceptor agonists to release internal calcium for contractions in the canine saphenous vein were evaluated. Contractions to these alpha-1 agonists at two different equieffective concentrations (concentrations that produced 50 and 80% of the maximum phenylephrine response, PE50 and PE80, respectively) were determined in both zero external calcium medium and in normal calcium medium containing 5 mM La . If a correlation exists between the efficacy of an agonist and its ability to release internal Ca++, the contractile response to each agonist should be similar under these conditions. The results indicated marked variations in mechanical responses elicited by these alpha-1 agonists at both PE50 and PE80 concentrations. The data suggest a lack of correlation between efficacy and the ability to release internal calcium to induce contractions for a series of full alpha-1 adrenoceptor agonists in venous smooth muscle.

Evidence that oxmetidine inhibits transmembrane-calcium flux in cardiac and vascular tissue.

Oxmetidine, at concentrations in excess of 1 X 10(-6)M, caused concentration-dependent negative inotropic and chronotropic responses in guinea-pig isolated heart preparations. Oxmetidine, at concentrations in excess of 1 X 10(-5)M, caused negative inotropic responses in guinea-pig papillary muscle preparations. The negative inotropic responses to oxmetidine were associated with shortening of the plateau phase of the action potential. Verapamil and nifedipine caused similar shortening of the plateau phase of the action potential at equivalent negative inotropic concentrations indicating that oxmetidine may also act as a calcium antagonist. In preparations partially depolarized by raising extracellular K+ concentration, oxmetidine also exhibited negative inotropic activity and reduced the calcium-dependent action potential. However, unlike verapamil and nifedipine, oxmetidine did not show voltage-dependent activity. Oxmetidine, at concentrations in excess of 1 X 10(-5)M, inhibited Ca2+-dependent contractions of dog saphenous vein preparations and inhibited 45Ca2+-uptake into veins depolarized by high extracellular K+. In vivo, these calcium antagonist actions of oxmetidine were demonstrated by vasodilatation, reduction in blood pressure, bradycardia and reduced cardiac output in anaesthetized cats. Oxmetidine, at concentrations of 1 X 10(-5)M and above, shows properties consistent with inhibition of transmembrane Ca2+ flux. This action can be distinguished from other calcium antagonists as the effects of oxmetidine are not voltage-dependent.

Role of extracellular calcium in contractions produced by activation of postsynaptic alpha-2 adrenoceptors in the canine saphenous vein.

Canine saphenous vein (CSV) has been shown to contain both postsynaptic alpha-1 and alpha-2 adrenoceptors. Our previous studies have shown that activation of postsynaptic alpha-1 adrenoceptors in this tissue utilizes both extracellular and intracellular Ca++ to produce contractions. In the present study, the source of calcium mobilized by activation of postsynaptic alpha-2 adrenoceptors in CSV was elucidated. Contractions of tissue rings to the supramaximal concentrations of three selective alpha-2 agonists, B-HT 920, M-7 and clonidine, were determined in the absence and presence of 5 mM La . In the presence of La , clonidine and M-7 produced small but statistically significant contractions (8-14% of control) which were abolished when the alpha-1 adrenoceptors were inactivated by phenoxybenzamine (10(-7) M, 30 min). In contrast, contractions to B-HT 920 were abolished completely in the presence of La . All the three alpha-2 agonists stimulated 45Ca++ uptake into CSV (0.3-0.4 mmol/kg wet weight, 10 min). 45Ca++ efflux studies demonstrated that the selective alpha-2 agonist, B-HT 920 (10(-5) M plus 10(-7) M phenoxybenzamine), did not induce an increase in the rate of 45Ca++ efflux. In contrast, an augmented 45Ca++ efflux rate was observed with the alpha-1 agonist, phenylephrine (10(-4) M plus 10(-7) M rauwolscine). These results suggest that activation of postsynaptic alpha-2 adrenoceptors in CSV utilizes primarily extracellular Ca++ to produce contractions.

Comparison of the calcium antagonistic activity of nifedipine and FR34235 in the canine saphenous vein.

The effects of FR34235 (FR) on contractions and 45Ca2+ uptake stimulated by depolarization (80 mM KCl) and activation of postsynaptic alpha 1-adrenoceptors in the canine saphenous vein (CSV) were studied and compared with those of nifedipine. The 45Ca2+ uptake and contractions evoked by KCl were inhibited in a dose-dependent fashion by FR [concentration producing 50% inhibition [(IC50 = 2.0 +/- 0.5 and 1.2 +/- 0.3 X 10(-8) M, respectively)] and nifedipine (IC50 = 6.0 +/- 1.0 and 4.0 +/- 0.9 X 10(-8) M, respectively). FR was a potent inhibitor of the 45Ca2+ uptake and contractions of CSV induced by the selective alpha 1-agonist, SK&F l-89748 (l-[1,2,3,4-tetrahydro-8-methoxy-5-(methylthio)-2-naphthalenamine] ) (IC50 = 7.2 +/- 0.7 and 7.0 +/- 1.8 X 10(-8) M, respectively). In contrast, the contractions and 45Ca2+ uptake elicited by SK&F l-89748 were much less sensitive to nifedipine (IC50 = 6.0 +/- 1.4 and 6.6 +/- 0.4 X 10(-6) M, respectively). The results suggest the following: (a) both FR and nifedipine are potent antagonists of the voltage-dependent Ca2+ channels in CSV; and (b) FR is a more effective antagonist of receptor-operated Ca2+ entry in the saphenous vein than nifedipine.

N-[2-hydroxy-5-[2-(methylamino)ethyl]phenyl]methanesulfonamide. A potent agonist which releases intracellular calcium by activation of alpha 1-adrenoceptors.

N-[2-Hydroxy-5-[2-(methylamino)ethyl]phenyl]methanesulfonamide (SK&F 102652) has been prepared and characterized pharmacologically. It is a potent agonist with an EC50 of 25 nM at alpha 1-adrenoceptors as determined in the isolated perfused rabbit ear artery. On the presynaptic alpha 2-adrenoceptors of the guinea pig atrium it was considerably weaker, demonstrating an EC50 for inhibition of neurotransmission of 1200 nM and thus an overall alpha 1/alpha 2 selectivity ratio of greater than or equal to 48. In the vascular smooth muscle of the canine saphenous vein an EC100 concentration of this agonist in the presence of zero external Ca2+ induced 37.9 +/- 1.4% of the maximal contractile response due to this agent while the endogenous ligand norepinephrine evoked only 14.5 +/- 0.4% of the maximum. In the presence of low (1 microM) external calcium, this agent produced 78.3 +/- 5.3% of maximum while norepinephrine gave 45.3 +/- 7.4%. This agent produces alpha 1-adrenoceptor-mediated contraction primarily by release of intracellular Ca2+ and should provide a useful tool for characterizing alpha 1-receptor subtypes.

Calcium utilization in the vasoconstriction to enantiomers of SK&F 89748-A.

The effects of calcium entry blockade on the vasoconstriction to the selective alpha-1 adrenoceptor agonists d- and I-SK&F 89748-A (1,2,3,4-tetrahydro-8-methoxy-[5-methylthiol] -2-naphthalenamine HCl) in pithed rats and in rat and guinea-pig isolated aortas were studied. The log-dose response curves for the increase in diastolic pressure in pithed rats to i.v. injections of both enantiomers of SK&F 89748-A were maximally shifted only 5-fold to the right after pretreatment of the animals with nifedipine (1 or 3 mg/kg i.a.) or l-verapamil (0.3 or 1 mg/kg i.a.), showing the relative insensitivity of the vasopressor responses to SK&F 89748-A to these calcium entry blockers. In the rat isolated aorta, the contractile responses to the l-enantiomer of SK&F 89748-A were significantly more susceptible to calcium entry blockade with l-verapamil, nifedipine and D600 than the d-isomer. The contractions of the guinea-pig isolated aorta to both isomers proved highly insensitive to calcium slow channel blockade by D600. These results indicate that the contractions of vascular smooth muscle in pithed rats in vivo and of guinea-pig isolated aortas in vitro initiated by the d-and l-isomers of SK&F 89748-A are largely dependent upon processes not requiring an influx of extracellular calcium. The differential sensitivity to calcium entry blockade of the contractile responses of rat isolated aortas to the d- and l-isomers of SK&F 89748-A may reflect different ways of interaction of both enantiomers with the alpha-l adrenoceptor on rat aorta.

Measurements of 45Ca2+ uptake and contractile responses after activation of postsynaptic alpha 1-adrenoceptors in the isolated canine saphenous vein: effects of calcium entry blockade.

The increase in 45Ca2+ content produced by the EC100 concentrations of a series of alpha 1-agonists in the canine saphenous vein (CSV) was determined in the presence of 10(-7) M rauwolscine and found to be correlated (r = 0.92) with the intrinsic activities of the alpha 1-agonists in CSV. The degree of inhibition of 45Ca2+ uptake by nifedipine (1 microM) was inversely correlated (r = -0.97) with the intrinsic activities of the alpha 1-agonists and also inversely correlated (r = -0.95) with the 45Ca2+ uptake induced by the agonists. In the presence of 5 mM LaCl3, there was no significant 45Ca2+ uptake elicited by KCl (80 mM) or activation of postsynaptic alpha 1-adrenoceptors in this tissue, and the alpha 1-agonists were found to have a varied ability to release internal Ca2+ for contractions. It is concluded that (1) activation of alpha 1-adrenoceptors in CSV utilizes both intracellular and extracellular Ca2+ for contractions; (2) the increase in 45Ca2+ content after activation of post-synaptic alpha 1-adrenoceptors in CSV is directly correlated with the intrinsic ability of the alpha 1-agonists to induce contractions; and (3) the sensitivity of the 45Ca2+ uptake to nifedipine is inversely related to the intrinsic ability of the alpha 1-agonists to translocate extracellular Ca2+.

Postsynaptic alpha adrenoceptors on vascular smooth muscle.

A heterogeneous population of alpha adrenoceptors mediates vasoconstriction in the canine saphenous vein (CSV). Studies with isolated strips of venous smooth muscle incubated with selective alpha-adrenoceptor agonists and antagonists revealed that both alpha 1 and alpha 2 adrenoceptors exist independently in this tissue and both subtypes mediate a contractile response. Measurement of contractile responses in reduced or zero external calcium conditions indicates that stimulation of alpha 1 adrenoceptors induces contractions by influx of extracellular calcium and release of calcium from internal stores. In contrast, 45Ca uptake studies suggest that activation of the postsynaptic alpha 2 adrenoceptor produces vasoconstriction dependent only on influx of extracellular calcium. The influx of calcium produced by the selective alpha 2-adrenoceptor agonist BHT-920 is inhibited by calcium entry blockers. Measurements of transmembrane potentials from smooth muscle cells of the CSV suggest that alpha 1-adrenoceptor activation produces depolarization and contraction (electromechanical coupling) whereas alpha 2-adrenoceptor stimulation does not result in concentration-dependent depolarization of the smooth muscle cells (pharmacomechanical coupling).