Effects of Arterial Stiffness on Cerebral WM Integrity in Older Adults: A Neurite Orientation Dispersion and Density Imaging and Magnetization Transfer Saturation Imaging Study.

BACKGROUND AND PURPOSE: Arterial stiffness is reported to be able to cause axonal demyelination or degeneration. The present study aimed to use advanced MR imaging techniques to examine the effect of arterial stiffness on the WM microstructure among older adults. MATERIALS AND METHODS: Arterial stiffness was measured using the cardio-ankle vascular elasticity index (CAVI). The high-CAVI (mean CAVI ≥ 9 points) and the low-CAVI groups (mean CAVI < 9 points) were created. The neuronal fiber integrity of the WM was evaluated by neurite orientation dispersion and density imaging and magnetization transfer saturation imaging. Tract-Based Spatial Statistics and the tracts-of-interest analysis were performed. Specific WM regions (corpus callosum, internal capsule, anterior thalamic radiation, corona radiata, superior longitudinal fasciculus, forceps minor, and inferior fronto-occipital fasciculus) were selected in the tracts-of-interest analysis. RESULTS: In Tract-Based Spatial Statistics, the high-CAVI group showed a significantly lower myelin volume fraction value in the broad WM and significantly higher radial diffusivity and isotropic volume fraction values in the corpus callosum, forceps minor, inferior fronto-occipital fasciculus, internal capsule, corona radiata, and anterior thalamic radiation than the low-CAVI group. In tracts-of-interest analysis using multivariate linear regression, significant associations were found between the mean CAVI and radial diffusivity in the anterior thalamic radiation and the corona radiata; isotropic volume fraction in the anterior thalamic radiation and the corona radiata; and myelin volume fraction in the superior longitudinal fasciculus (P < .05). Additionally, partial correlation coefficients were observed for the significant associations of executive function with radial diffusivity and myelin volume fraction (P < .05). CONCLUSIONS: Arterial stiffness could be associated with demyelination rather than axonal degeneration.

[Recurrent right atrium thrombus in long term follow-up after repair of atrial septal defect; report of a case].

A 76-year-old woman with recurrent ball-like thrombus in right atrium after primary repair of atrial septal defect (ASD) and tricuspid annuloplasty was successfully treated by surgical resection and strict management of anticoagulation and antiarrhythmics. A routine follow-up echocardiography, 27 months after initial operation, showed a swinging ball mass looks like a myxoma in the right atrium. Intra-operative findings showed the mass attached the free wall of right atrium with a 5 mm stalk, which was far from the ASD patch, initial suture lines, and the tricuspid annulus. Histological examination revealed the round and smooth mass was thrombus. She was successfully discharged 13 days after the 2nd operation without any complaint. A postoperative laboratory check demonstrated normal coagulability. Despite the patient was prescribed warfarin potassium and aspirin, the follow-up echocardiography at 3 months showed a recurrent thrombus in the right atrium. However the strict anticoagulation therapy with warfarin potassium and aspirin induced thrombolysis and prevent any embolic event, 1 month later. It is important to continue a strict anticoagulant therapy and prevent arrhythmia to avoid recurrence thrombus.

Effect of YM087, a potent nonpeptide vasopressin antagonist, on vasopressin-induced protein synthesis in neonatal rat cardiomyocyte.

OBJECTIVE: Hypertrophy of cardiomyocytes may play an important role in the pathogenesis of cardiac hypertrophy associated with various cardiovascular diseases such as congestive heart failure. The aim of this study was to investigate whether vasopressin (AVP) induces protein synthesis in cultured neonatal rat cardiomyocytes through its specific receptor and whether YM087, a newly synthesized nonpeptide AVP receptor antagonist, inhibits AVP-induced protein synthesis in vitro. METHODS: AVP receptors on cardiomyocytes were characterized using the radioligand [3H] AVP. The effects of AVP and YM087 on intracellular free calcium concentration ([Ca2+]i), mitogen-activated protein (MAP) kinase and [3H]-leucine incorporation were investigated in cultured neonatal rat cardiomyocytes. RESULTS: In cardiomyocytes, Scatchard analysis showed a single population of high-affinity binding sites with the expected AVP V1A receptor subtype profile. YM087 showed high affinity for cardiomyocyte V1A receptors with a Ki value of 0.63 nM. In these same cells, YM087 potently inhibited AVP-induced increases in [CA2+]I and activation of MAP kinase in a concentration-dependent manner. In addition, AVP concentration-dependently stimulated the synthesis of protein without changing the rate of DNA synthesis, and YM087 prevented AVP-induced protein synthesis in a concentration-dependent manner. CONCLUSIONS: These results suggest that AVP directly causes protein synthesis and YM087 is a potent inhibitor of AVP-induced protein synthesis of cardiomyocytes and thus may have beneficial effects in the development and regression of cardiomyocytic hypertrophy.

A novel anionic conductance affects action potential duration in isolated rat ventricular myocytes.

Effects of extracellular anions were studied in electrophysiological experiments on freshly isolated rat ventricular myocytes. Under current-clamp, action potential duration (APD) was prolonged by reducing the extracellular Cl(-) concentration and shortened by replacement of extracellular Cl(-) with I(-). Under voltage-clamp, membrane potential steps or ramps evoked an anionic background current (I(AB)) carried by either Cl(-), Br(-), I(-) or NO(3)(-). Activation of I(AB) was Ca(2+)- and cyclic AMP-independent, and was unaffected by cell shrinkage. I(AB) was insensitive to stilbene and fenamate anion transport blockers at concentrations that inhibit Ca(2+)-, cyclic AMP- and swelling-activated Cl(-) currents in ventricular cells of other mammals. These results suggest that I(AB) may be carried by a novel class of Cl(-) channel. Correlation of anion substitution experiments on membrane current and action potentials revealed that I(AB) could play a major role in controlling rat ventricular APD. These findings have important implications for those studying cardiac Cl(-) channels as potential targets for novel antiarrythmic agents.

Pharmacologic characterization of the oxytocin receptor in human uterine smooth muscle cells.

[(3)H]-oxytocin was used to characterize the oxytocin receptor found in human uterine smooth muscle cells (USMC). Specific binding of [(3)H]-oxytocin to USMC plasma membranes was dependent upon time, temperature and membrane protein concentration. Scatchard plot analysis of equilibrium binding data revealed the existence of a single class of high-affinity binding sites with an apparent equilibrium dissociation constant (K(d)) of 0.76 nM and a maximum receptor density (B(max)) of 153 fmol mg(-1) protein. The Hill coefficient (n(H)) did not differ significantly from unity, suggesting binding to homogenous, non-interacting receptor populations. Competitive inhibition of [(3)H]-oxytocin binding showed that oxytocin and vasopressin (AVP) receptor agonists and antagonists displaced [(3)H]-oxytocin in a concentration-dependent manner. The order of potencies for peptide agonists and antagonists was: oxytocin>[Asu(1,6)]-oxytocin>AVP= atosiban>d(CH(2))(5)Tyr(Me)AVP>[Thr(4),Gly(7)]-oxytocin>dDAVP, and for nonpeptide antagonists was: L-371257>YM087>SR 49059>OPC-21268>SR 121463A>OPC-31260. Oxytocin significantly induced concentration-dependent increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) and hyperplasia in USMC. The oxytocin receptor antagonists, atosiban and L-371257, potently and concentration-dependently inhibited oxytocin-induced [Ca(2+)](i) increase and hyperplasia. In contrast, the V(1A) receptor selective antagonist, SR 49059, and the V(2) receptor selective antagonist, SR 121463A, did not potently inhibit oxytocin-induced [Ca(2+)](i) increase and hyperplasia. The potency order of antagonists in inhibiting oxytocin-induced [Ca(2+)](i) increase and hyperplasia was similar to that observed in radioligand binding assays. In conclusion, these data provide evidence that the high-affinity [(3)H]-oxytocin binding site found in human USMC is a functional oxytocin receptor coupled to [Ca(2+)](i) increase and cell growth. Thus human USMC may prove to be a valuable tool in further investigation of the physiologic and pathophysiologic roles of oxytocin in the uterus. British Journal of Pharmacology (2000) 129, 131 - 139

Pharmacological characterization of the human vasopressin receptor subtypes stably expressed in Chinese hamster ovary cells.

Three subtypes of human (h) arginine vasopressin (AVP) receptors, hV1A, hV1B and hV2, were stably expressed in Chinese hamster ovary (CHO) cells and characterized by [3H]-AVP binding studies. In addition, the coupling of the expressed receptor protein to a variety of signal transduction pathways was investigated. Scatchard analysis of saturation isotherms for the specific binding of [3H]-AVP to membranes, prepared from CHO cells transfected with hV1A, hV1B and hV2 receptors, yielded an apparent equilibrium dissociation constant (Kd) of 0.39, 0.25 and 1.21 nM and a maximum receptor density (Bmax) of 1580 fmol mg(-1) protein, 5230 fmol mg(-1) protein and 7020 fmol mg(-1) protein, respectively. Hill coefficients did not differ significantly from unity, suggesting binding to homogenous, non-interacting receptor populations. Pharmacological characterization of the transfected human AVP receptors was undertaken by measuring the relative ability of nonpeptide AVP receptor antagonists, YM087, OPC-21268, OPC-31260, SR 49059 and SR 121463A, to inhibit binding of [3H]-AVP. At hV1A receptors, the relative order of potency was SR49059>YM087>OPC-31260>SR 121463A> >OPC-21268 and at hV2 receptors, YM087=SR 121463A>OPC-31260>SR 49059> >OPC-21268. In contrast, the relative order of potency, at hV1B receptors, was SR 49059> >SR 121463A=YM087=OPC-31260=OPC-21268. In CHO cells expressing either hV1A or hV1B receptors, AVP caused a concentration-dependent increase in intracellular Ca2+ concentration ([Ca2+]i) with an EC50 value of 1.13 nM and 0.90 nM, respectively. In contrast, stimulation of CHO cells expressing hV2 receptors resulted in an accumulation of cyclic AMP with an EC50 value of 2.22 nM. The potency order of antagonists in inhibiting AVP-induced [Ca2+]i or cyclic AMP response was similar to that observed in radioligand binding assays. In conclusion, we have characterized the pharmacology of human cloned V1A, V1B and V2 receptors and used these to determine the affinity, selectivity and potency of nonpeptide AVP receptor antagonists. Thus they may prove to be a valuable tool in further examination of the physiological and pathophysiological roles of AVP.

Effects of YM471, a nonpeptide AVP V(1A) and V(2) receptor antagonist, on human AVP receptor subtypes expressed in CHO cells and oxytocin receptors in human uterine smooth muscle cells.

YM471, (Z)-4'-[4,4-difluoro-5-[2-(4-dimethylaminopiperidino)-2-oxoethylidene]-2,3,4,5-tetrahydro-1H-1-benzoazepine-1-carbonyl]-2-phenylbenzanilide monohydrochloride, is a newly synthesized potent vasopressin (AVP) receptor antagonist. Its effects on binding to and signal transduction by cloned human AVP receptors (V(1A), V(1B) and V(2)) stably expressed in Chinese hamster ovary (CHO) cells, and oxytocin receptors in human uterine smooth muscle cells (USMC) were studied. YM471 potently inhibited specific [(3)H]-AVP binding to V(1A) and V(2) receptors with K(i) values of 0.62 nM and 1.19 nM, respectively. In contrast, YM471 exhibited much lower affinity for V(1B) and oxytocin receptors with K(i) values of 16.4 microM and 31.6 nM, respectively. In CHO cells expressing V(1A) receptors, YM471 potently inhibited AVP-induced intracellular Ca(2+) concentration ([Ca(2+)](i)) increase, exhibiting an IC(50) value of 0.56 nM. However, in human USMC expressing oxytocin receptors, YM471 exhibited much lower potency in inhibiting oxytocin-induced [Ca(2+)](i) increase (IC(50)=193 nM), and did not affect AVP-induced [Ca(2+)](i) increase in CHO cells expressing V(1B) receptors. Furthermore, in CHO cells expressing V(2) receptors, YM471 potently inhibited the production of cyclic AMP stimulated by AVP with an IC(50) value of 1.88 nM. In all assays, YM471 showed no agonistic activity. These results demonstrate that YM471 is a potent, nonpeptide human V(1A) and V(2) receptor antagonist which will be a valuable tool in defining the physiologic and pharmacologic actions of AVP.

Binding characteristics of YM087, an AVP receptor antagonist, in rhesus monkey liver and kidney membranes.

The binding characteristics of YM087, a nonpeptide vasopressin (AVP) V1A and V2 receptor antagonist, were studied using 3H-AVP binding to rhesus monkey liver and kidney membrane preparations. Both membrane preparations exhibited one class of high-affinity binding sites. However each membrane's receptors were different, with Kd values of 0.57 and 1.11 nM, Bmax values of 59.6 and 147 fmol/mg protein for liver and kidney, respectively. AVP receptor agonist or antagonist binding inhibition studies confirmed that these receptors belong to the V1A (liver) and V2 (kidney) subtypes. YM087 showed high affinity for both liver V1A and kidney V2 receptors with Ki values of 26.3 and 9.89 nM, respectively. These results show that YM087 is a potent, nonpeptide dual AVP V1A and V2 receptor antagonist, and would be a powerful tool for understanding the physiologic roles of AVP.

Characterization of rodent liver and kidney AVP receptors: pharmacologic evidence for species differences.

Radioligand binding studies with [3H]vasopressin (AVP) were used to determine the affinities of AVP receptor agonists and antagonists for mouse liver and kidney plasma membrane preparations. Both membrane preparations exhibited one class of high-affinity binding site. AVP ligand binding inhibition studies confirmed that mouse liver binding sites belong to the V1A subtype while kidney binding sites belong to the V2 receptor subtype. The affinity of each ligand for mouse V1A receptors was very similar to that for rat V1A receptors, showing differences in Ki values of less than 3-fold. In contrast, several peptide (d(CH2)5Tyr(Me)AVP) and nonpeptide (OPC-21268 and SR 49059) ligands had different affinities for mouse and rat kidney V2 receptors, with differences in Ki values ranging from 14- to 17-fold. These results indicate that mouse and rat kidney V2 receptors show significant pharmacologic differences.

Characterization of vasopressin receptor in rat lung.

This study characterized rat lung membrane arginine vasopressin (AVP) receptors in detail. Specific binding of [3H]AVP to rat lung membranes was dependent upon time, temperature and membrane protein concentration. Scatchard plot analysis of equilibrium binding data revealed the existence of a single class of high-affinity binding sites with a Kd of 0.45 nM and a Bmax of 76.6 fmol/mg protein. Competitive inhibition of [3H]AVP binding showed that neurohypophysial hormones as well as their synthetic analogues displaced [3H]AVP in a concentration-dependent manner. The order of potencies for the native peptides was: AVP > lysine vasopressin = arginine vasotocin > oxytocin. Furthermore, potent V1A receptor antagonists, d(CH2)5Tyr(Me)AVP and dPTyr(Me)AVP, showed high affinity for lung membranes. In contrast, the V2 receptor agonist, dDAVP, and the specific oxytocin receptor agonist, [Thr4,Gly7]oxytocin, did not affect AVP binding. These results suggest that the lung contains the V1A receptor subtype. The lung membrane AVP receptor characterized in this study may play an important role in mediating the physiological effects of AVP in the lung.

Presence of presynaptic inhibitory alpha 1-adrenoceptors in the cardiac sympathetic nerves of the dog: effects of prazosin and yohimbine on sympathetic neurotransmission to the heart.

The effects of prazosin and yohimbine on sympathetic neurotransmission to the heart were investigated in perfused dog hearts in situ in an attempt to determine whether alpha 1-adrenoceptors are located presynaptically in the cardiac sympathetic nerves. Intra-arterial injections of prazosin (1-30 micrograms) and yohimbine (0.3-10 micrograms) into the right coronary artery during cardiac sympathetic nerve stimulation further increased the tachycardia resulting from the stimulation. Continuous infusions of methoxamine (20-40 micrograms/min) and of clonidine (2-4 micrograms/min) into the right coronary artery during cardiac sympathetic nerve stimulation caused sustained reduction of the tachycardia. Prazosin under methoxamine infusion enhanced the tachycardia to a greater extent than in the absence of methoxamine. Prazosin under clonidine infusion enhanced the tachycardia to the same extent as it did in the absence of clonidine. These results suggest that prazosin antagonizes the effect of methoxamine but does not antagonize that of clonidine. The results obtained with yohimbine were in contrast to the effects of prazosin, showing the antagonism of clonidine by yohimbine. Prazosin and yohimbine both had little effect on the heart rate during either the resting state or the infusion of norepinephrine. These results suggest that the prazosin- and yohimbine-induced enhancement of the tachycardia resulting from cardiac sympathetic nerve stimulation is due to a presynaptic effect. However, the presynaptic effect of prazosin appears to differ from that of yohimbine. The presence of presynaptic alpha 1-adrenoceptors regulating norepinephrine release, as well as of alpha 2-adrenoceptors, is suggested in the cardiac sympathetic nerves of the dog.

Cardiovascular effects of YM099, a novel K+ channel opener, in anesthetized and conscious dogs.

Cardiovascular effects of a newly synthesized benzoxadiazol derivative K+ channel opener, YM099, 2-(7,8-dihydro-6,6-dimethyl-6H-[1,4]oxazino[2,3- f][2,1,3]benzoxadiazol-8-yl) pyridine N-oxide, were evaluated in dogs. In pentobarbital-anesthetized dogs, YM099 (1-10 micrograms/kg i.v.), similarly to levcromakalim (1-10 micrograms/kg i.v.), dose dependently increased coronary artery blood flow, max.d p/dt and cardiac output, and decreased total peripheral resistance and mean blood pressure, with a small increase in heart rate. These vasodilator effects were antagonized by glibenclamide (3 mg/kg i.v.). Interestingly, YM099 selectively increased coronary artery blood flow, although it increased carotid, coronary, mesenteric and renal artery blood flows and cardiac output. In addition, YM099 (1-10 micrograms/kg i.v.) increased large conductive coronary artery vessel diameter as well as coronary artery blood flow. In conscious dogs, YM099 (100 micrograms/kg i.v.) also increased the diameter of large conductive and small resistive coronary arteries. In conclusion, YM099 is a potent vasodilator agent, with particularly pronounced effects on the coronary artery. These effects of YM099 may be mediated by the opening of ATP-sensitive K+ channels.

Pharmacological profile of YM087, a novel nonpeptide dual vasopressin V1A and V2 receptor antagonist, in dogs.

The pharmacological profile of YM087 (4'-[(2-methyl-1,4,5,6-tetrahydroimidazo[4,5-d][1]benzazepin -6-yl) carbonyl]-2-phenylbenzanilide monohydrochloride) was investigated in dogs. YM087 showed high affinity for vasopressin V1A and V2 receptors in radioligand receptor binding studies with dog platelets (V1A) and kidney (V2). Intravenously injected YM087 (3-100 micrograms/kg) dose dependently inhibited the pressor response to exogenous vasopressin in anesthetized dogs. Intravenous (10-100 micrograms/kg) and oral (30-300 micrograms/kg) administration of YM087 dose dependently increased urine flow with little effect on urinary sodium and potassium excretion in normally hydrated conscious dogs. Concomitantly, the urine osmolality dropped below the plasma osmolality (300 mOsm/kg H2O). In contrast, intravenously injected furosemide (300 micrograms/kg) increased urine flow with marked increases in urinary sodium and potassium excretion. These results indicate that YM087 is the first orally effective dual vasopressin V1A and V2 receptor antagonist and that it will be a new tool in the investigation of the physiological and pathophysiological role of vasopressin in the cardiovascular system and kidney. YM087 may be useful for the treatment of patients with congestive heart failure, renal diseases and water-retaining diseases.

Antihypertensive activity of a nonpeptide angiotensin II receptor antagonist, YM358, in rats and dogs.

The antihypertensive activity of YM358, 2,7-diethyl-5-[[2'-(1 H-tetrazol-5-yl)biphenyl-4-yl]methyl]-5H-pyrazolo[1,5-b][1,2,4]tri azole potassium salt monohydrate, a new nonpeptide angiotensin II receptor antagonist, was characterized in rats and dogs. In conscious rats, YM358 after a single oral administration (1-30 mg/kg) lowered blood pressure. The rank order of hypotensive potency of YM358 in conscious rats was 2-kidney, 1-clip renal hypertensive rats > spontaneously hypertensive rats > normotensive rats on the basis of maximum hypotension. YM358 also caused decreases in blood pressure in 2-kidney, 1-clip renal hypertensive dogs and furosemide-treated dogs. Repeated administration of YM358 to 2-kidney, 1-clip renal hypertensive rats for 28 days produced a stable and long-lasting antihypertensive effect without influencing circadian blood pressure and heart rate rhythms. No reflex tachycardia was observed in any animals of either species treated with YM358. Therefore, the pharmacological profile of this compound indicates that YM358 has potential as a useful antihypertensive agent.

Pharmacological profile of YM358, a novel nonpeptide angiotensin AT1 receptor antagonist.

The pharmacological profile of YM358, 2,7-diethyl-5-[[2'-(1 H-tetrazol-5-yl)biphenyl-4-yl]methyl]-5H-pyrazolo[1,5-b][1,2,4]tri azole potassium salt monohydrate, a novel non-peptide angiotensin AT1 receptor antagonist, was studied in vitro and in vivo. YM358 competed with [125I][Sar1, Ile8]angiotensin II for angiotensin AT1 receptors in rat liver membranes. YM358 displayed competitive kinetics and the pKi value was calculated as 8.79. In contrast, YM358 had little effect on the binding of [125I][Sar1, Ile8]angiotensin II to the angiotensin AT2 receptor in bovine cerebellum. In isolated rabbit aorta, YM358 produced a parallel rightward shift in the concentration-response curve for angiotensin II with a pA2 value of 8.82. YM358 had no effect on the contraction induced by KCl, norepinephrine, serotonin, histamine, prostaglandin F2alpha or endothelin-1 even at 10(-5) M. On the basis of pKi values in the binding assay and pA2 values in the isolated tissues, YM358 was approximately 3-10 times more potent than losartan in antagonizing angiotensin AT1 receptors. In pithed rats, intravenous administration of YM358 inhibited an increase in mean blood pressure induced by intravenous infusion of angiotensin II in a dose-dependent manner. In conscious normotensive rats, YM358 at 3-30 mg/kg p.o. inhibited the angiotensin II-induced pressor response in a dose-dependent manner. YM358 at 30 mg/kg caused maximum and complete inhibition 30 min after dosing, and inhibition lasted more than 24 h. These results demonstrate that YM358 is a potent, AT1-selective and competitive nonpeptide angiotensin receptor antagonist. Moreover, YM358 is both orally active and long-lasting. This pharmacological profile suggests that YM358 would be suitable for the treatment of cardiovascular disorders such as hypertension and chronic heart failure.

Comparison of vasopressin binding sites in human uterine and vascular smooth muscle cells.

Several studies indicate that oxytocin and vasopressin receptors in the human uterus are heterogeneous. We have investigated whether oxytocin and vasopressin bind to separate receptors or one class of receptors in human uterine smooth muscle cells. [3H]d(CH2)5Tyr(Me)AVP, the vasopressin V1A receptor selective radioligand, was used for comparison of vasopressin binding sites in human uterine and vascular smooth muscle cell membranes. Both membrane preparations exhibited one class of high-affinity binding sites with Kd values of 6.44 and 0.47 nM, Bmax values of 166 and 34.8 fmol/mg protein for uterine and vascular smooth muscle cells, respectively. In vascular preparations, the selective vasopressin V1A receptor antagonist, SR 49059 ((2S) 1-[(2R 3S)-(5-chloro-3-(2-chlorophenyl)- -(3.4-dimethoxybenzenesulfonyl)-3-hydroxy-2,3-dihydro-1H-indole-2- carbonyl]-pyrrolidine-2-carboxamide), showed high affinity with Ki value of 0.98 nM, confirming that these receptors belong to the vasopressin V1A receptor subtype. On the contrary, in uterine preparations, binding of [3H]d(CH2)5Tyr(Me)AVP was more effectively displaced by oxytocin and the oxytocin receptor selective antagonist, L-371257, (1-[1-[4-[ N-Acetyl-4-piperidinyl)oxy]2-methoxybenzoyl]piperidin-4-yl]- 4H-3,1-benzoxazin-2(1H)-one), than vasopressin and SR 49059, suggesting that binding may be due to cross-reaction with the oxytocin receptors. These results suggest that human uterine smooth muscle cells express only a high density of oxytocin receptors.

Vasopressin increases vascular endothelial growth factor secretion from human vascular smooth muscle cells.

Vascular endothelial growth factor (VEGF) is a potent and specific mitogen of vascular endothelial cells which promotes neovascularization in vitro. To determine whether vasopressin induces VEGF secretion in human vascular smooth muscle cells, we performed enzyme-linked immunosorbent assays. Vasopressin potently induced a time-dependent and concentration-dependent (maximal, 10(-7) M) increase in VEGF secretion by human vascular smooth muscle cells that was maximal after 24 h. Furthermore, vasopressin also concentration-dependently caused mitogenic effect, as reflected by total protein content of cells per culture well. These vasopressin-induced VEGF secretion increase and mitogenic effect of these cells were potently inhibited by vasopressin V1A receptor antagonists, confirming this is a vasopressin V1A receptor-mediated event. These results indicate that vasopressin increases VEGF secretion in human vascular smooth muscle cells, the magnitude of VEGF secretion being temporally related to the mitogenic effect of vascular smooth muscle cells and the potency of the growth-promoting stimulus. Vasopressin-induced VEGF secretion by proliferating vascular smooth muscle cells could act as a paracrine hormone to powerfully influence the permeability and growth of the overlying vascular endothelium, vasopressin play a more fundamental role in the regulation of vascular function than has previously been recognized.

Cardiovascular and renal effects of conivaptan hydrochloride (YM087), a vasopressin V1A and V2 receptor antagonist, in dogs with pacing-induced congestive heart failure.

The systemic hemodynamic and renal responses to conivaptan hydrochloride (YM087; 4'-(2-methyl-1,4,5,6-tetrahydroimidazo[4,5-d][1]benzoazepine -6-carbonyl)-2-phenylbenzanilide monohydrochloride), a vasopressin V1A and V2 receptor antagonist, were determined in pentobarbital-anesthetized dogs after 2 to 3 weeks of rapid right ventricular pacing. Congestive heart failure, characterized by decreases in first derivative of left ventricular pressure (left ventricular d P/dt(max)) and cardiac output, and increases in left ventricular end-diastolic pressure and total peripheral vascular resistance, was induced by chronic rapid right ventricular pacing at 260-280 beats/min. Intravenous administration of conivaptan (0.1 mg/kg) significantly increased left ventricular dP/dt(max) and cardiac output and significantly decreased left ventricular end-diastolic pressure and total peripheral vascular resistance. Conivaptan also increased urine flow and reduced urine osmolality by markedly increasing free water clearance. These results indicate that conivaptan produced hemodynamic improvement and marked aquaresis in dogs with congestive heart failure. Therefore, conivaptan may find clinical use in treating patients with congestive heart failure.

Renal effect of YM435, a new dopamine D1 receptor agonist, in anesthetized dogs.

The renal effects of YM435 ((-)-(S)-4-(3,4-dihydroxyphenyl)-7,8-dihydroxy -1,2,3,4-tetrahydroisoquinoline hydrochloride hydrate), a dopamine D1 receptor agonist, were investigated in anesthetized dogs. Intravenous infusion of YM435 (0.1-3 micrograms/kg per min) increased renal blood flow and decreased mean blood pressure in a dose-dependent manner with little effect on heart rate. Glomerular filtration rate, urine flow and urinary sodium excretion were concomitantly increased. The renal effect of YM435 by intravenous infusion at 0.3 microgram/kg per min was completely blocked by treatment with the selective dopamine D1 receptor antagonist SCH 23390 (7-chloro-8-hydroxy-3-methyl-1-phenyl-2,3,4,5-tetrahydro-(1H)-3-benzazep ine hydrochloride). Furthermore, intravenous infusion of YM435 (0.3 microgram/kg per min) reversed the angiotensin II-induced decreases in renal blood flow, glomerular filtration rate, urine flow and urinary sodium excretion, and prevented the decrease in renal blood flow, glomerular filtration rate and urine flow induced by renal nerve stimulation and platelet-activating factor (PAF). These results suggest that intravenous administration of YM435 produces renal vasodilating and diuretic/natriuretic effects by stimulation of dopamine D1 receptors, and demonstrate that YM435 can inhibit angiotensin II-, renal nerve stimulation- and PAF-induced renal dysfunction.

The role of extracellular Ca2+ in carbachol-induced tonic contraction of the pig detrusor smooth muscle.

The role of extracellular Ca2+ in the tonic-contractile response to muscarinic receptor stimulation was investigated in isolated detrusor smooth muscle from the pig urinary bladder. Carbachol (10(-8)-10(-5) M) produced a concentration-dependent contractile response in isolated pig detrusor smooth muscle strips consisting of an initial phasic component followed by a tonic component. During the plateau of the tonic contractions induced by carbachol at the submaximal concentration of 10(-6) M, the inhibiting effects of atropine, EGTA, nifedipine (a voltage-dependent calcium channel antagonist), H-7 [a protein kinase C (PKC) inhibitor] and YM934 (a potassium channel opener) on the contractions were evaluated. Atropine (10(-10)-3 x 10(-8) M) concentration-dependently inhibited the tonic contractions induced by carbachol. In the same experimental conditions, EGTA (4 mM) and nifedipine (10(-9)-3 x 10(-7) M) depressed the tonic contractions in a concentration-dependent manner as did H-7 (10(-5)-3 x 10(-5) M) and YM934 (10(-8)-10(-6) M). However, H-7 (10(-5)-3 x 10(-5) M) and YM934 (10(-6) M) were very weak in inhibiting the contractions induced by KCl (50 mM) in isolated pig detrusor smooth muscle strips. These results suggest that the tonic-contractile response induced by carbachol in pig detrusor smooth muscle strips is dependent mainly on depolarization of the cell membranes and an influx of extracellular Ca2+, and also suggest that this depolarizing response may be due to inactivation of ATP-sensitive potassium channels through muscarinic activation of PKC.