AT(1)-receptor blockade and sympathetic neurotransmission in cardiovascular disease.

1. The present survey is dealing with the interactions between the renin-angiotensin-aldosterone system (RAAS) and the sympathetic nervous system (SNS) in various organs and tissues, with an emphasis on the angiotensin AT-receptors located at the sympathetic nerve endings. 2. Angiotensin II, the main effector of the RAAS is known to stimulate sympathetic nerve traffic and its sequelae in numerous organs and tissues, such as the central nervous system, the adrenal medulla, the sympathetic ganglia and the sympathetic nerve endings. These stimulatory effects are mediated by AT(1)-receptors and counteracted by AT(1)-receptor antagonists. 3. Sympatho-inhibition at the level of the sympathetic nerve ending appears to be a class effect of the AT(1)-receptor blockers, mediated by presynaptic AT(1)-receptors. With respect to the ratio pre-/postsynaptic AT(1)-receptor antagonism important quantitative differences between the various compounds were found. 4. Both the pre- and postjunctional receptors at the sympathetic nerve endings belong to the AT(1)-receptor population. However, the presynaptic receptors belong to the AT(1B)-subtype, whereas the postjunctional receptors probably belong to a different AT(1)-receptor subpopulation. 5. Sympatho-inhibition is a class effect of the AT(1)-receptor antagonists. In conditions in which the SNS plays a pathophysiological role, such as hypertension and congestive heart failure, this property may well be of therapeutic relevance.

The effects of hypochlorite-induced oxidative stress on presynaptic M2-receptors at sympathetic nerve endings in the rat tail artery.

1 It was shown recently that stimulation of cardiac muscarinic M2-receptors revealed an enhanced negative inotropic response in isolated rat left atria after exposure to hypochlorite-induced oxidative stress. This phenomenon was not observed after stimulation of the cardiac A1-receptor, which like the M2-receptor is coupled to Gi-proteins. Since even the contractile response to M3-receptor stimulation was not amplified in the rat portal vein, we hypothesized a M2-receptor specificity of this hypochlorite-induced enhancement. 2 The present study was performed in order to investigate whether the sympathoinhibitory response to presynaptically located M2-receptor stimulation would also be modified after exposure to hypochlorite in the rat tail artery. We applied electrical field stimulation (EFS) in order to mimic sympathetic neurotransmission. 3 EFS increased the vascular tone frequency-dependently (0.3-4 Hz). EFS-induced vasoconstriction could be attenuated by acetylcholine (30 nM-1 microM) in a concentration-dependent manner. Hypochlorite (10 and 100 microM) did not affect the sympathoinhibitory effect of acetylcholine (100 nM). 4 In conclusion, in contrast to cardiac M2-receptors, hypochlorite did not amplify the sympathoinhibitory effects of presynaptic M2-receptors. The different responsiveness between neuronal and cardiac M2-receptors to hypochlorite may be explained by the different G-protein subunits involved in the activation of the underlying signalling cascade.

Inhibition of facilitation of sympathetic neurotransmission and angiotensin II-induced pressor effects in the pithed rat: comparison between valsartan, candesartan, eprosartan and embusartan.

BACKGROUND: In the pithed rat model, endogenously generated angiotensin (Ang) II can enhance sympathetic neurotransmission by acting on Ang II type 1 (AT1) receptors that are located on sympathetic nerve terminals. OBJECTIVE: To compare the inhibitory potency of candesartan, valsartan, eprosartan and embusartan in blocking presynaptically and postsynaptically located AT1 receptors. DESIGN: To investigate blockade of presynaptic AT1 receptors, we studied the effect of AT1 receptor blockade on the sequelae of electrical stimulation of the thoracolumbar sympathetic outflow (0.25-8 Hz). To investigate the interaction between postsynaptic AT1 blockers and alpha-adrenoceptors, the effects of these compounds on pressor responses to exogenous noradrenaline were determined. To investigate blockade of postsynaptic AT1 receptors, we studied the effect of the AT1 antagonists on dose-response curves elicited by exogenous Ang II. RESULTS: The stimulation-induced increase in diastolic blood pressure (DBP) and the Ang II-elicited DBP response were dose-dependently reduced by all AT1 receptor blockers. Interestingly, the greatest doses of the AT1 antagonists caused less than maximal reduction in the stimulation-induced increase in DBP, resulting in a U-shaped dose-response relationship. To compare sympathoinhibitory potencies, the doses that, at 2 Hz, reduced the change in DBP by 20 mmHg (ED20 values, expressed as -log mol/kg) were calculated; they were 5.50 +/- 0.12, 5.77 +/- 0.10, 6.32 +/- 0.12 and 5.62 +/- 0.13 for valsartan, candesartan, eprosartan and embusartan, respectively. The order of potency, therefore, was eprosartan> valsartan = candesartan = embusartan (where > signifies P < 0.05). To compare the order of potency for inhibition of the Ang II-induced increase in DBP, we calculated pA2 values (the X intercept in Schild regression). They were 7.20 +/- 0.17, 8.01 +/- 0.01, 7.20 +/- 0.03 and 7.25 +/- 0.16, for valsartan, candesartan, eprosartan and embusartan, respectively. Accordingly, the order of potency for inhibition of the direct pressor effects of Ang II was candesartan> valsartan = eprosartan = embusartan (where > signifies P < 0.05). CONCLUSION: In the pithed rat, the effects on DBP of stimulation of the thoracolumbar spinal cord are partly dependent on endogenously formed Ang II. These effects can be counteracted by blockade of presynaptically located AT1 receptors. No interaction was found between postsynaptically located AT1 receptors and alpha-adrenoceptors. The order of potency of the agents tested for sympathoinhibition clearly differed from that for inhibition of the direct pressor effects of Ang II. These findings suggest considerable differences in affinity of the various AT1 blockers for pre- and postsynaptic AT1 receptors.

Effect of the AT1-receptor antagonists losartan, irbesartan, and telmisartan on angiotensin II-induced facilitation of sympathetic neurotransmission in the rat mesenteric artery.

SUMMARY: The effect of the AT1-receptor antagonists losartan, irbesartan, and telmisartan on angiotensin II (Ang II)-induced facilitation of noradrenergic neurotransmission was investigated in the isolated rat mesenteric artery under isometric conditions. Electrical field stimulation (2, 4, and 8 Hz) caused a frequency-dependent increase of contractile force. At stimulation frequencies of 2, 4, and 8 Hz, Ang 11 (10 nM) increased the stimulation-induced vasoconstrictor responses by a factor 4.8 +/- 0.9, 2.9 +/- 0.7, and 1.3 +/- 0.1, respectively (p < 0.05 compared with control for all frequencies). The enhancement could be concentration-dependently antagonized by losartan (1 nM-1 microM), irbesartan (0.1 nM-0.1 microM), and telmisartan (0.01 nM-0.01 microM). At a stimulation frequency of 2 Hz, the relation between stimulation-induced vasoconstrictor responses (in presence of Ang II 10 nM) and the concentration of the AT1-antagonists used could be described by linear regression. The order of potency concerning sympathoinhibition was telmisartan > irbesartan > losartan (p < 0.05 between linear regression lines). Contractile responses to exogenous noradrenaline were unaltered in the presence of Ang II 10 nM. We conclude that the facilitating effect of Ang II on noradrenergic neurotransmission is mediated by presynaptically located AT1-receptors. Conversely, this facilitating effect can be dose-dependently counteracted by blockade of these receptors. Sympathoinhibitory properties are likely to contribute to the therapeutic effect of AT1-blockers, in particular in conditions in which the sympathetic nervous system is activated, such as congestive heart failure and hypertension.

Inhibition of angiotensin II-induced facilitation of sympathetic neurotransmission in the pithed rat: a comparison between losartan, irbesartan, telmisartan, and captopril.

OBJECTIVE: Numerous studies have shown that angiotensin II enhances sympathetic nervous transmission. The objective of the present study was to quantify the inhibitory effect of the angiotensin II type 1 (AT1) receptor blockers losartan, irbesartan and telmisartan and the angiotensin converting enzyme (ACE) inhibitor captopril on sympathetic neurotransmission and to compare the potency of these agents both at the presynaptic and the postsynaptic levels. DESIGN: In the male, normotensive pithed rat model, we studied the effect of losartan (1, 3, 10 and 30 mg/kg), irbesartan (3, 10, 30 and 60 mg/kg), telmisartan (0.3, 1, 3 and 10 mg/kg) and captopril (1.5, 5, 15 and 45 mg/kg) on electrical stimulation of the thoraco-lumbar spinal cord. To investigate the interaction between postsynaptic AT1-receptors and alpha-adrenoceptors, the effects of these compounds on pressor responses to exogenous noradrenaline were studied. RESULTS: Stimulation of the thoracolumbar spinal cord caused a stimulation-frequency dependent rise in diastolic blood pressure (DBP) that could be dose-dependently reduced by both AT1 receptor blockade and ACE inhibition. Interestingly, the highest doses of the AT1 antagonists caused less than maximal reduction in the rise in DBP. This phenomenon was not observed after ACE inhibition by captopril. In experiments with exogenous noradrenaline, no effect of AT1 blockade or ACE inhibition on alpha-adrenoceptor-mediated blood pressure responses was seen. CONCLUSION: We conclude that, in the pithed rat model, the effects of stimulation of the thoraco-lumbar spinal cord on DBP are counteracted by blockade of presynaptically located AT1 receptors. The order of potency concerning sympatico-inhibition is telmisartan > losartan > irbesartan. Regarding the inhibition of angiotensin II-induced facilitation of sympathetic neurotransmission, marked differences were observed between selective AT1 blockade and ACE inhibition. The finding that all three AT1 blockers cause less than maximal inhibition in their highest doses, as opposed to captopril, suggests that this is a class effect of the AT1 antagonists.

Beta-adrenergic responses are significantly enhanced in rat carotid artery with intimal hyperplasia.

The objective of the present study was to investigate the influence of balloon injury and subsequent neointima formation in the rat carotid artery on the beta-adrenoceptor function. Rat left common carotid artery was subjected to balloon injury with an arterial embolectomy catheter; the contralateral artery was sham-operated. Immediately, and at 2, 8 and 16 weeks post-injury, both the injured and the sham-operated carotid arteries were isolated and mounted in an isometric wire-myograph set-up. Subsequently, concentration-response curves (CRCs) were constructed for the beta-adrenoceptor agonist isoprenaline after precontraction with the thromboxane A2 (TP)-receptor agonist U46619 (30 nM) of the injured and sham-operated artery preparations. To evaluate the involvement of the beta1- and the beta2-adrenoceptor subtypes, CRCs were constructed in the presence of CGP 20712A (0.1 nM, a beta1-adrenoceptor-selective antagonist) and ICI 118,551 (10 nM, a beta2-adrenoceptor-selective antagonist). L-NAME (100 microM) and indomethacine (10 microM) were used to evaluate the influence of nitric oxide (NO) or prostanoids, respectively. Immediately post-injury, isoprenaline-induced vasorelaxation was impaired in the injured carotid artery preparations: Emax=19.6 +/- 2.2% vs. 64.0 +/- 4.6%, injured vs. sham, n=8, P<0.05. However, from 2 weeks post-injury onwards, this response appeared enhanced in the injured preparations: Emax, 2 weeks= 86.4 +/- 2.2% vs. 49.7 +/- 5.7%, injured vs. sham, n=5, P<0.05. In addition, the sensitivity for isoprenaline was increased in these preparations: pD2, 2 weeks=7.48 +/- 0.08 vs. 6.88 +/- 0.10, injured vs. sham, n=5, P<0.05. The beta-adrenoceptor population in both types of preparations consisted mainly of the beta2-adrenoceptor subtype, although at 8 and 16 weeks post-injury, the beta1-adrenoceptor subtype appeared to be present as well in the injured artery preparations. Inhibition of NO synthesis led to significant decreases of beta-adrenoceptor-mediated vasorelaxation both in injured and in sham-operated artery preparations for all time points, except at 16 weeks. Cyclo-oxygenase inhibition had no influence on isoprenaline-induced vasorelaxation in injured and sham-operated preparations. From this, it is concluded that beta-adrenoceptor-mediated vasorelaxation in rat carotid artery is partially NO-dependent and occurs mainly via activation of the beta2-adrenoceptor subtype. Balloon injury and subsequent neointima formation in the rat carotid artery lead initially to an impairment, but subsequently to an enhancement of the beta-adrenoceptor-mediated vasorelaxation. The impairment is attributable to the removal of endothelium, whereas the enhanced beta-adrenoceptor-mediated function may be related to the occurrence of an NO system in the neointimal smooth muscle cells.

Evidence for a sympatholytic effect of mibefradil in the pithed rat preparation.

OBJECTIVE: The T-type prevalent calcium channel blocker mibefradil (MIB) was shown to possess N-type calcium channel blocking properties. As this particular type of calcium channel is known to be crucially involved in the neuronal release of noradrenaline, we have investigated whether MIB could be a sympatholytic drug. METHODS: To evaluate the sympathoinhibitory action, the effects of 3 and 10 micromol/kg MIB on the tachycardic effect of electrical stimulation of the preganglionic cardioaccelerator nerves in the pithed rat were investigated. The effect of MIB on the dose-response curve of externally applied noradrenaline was also studied. To compare the results with a classic L-type calcium channel blocker, the experiments were repeated with 3 and 10 micromol/kg verapamil (VER). RESULTS: The maximal increase in heart rate in response to electrical nerve stimulation was 96 +/- 7 bpm (control, n = 6), 70 +/- 6 bpm (3 micromol/kg MIB, n = 8), 57 +/- 6 bpm (10 micromol/kg MIB, n = 5), 93 +/- 5 bpm (3 micromol/kg VER, n = 6) and 46 +/- 7 bpm (10 micromol/kg VER, n = 5). The tachycardic response to electrical stimulation at 1, 5 and 10 Hz was completely blocked by 5 mg/kg intravenous guanethidine. The maximal increase in heart rate in response to noradrenaline was 96 +/- 4 bpm (control, n = 6), 103 +/- 6 (3 micromol/kg MIB, n = 6), 42 +/- 9 bpm (10 micromol/kg MIB, n = 5), 73 +/- 5 bpm (3 micromol/kg VER, n = 5) and 40 +/- 7 bpm (10 micromol/kg VER, n = 6). Under control conditions and in the presence of 3 micromol/kg MIB and VER the maximal effect of noradrenaline was reached at 0.1 micromol/kg whereas in the presence of 10 micromol/kg MIB and VER it was reached at a dose of 1 micromol/kg. MIB at a dose of 3 micromol/kg was significantly more effective in reducing the chronotropic response to electrical stimulation compared with externally applied noradrenaline. For VER the opposite holds true. These differences were not observed with doses of 10 micromol/kg MIB and VER. CONCLUSION: Mibefradil, besides its direct effect on cardiac T- and L-type calcium channels, reduces the release of noradrenaline from sympathetic nerve endings, most probably by inhibition of presynaptic N-type calcium channels. In the model used this effect is only observable at relatively low concentrations, most probably because of the direct cardiodepressant action of MIB provoked by L-type channel blockade.

Reactive oxygen species-induced aortic vasoconstriction and deterioration of functional integrity.

Oxygen derived free radicals and other reactive oxygen species (ROS) are involved in a variety of disease states, which can have cardiac and vascular implications. The present study was performed to investigate the mechanism of ROS-induced vasoconstriction and the influence of ROS on the functional integrity of isolated rat thoracic aorta. ROS were generated by means of electrolysis (30 mA, during 0.5, 1, 2 or 3 min) of the organ bath fluid. ROS induced a transient (approximately 60 min) vasoconstriction and the maximally induced contraction was dependent on the duration of electrolysis. Dimethyl sulfoxide (DMSO) diminished the ROS-induced vasoconstriction almost completely, indicating a major influence of hydroxyl radicals on contractility. The dual cyclooxygenase/lipoxygenase inhibitor, meclofenamate, completely prevented the ROS-induced vasoconstriction. The phospholipase A2 (PLA2) inhibitor, oleyloxyethyl phosphorylcholine, was able to reduce the vasoconstriction elicited by ROS by approximately 70%. Conversely, the specific cytoplasmic PLA2 inhibitor arachidonyl trifluoromethylketone proved ineffective in this respect. By using the specific mitogen-activated protein kinase (MAPkinase) kinase inhibitor PD98059, it was shown that the activation of extracellular-regulated kinase (ERK) MAPkinase contributes to the ROS-induced vasoconstriction. The effects of ROS on the functional integrity of the aortae were investigated, in particular with respect to receptor (alpha1-adrenoceptor) and non-receptor-mediated contractile responses (high potassium solution). In addition, both the endothelium dependent (methacholine) and endothelium independent (sodium nitroprusside) vasorelaxation were investigated before and after ROS exposure. Electrolysis periods of 0.5 and 1 min induced a modest leftward shift of the concentration response curves for the alpha1-adrenoceptor agonist methoxamine. Longer electrolysis periods of 2 and 3 min additionally decreased the maximal response to (alpha1-adrenoceptor stimulation. Methacholine-induced vasorelaxation proved diminished in aortae subjected to electrolysis (0.5, 1, 2 and 3 min), whereas relaxation to sodium nitroprusside was nearly complete in all groups. KCl-induced contractions proved attenuated only after longer electrolysis periods of 2 and 3 min. This ROS-induced deterioration of functional integrity was almost completely prevented by 0.6% DMSO. From these results we may conclude that ROS induce an eicosanoid and ERK MAPkinase-mediated vasoconstriction in isolated rat thoracic aorta. In addition, exposure to ROS leads to a deterioration of functional integrity characterized by endothelial dysfunction and decreased contractile function.

The influence of chronic inhibition of nitric oxide synthesis on contractile and relaxant properties of rat carotid and mesenteric arteries.

Balloon denudation of the rat carotid artery leads to an immediate decrease in beta-adrenoceptor-medi-ated vasodilator response. However, this arterial function becomes significantly enhanced during subsequent formation of neointima with the endothelial cell lining still being absent. It was therefore hypothesized that chronic suppression of endothelium-dependent nitric oxide (NO) synthesis may eventually upregulate the beta-adrenoceptor system on vascular smooth muscle. To investigate this hypothesis, male Wistar rats were treated chronically with the L-arginine analogue NG-nitro-L-arginine methyl ester (L-NAME) to inhibit the synthesis of NO (i.e. 15 mg/kg per day or 0.06 mmol/kg per day for 6 weeks p.o.). Prior to the experiments the mean arterial blood pressure (MAP) was significantly elevated in the L-NAME-treated rats (i.e. 128.4+/-3.4 mmHg vs. 100.0+/-2.9 mmHg, L-NAME vs. control, n=4, P<0.05). The functional properties of the isolated vessel preparations were established by isometric force measurement in a myograph setup, in the absence of L-NAME. The maximal contractile responses to high potassium chloride solution (100 mM), to the alpha1-adrenoceptor agonist phenylephrine, and to the thromboxane A2-agonist U46619, were not influenced by chronic L-NAME-treatment in the carotid and mesenteric artery preparations. The vasodilator responses to the cholinergic agonist methacholine were significantly impaired in the carotid arteries of L-NAME-treated animals: 30.9+/-7.9% vs. 64.6+/-2.0%, P<0.05, L-NAME vs. control, n=10. However, these responses appeared not to be modulated in the mesenteric artery preparations after chronic L-NAME treatment. Separate experiments showed that these responses could be blocked both in the rat carotid and mesenteric artery with L-NAME (10 mM) in vitro. Addition of the NO synthase substrate L-arginine could partially but significantly reverse this blockade. Chronic inhibition of NO synthesis caused significant deterioration of beta-adrenoceptor-mediated vasodilator responses. For carotid arteries: Emax=36.1+/-9.4% vs. 65.9+/-6.0%, P<0.05, L-NAME vs. control, n=5; and pD2=6.7+/-0.2 and 7.4+/-0.1, respectively, P<0.05, n=5. For mesenteric arteries: pD2=7.7+/-0.0 and 8.0+/-0.0, respectively, P<0.05, n=5. From these data, it may be concluded that chronic L-NAME treatment results in a stable impairment of the endothelium-dependent NO system in the rat carotid but not mesenteric arteries. The stated hypothesis fails as the beta-adrenoceptor-induced vasorelaxation of carotid and mesenteric arteries became significantly impaired, rather than enhanced. Taken together, the beta-adrenoceptor function in the rat carotid artery is apparently more dependent on endothelial NO synthesis than that in the rat mesenteric artery.

Heterogeneity in morphological characteristics of coronary arteries and aortae in various models of hypertension.

OBJECTIVE: Hypertension is associated with important structural and functional changes in vascular smooth muscle. The question arises whether different vascular beds and different models of hypertension react similarly or in a heterogeneous manner to the hypertensive state. METHODS: We quantitatively investigated the morphology of the coronary arteries and thoracic aortae taken from 4, 30 and 52-week-old spontaneously hypersensitive rats (SHR) and Wistar Kyoto rats (WKY), respectively, and from hypertensive rats with aortic stenosis (ASR). RESULTS: In coronary arteries taken from the SHR the media area was slightly increased compared with those obtained from the age-matched WKY. Increasing age caused a significant increase in media area of the SHR vessels, but not in WKY tissues. No differences were found in the media area values of the coronary arteries obtained from the ASR compared with those obtained from SHAM (operated control rats) rats. The media area of the aortae taken from SHR was increased when compared with those from the age-matched WKY rats. In addition, the media area of the aortae from the surgically induced aortic stenosis rats (ASR) when compared with tissues from SHAM rats was significantly increased. CONCLUSIONS: Different animal models of hypertension appear to develop largely heterogeneous profiles of vascular hypertrophy, with different morphometric characteristics.

Vascular responsiveness in isolated perfused kidneys of diabetic hypertensive rats.

OBJECTIVE: The aim of this study was to investigate whether diabetes and hypertension cause additive effects in the responses to various vasoconstrictor and vasodilator agents, in isolated perfused kidneys obtained from streptozotocin (STZ)-diabetic Wistar-Kyoto (WKY) rats and from diabetic spontaneously hypertensive rats (SHR). METHODS: SHR and WKY rats were administered STZ 55 mg/kg by intravenous injection into a lateral tail vein at age 12 weeks. Eight weeks later the kidneys were isolated and perfused via the left renal artery with a physiological salt solution. Renal perfusion pressure was measured continuously. Concentration response curves were plotted for various vasoconstrictor and vasodilator agents. RESULTS: Both the diabetic and the hypertensive state were associated with an increased wet kidney weight. The contractile responses of the renal arterial system to phenylephrine (PhE), serotonin (5-HT) and angiotensin II (Ang II) in terms both of the maximal rise in perfusion pressure (mmHg) and of the sensitivity (log EC50) were the same in preparations from diabetic WKY rats and in those from normoglycaemic WKY rats. The maximal contractile responses both to PhE and to Ang II were enhanced in kidneys from SHR compared with those in kidneys from their normotensive controls, whereas simultaneously occurring diabetes impaired this sensitization. After precontraction with 3 x 10(-6) mol/l PhE both endothelium-dependent (methacholine) and endothelium-independent (sodium nitroprusside) vasodilator drugs caused the same vasodilator response in the preparations taken from the four groups of animals. CONCLUSION: In isolated perfused kidneys obtained from STZ-diabetic WKY rats and SHR, the isolated diabetic state did not influence the vasoconstriction caused by various agonists. However, the enhanced vascular reactivity in the hypertensive state was blunted by simultaneously occurring diabetes mellitus. Endothelium-dependent and -independent vasorelaxation in this model was not affected neither by the hypertensive nor by the diabetic state.

Influence of nifedipine on pressor responses induced by different alpha-adrenoceptor agonists and angiotensin II in pithed diabetic hypertensive rats.

OBJECTIVE: Both intracellular and extracellular sources of calcium are involved in the activation of contraction in vascular smooth muscle. In the diabetic or hypertensive state, or both, changes induced in calcium handling by various types of agonists may vary considerably. METHODS: We investigated in which manner L-type calcium-channel blockade with nifedipine influences the pressor effects of the alpha 1-adrenoceptor agonists cirazoline and ST 587, the alpha 2-adrenoceptor agonist UK 14.304 and angiotensin II, all exerting a differential influence on calcium homeostasis, in pithed spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats rendered diabetic by an injection of 55 mg/kg streptozotocin. RESULTS: In diabetic WKY rats and SHR, the maximal pressor response was impaired for all agonists. The hypertensive state enhanced the maximal pressor response to all agonists. No difference was found in the nifedipine-induced depression of the pressor response to cirazoline and angiotensin II in the four groups of rats. The maximal pressor responses to ST 587 and UK 14.304 were more effectively depressed by administration of 0.3 mg/kg nifedipine both in diabetic WKY rats and in diabetic SHR than they were in their non-diabetic controls. CONCLUSIONS: Hypertension was associated with enhanced pressor response, whereas the diabetic state counteracted this effect. The pressor responses in pithed diabetic and diabetic hypertensive rats were clearly more dependent on the nifedipine-sensitive calcium influx than were those in their non-diabetic controls.

Opposite influences of hypertension and diabetes mellitus on pressor responses induced by different alpha-adrenoceptor agonists and angiotensin II in pithed rats.

Hypertension is often present in the diabetic individual and it is known to aggravate the vascular complications associated with diabetes. The pressor responses to two alpha 1-adrenoceptor agonists (ST587 and cirazoline), two alpha 2-adrenoceptor agonists (azepexole dihydrochloride (B-HT933) and UK14.304) and angiotensin II were investigated in pithed spontaneously hypertensive rats (SHR) and in pithed normotensive Wistar Kyoto rats (WKY) made diabetic by a single i.v. injection (55 mg/kg) of streptozotocin (STZ). Two months after diabetes was induced, the effect of the agonists on basal diastolic blood pressure (DBP) was determined. In pithed diabetic WKY and SHR, the maximal pressor response was impaired for all agonists. The dose response curves were shifted to the right when compared with their non-diabetic controls. The hypertensive state enhanced the maximal pressor response to all agonists compared with non-hypertensive animals. Additional diabetes blunted this increase in the effects of ST587, B-HT933 and angiotensin II, but not in those of cirazoline and UK14.304. Hypertension caused a leftward shift of the dose response curve for ST587 when compared with the non-hypertensive state. However, this effect was not observed when diabetes was present as well. In conclusion, hypertension resulted in an enhanced pressor effect, possibly caused by vascular hypertrophy, whereas the diabetic state counteracted this effect.

Reduced muscarinic cholinoceptor density and sensitivity in various models of experimental cardiac hypertrophy.

1. In the present study we investigated functional and binding characteristics of muscarinic receptors in experimental cardiac hypertrophy. 2. As models of cardiac hypertrophy we used hearts of spontaneously hypertensive rats (SHR) and Wistar rats with surgically induced abdominal aorta stenosis (ASR). Wistar Kyoto rats (WKY) and sham operated Wistar rats were used as respective controls. 3. The hypertrophy was more pronounced in hearts of ASR compared to SHR, although the mean arterial pressure was found to be lower. 4. Isolated, perfused Langendorff heart preparations (paced with 5 Hz) from both groups of hypertrophied hearts were less sensitive to the muscarinic agonists oxotremorin and methacholine (P < 0.05, all n = 6) when compared with control organs. The maximal reduction in contractile force induced by methacholine was 59.3 +/- 4.5% in SHR and 41.6 +/- 3.4% in ASR hearts versus 26.4 +/- 4.1% and 25.0 +/- 2.6% in control organs, respectively. 5. The density (fmol/mg protein-1) of muscarinic receptors in membrane homogenates of hearts from SHR (127.6 +/- 11.5) was unchanged, whereas in hearts from ASR (221.0 +/- 8.9) it was found to be reduced (P < 0.05) when compared to the respective controls (142.5 +/- 14.7 and 308.8 +/- 16.1, respectively, all n = 6). 6. From the present data we conclude that cardiac hypertrophy results in a loss of sensitivity towards muscarinic receptor stimulation. A corresponding reduction of left ventricular receptor density could only be demonstrated in massively hypertrophied hearts of ASR.

Streptozotocin-induced diabetes mellitus in spontaneously hypertensive rats: a pathophysiological model for the combined effects of hypertension and diabetes.

The present study was undertaken to investigate the combined effects of hypertension and streptozotocin-induced diabetes mellitus in the rat. Accordingly, four groups of rats were studied: Wistar Kyoto rats (WKY), diabetic WKY, spontaneously hypertensive rats (SHR) and diabetic SHR, respectively. The mean arterial blood pressure was increased in hypertensive animals compared to normotensive animals. The base excess in the diabetic rats was higher than that of normoglycemic animals. An elevated glucose concentration was found in the blood and urine of streptozotocin-treated rats. Ketone bodies were detected in the urine and blood of the diabetic rats. Mortality rates after treatment were not different among the four groups. In separate experiments, isolated working hearts of the various groups were set up and analyzed. For the maximal left ventricular pressure (mm Hg) the following values were formed: 110.0 +/- 2.6, 93.6 +/- 2.7, 93.4 +/- 3.0, and 87.5 +/- 2.4, respectively. The wet heart weights, dry heart weights, and body weights of the diabetic rats were lower than those of normoglycemic animals. The wet heart weight/body weight ratio, however, was increased by diabetes and hypertension (0.43 +/- 0.01, 0.47 +/- 0.01, 0.47 +/- 0.01, and 0.54 +/- 0.02, respectively). There were no significant differences between the water content of the hearts from the four different groups. Pathologic examination of the hearts showed myocardial hypertrophy and medial hypertrophy of coronary arteries in diabetic and hypertensive animals. There was no difference in relative collagen content in the hearts of the four groups.(ABSTRACT TRUNCATED AT 250 WORDS)

In vitro effects of nifedipine, nisoldipine, and lacidipine on rat isolated coronary small arteries.

We investigated the effect of 1,4-dihydropyridine calcium antagonists (nifedipine, nisoldipine, and lacidipine) on serotonin (5-HT)- and KCl (120 mM)-induced contractions of rat isolated septal coronary artery. The preparations showed the well-known biphasic response to KCl depolarization. All three calcium antagonists were more potent in inhibiting the second, tonic phase compared to the first, transient phase, with pIC50 values of 7.17 +/- 0.12/8.27 +/- 0.07 (nifedipine), 7.93 +/- 0.21/8.96 +/- 0.06 (nisoldipine), and 8.28 +/- 0.07/9.79 +/- 0.04 (lacidipine) for suppressing the first and the second phase, respectively. Furthermore, the influence of the calcium antagonists on the 5-HT concentration-response curve was investigated. Nifedipine, nisoldipine, and lacidipine concentration dependently depressed the maximum effect of 5-HT on the coronary artery preparations without influencing the pEC50 of the 5-HT concentration-response curve. In conclusion, all three dihydropyridine calcium antagonists are potent and effective inhibitors of depolarization- or 5-HT-induced coronary artery contractions. Lacidipine, a new lipophilic compound, was the most potent one in inhibiting the depolarization-induced contraction, thereby showing a marked selectivity for the tonic phase compared to the initial phasic response.

Effects of manidipine and other calcium antagonists on rat renal arcuate arteries.

We investigated the effect of 1,4-dihydropyridine calcium antagonists (nifedipine, nisoldipine, and manidipine) on serotonin (5-hydroxytryptamine [5-HT])- and KCl (120 mmol/L)-induced contractions of rat isolated renal arcuate arteries. The preparation showed the well-known biphasic response to KCl-induced depolarization. All three calcium antagonists were more potent in inhibiting the second phase (tonic) compared with the first phase (transient) of the effect. The inhibitory concentration of 50% (pIC50) values were 6.92 +/- 0.24/8.51 +/- 0.08 (nifedipine, n = 32), 7.61 +/- 0.10/9.33 +/- 0.03 (nisoldipine, n = 28), and 7.61 +/- 0.13/9.07 +/- 0.06 (manidipine, n = 32) for the suppression of the first and second phases, respectively. In small coronary and renal arteries maximally activated with KCl solution, nifedipine and manidipine concentrations dependently inhibited the calcium concentration response curves. Manidipine was more potent than nifedipine (pIC50 coronary artery: 9.26 +/- 0.14 vs 7.93 +/- 0.22; pIC50 renal artery: 9.14 +/- 0.14 vs 7.77 +/- 0.21), but both compounds showed the same potency in the two different preparations. Furthermore, the influence of the calcium antagonists on the 5-HT concentration-response curve was investigated. Marked differences were found in the ability of the three compounds to inhibit the 5-HT-induced vasoconstriction of isolated renal arteries. For manidipine, a 93% +/- 3% reduction of the maximal 5-HT-induced contraction was observed, which was a significantly stronger inhibition compared with nifedipine (43% +/- 4.12%) or nisoldipine (26% +/- 0.37%). The pIC50 values were 7.96 +/- 0.15 (nifedipine), 8.17 +/- 0.14 (nisoldipine), and 7.84 +/- 0.12 (manidipine).(ABSTRACT TRUNCATED AT 250 WORDS)

Cardioprotection by nifedipine in isolated working hearts: a comparative study on three different types of experimental ischemia.

Cardiac ischemia can be provoked by different methods in animal models and in isolated organs. Accordingly, three different procedures were followed to find the most sensitive model for the analysis of the anti-ischemic activity of calcium antagonists. The experiments were performed in the isolated working heart preparation of the rat, paced at the frequency of 5 Hz and perfused with Tyrode solution at 37 degrees C. Global ischemia was achieved by closing off the supply of the perfusion medium and surrounding the heart with Tyrode solution of 37 degrees C gassed with N2; low-flow ischemia was achieved by reducing the cardiac afterload from 51.5 to 11.0 mm Hg; ligation of the left descending coronary artery was performed in order to provoke regional ischemia. Nifedipine was applied in a concentration (EC50) known to reduce the contractile force by one-half of its basal value. The following parameters were determined after 15 min of nifedipine pretreatment and at the end of the experiment: LVP (left ventricular pressure),+dP/dtmax (LVP's first derivative), AO (aortic output), CF (coronary flow), and CO (cardiac output). From the data obtained, the percentages of recovery were calculated. Nifedipine caused a significant improvement in the functional recovery of most of the parameters studied. This improvement, however, was much more pronounced in the model of the low-flow ischemia, which is obviously more sensitive to the anti-ischemic activity of calcium antagonists than the other experimental procedures studied. Low-flow ischemia appears to be preferable to other procedures for the screening of the potential anti-ischemic activity of calcium antagonists and other drugs.

Depressed inotropic response to beta-adrenoceptor agonists in the presence of advanced cardiac hypertrophy in hearts from rats with induced aortic stenosis and from spontaneously hypertensive rats.

OBJECTIVE: To study the inotropic response to beta-adrenoceptor stimulation in isolated hypertrophied hearts from hypertensive rats. DESIGN AND METHODS: Cardiac hypertrophy was induced in Wistar rats by stenosing the abdominal aorta. Functional responses to isoprenaline, dobutamine, terbutaline and salbutamol were measured in paced (5 Hz), aortically stenosed hearts (18-20 and 32-34 weeks of age) and compared with those of sham-operated spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats. RESULTS: Following aortic stenosis, which was accompanied by less hypertension than that sustained by SHR, the Wistar rat hearts showed more pronounced cardiac hypertrophy. An initially equal inotropic response to the beta-adrenoceptor agonists (18-20 weeks) was reduced to 45% at 32-34 weeks in SHR but not in WKY rat hearts. The response to beta-adrenoceptor stimulation in the hypertrophied Wistar rat hearts was reduced at 18-20 weeks to 30% and at 32-34 weeks to 10% of controls, respectively. The response by all hypertrophied hearts to forskolin and N,2'-O-dibutyryladenosine 3':5' monophosphate was also diminished. CONCLUSIONS: The impaired contractile response to beta-adrenoceptor agonism is more clearly related to cardiac hypertrophy than to hypertension.

Differential effects of alpha 1- and alpha 2-adrenoceptor agonists on peripheral vasoconstriction in pithed diabetic rats.

The pressor responses to selective alpha 1-adrenoceptor agonists (cirazoline and methoxamine) and to alpha 2-adrenoceptor agonists (UK-14,304 and B-HT 933) were investigated in pithed, streptozotocin-induced diabetic rats and age-matched control animals. Three months after induction of diabetes, the basal values of diastolic blood pressure (DBP) were the same in pithed diabetic and control rats (34.3 +/- 4.4 vs. 32.8 +/- 4.4 mm Hg, p greater than 0.05, n = 30). In pithed diabetic rats, dose-response curves for the vasoconstrictor effects of cirazoline and methoxamine were shifted to the right with a slight decrease in the maximum response as compared with those in control animals. In contrast, no shift in dose-response curves for the alpha 2-adrenoceptor-mediated vasoconstrictor effects was observed, as reflected by similar pD2 values. A pronounced decrease in the maximum pressor response to the alpha 2-adrenoceptor agonists could be demonstrated. This reduction was significantly greater than that of the alpha 1-adrenoceptor agonists. In conclusion, the functional role of alpha 1- and alpha 2-adrenoceptors in the peripheral resistance vessels appears to be differentially influenced by the diabetic state.