RESUMO
The experimental anticancer drug cyclopentenyl cytosine (CPEC) was associated with cardiotoxicity in a phase I study. The aim of the present study was twofold; first we investigated whether the observed effects could be reproduced in in-vitro and in-vivo rat models. Second, we intended to investigate the underlying mechanism of the possible cardiotoxicity of CPEC. Effects on frequency and contractility were studied on the isolated atria of 18 male Wistar rats. Atria were incubated with 0.1 mmol L(-1) (n = 6) or 1 mmol L(-1) (n = 6) CPEC for 1.5 h and compared with control atria (incubation with buffer solution, n = 6). The cardiac apoptosis-inducing potential was studied in-vivo on 66 rats by 99mTc-Annexin V scintigraphy, followed by postmortem determination of radioactivity in tissues, histological confirmation with the TUNEL assay (late-phase apoptosis), and immunohistochemical staining for cleaved caspase 3 and cytochrome C (early-phase apoptosis). Serum levels of the necrotic cardiomyopathy marker troponin T were also determined. No effect on heart frequency was found in the isolated atria after CPEC treatment. A trend towards a decrease of contraction force was observed. However, the differences were not statistically significant. 99mTc-Annexin V scintigraphy showed no increase in cardiac uptake ratio upon CPEC treatment in the in-vivo rat model, which was confirmed by determination of radioactivity in heart versus blood ratios. At each section a few individual isolated late apoptotic cells (< 5) could be identified by the TUNEL assay in the highest CPEC dose group (90 mg kg(-1)) but not in controls or in rats treated with 60 mg kg(-1) CPEC. Staining for the early apoptosis markers cleaved caspase 3 and cytochrome C did not reveal any significant differences between treated and control rats. Cardiac troponin T levels were not increased after CPEC treatment. CPEC does not affect heart frequency or contraction force in our cardiotoxicity models. Moreover, we did not find an indication of CPEC-induced apoptosis in heart tissue.
Assuntos
Antineoplásicos/toxicidade , Cardiomiopatias/induzido quimicamente , Citidina/análogos & derivados , Coração/efeitos dos fármacos , Animais , Anexina A5/metabolismo , Apoptose/efeitos dos fármacos , Cardiomiopatias/sangue , Cardiomiopatias/patologia , Citidina/toxicidade , Modelos Animais de Doenças , Coração/diagnóstico por imagem , Átrios do Coração/diagnóstico por imagem , Átrios do Coração/efeitos dos fármacos , Átrios do Coração/patologia , Átrios do Coração/fisiopatologia , Marcação In Situ das Extremidades Cortadas , Masculino , Contração Miocárdica/efeitos dos fármacos , Miocárdio/patologia , Compostos de Organotecnécio , Ratos , Ratos Wistar , Tomografia Computadorizada de Emissão de Fóton Único , Troponina T/sangueRESUMO
Sympathetic neurotransmission in tissues with intact sympathetic nerve arborization is extensively dependent on calcium influx via N-type calcium-channels. It was the objective of the present study to assess and compare the claimed sympatholytic effect of the 1,4-dihydropyridine compound cilnidipine with other voltage-dependent calcium-channel (VDCC) antagonists. We studied these compounds by means of three different models. In the rabbit isolated thoracic aorta, the alleged sympatholytic properties displayed by these compounds were evaluated in the noradrenaline spillover model. Additionally, the influence of cilnidipine on stimulation-induced constrictor responses was studied in the rat isolated tail artery (male Wistar rats, 250-300 g) in addition to its effect on noradrenaline-induced contractions. Finally, we studied the influence of cilnidipine and other calcium-channel blockers on stimulation-induced chronotropic responses, in order to address N- or L-type selectivity, in the pithed rat model (male Wistar rats, 260-320 g). Furthermore, we evaluated their effect on noradrenaline-induced tachycardia. In the isolated rabbit thoracic aorta preparation omega-conotoxin GVIA (0.1 microM) nearly abolished the sympathetic outflow caused by stimulation, whereas nifedipine (0.1 microM) and amlodipine (1 microM) did not influence the evoked noradrenaline release. Cilnidipine (1 microM) significantly attenuated the response by nearly 18% and mibefradil (1 microM) by c. 42%. The stimulation-induced constrictor response (prejunctional effect) in the rat isolated tail artery could be blocked by omega-conotoxin GVIA (0.5 and 1 microM). Cilnidipine (10 nm and 0.1 microM) significantly attenuated responses to stimulation by maximally 20%, whereas it did not influence the constrictor response to noradrenaline (postjunctional effect). The mean heart rate in the pithed rat model amounted to 309.3 +/- 3.6 beats/min (bpm). Electrical stimulation of the cardio-accelerator nerves (C7-Th1) resulted in an increase by 106.7 +/- 2.2 bpm. All antagonists studied, except for nifedipine, attenuated the chronotropic response to stimulation (P < 0.05). The rank order of sympatholytic efficacy was: omega-conotoxin GVIA (84.8%), mibefradil (75.1%), cilnidipine (43.0%) and amlodipine (34.8%). Noradrenaline (10 nmol/kg) increased the heart rate by 117.8 +/- 2.7 bpm. This chronotropic response was influenced equally well by the calcium-channels blockers as observed in the stimulation (prejunctional) experiment. In conclusion, the N-type channel blocking properties and thus sympatholytic effect of cilnidipine could be demonstrated in some (vascular) but not all (cardiac) models studied. At the level of the vasculature cilnidipine reduced the neurotransmitter release to electrical stimulation in both the noradrenaline spillover model and in the model of the rat isolated tail artery, respectively. For amlodipine and nifedipine no sympatholytic activity could be demonstrated. In the pithed rat model, we were unable to demonstrate a selective N-type blocking effect for the VDCC-antagonists.
Assuntos
Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio Tipo N/efeitos dos fármacos , Di-Hidropiridinas/farmacologia , Simpatolíticos/farmacologia , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/fisiologia , Artérias/efeitos dos fármacos , Artérias/fisiologia , Estado de Descerebração , Estimulação Elétrica , Coração/inervação , Frequência Cardíaca/efeitos dos fármacos , Técnicas In Vitro , Masculino , Norepinefrina/antagonistas & inibidores , Norepinefrina/metabolismo , Norepinefrina/farmacologia , Coelhos , Ratos , Ratos Wistar , Sistema Nervoso Simpático/fisiologia , Vasoconstrição/efeitos dos fármacosRESUMO
OBJECTIVE: It was the objective of this study to investigate whether a facilitatory role of vasopressin (AVP) on sympathetic nerve activity can be demonstrated in humans at the peripheral level. METHODS: Eight subjects (32 +/- 2.3 years) participated in this study. Forearm blood flow (FABF) was measured using the venous occlusion plethysmography model. Each session was performed in the presence of a continuous infusion (into the brachial artery) of AVP in sub-pressor dosage of 0.008 ng/kg per min, or NaCl 0.9%. Using lower-body negative pressure (LBNP) (-10, -20 and -30 mmHg) the combined pre- and postsynaptic action of AVP on the sympathetic nervous system was investigated. This was followed by a second protocol in which the possible postsynaptic effects of AVP were evaluated with intra-arterial infused norepinephrine (NE). RESULTS: The baseline FABF was 5.2 +/- 0.6 ml/100 ml per min. After infusion of AVP (0.008 ng/kg per min), the FABF remained unchanged at a flow of 5.5 +/- 0.6 ml/100 ml per min (P = 0.26). LBNP caused a pressure-dependent decrease in FABF (25.6 +/- 4.4, 29.0 +/- 6.1 and 38.6 +/- 6.9%, for -10, -20 and -30 mmHg, respectively). AVP significantly enhanced the FABF responses to lower-body negative pressures (38.0 +/- 8.6, 49.3 +/- 5.1 and 58.9 +/- 6.3%, respectively (P = 0.014). NE caused a dose-dependent vasoconstriction by 3.1 +/- 4.6, 17.0 +/- 4.3 and 23.2 +/- 4.9%, at dosages of 10, 20 and 40 pg/min, respectively, unaffected by AVP (P = 0.91). CONCLUSIONS: We conclude that AVP can facilitate vasoconstriction mediated by the peripheral sympathetic nervous system at the presynaptic level in humans.
Assuntos
Arginina Vasopressina/administração & dosagem , Terminações Pré-Sinápticas/efeitos dos fármacos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Sistema Nervoso Simpático/efeitos dos fármacos , Vasoconstritores/administração & dosagem , Adulto , Artéria Braquial/inervação , Artéria Braquial/fisiologia , Feminino , Humanos , Pressão Negativa da Região Corporal Inferior , Masculino , Norepinefrina/administração & dosagem , PletismografiaRESUMO
The present study was designed to investigate the influence of maturation (young versus adult) on the angiotensin II-mediated facilitation of sympathetic nerve traffic (prejunctional AT1-receptor) as well as on the angiotensin II-mediated vasoconstriction (postjunctional AT1-receptor). Additionally, we investigated the inhibitory effect of the selective AT1-receptor antagonist eprosartan on angiotensin II-mediated responses at both sites during maturation. Male New Zealand White rabbits, aged 12 to 14 and 35 to 38 weeks (young versus adult, respectively), were used. To study angiotensin II at the neuronal AT1-receptor we investigated its influence on electrical field stimulation (EFS)-evoked sympathetic neurotransmission in the isolated thoracic aorta in a noradrenaline spillover model. To study the effects of angiotensin II at the level of the vasculature concentration-response curves for angiotensin II were constructed. In both models the influence of eprosartan on angiotensin II-mediated responses was studied. Angiotensin II (0.01 nM-0.1 microM) concentration-dependently enhanced the EFS-evoked noradrenaline release in both groups. No differences concerning the relative (approximately 100%, P > 0.05) and absolute facilitation were observed between groups, although concentrations required in adult rabbits exceeded those in young animals by 1 unity log M increment. Eprosartan concentration-dependently attenuated the angiotensin II-enhanced (10 nM) sympathetic outflow. The inhibitory potency differed approximately by a factor ten between both groups (young; pIC50 7.91 +/- 0.12 and adult; pIC50 8.81 +/- 0.31, respectively, P < 0.05). Angiotensin II (1 nM-0.3 microM) caused a concentration-dependent increase in contractile force (young rabbits; Emax 20.62 +/- 2.24 mN, pD2 8.16 +/- 0.04, n = 10 and adult rabbits; Emax 21.64 +/- 3.86 mN, pD2 7.63 +/- 0.02, n = 7). We observed approximately a 0.5 unity log M increment difference in potency, although the maximal absolute contraction was similar in both groups. Eprosartan (0.1 nM-0.1 microM) inhibited the angiotensin II-mediated contractions in a competitive manner in preparations from young rabbits (pA2 8.90 +/- 0.11, n = 24), whereas a mixed form of antagonism, in the same concentration range, was observed in tissues from adult rabbits. One possible explanation concerning these experiments is that maturation influences the AT1-receptor density negatively, although further studies are necessary to test this question. In addition, the decreased AT1-receptor density offers a potential explanation for the discrepancy in the profile of antagonism displayed by eprosartan in young compared with adult rabbits.
Assuntos
Acrilatos/farmacologia , Envelhecimento/fisiologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II , Antagonistas de Receptores de Angiotensina , Anti-Hipertensivos/farmacologia , Imidazóis/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Sistema Renina-Angiotensina/efeitos dos fármacos , Tiofenos , Animais , Desipramina/farmacologia , Eletrofisiologia , Masculino , Norepinefrina/farmacologia , Coelhos , Receptor Tipo 1 de Angiotensina/fisiologia , Receptores de Angiotensina/fisiologia , Sistema Renina-Angiotensina/fisiologia , Tetrodotoxina/farmacologia , Vasoconstrição/efeitos dos fármacosRESUMO
A series of iodinated thiadiazolyltetrahydro-1-methyl-pyridine (TZTP) compounds was synthesized and evaluated in vitro and in vivo as potential radioligands for imaging of the muscarinic M2 receptor subtype with SPET. One of these compounds, 5-(E)-iodopentenylthio-TZTP, has high in vitro affinity (Ki = 4.9 nM) and moderate selectivity for the muscarinic M2 receptor subtype. Although the uptake pattern in the biodistribution studies in rats is consistent with muscarinic M2 receptor disribution, specific in vivo binding to these receptors could not be demonstrated. The usefulness of this tracer in human SPET imaging may therefore be limited.
Assuntos
Piridinas/farmacocinética , Receptor Muscarínico M2/metabolismo , Tiadiazóis/farmacocinética , Animais , Estudos de Viabilidade , Masculino , Taxa de Depuração Metabólica , Especificidade de Órgãos , Piridinas/química , Compostos Radiofarmacêuticos/síntese química , Compostos Radiofarmacêuticos/farmacocinética , Ratos , Ratos Wistar , Tiadiazóis/química , Distribuição TecidualRESUMO
To investigate the involvement of the mitogen-activated protein kinase (MAPK) family of extracellular signal-regulated kinase (ERK) 1 and 2 (MAPKerk1/2) in the vasopressin-mediated vasoconstriction in the rat aorta. Vasopressin-induced vasoconstriction was measured in isolated rat thoracic aortae in the presence or absence of MAPKerk1/2 kinase (MKKmek1/2) inhibitors. Thereafter the MAPKerk1/2 phosphorylation in the rat aorta was quantified using Western blot analysis. Vasopressin (1-300 nm) induced a concentration-dependent vasoconstriction, which could be inhibited concentration dependently by the selective MKKmek1/2 inhibitors, PD 98059 (10 and 100 microm) and U 0126 (10 and 100 microm). Western blot analysis revealed a 2.7 +/- 0.6-fold increase in the MAPKerk1/2 phosphorylation induced by vasopressin (300 nm). This phosphorylation could be dose dependently prevented by both PD 98059 (100 microm) and U 0126 (10 and 100 microm). These results indicate that vasoconstriction induced by vasopressin is partly regulated by the MAPKerk1/2 pathway.
Assuntos
Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Vasoconstritores/farmacologia , Vasopressinas/farmacologia , Animais , Aorta Torácica/efeitos dos fármacos , Western Blotting , Butadienos/farmacologia , Inibidores Enzimáticos/farmacologia , Flavonoides/farmacologia , Técnicas In Vitro , Masculino , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Contração Muscular/efeitos dos fármacos , Músculo Liso Vascular/efeitos dos fármacos , Contração Miocárdica/efeitos dos fármacos , Nitrilas/farmacologia , Fosforilação , Potássio/farmacologia , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Angiotensin (AT) II is known to enhance responses to electrical field stimulation (EFS) via AT1 receptors located on sympathetic nerve terminals. Differences in potency exist between AT1 receptor antagonists regarding the inhibition of the prejunctional and postjunctional AT1 receptors. It is hypothesized that prejunctional AT1 receptors might belong to the AT1B receptor subtype. Accordingly, the authors investigated whether AT1B receptor inhibition by high concentrations of PD123319 could suppress ATII-augmented noradrenergic transmission (prejunctional) in the rabbit thoracic aorta by means of a noradrenaline spillover model. Additionally, the influence of PD123319 on ATII-enhanced constrictor responses to electrical field stimulation was investigated in the isolated rabbit mesenteric artery. Furthermore, the authors investigated whether PD123319 could influence the constrictor responses (postjunctional) to ATII in both preparations. In the thoracic aorta, ATII (10 nM) caused a significant enhancement of EFS-evoked [3H]-noradrenaline release by a factor of 2.0 +/- 0.1. This reinforcement could be inhibited by PD123319 (0.1, 1, and 10 microM). The constrictor response to ATII was unaffected by PD123319. In the mesenteric artery, ATII (0.5 nM) caused a significant enhancement of constrictor responses to EFS by factors of 2.9 +/- 0.3, 2.3 +/- 0.3, and 1.6 +/- 0.1 at 1, 2, and 4 Hz, respectively. This enhancement could be attenuated by PD123319 (1 and 10 microM). The constrictor response to ATII was unaffected by PD123319. It is concluded that the prejunctional AT1 receptors belong to the AT1B subtype whereas postjunctional AT1 receptors do not.
Assuntos
Angiotensina II/farmacologia , Imidazóis/farmacologia , Piridinas/farmacologia , Receptor Tipo 1 de Angiotensina/efeitos dos fármacos , Vasoconstritores/farmacologia , Animais , Interações Medicamentosas , Estimulação Elétrica , Masculino , Músculo Liso Vascular , Coelhos , Receptor Tipo 1 de Angiotensina/classificaçãoRESUMO
Congestive heart failure (CHF) is characterised by activation of the renin-angiotensin-aldosterone system (RAAS) and the sympathetic nervous system (SNS). Both systems are known to interact and to potentiate each other s activities. We recently demonstrated that angiotensin II (Ang II) enhances sympathetic nerve traffic via prejunctionally-located AT1-receptors. At present, little is known about the effects of Ang II at the level of the sympathetic neurones in CHF. Accordingly, we investigated the effect of Ang II in the presence and absence of the AT1-receptor antagonist, eprosartan, on stimulation-induced nerve traffic in isolated thoracic aorta preparations obtained from rabbits suffering from experimentally-induced CHF. Control-preparations were obtained from age-matched animals. Sympathetic activity was assessed by a [3H]noradrenaline spill-over model. Additionally, Ang II constrictor responses were compared between CHF and control vessels in the presence and absence of eprosartan. Additionally, to study postjunctional facilitation, the effects of Ang II on postsynaptic a-adrenoceptor-mediated responses were studied using noradrenaline. Stimulation-evoked SNS-neurotransmission was similar in both groups (CHF versus control). Ang II (0.1 nM 0.1 M) caused a concentration-dependent increase of the stimulation-evoked sympathetic outflow in both groups, with a maximum at 10 nM (control [n=7], FR2/FR1 2.03+0.11 and CHF- preparations [n=7], FR2/FR1 1.71+0.07). The enhancement by Ang II was decreased in CHF- preparations compared with controls (p<0.05). Eprosartan concentration-dependently attenuated the Ang II-enhanced (10 nM) sympathetic outflow in both CHF- and control preparations. The sympatho-inhibitory potency of eprosartan was similar in both groups (control pIC50 8.81 0.31; CHF 8.65+0.42). Ang II (1 nM 0.3 M) concentration-dependently increased the contractile force in control preparations (Emax 21.64+3.86 mN, pD2 7.63+0.02, n=7). Eprosartan (1 nM 0.1 M) influenced the Ang II- contractions via a mixed form of antagonism. In CHF-preparations, Ang II caused impaired vascular contraction. The KCl-induced contraction was decreased in the CHF- compared with control preparations (13.02+0.64 mN versus 30.40+0.89 mN). The relative Ang II contraction (% of KCl) was also decreased (2.3% vs. 58.0%). Concentration-response curves to noradrenaline (%KCl) were similar (control pD2 6.93+0.05, Emax 131.0+2.7; CHF pD2 7.00+0.05, Emax 136.7+2.6) (p>0.05) and were not affected by Ang II. We conclude that Ang II-enhanced sympathetic neurotransmission is mediated by the prejunctional AT1-receptor in both control and CHF-preparations. The decreased facilitation of SNS effects by Ang II may be explained by down-regulation or desensitisation of the neuronal AT1-receptor. Additionally, the aortic contractile capacity in heart failure rabbits appears to be decreased, probably as a result of heart failure-associated neuroendocrine and functional changes.
Assuntos
Angiotensina II/farmacologia , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/fisiopatologia , Insuficiência Cardíaca/fisiopatologia , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/fisiopatologia , Tiofenos , Acrilatos/administração & dosagem , Acrilatos/farmacologia , Angiotensina II/administração & dosagem , Bloqueadores do Receptor Tipo 1 de Angiotensina II , Animais , Aorta Torácica/inervação , Relação Dose-Resposta a Droga , Estimulação Elétrica , Imidazóis/administração & dosagem , Imidazóis/farmacologia , Técnicas In Vitro , Masculino , Concentração Osmolar , CoelhosRESUMO
OBJECTIVES: To identify and quantify the stimulatory and inhibitory activity of endothelial factors on Arginine vasopressin (AVP)-induced contractions. METHODS: In a standard organ bath set-up for isometric force recording, rabbit isolated renal artery rings were exposed to cumulative concentrations of AVP. Experiments were performed in the presence or absence of functional endothelium, or in the presence of N-Nitro-L-Arginine 10 microM (L-NNA) (NO-synthase inhibitor). RESULTS: Arginine vasopressin induced a maximal contractile response of 6.5 +/- 0.1 mN in vessels with and 6.3 +/- 0.3 mN in vessels without endothelium. The preincubation with l-NNA resulted in an enhanced response to AVP of 12.6 +/- 0.8 mN (P < 0.05). The augmentation of the AVP induced contractile response by NOS inhibition, which was not seen in preparations after the removal of the endothelium, suggests an endothelium dependent factor that is co-released with NO. The unknown nature of this endothelium dependent contractile factor was not influenced by indomethacin 100 microM (cyclooxygenase inhibitor), meclofenamic acid 20 microM (cyclooxygenase and lipoxygenase inhibitor), or bosentan 100 microM (endothelin antagonist). Charybdotoxin 0.1 microM (inhibitor of Ca2+ -activated K+ channels) specifically increased the contractile force in preparations with and without endothelium, or in the presence of l-NNA to 11.2 +/- 0.4 mN, 14.0 +/- 0.8 mN, and 19.0 +/- 0.8 mN, respectively (P < 0.05 compared with the experiments without charybdotoxin). SR 49059 (vasopressin 1 receptor (V1) antagonist) antagonized the effects of AVP, whereas SR 121463 B (V2 antagonist) was ineffective. In contrast to the results obtained with AVP, desmopressin (V2 agonist) showed no effect. CONCLUSION: The completely V1 dependent AVP-induced contraction is partly inhibited by the stimulated release of NO. This was only demonstrable in endothelium intact vessels in the presence of l-NNA and not after removal of the endothelium. This strongly suggests the involvement of an unknown endothelium V1 receptor dependent contractile factor that is not influenced by inhibition of the prostaglandin, lipoxygenase, or endothelin pathway, or by blockade of the V2 receptor.
Assuntos
Arginina Vasopressina/farmacologia , Endotélio Vascular/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Nitroarginina/farmacologia , Vasoconstrição/efeitos dos fármacos , Animais , Arginina Vasopressina/antagonistas & inibidores , Arginina Vasopressina/fisiologia , Endotélio Vascular/fisiologia , Óxido Nítrico Sintase/antagonistas & inibidores , Coelhos , Artéria Renal/efeitos dos fármacosRESUMO
Angiotensin II (Ang II) enhances sympathetic neurotransmission via AT(1)-receptors located on sympathetic nerve terminals. We recently demonstrated that inhibition of Ang II-mediated facilitation in the pithed rat by irbesartan resulted in a U-shaped dose response curve, which was not observed when PD 123319, at a concentration that selectively blocks the AT(2)-receptor, was co-administered. Hence, the irbesartan-mediated upstroke might be explained by the involvement of the AT(2)-receptor after AT(1) blockade with high-dose irbesartan. In the present study, we further investigated the possible role of the AT(2)-receptor in Ang II-mediated facilitation in vitro. We studied the effect of the AT(2)-receptor antagonist PD 123319 (10 nM) on Ang II-enhanced sympathetic outflow evoked by electrical field stimulation (EFS) in the rat isolated inferior vena cava. Additionally, we investigated the effect of the AT(1)-receptor blocker irbesartan (0.1 nM 1 M) on the sequelae of Ang II-enhanced, EFS-evoked sympathetic nerve traffic in the presence or absence of PD 123319 (10 nM). PD 123319 did not influence Ang II-enhanced sympathetic outflow. Irbesartan dose-dependently attenuate Ang II-augmented transmitter release (pIC50 7.99+0.03), whereas no U-shaped concentration-response relationship for irbesartan was observed. Co-administration of PD 123319 with irbesartan proved unable to influence Ang II-mediated facilitation differently compared with irbesartan alone. The experimental observations indicate that the AT(2)-receptor is not involved in Ang II-mediated enhancement of sympathetic nerve traffic in the present in vitro study.
Assuntos
Angiotensina II/farmacologia , Receptores de Angiotensina/metabolismo , Sistema Nervoso Simpático/fisiologia , Transmissão Sináptica/fisiologia , Vasoconstritores/farmacologia , Antagonistas de Receptores de Angiotensina , Animais , Anti-Hipertensivos/farmacologia , Compostos de Bifenilo/farmacologia , Sinergismo Farmacológico , Estimulação Elétrica , Imidazóis/farmacologia , Técnicas In Vitro , Irbesartana , Masculino , Piridinas/farmacologia , Ratos , Ratos Wistar , Receptor Tipo 2 de Angiotensina , Sistema Nervoso Simpático/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Tetrazóis/farmacologia , Trítio , Veia Cava Inferior/inervaçãoRESUMO
The aim of the present study was to determine the inhibitory potency of two selective angiotensin AT(1) receptor antagonists, eprosartan and candesartan, at the level of the sympathetic nerve terminal and the vascular smooth muscle. Male New Zealand White rabbits, weighing 2100-2550 g, were used. To study eprosartan and candesartan at the neuronal angiotensin AT(1) receptor, we investigated their influence on the angiotensin II-enhanced, electrical field stimulation-evoked sympathetic transmission in the rabbit isolated thoracic aorta in a noradrenaline spillover model. To study both antagonists at the vascular angiotensin AT(1) receptor, concentration-response curves for angiotensin II were constructed in the presence or absence of the two angiotensin AT(1) receptor antagonists. Angiotensin II (10 nM) caused a significant increase by 107+/-11.1% of the stimulation-evoked sympathetic outflow, which was concentration-dependently inhibited by both eprosartan (pIC(50) 7.91+/-0.12) and candesartan (pIC(50) 10.76+/-0.13). Angiotensin II (1 nM-0.3 microM) caused a concentration-dependent increase in contractile force (E(max) 20.62+/-2.24 mN, pD(2) 8.16+/-0.04). Both eprosartan (pA(2) 8.90+/-0.11, pIC(50) 8.87+/-0.12 (10 nM angiotensin II)) and candesartan (pD(2)' 10.80+/-0.13) counteracted the contractions evoked by cumulative concentrations of angiotensin II. Candesartan proved a more potent antagonist than eprosartan at both the pre- and postjunctional angiotensin AT(1) receptor. For eprosartan, vascular inhibitory concentrations were 10-fold lower than sympatho-inhibitory concentrations, whereas for candesartan, inhibitory concentrations at both sites were similar. The results may be explained by differences between the pre- and postjunctional angiotensin AT(1) receptor subtype.
Assuntos
Acrilatos/farmacologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II , Benzimidazóis/farmacologia , Imidazóis/farmacologia , Terminações Pré-Sinápticas/efeitos dos fármacos , Tetrazóis/farmacologia , Tiofenos , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/fisiologia , Compostos de Bifenilo , Relação Dose-Resposta a Droga , Técnicas In Vitro , Masculino , Terminações Pré-Sinápticas/fisiologia , Coelhos , Receptor Tipo 1 de Angiotensina/fisiologia , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologiaRESUMO
Reactive oxygen species (ROS) play a role in cardiovascular diseases such as heart failure and hypertension. Furthermore, increasing evidence has accumulated suggesting that ROS can also be formed subsequent to the stimulation of various receptors, thus functioning as second messengers. The objective of the present study was to elucidate the role of intracellular-generated ROS in the inotropic and chronotropic effects of the alpha1- and beta-adrenoceptor and the ET-receptor stimulation in isolated rat atria. In addition, we investigated whether the MAPKerk pathway is involved in the ROS-provoked rise of contractile force. For this purpose hydrogen peroxide was applied, which is known to serve several endogenous functions as a second messenger. Moreover, hydrogen peroxide readily crosses cell membranes, which thus allows to mimic the intracellular formation. Preincubation of atria with EUK 8 (400 microM), a cell permeable superoxide dismutase- and catalase-mimetic, reduced the positive inotropic effect upon alpha1-adrenoceptor and ET-receptor stimulation. The responsiveness to beta-adrenoceptor stimulation remained unaffected by this pretreatment. The chronotropic effects were not altered by preincubation with EUK 8. In contrast to the MAPK(p38) inhibitor SB203580 (2 and 10 microM), the two MKKmek inhibitors PD98059 (30 and 100 microM) and U0126 (10 microM) significantly attenuated the positive inotropic response to hydrogen peroxide in isolated rat left atria. In addition, inhibition of the Na+/H+ exchange (NHE) by cariporide (1 microM) counteracted ROS-provoked increase of contractile force. From the present study we conclude that the inotropic responses to alpha1-adrenoceptor and ET-receptor stimulation are, at least partially, caused by intracellular-formed ROS, that subsequently may activate the MAPKerk pathway and the NHE.
Assuntos
Frequência Cardíaca/fisiologia , Contração Miocárdica/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Espécies Reativas de Oxigênio/farmacologia , Receptores Adrenérgicos/metabolismo , Receptores de Endotelina/metabolismo , Animais , Cardiotônicos/farmacologia , Átrios do Coração/efeitos dos fármacos , Átrios do Coração/metabolismo , Frequência Cardíaca/efeitos dos fármacos , Masculino , Contração Miocárdica/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
1. Reactive oxygen species (ROS) are known to be involved in the progression of various cardiovascular diseases. One source of ROS is activated neutrophils, which can release superoxide anion radicals and hydrogen peroxide by membrane-bound NAD(P)H oxidases. These ROS not only destroy bacteria, but may also affect mammalian tissue. In addition, hydrogen peroxide serves as a substrate for myeloperoxidase, an enzyme that is released by activated neutrophils during inflammatory processes, as seen, for instance, in reperfusion injury and atherosclerosis. Myeloperoxidase catalyses the oxidation of chloride by hydrogen peroxide, yielding hypochlorite, an extremely potent oxidant. 2. The purpose of the present study was to evaluate the effects of hypochlorite on a variety of receptor-dependent processes in rat isolated left atria and rat thoracic aorta and to compare these results with the phenomena observed after incubation with hydrogen peroxide. 3. In the presence of hypochlorite (300 micro mol/L), the positive inotropic response of alpha1-adrenoceptor stimulation by methoxamine (300 micro mol/L) was converted into a negative inotropic response. In contrast, the positive inotropic effects of the beta1/beta2-adrenoceptor agonist isoprenaline (3 micro mol/L) and endothelin (ET)-1 (100 nmol/L) remained largely unaffected. 4. The inversion of alpha1-adrenoceptor-mediated inotropy was not obtained in the presence of hydrogen peroxide (500 micro mol/L). Hydrogen peroxide did not affect the positive inotropic response of isoprenaline, but it completely abolished the inotropic effect of ET-1. 5. The effect of cardiac M2-receptor stimulation was studied in the presence of hypochlorite and hydrogen peroxide. The negative inotropic response to acetylcholine (ACh) was significantly enhanced after hypochlorite incubation compared with control. 6. In the rat thoracic aorta, endothelial function, evaluated by means of ACh-induced vasodilation, was completely abolished in the presence of hypochlorite (100 micro mol/L), but remained unaffected by treatment with the same concentration of hydrogen peroxide. 7. From these data, we conclude that hypochlorite exerts more toxic properties than its precursor hydrogen peroxide, leading to substantial physiological alterations in cardiac and vascular tissue.
Assuntos
Endotélio Vascular/efeitos dos fármacos , Peróxido de Hidrogênio/farmacologia , Ácido Hipocloroso/farmacologia , Oxidantes/farmacologia , Receptor Muscarínico M2/efeitos dos fármacos , Receptores Adrenérgicos alfa 1/efeitos dos fármacos , Acetilcolina/farmacologia , Agonistas alfa-Adrenérgicos/farmacologia , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/fisiologia , Agonistas Colinérgicos/farmacologia , Depressão Química , Endotélio Vascular/fisiologia , Átrios do Coração/efeitos dos fármacos , Peróxido de Hidrogênio/metabolismo , Técnicas In Vitro , Masculino , Metoxamina/farmacologia , Contração Muscular/efeitos dos fármacos , Contração Muscular/fisiologia , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/fisiologia , Ratos , Ratos Wistar , Receptor Muscarínico M2/fisiologia , Receptores Adrenérgicos alfa 1/fisiologiaRESUMO
Angiotensin II (Ang II) can enhance sympathetic neurotransmission by acting on (AT1) receptors that are located on sympathetic nerve terminals. We investigated presynaptic blockade by the selective AT1-receptor antagonist irbesartan in pithed spontaneously hypertensive rats and normotensive Wistar-Kyoto rats (WKY). We compared the presynaptic inhibitory dose with that required for the blockade of AT1-receptors on vascular smooth muscle in both strains. To investigate blockade of presynaptic AT1-receptors, we studied the effect of irbesartan on the sequelae of electric stimulation of the thoraco-lumbar sympathetic outflow (0.25-8 Hz). To study the interaction between postsynaptic AT1-blockers and alpha-adrenoceptors, the effects of irbesartan on pressor responses to exogenous noradrenaline (NA) were established. Additionally, we studied the effect of irbesartan on dose-response curves for the vasoconstriction induced by exogenous Ang II. Pressor responses to electrical stimulation of thoracolumbar sympathetic neurones, to exogenous Ang II, as well as to (NA) were enhanced in spontaneously hypertensive rats (SHR) compared with WKY. The stimulation-induced rise in DBP could be dose-dependently reduced by irbesartan (0.3-10 mg/kg) in both SHR and WKY. The pIC50 values (doses which suppress the rise in DBP by 50% compared with control) were 5.60 +/- 0.09 and 5.72 +/- 0.08 for SHR and WKY, respectively (P > 0.05). In SHR, no effect of irbesartan (3 mg/kg) on pressor responses to exogenous NA was observed. In contrast, in WKY, irbesartan (3 mg/kg) caused a rightward shift of the dose-response curve to exogenous NA. Irbesartan (0.3-3 mg/kg) caused a depression of E(max) values and a rightward shift of the dose-response curves to exogenous Ang II in a similar fashion in both SHR and WKY. From these results we conclude that both in SHR and in WKY, Ang II exerts a facilitatory effect on sympathetic neurotransmission, which is mediated by prejunctional AT1-receptors in both strains. Irbesartan displays comparable sympatho-inhibitory potency in the normotensive and hypertensive pithed rat preparations. A facilitatory effect via postsynaptically located AT1-receptors on alpha-adrenoceptor-mediated responses exists in WKY, but not in SHR. In both strains the required dose to inhibit presynaptic effects is somewhat higher than the dose required to inhibit postsynaptic effects. No differences, therefore, seem to exist between the two strains regarding the affinity of irbesartan for pre- and postjunctional AT1-receptors, respectively.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/farmacologia , Compostos de Bifenilo/farmacologia , Hipertensão/tratamento farmacológico , Sistema Nervoso Simpático/efeitos dos fármacos , Simpatolíticos/farmacologia , Tetrazóis/farmacologia , Agonistas alfa-Adrenérgicos/administração & dosagem , Agonistas alfa-Adrenérgicos/farmacologia , Angiotensina II/metabolismo , Angiotensina II/farmacologia , Antagonistas de Receptores de Angiotensina , Inibidores da Enzima Conversora de Angiotensina/administração & dosagem , Animais , Compostos de Bifenilo/administração & dosagem , Pressão Sanguínea/efeitos dos fármacos , Estado de Descerebração , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Estimulação Elétrica , Frequência Cardíaca/efeitos dos fármacos , Hipertensão/fisiopatologia , Irbesartana , Masculino , Norepinefrina/administração & dosagem , Norepinefrina/farmacologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Receptor Tipo 1 de Angiotensina , Especificidade da Espécie , Sistema Nervoso Simpático/fisiopatologia , Tetrazóis/administração & dosagemRESUMO
Both in human and in experimental heart failure (HF), the renin-angiotensin system and the sympathetic nervous system are activated. In a previous study a facilitatory action of angiotensin II (Ang II) was shown in the rabbit mesenteric artery, which was mediated via prejunctionally located Ang II type 1 (AT ) receptors. Very little is known about the effects of Ang II on sympathetic neurotransmission at the peripheral level in congestive heart failure (CFH). Accordingly, in the isolated mesenteric arteries obtained from rabbits with experimentally induced CHF, as well as in age-matched control rabbits, the effect of Ang II on contractions provoked by electrical field stimulation was investigated in the presence and absence of the AT receptor antagonist eprosartan. Additionally, to investigate a possible postjunctional facilitation, the effects of Ang II on alpha-adrenoceptor-mediated responses were studied using noradrenaline (NA). Lastly, the vasoconstrictor effects of Ang II were compared between HF rabbits and controls, by constructing concentration-response curves to Ang II. In control rabbits, Ang II 0.5 n caused an enhancement of stimulation-induced responses by a factor 3.2 +/- 0.5, 2.4 +/- 0.3, and 1.5 +/- 0.08, at 1, 2, and 4 Hz, respectively ( < 0.05 at all frequencies compared with vehicle). In rabbits with HF, the enhancement by Ang II (0.5 n ) amounted to a factor 2.1 +/- 0.2, 1.7 +/- 0.1, and 1.2 +/- 0.04, at 1, 2, and 4 Hz, respectively ( < 0.05 compared with vehicle at all frequencies). Accordingly, the enhancing effect of Ang II was more pronounced in the control group compared with rabbits with HF ( < 0.05 at each frequency). Eprosartan (1 nM -0.1 microM) could inhibit the facilitatory effects of Ang II in arteries from HF as well as from control rabbits. Contractile responses to exogenous NA (3 n -0.1 m ) were the same in HF rabbits and controls, and they were unaltered in the presence of Ang II 0.5 n Ang II (0.1 nM -1 microM) caused a concentration-dependent increase in contractile force, which was the same in HF rabbits and controls. From these findings it can be concluded that in rabbits with CHF as well as in control animals, Ang II facilitates the stimulation-induced vasoconstrictor responses via prejunctionally located AT receptors. The facilitating effect was decreased in vessels obtained from rabbits with CHF, whereas responses to exogenous Ang II were unchanged. These findings may be explained by downregulation or uncoupling of the prejunctional AT receptor.
Assuntos
Angiotensina II/farmacologia , Insuficiência Cardíaca/fisiopatologia , Artérias Mesentéricas/efeitos dos fármacos , Norepinefrina/farmacologia , Transmissão Sináptica/efeitos dos fármacos , Antagonistas de Receptores de Angiotensina , Animais , Doença Crônica , Estimulação Elétrica/métodos , Técnicas In Vitro , Masculino , Artérias Mesentéricas/fisiologia , Inibição Neural/efeitos dos fármacos , Inibição Neural/fisiologia , Coelhos , Receptor Tipo 1 de Angiotensina , Receptores de Angiotensina/agonistas , Receptores de Angiotensina/fisiologia , Transmissão Sináptica/fisiologia , Vasoconstrição/efeitos dos fármacos , Vasoconstrição/fisiologiaRESUMO
T-type calcium channels may be involved in the maintenance of myogenic tone. We tested their role in isolated rat cremaster arterioles obtained after CO(2) anesthesia and decapitation. Total RNA was analyzed by RT-PCR and Southern blotting for calcium channel expression. We observed expression of voltage-operated calcium (Ca(V)) channels Ca(V)3.1 (T-type), Ca(V)3.2 (T-type), and Ca(V)1.2 (L-type) in cremaster arterioles (n = 3 rats). Amplification products were observed only in the presence of reverse transcriptase and cDNA. Concentration-response curves of the relatively specific L-type blocker verapamil and the relatively specific T-type blockers mibefradil and nickel were made on cannulated vessels with either myogenic tone (75 mmHg) or a similar level of constriction induced by 30 mM K(+) at 35 mmHg. Mibefradil and nickel were, respectively, 162-fold and 300-fold more potent in inhibiting myogenic tone compared with K(+)-induced constriction [log(IC(50), M): mibefradil, basal -7.3 +/- 0.2 (n = 9) and K(+) -5.1 +/- 0.1 (n = 5); nickel, basal -4.1 +/- 0.2 (n = 5) and K(+) -1.6 +/- 0.5 (n = 5); means +/- SE]. Verapamil had a 17-fold more potent effect [log(IC(50), M): basal -6.6 +/- 0.1 (n = 5); K(+) -5.4 +/- 0.3 (n = 4); all log(IC(50)) P < 0.05, basal vs. K(+)]. These data suggest that T-type calcium channels are expressed and involved in maintenance of myogenic tone in rat cremaster muscle arterioles.
Assuntos
Arteríolas/fisiologia , Canais de Cálcio Tipo T/metabolismo , Tono Muscular/fisiologia , Músculo Esquelético/irrigação sanguínea , Resistência Vascular/fisiologia , Animais , Arteríolas/efeitos dos fármacos , Arteríolas/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio Tipo L/efeitos dos fármacos , Canais de Cálcio Tipo L/genética , Canais de Cálcio Tipo L/metabolismo , Canais de Cálcio Tipo T/efeitos dos fármacos , Canais de Cálcio Tipo T/genética , Técnicas In Vitro , Masculino , Tono Muscular/efeitos dos fármacos , Potássio/farmacologia , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Grau de Desobstrução Vascular/efeitos dos fármacosRESUMO
OBJECTIVE: To evaluate the facilitating effect of angiotensin II on sympathetic neurotransmission to quantitatively compare the sympatho-inhibitory potencies of the selective AT1 -receptor antagonists losartan, irbesartan and telmisartan in the isolated rabbit thoracic aorta. DESIGN: To investigate the influence of pharmacological compounds on pre-junctional sympathetic transmission, the quantification of sympathetic transmitter release is the most straightforward approach. METHODS: To investigate the sympatholytic properties of AT1 -blockers, we studied their effects on the enhancement by angiotensin II of electrical field stimulation (EFS)-evoked (2 Hz) sympathetic transmission in a modified spillover model. RESULTS: Angiotensin II (0.01 nmol/l-0.1 micromol/l) caused a concentration-dependent enhancement of EFS-evoked noradrenaline release (control versus concentrations 0.1 nmol/l-0.1 micromol/l, P<0.05). The maximal augmentation, by almost 100%, was observed at a concentration of 1 nmol/l (FR2/FR1, 2.03 +/- 0.11 versus control, 0.99 +/- 0.03). Higher concentrations (up to 0.1 micromol/l) produced less than maximal facilitation. The AT1 -receptor antagonists losartan (0.1 nmol/l-0.1 micromol/l), telmisartan (0.01-10 nmol/l) and irbesartan (0.1 nmol/l-0.1 micromol/l) concentration dependently attenuated the angiotensin II-mediated (1 nmol/l) enhancement of EFS-evoked sympathetic outflow. The concentrations that reduced the enhancement by 50% (IC50 values, expressed as -log mol/l +/- SEM) were 9.05 +/- 0.16 losartan, 10.28 +/- 0.20 telmisartan and 9.20 +/- 0.23 irbesartan. Accordingly, the order of potency with respect to sympatho-inhibition proved telmisartan> irbesartan = losartan (where > signifies P<0.05). CONCLUSIONS: The facilitating effect of angiotensin II on the sequelae of neuronal stimulation appears to be mediated by pre-synaptically located AT1 -receptors. Facilitation can be concentration dependently attenuated by AT1 -blockade. The order of potency with respect to sympatho-inhibition is telmisartan irbesartan = losartan. These differences may be explained by differences in affinity for the pre-synaptic AT1 -receptor.
Assuntos
Antagonistas de Receptores de Angiotensina , Aorta Torácica/efeitos dos fármacos , Simpatolíticos/farmacologia , Angiotensina II/farmacologia , Animais , Aorta Torácica/inervação , Benzimidazóis/farmacologia , Benzoatos/farmacologia , Compostos de Bifenilo/farmacologia , Relação Dose-Resposta a Droga , Estimulação Elétrica , Técnicas In Vitro , Irbesartana , Losartan/farmacologia , Norepinefrina/metabolismo , Concentração Osmolar , Coelhos , Receptor Tipo 1 de Angiotensina , Sistema Nervoso Simpático/fisiologia , Transmissão Sináptica/efeitos dos fármacos , Telmisartan , Tetrazóis/farmacologiaRESUMO
It is well known that angiotensin II (Ang II) can facilitate the effects of sympathetic neurotransmission. In the present study, using various experimental models, we investigated the inhibitory effects of several Ang II subtype 1 receptor (AT1) antagonists on this Ang II-induced facilitation. We compared the sympatho-inhibitory potencies of the AT1 blockers with their respective potencies regarding inhibition of the direct vasoconstrictor effects of Ang II. In the isolated mesenteric artery, we investigated the effects of Ang II in the presence and absence of losartan, irbesartan and telmisartan on stimulation-induced vasoconstrictor responses. In the pithed rat, we studied the effect of AT1 blockade on the sequelae of electrical stimulation of the thoracolumbar sympathetic outflow (presynaptic AT1 blockade) as well as on dose-response curves elicited by exogenous Ang II (postsynaptic AT1 blockade). Additionally, we compared the sympatho-inhibitory of irbesartan in spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats. In the isolated mesenteric artery, Ang II (10 nM) significantly enhanced stimulation-induced vasoconstrictor responses. The enhancement could be antagonized in a concentration-dependent manner by losartan (1 nmol/l to 1 mumol/l), irbesartan (0.1 nmol/l to 0.1 mumol/l) and telmisartan (0.01 nmol/l to 0.01 mumol/l). The sympatho-inhibitory potency was telmisartan > irbesartan > losartan. In the pithed normotensive rat, the stimulation-induced increase in diastolic blood pressure (DBP) as well as the Ang II-elicited DBP response were dose-dependently reduced by all the AT1 receptor blockers investigated. The order of potency with respect to sympatho-inhibition was eprosartan > valsartan = candesartan = embusartan = telmisartan > losartan > irbesartan (comparison of doses which at 2 Hz reduced delta DBP by 20 mmHg, differences significant at P < 0.05). The order of potency regarding inhibition of the Ang II-induced DBP increase was candesartan > embusartan = valsartan = eprosartan = telmisartan > irbesartan > losartan (comparison of the antagonist concentration, in the presence of which twice the agonist concentration, in the presence of which twice the agonist concentration is needed to cause the same effect [pA2 values], differences significant at P < 0.05). In the pithed SHR and the normotensive WKY rat the sympatho-inhibitory potency of irbesartan did not differ significantly between both strains. It can be concluded that all AT1 receptor antagonists appear to possess sympatho-inhibitory properties, which may be of potential interest in the treatment of hypertension and heart failure. Our findings suggest differences in pre- and postsynaptic inhibition between the various compounds, since for eprosartan and losartan the sympatholytic doses and postsynaptic inhibitory doses differed far less than for the other AT1 receptor antagonists.
Assuntos
Antagonistas de Receptores de Angiotensina , Anti-Hipertensivos/farmacologia , Simpatolíticos/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Relação Dose-Resposta a Droga , Estimulação Elétrica , Frequência Cardíaca/efeitos dos fármacos , Masculino , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/fisiologia , Ratos , Ratos Endogâmicos SHR , Ratos Endogâmicos WKY , Ratos Wistar , Receptor Tipo 1 de AngiotensinaRESUMO
The activation of mitogen-activated protein kinase (MAPK) pathways in the heart, for instance by alpha(1)-adrenoceptor agonists and endothelin-1, has primarily been associated with cellular growth regulation. Here we have investigated a possible role of MAPK pathways in the inotropic and chronotropic effects of adrenoceptor agonists and endothelin-1 in isolated rat left and right atria. Inotropic and chronotropic responses of the isolated atria to methoxamine, isoprenaline and endothelin-1 were measured in the absence and presence of inhibitors of MAPK pathways. The MAPK kinase (MKK(mek)) inhibitors PD98059 (100 microM) and U0126 (10 microM) significantly inhibited the inotropic responses to the alpha(1)-adrenoceptor agonist methoxamine (300 microM) and endothelin-1 (50 nM), but not the chronotropic responses to these agonists. U0126 but not PD98059 inhibited the inotropic response to 3 microM isoprenaline. None of the aforementioned inotropic and chronotropic effects were inhibited by the MAPKP(p38) inhibitor SB203580 (2 microM). We conclude that activation of the PD98059/U0126-sensitive MAPK pathway is essential for the inotropic but not chronotropic actions of adrenoceptor agonists and endothelin-1.