Positive inotropic action of angiotensin II in the pithed rat

Summary The cardiovascular effects of angiotensin II were examined in aortic blood pressure-controlled and-uncontrolled pithed rats. Angiotensin II induced a dose-dependent increase in diastolic blood pressure, left ventricular pressure (LVP), dP/dt (the first derivative of LVP) and heart rate in pithed rats. The maximal responses for these parameters were similar to those to noradrenaline, except for the rise in diastolic blood pressure, where noradrenaline caused a greater increase than angiotensin II. After treatment with propranolol, the positive chronotropic effect of angiotensin II was abolished. Angiotensin II produced a dose-dependent increase in diastolic blood pressure, which was similar to that of vasopressin, and an increase in dP/dt_max, which proved much greater than that of vasopressin. When aortic blood pressure was controlled and the -receptors were blocked by propranolol, angiotensin II caused a dose-dependent increase in dP/dt_max without affecting the left ventricular enddiastolic pressure. The same results were obtained after both - and -adrenoceptors were blocked by propranolol and phentolamine. Losartan but not PD 123177 caused parallel rightward shifts of the dose-response curve of angiotensin II for dP/dt_max in the aortic blood pressure controlled pithed rat without altering the maximal response. It is concluded that in the pithed rat angiotensin II produced an increase in myocardial contractile force which is not mediated by - or -adrenoceptors. The inotropic effect appears to be mediated by angiotensin receptors, of the AT₁-subtype.Correspondence to J. Zhang at the above address     

Chronic β1-adrenoceptor blockade sensitises the H1 and H2 receptor systems in human atrium: role of cyclic nucleotides

We have reported that chronic treatment of patients with ₁-adrenoceptor blockers sensitises isolated atrial preparations to adrenaline, noradrenaline and 5-HT. We have now examined the effect of chronic treatment with -adrenoceptor blockers on responses to histamine of human right atrial appendages. We compared the effects of histamine on contractile force, cyclic AMP and cyclic GMP levels as well as cyclic AMP-dependent protein kinase (PKA) activity and explored the arrhythmogenic effects of histamine in preparations obtained from patients chronically treated or not treated with -adrenoceptor blockers.Histamine increased contractile force in paced preparations; the effects were blocked by the H₂ receptor antagonist famotidine (0.1–30 mol/1). The maximum inotropic response to histamine was doubled and the inotropic potency of histamine 0.4 log units greater in atria from -adrenoceptor blocker-treated compared to non -adrenoceptor blocker-treated patients. Histamine elicited frequency-dependent arrhythmias that were blocked by famotidine (30 mol/1) but not by mepyramine (1 mol/1). The incidence of arrhythmias was higher in atria from -adrenoceptor blocker-treated compared to untreated patients. Histamine increased both cyclic AMP and cyclic GMP levels, as well as PKA activity, significantly more in atria from -adrenoceptor blocker-treated compared to those from untreated patients. Mepyramine 1 mol/l prevented the histamine-evoked increase in cyclic GMP levels, reduced the inotropic hyperresponsiveness and abolished the hyperresponsiveness in cyclic AMP levels and PKA activity observed in patients chronically treated with blockers. Sodium nitroprusside 10 mol/l caused smaller increases of cyclic GMP levels than histamine and restored the contracile force depressed by mepyramine to its original level in atria from -adrenoceptor blocker-treated patients.The evidence is consistent with sensitisation of both the histamine H₁ and histamine H₂ receptor systems by chronic ₁-adrenoceptor blockade. H₁ receptor-mediated increases in cyclic GMP, enhanced through an as yet unknown mechanism by chronic ₁-adrenoceptor blockade, may inhibit phosphodiesterase 3 activity, thereby causing enhanced histamine-evoked increases in cyclic AMP levels and PKA activity, and accounting partially for the increased inotropic responses to histamine through H₂ receptors.     

MDL 72222: a potent and highly selective antagonist at neuronal 5-hydroxytryptamine receptors

Summary The properties of MDL 72222 (1H,3,5H-tropan-3-yl-3,5-dichlorobenzoate), a novel compound with potent and selective blocking actions at certain excitatory 5-hydroxytryptamine (5-HT) receptors on mammalian peripheral neurones, are described.On the rabbit isolated heart, MDL 72222 was a potent antagonist of responses mediated through the receptors for 5-HT present on the terminal sympathetic fibres. The threshold for antagonism was approximately 0.1 nM and the negative logarithm of the molar concentration of MDL 72222 which reduced the chronotropic response of the isolated rabbit heart to twice an ED₅₀ of 5-HT to that of the ED₅₀ was 9.27. MDL 72222 was also highly selective since responses to the nicotine receptor agonist, dimethylphenylpiperazinum iodine (DMPP), were inhibited only at concentrations more than 1000 times those necessary to inhibit 5-HT.In the anaesthetised rat, MDL 72222 produced marked blockade of the Bezold-Jarisch effect of 5-HT. Again, inhibition was selective since much higher doses of MDL 72222 failed to alter the response to electrical stimulation of the efferent vagus nerves. In contrast, MDL 72222 proved only a weak and essentially non-selective antagonist of responses mediated by the 5-HT M-receptor present on the cholinergic nerves of the guinea-pig ileum.MDL 72222 does not block smooth muscle contractile responses elicited by oxytocin or mediated through 5-HT D-receptors, muscarinic or nicotinic cholinoceptors or histamine H₁-receptors except at relatively high concentrations. Similarly, in a number of radioligand binding assays carried out using brain tissue membranes, the displacing effects of MDL 72222 were absent or weak at sites identifying compounds with activity at ₁, ₂ or -adrenoceptors, 5-HT₁ or 5-HT₂ receptors, benzodiazepine receptors or histamine H₁-receptors.MDL 72222 is the first reported selective and potent antagonist of responses mediated through the 5-HT receptors present on the terminal sympathetic neurones of the rabbit heart and on the neurones subserving the afferent limb of the Bezold-Jarisch reflex. The compound should provide a useful means by which responses mediated through such sites can be distinguished.     

Pharmacological characterization of cardiac histamine receptors: sensitivity to H1-receptor antagonists

The ability of five prototypic antihistamines (diphenhydramine, promethazine, cyclizine, chlorpheniramine and tripelennamine) to modify the cardiac actions of histamine was studied in order to determine the reactivity of cardiac histamine receptors. Histamine, as a function of dose, increases the rate, the force of contraction, the coronary flow and the time of atrioventricular conduction of the isolated guinea pig heart. Each of the five prototypic antihistamines failed to inhibit histamine-induced stimulation of cardiac rate and contractility. at the same time these antihistamines did inhibit the negative dromotropic effect of histamine. Chlorpheniramine and diphenhydramine antagonized the coronary dilating effect of histamine.These results suggest that two types of histamine receptors may be present in the guinea pig heart: H₁-receptors, at the atrioventricular node and coronary vessels; H₂-receptors at the sinoatrial node and on the ventricular fibers. The activity of these prototypic antihistamines is consistent with their action exclusively on H₁-receptors.     

Calcium dependency of the AT1-receptor mediated effects in the rat portal vein: influence of calcium antagonists

The calcium dependency of AT₁-receptor mediated contractions was studied in isolated rat portal vein preparations.The spontaneous phasic contractile force of the rat portal vein was increased (ED₅₀ = 1.76 mmol/l) and the frequency of contractions decreased by raising the extracellular calcium concentration. The Ang 11-induced rise in phasic contractile force (mediated by AT₁-receptors, Zhang et al. 1993) proved most pronounced at 0.9 mmol/l of calcium chloride, but it was weaker at either lower or higher calcium concentrations. The maximal increases in the phasic contractile force induced by Ang II were 2.4±0.4, 14.8±0.9 and 5±0.5 mN at calcium concentrations of 0.5, 0.9 and 2.5 mmol/l, respectively. Calcium antagonists reduced at the lower and abolished at the higher concentrations (nifedipine 2×10^–8 or 10^–7 mol/l; verapamil 10^–7 or 5 × 10^–7 mol/l; diltiazem 3 × 10^–7 or 10–6 mol/l) the spontaneous contractile force. All of these calcium antagonists caused a strong inhibition or suppression of the phasic contractions induced by Ang II.The rank order of potency was nifedipine >verapamil > diltiazem. Ang II (10^–6 mol/l) elicited a tonic contraction which was abolished by the AT₁-receptor antagonist losartan 10-6 mol/l but not by the AT₂-receptor antagonist PD 123177 (10–5 mol/l). Very high concentrations of nifedipine (10–6 mol/l), verapamil (5 × 10-6 mol/l) and diltiazem (5 × 10^–6 mol/l) almost suppressed the tonic effect evoked by the activation of AT₁-receptors.In a nominally Ca²⁺ free, EGTA-containing solution, a single supra-maximal concentration of Ang II (10^–6 mol/l) caused a transient contraction, also mediated by AT₁-receptors. This finding suggests the existence of Ang II-sensitive intracellular calcium stores in this preparation. The depletion of such stores proved complete after 4–6 min of perfusion in a Ca²⁺ free, EGTA-containing solution.In conclusion, various types of contractions (a transient contraction in a Ca²⁺-free medium, phasic and tonic contractions) induced by Ang II in the rat portal vein proved to be mediated by AT₁-receptors. These contractions were clearly modified by changes in the availability of extra- and possibly intracellular calcium ions. The calcium movements elicited by stimulation of AT₁-receptors in a calcium containing solution were inhibited by the three calcium antagonists investigated. Correspondence to: J. S. Zhang at the above address     

Positive chronotropic activity of angiotensin II in the pithed normotensive rat is primarily due to activation of cardiac β1-adrenoceptors

Summary In the pithed normotensive rat the intravenous administration of increasing doses of angiotensin II caused an increase in heart rate by a maximum of 93 ± 6 beats/min. This increase in heart rate was diminished dose-dependently by the concomitant infusion of the angiotensin II analogue saralasin. It was not affected by acute bilateral adrenalectomy but strongly reduced by pretreatment with reserpine. By using the selective antagonists of ₁-, ₂-, ₁- or ₂-adrenoceptors prazosin, yohimbine, atenolol and ICI 118,551, respectively, it was shown that this tachycardia was mediated by stimulation of cardiac ₁-adrenoceptors. Moreover, the high endogenous angiotensin II level following pithing contributed to the basal heart rate in the pithed rat model. From our experiments it may be concluded that angiotensin II induces tachycardia in the pithed rat primarily by stimulating the sympathetic ganglia leading to the release of noradrenaline, which subsequently activates cardiac ₁-adrenoceptors. Send offprint requests to P. A. van Zwieten at the above address     

A 5-HT4-like receptor in human left atrium

Summary The effects of 5-hydroxytryptamine (5-HT) on left atrial preparations obtained from 5 patients with terminal heart failure who were undergoing heart transplant surgery were investigated. The preparations were paced under isometric conditions. In the presence of (–)-pindolol I pmol/l (to block -adrenoceptors) and cocaine 6 gmol/l (to block tissue uptake of 5-HT) 5-HT increased contractile force with a pEC₅₀ of 7.0. The maximum effect of 5-HT amounted to 24.5% of that caused by a maximally effective concentration of (–)-isoprenaline (200 mol/l) and 25% of that caused by 6.75 mmol/l CaCl₂. The, effects of 5-HT were competitively antagonised by 3-tropanyl-1H-indole-3-carboxylate (ICS 205–930) with a pK_B of 6.8. The effects of 5-HT on cyclic AMP levels and cyclic AMP-dependent protein kinase activity were also studied using left atrial tissues from one of the patients; 5-HT increased the cyclic AMP content and stimulated the kinase. The results are consistent with the existence of a human left atrial 5-HT receptor which is similar to the recently identified human right atrial 5-HT receptor that resembles the 5-HT₄ receptor. The left atrial 5-HT₄-like receptor is functional in tissues obtained from patients with terminal heart failure. Send offprint requests to A. J. Kaumann at the above address     

Effects of intravenous propranolol and metoprolol and their interaction with isoprenaline on pulmonary function,heart rate and blood pressure in asthmatics

Summary The effects of propranolol (0.06 mg/kg i.v.), the selective ₁-receptor antagonist metoprolol (0.12 mg/kg i.v.) and a placebo on pulmonary function, heart rate and blood pressure have been compared in asthmatics. The interaction of these drugs with increasing doses of isoprenaline on the same variables was also studied. The two-blockers reduced resting heart rate to the same extent, indicating the same degree of blockade of cardiac-receptors. Both-blockers reduced the basal forced expiratory volume in one second (FEV₁), and the effect tended to be more pronounced after propranolol. Isoprenaline caused a dose-dependent increase in FEV₁ and vital capacity (VC). These effects were almost completely blocked by propranolol, whereas after metoprolol the changes approached that of the placebo. The isoprenaline-induced increase in heart rate and fall in diastolic blood pressure was also inhibited to a considerably greater extent by propranolol than by metoprolol. The results show a selectivity of metoprolol for so-called ₁-receptors and indicate that metoprolol may be used in asthmatics provided that it is combined with ₂-receptor-stimulating drugs.     

Dissociation of phosphoinositide hydrolysis and positive inotropic effect of histamine mediated by H1-receptors in guinea-pig left atria

Summary The present study was undertaken to determine whether the phosphoinositide hydrolysis is responsible for the positive inotropic effect of histamine in guinea-pig left atria. Histamine induced hydrolysis of phosphoinositides and a positive inotropic effect in a concentration-dependent manner. These effects were antagonized by chlorpheniramine (0.1 mol/l) but not by cimetidine (10 mol/l). At a concentration of 1 mol/l histamine produced a dual-component positive inotropic response composed of an initial increasing phase and a second and late developing, greater positive inotropic phase. Histamine (10 mol/l) caused a gradual increase in the formation of [³H]inositol trisphosphate (IP3) and a significant increase in the [³H]IP₃ level was detected 10 min after the stimulation. Thus, the increase in IP₃ did not precede the increase in force of contraction. The phospholipase C inhibitors 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate (100 mol/l) and neomycin (100 mol/l) significantly reduced the histamine-induced [³H]inositol monophosphate accumulation. However, pretreatment with the phospholipase C inhibitors did not affect the positive inotropic effect of histamine, either in its extent or in its pattern. The phorbol esters 12-O-tetradecanoylphorbol-13-acetate (TPA) (100 nmol/l) and phorbol-12,13-dibutyrate (PDBu) (100 nmol/l) also significantly inhibited the phosphoinositide hydrolysis induced by histamine. The inhibitory effect of the phorbol esters on the phosphoinositide response was completely abolished in the presence of 10 mol/l 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7), a protein kinase C inhibitor. TPA significantly attenuated the positive inotropic effect of histamine without changing the dual-component pattern, whereas PDBu merged two distinct components of the histamine inotropic response into one and potentiated the early part of the positive inotropic effect. However, neither of the changes which the phorbol esters produced in the positive inotropic response to histamine was blocked by H-7. In addition, H-7 itself failed to modify the positive inotropic effect of histamine. These results indicate that histamine induces hydrolysis of phosphoinositides in guinea-pig left atria that is mediated by H₁-receptors, but this biochemical event does not appear to contribute to the H₁-receptor-mediated positive inotropic action. Send offprint requests to Y. Hattori at the above address     

Resistance of adrenergic neurotransmission in the toad heart to adrenoceptor blockade

Summary Stimulation of sympathetic nerves to the toad heart produced increases in both the rate and force of cardiac beat. Although these responses were abolished by treatment with bretylium (10^–6 mol ·l^–1) or 6-hydroxydopamine (100 mg·kg^–1), and surgical sympathetic denervation, they were not abolished by treatment with propranolol (10^–6 mol·l^–1) and phentolamine (3×10^–6 mol·l^–1), either alone or in combination. The responses remaining after adrenoceptor blockade could not be ascribed to the effects of neurally released dopamine, ATP, adenosine, histamine or a variety of neuropeptides, although the participation of an as yet unidentified co-transmitter cannot be ruled out. Quantitative analysis of the interactions between propranolol and adrenaline on cardiac adrenoceptors, after blockade of -receptors and amine uptake mechanisms, revealed that these interactions do not comply with the conditions for simple competitivity. Therefore, in addition to its action on -receptors, adrenaline seems to be producing excitation of the toad heart by acting on adrenoceptors which cannot be classified as either -or -receptors. These results, together with the existence of close neuromuscular gaps (<50nm) in the toad heart, are consistent with the hypothesis that sympathetic excitation of the toad heart is mediated by both extra-junctionl -adrenoceptors, and junctional adrenoceptors which are neither -nor -receptors.     

5-Hydroxytryptamine-induced increase in left ventricular dP/dtmax does not suggest the presence of ventricular 5-HT4 receptors in the pig

Summary Although 5-hydroxytryptamine (5-HT) increases porcine atrial force and rate via 5-HT₄ receptors, its effect on left ventricular contractility is not known. Therefore, using the maximum rate of rise of left ventricular pressure (LVdP/dt_max) as an index of cardiac contractility, we have attempted to analyze the possible role of ventricular 5-HT₄ receptors in the anaesthetized pig. The full agonists at 5-HT₄ receptors, 5-HT and 5-methoxytryptamine (each 3, 10 and 30 g · kg^–1), and the -adrenoceptor agonist, isoprenaline (0.01, 0.03 and 0.1 g · kg^–1), increased heart rate, LVdP/dt_max and cardiac output. For a given degree of tachycardia, the increase in LVdP/dt_max by isoprenaline was substantially more than that observed with either 5-HT or 5-methoxytryptamine. The 5-HT₄ receptor partial agonist, renzapride (3, 10, 30, 100 and 300 g · kg^–1), also increased heart rate and LVdP/dt_max dose-dependently. When the heart was paced at 150 beats · min^–1, increases in LVdP/dt_max as well as cardiac output (except with the highest doses) by 5-HT, 5-methoxytryptamine and isoprenaline were clearly attenuated. However, the magnitude of attenuation of LVdP/dt_max responses by cardiac pacing was more marked in the case of 5-HT and 5-methoxytryptamine than with isoprenaline.The effects of renzapride (300 g · kg^–1) and tropisetron (0.3 and 3 mg · kg^–1) on increases in heart rate and LVdP/dt_max by 5-HT, 5-methoxytryptamine and isoprenaline were also studied. In the absence of atrial pacing, both renzapride and tropisetron (3 mg · kg^–1) effectively antagonized the responses to 5-HT and 5-methoxytryptamine; except for some decrease in the LVdP/dt_max response by tropisetron, the effect of isoprenaline remained essentially unchanged after the antagonists. During atrial pacing, renzapride significantly antagonized the responses to the first two doses of 5-HT, but the responses to the highest 5-HT dose and to 5-methoxytryptamine remained unaffected. Though, particularly after its higher dose, tropisetron reduced the responses to 5-HT and 5-methoxytryptamine, isoprenaline responses were also affected.The above results show that a significant part of the increase in LVdP/dt_max by 5-HT receptor agonists in the anaesthetized pig is a consequence of tachycardia elicited by these compounds via 5-HT₄ receptors. Since the increase in LVdP/dt_max, compared to tachycardia, was much less with 5-HT and 5-methoxytryptamine than with isoprenaline, and since the antagonism by renzapride and tropisetron against 5-HT and 5-methoxytryptamine during atrial pacing was relatively weaker and/or unspecific, it appears unlikely that the increase in LVdP/dt_max, during atria] pacing is mediated by ventricular 5-HT₄ receptors. This view is substantiated by our recent in vitro experiments where 5-HT (0.01 to 100 mol/l) failed to significantly increase contractions of porcine left ventricular trabeculae.Correspondence to P. R. Saxena at the above address     

Facilitatory presynaptic angiotensin receptors on the sympathetic nerves of the human saphenous vein and pulmonary artery. Potential involvement in β-adrenoceptor-mediated facilitation of noradrenaline release

Summary Spirally cut strips of human saphenous vein and pulmonary artery preincubated with ³H-noradrenaline were superfused in the presence of corticosterone and desipramine or cocaine. In the saphenous vein angiotensin I, angiotensin II and angiotensin III concentration-dependently increased the electrically (2 Hz) evoked tritium overflow (relative order of potency: angiotensin II > angiotensin I > angiotensin III). The angiotensin receptor antagonist saralasin displaced the concentration-response curve of angiotensin II to the right, and also blocked the facilitatory effect of angiotensin III. Captopril, an inhibitor of angiotensin converting enzyme, did not modify the concentration-response curve of angiotensin I and did not significantly diminish the release-increasing effect of the nonselective -adrenoceptor agonist isoprenaline, whereas saralasin attenuated the facilitatory effect of the ₂-adrenoceptor agonist procaterol. In the pulmonary artery the angiotensin receptor agonist Val⁵-angiotensin II-Asp¹--amide also increased the electrically evoked tritium overflow in a concentration-dependent manner. It is concluded that the sympathetic nerve fibres of the human saphenous vein (and probably of the human pulmonary artery as well) are endowed with facilitatory presynaptic angiotensin receptors. Angiotensin I exerted its facilitatory effect in the saphenous vein probably via direct stimulation of angiotensin receptors but not by conversion to angiotensin II. Furthermore, the ₂-adrenoceptor-induced facilitation of noradrenaline release may in part be mediated by local stimulation of angiotensin II synthesis, which may occur by increased formation or activation of renin and/or increased availability of angiotensinogen.This study was supported by a grant of the Deutsche Forschungsgemeinschaft Send offprint requests to M. Göthert at the above address     

Desensitization of kitten atria to chronotropic,inotropic and adenylyl cyclase stimulating effects of (-)isoprenaline

Summary Desensitization of kitten atria with 30 M (-)isoprenaline resulted in a 6-fold and 15-fold increase in the EC₅₀'s of (-)isoprenaline for its positive chronotropic effects (sinus pacemakers) and positive inotropic effects (left atria), respectively, but only in a 2-fold increase of the EC₅₀ of (-)isoprenaline for adenylyl cyclase stimulation in membrane particles from atria. However, maximum cyclase stimulation by (-)isoprenaline was decreased to 1/2 in membranes from (-)isoprenaline-treated atria, whereas maximum increases in rate of sinus pacemakers and force of left atria were unchanged and reduced by 15%, respectively. The high affinity -adrenoceptor blocker (-)bupranolol antagonized the adenylyl cyclase stimulation by (-)isoprenaline to similar extent in membranes from (-)isoprenaline and untreated atria, suggesting that the apparent affinity of -adrenoceptors for ligands is unchanged by desensitization. The evidence is compatible with the concept that desensitization is associated with decreased availability of receptors and with the view that near maximal positive chronotropic effects of catecholamines may be caused by only threshold increases in membrane adenylyl cyclase activity.     

Angiotensin II generation in the rat vena cava: Stimulation of local synthesis by β-adrenoceptor activation

Summary There is considerable evidence to suggest that isolated tissues have the capacity to generate angiotensin II and that angiotensin 11 thus generated may enhance noradrenergic neurotransmission. In the present study the possibility that there may be local formation of angiotensin II within the rat vena cava and its consequence to noradrenergic transmission has been investigated. The angiotensin II precursors, angiotensin I and a synthetic tetradecapeptide renin substrate, each enhanced the stimulation-induced efflux of radioactivity from tissues previously incubated with ³H-noradrenaline. The effects of angiotensin I were blocked by the converting enzyme inhibitor captopril and the receptor antagonist saralasin, indicating local conversion to angiotensin II. The effect of the tetradecapeptide was blocked by saralasin, but not by captopril, suggesting either a direct effect of this peptide or conversion to angiotensin II by a pathway not involving angiotensin converting enzyme. The -adrenoceptor agonist isoprenaline enhanced noradrenergic transmission in the rat vena cava, relaxed guinea-pig trachea precontracted with carbachol and increased heart rate in rat isolated atria. Captopril and saralasin blocked the effect of isoprenaline in the vena cava, but did not alter its effects in the atria or trachea. This suggests that in the rat vena cava the facilitation of noradrenergic transmission by isoprenaline may involve stimulation of local angiotensin II production. Send offprint requests to D. F. Story at the above address     

Biochemical effects and drug levels in rats after long-term treatment with the specific 5-HT-uptake inhibitor,citalopram

The effects in rats of long-term administration of the potent, specific 5-HT uptake inhibitor citalopram have been investigated. Citalopram hydrobromide (MW=405) was given in the diet, 99 or 25 mol/kg daily, for 13 days or orally, 49 mol/kg twice a day, for 14 days. High plasma and brain levels of citalopram were found during the treatment period, whereas negligible amounts were found 24 h after withdrawal. The 5-HT uptake mechanism in blood platelets was completely blocked, since levels of whole blood 5-HT during and shortly (2 days) after treatment were decreased by 75–90%. The drug load after the two highest doses in terms of plasma drug levels was the same as in depressed patients treated with citalopram. Receptor binding technique ex vivo was applied to different brain parts to measure receptor parameters for several neurotransmitters. All data were evaluated by Eadie-Hoffstee analysis. No changes were seen in B _max and K _d for -receptors (³H-dihydroalprenolol) in frontal cortex, occipital+temporal cortex, whole cortex and limbic structures, 5-HT₂ receptors (³H-spiroperidol) in frontal and whole cortex, ₁-receptors (³H-prazosin) in rest of brain and DA D-2 receptors (³H-spiroperidol) in corpus striatum and limbic structures. The uptake mechanism for 5-HT as well as the inhibitory effect of citalopram on this uptake remained unaffected in brain synaptosomes derived from control and from citalopram (99 mol/kg)-treated rats. Thus long-term treatment with citalopram does not induce changes in neurotransmitter receptors as seen with most tricyclic as well as newer atypical antidepressants. Most striking is the lack of - and 5-HT₂ receptor down-regulation. Since citalopram clinically shows clear antidepressant activity, this down-regulation does not seem to be a prerequisite of antidepressant activity.     

Phentolamine and hypoxia: modulation of contractility and α1-adrenoceptors in isolated rat atria

The hypoxia-induced effects on the binding sites and affinity constant of adrenoceptors, in the presence and absence of phentolamine, were determined for atrial membranes of hearts from normal and genetically hyperlipidaemic Yoshida (YOS) rats. Atrial function was also measured during normoxia and hypoxia, in the presence and absence of phentolamine.Hypoxia increased a1-adrenoceptor density in atrial membranes of normal rats (B_max 10.6 to 26.7 fmoles/mg protein). Phentolamine prevented the increase in the B_max of ₁-adrenoceptors and increased the equilibrium dissociation constant of these receptors (K _D 0.17 to 0.53 nmol/l). Beta-adrenoceptors did not change during hypoxia, but the B_max was slightly increased (26%) in the presence of phentolamine. Thus, the ₁/\ ratio increased from 0.40 in normoxia to 1.06 in hypoxia. In normoxic atria from YOS rats, the ₁/\ ratio was already elevated (0.86) in comparison to control rats (mainly due to a higher density of at-adrenoceptors in atrial membranes from YOS rats). This ratio was not modified by hypoxia (0.84), but decreased when phentolamine was present (0.30).Hypoxia reduced the force of contraction and increased diastolic tension of atria of normal rats, while the sinus rate was not significantly modified. Phentolamine abolished the increase in diastolic tension and reduced the negative effect of hypoxia on contractile force. In YOS rat atria, functional parameters were modified by hypoxia in a qualitatively similar way to that of normal rat atria.The observed increase in ₁-adrenoceptor density during hypoxia is in accordance with the results of experiments with animal models of the ischaemic heart and with findings in human heart failure. The possible therapeutic significance of these data is considered. Correspondence to: G. Fassina at the above address     

Effects of Ca2+ channel antagonists and ryanodine on H1-receptor mediated electromechanical response to histamine in guinea-pig left atria

Summary Effects of organic Ca²⁺ channel antagonists, Ni²⁺ and ryanodine on the electrophysiological and positive inotropic responses to histamine were examined in isolated guinea-pig left atria. Histamine increased force of contraction, prolonged action potential duration (APD) and hyperpolarized the membrane in a concentration-dependent manner. Histamine at a concentration of 1 mol/l produced a dual-component positive inotropic response composed of an initial increasing phase (initial component) and a second an late developing, greater positive inotropic phase (second component), whereas causing monophasic changes in APD and resting potential. The electrophysiological and dual-component positive inotropic effects induced by histamine were antagonized by chlorpheniramine (1 mol/l) but not by cimetidine (10 mol/l), indicating that both effects are exclusively mediated by H₁-receptors. The positive inotropic response to 1 mol/l histamine was changed by the pretreatment with nifedipine (1 mol/l) and nisoldipine (1 mol/l). In the presence of these dihydropyridines, the second component was almost completely abolished, while the initial component was hardly affected. On the other hand, verapamil (3 mol/l) and diltiazem (10 mol/l) failed to modify the multiphasic inotropic response to histamine. None of the Ca²⁺ channel antagonists affected the histamine-induced APD prolongation. In the presence of Ni²⁺ at a concentration of 0.3 mmol/l, at which it produced no negative inotropic action, the second component of the positive inotropic effect of histamine was specifically suppressed whereas the histamine-induced APD prolongation was unaffected. Preferential attenuation of the second component was also observed in the presence of 30 nmol/l ryanodine. However, the electrophysiological alterations caused by histamine remained unchanged. These results suggest that in guinea-pig left atria the H₁-receptor-mediated prolongation of APD seems unlikely to be due to enhancement of the slow inward Ca²⁺ current. Conversely, the increased Ca²⁺ influx as a result of the APD prolongation may contribute to the second component of the positive inotropic effect of histamine. Dihydropyridine Ca²⁺ channel antagonists, Ni²⁺ and ryanodine are all capable of inhibiting the second component, but do so possibly via different mechanisms, implying the complicated mechanisms underlying the H₁-receptor-mediated positive inotropic effect.Send offprint requests to Y. Hattori     

A postsynaptic effect of tyramine in ventricular muscle

Summary Tachyphylaxis and susceptibility to pretreatment with reserpine demonstrate that tyramine increases contractility of the guinea-pig papillary muscle in concentrations up to 10^–4 M by means of an indirect sympathomimetic effect.In higher concentrations (up to 3×10^–3 M) tyramine causes an increase in force of contraction which is characterized by a slowing of relaxation of the isometric contraction curve. Pretreatment with reserpine reveals that this positive inotropic effect is composed of an indirect sympathomimetic and a direct post-synaptic effect; the latter is not subject to tachyphylaxis and produces by itself (at 3×10^–3 M) an increase in force of contraction which amounts to about 50% of the maximum of the indirectly produced inotropic effect.The postsynaptically-induced positive inotropic effect of tyramine is not antagonized by propranolol (5×10^–6 M). It is unlikely, therefore, that it is brought about by stimulation of adrenergic -receptors.The height of the isometric contraction curve in reserpine-pretreated muscles is increased by tyramine as a result of an increase in the rate of force development despite a marked concentration-dependent increase in relaxation time (t ₂).The direct inotropic effect of tyramine is correlated with a prolongation of the duration of the action potential (AP) which amounts to 70% at 3×10^–3 M. In addition to its influence on the duration of the AP, tyramine slows the rate of depolarization and decreases the membrane resting potential.     

β 1- andβ 2-adrenoceptor binding and functional response in right and left atria of rat heart

Summary The properties of ₁- and ₂-adrenoceptors in right and left atria of rat heart, and their roles in mediating chronotropic and inotropic responses to-adrenoceptor agonists were examined. [¹²⁵I](-)-pindolol (¹²⁵IPIN) bound saturably and specifically to a single class of high affinity sites in homogenates of both right and left atria. Thek ₁'s for association in right and left atria were 6.5×10⁹ l/mol-min and 2.3×10⁹ l/mol-min respectively, while thek _–1's for dissociation were 0.20 min^–1 and 0.17 min^–1. The kinetically determinedK _D's were 75 pmol/l in right and 30 pmol/l in left atria and were similar to the equilibriumK _D's determined from Scatchard analysis of saturation isotherms of specific¹²⁵IPIN binding. Inhibition of¹²⁵IPIN binding by-adrenoceptor antagonists was stereoselective and the order of potency was timolol > 1-propranolol > d-propranolol > sotalol. Inhibition by ₁- and ₂-adrenoceptor subtype selective antagonists yielded flat displacement curves with low Hill coefficients. Nonlinear regression analysis of displacement by ₁-selective (practolol, atenolol and metoprolol) and ₂-selective (ICI 118,551) antagonists gave estimates of the proportion of ₁- and ₂-adrenoceptors present in rat atria. Right atria contained 67±4.2% ₂-adrenoceptors and 33±4.2% ₂-adrenoceptor, while left atria contained 67±2.8% ₁- and 33±2.8% ₂-adrenoceptors. Increases in the rate of spontaneously beating right atria and the force of electrically driven left atria caused by-adrenoceptor agonists were also measured. pA₂ values for non-subtype selective-adrenoceptor antagonists in inhibiting isoprenaline-induced increases in rate and force were highly correlated withK _D values determined for specific¹²⁵IPIN binding. pA₂ values for ₁- and ₂-selective antagonists in inhibiting isoprenaline-induced increases in rate and force correlated well with the pK _D values of these drugs in binding to ₁-adrenoceptors, but not with the pK _D values in binding to ₂-adrenoceptors. Dose-response curves for stimulation of both rate and force by the ₂-selective agonists procaterol and zinterol were shifted to a much greater extent by selective blockade of ₁-adrenoceptors with metoprolol than by selective blockade of ₂-adrenoceptors with ICI 118,551, suggesting that these compounds caused their effects by activating ₁-adrenoceptors. These results suggest that ₁- and ₂-adrenoceptors coexist in both left and right atria of rat heart in approximately a 21 ratio, however only ₁-adrenoceptors mediate the chronotropic and inotropic effects of-adrenoceptor agonists.Supported by a grant from the American Heart Association — Georgia Affiliate     

Biochemical effects of the antidepressant paroxetine,a specific 5-hydroxytryptamine uptake inhibitor

Paroxetine was shown to be a potent (K _i =1.1 nM) and specific inhibitor of [³H]-5-hydroxytryptamine (5-HT) uptake into rat cortical and hypothalamic synaptosomes in vitro. Lineweaver-Burk kinetic analysis determined that this inhibition was competitive in nature, implying a direct interaction with the 5-HT uptake transporter complex. Oral administration of paroxetine produced a dose-related inhibition of [³H]-5-HT uptake (ED ₅₀=1.9 mg/kg) into rat hypothalamic synaptosomes ex vivo with little effect on [³H]-l-noradrenaline (NA) uptake (ED ₅₀>30 mg/kg). This selectivity for 5-HT uptake was maintained after oral dosing for 14 days. Paroxetine (ED ₅₀ 1–3 mg/kg PO) prevented the 5-HT depleting effect of p-chloroamphetamine (PCA) in rat brain, demonstrating 5-HT uptake blockade in vivo. Radioligand binding techniques in rat brain in vitro showed that paroxetine has little affinity for ₁, ₂ or adrenoceptors, dopamine (D₂), 5-HT₁, 5-HT₂ or histamine (H₁) receptors at concentrations below 1000 nM. Paroxetine demonstrated weak affinity for muscarinic receptors (K _i =89 nM) but was at least 15 fold weaker than amitriptyline (K _i =5.1 nM). Paroxetine, therefore, provides a useful pharmacological tool for investigating 5-HT systems and furthermore should be an antidepressant with reduced tricyclic-like side-effects.