Increase In Developed Tension Research Articles

Effects of the thromboxane receptor antagonist S18886 in the porcine coronary circulation

Thromboxane A2 (TxA 2 ) is a potent coronary vasoconstrictor that has been implicated in promoting decreases in myocardial perfusion in a variety of (patho)-physiologic conditions. S18886 is a promising orally-active TxA 2 receptor antagonist currently approved for investigational clinical use. However, the coronary vascular effects of S18886 are unknown and its specificity and affinity for the thromboxane receptor in the coronary circulation remain unclear. We tested the hypothesis that administration of S18886 dose-dependently attenuates coronary vasoconstriction to the TxA 2 mimetic U46619 without influencing coronary responses to prostaglandin F 2α , acetylcholine, or smooth muscle depolarization (K + ). Experiments to test this hypothesis were performed in male (n = 5) and female (n = 6) domestic swine. Hearts were excised and the left circumflex coronary artery isolated, cleaned of periadventitial fat, and cut into 3 mm rings. Isometric tension of coronary artery rings was measured in response to log order increments of U46619 (1 nM to 1 μM) with and without S18886 (0.1-100 nM). Similar isometric studies were conducted with prostaglandin F 2α (10 nM-10 μM), acetylcholine (0.1-10 μM), and KCl (5-90 mM). U46619 induced concentration dependent increases in tension development of isolated coronary artery rings (average EC50 of 42 ± 19 nM). Incubation of coronary arteries with S18886 (1 nM) significantly attenuated coronary vasoconstriction to U46619 resulting in a rightward shift of the EC50 to 187 ± 38 nM (P < 0.02). Vehicle had no effect on U46619-induced contractions. Higher concentrations of S18886 (10nM and 100nM) dose-dependently reduced U46619-induced contractions by 77% ± 3 and 93% ± 6 respectively. S18886 (1 nM) antagonized coronary vasoconstriction of prostaglandin F2α (10 μM) by 68% ± 5 (P < 0.0001) but had no effect on either acetylcholine or KCl-induced contraction. Data from this investigation indicate that S18886 is an effective antagonist of U46619-induced vasoconstriction in the porcine coronary circulation. While S18886 does not influence coronary smooth muscle response to either acetylcholine or activation of L-type Ca 2+ channels, attenuation of prostaglandin F 2α suggests the antagonists specificity may extend beyond TxA 2 receptor signaling. NIH R01HL158723 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

New Thiazole Acetic Acid Derivatives: A Study to Screen Cardiovascular Activity Using Isolated Rat Hearts and Blood Vessels.

Cardiovascular diseases are one of the major causes of mortalities worldwide. In the present research, new synthetic derivatives of thiazole were studied using isolated hearts and blood vessels of rats. The heart and thoracic aorta were tested with six new synthesized thiazole acetic acid derivatives (SMVA-10, SMVA-35, SMVA-40, SMVA-41, SMVA-42 and SMVA-60), and the data obtained were statistically analyzed and compared. Isolated rat hearts were used to record the changes in developed tension and heart rate, while thoracic aortas were used to measure the contractile response, before and after treatments. Analysis of the results indicated a significant (p < 0.01) increase in developed tension with the addition of SMVA-35, SMVA-40, SMVA-41 and SMVA-42, which was augmented in the presence of adrenaline without affecting the heart rate. On the other hand, acetylcholine significantly decreased the developed tension, which was significantly reversed (p < 0.01) in the presence of compounds (SMVA-35 and SMVA-60). However, in the presence of SMVA-35 and SMVA-40, acetylcholine-induced bradycardia was significantly (p < 0.01) reduced. Furthermore, only SMVA-42 induced a dose-dependent contractile response in the isolated blood vessel, which was abolished in the presence of prazosin. Therefore, it can be concluded that some of the new synthesized thiazole derivatives exhibited promising results by raising the developed tension without changing the heart rate or blood vessel function, which could be helpful in failing heart conditions. However, more research is required to fully comprehend the function, mechanism and effectiveness of the compounds.

Concomitant excitation and tension development are required for myocellular gene expression and protein synthesis in rat skeletal muscle.

Loading-induced tension development is often assumed to constitute an independent cue to initiate muscle protein synthesis following resistance exercise. However, with traditional physiological models of resistance exercise, changes in loading-induced tension development also reflect changes in neural activation patterns, and direct evidence for a mechanosensitive mechanism is therefore limited. Here, we sought to examine the importance of excitation and tension development per se on initiation of signalling, gene transcription and protein synthesis in rat skeletal muscle. Isolated rat extensor digitorum longus muscles were allocated to the following interventions: (a) Excitation-induced eccentric contractions (ECC); (b) Passive stretching without excitation (PAS); (c) Excitation with inhibition of contractions (STIM+IMA ) and; (d) Excitation in combination with both inhibition of contractions and PAS (STIM+IMA +PAS). Assessment of transcriptional and translational signalling, gene transcription and acute muscle protein synthesis was compared in stimulated vs contra-lateral non-stimulated control muscle. Protein synthesis increased solely in muscles subjected to a combination of excitation and tension development (ECC and STIM+IMA +PAS). The same pattern was true for p38 mitogen-activated protein kinase signalling for gene transcription as well as for gene transcription of immediate early genes FOS and JUN. In contrast, mechanistic target of rapamycin Complex 1 signalling for translation initiation increased in all muscles subjected to increased tension development (ECC and STIM+IMA +PAS as well as PAS). The current study suggests that exercise-induced increases in protein synthesis as well as transcriptional signalling is dependent on the concomitant effect of excitation and tension development, whereas signalling for translation initiation is only dependent of tension development per se.

Importance of SERCA2a on early isolated diastolic dysfunction induced by supravalvular aortic stenosis in rats

Cardiac remodeling is defined as changes in shape and function of the heart in response to aggression (pressure overload). The sarcoplasmic reticulum calcium ATPase cardiac isoform 2a (SERCA2a) is a known factor that influences function. A wide spectrum of studies report a decrease in SERCA2a in heart failure, but none evaluate it's the role in early isolated diastolic dysfunction in supravalvular aortic stenosis (AoS). Our hypothesis was that SERCA2a participates in such dysfunction. Thirty-day-old male Wistar rats (60-80 g) were divided into AoS and Sham groups, which were submitted to surgery with or without aorta clipping, respectively. After 6 weeks, the animals were submitted to echocardiogram and functional analysis by isolated papillary muscle (IPM) in basal condition, hypoxia, and SERCA2a blockage with cyclopiazonic acid at calcium concentrations of 0.5, 1.5, and 2.5 mM. Western-blot analyses were used for SERCA2a and phospholamban detection. Data analysis was carried out with Student's t-test and ANOVA. AoS enhanced left atrium and E and A wave ratio, with preserved ejection fraction. Basal condition in IPM showed similar increases in developed tension (DT) and resting tension (RT) in AoS, and hypoxia was similar between groups. After cyclopiazonic acid blockage, final DT was equally decreased and RT was similar between groups, but the speed of relaxation was decreased in the AoS group. Western-blot was uniform in all evaluations. The hypothesis was confirmed, since functional parameters regarding SERCA2a were changed in the AoS group.

PKC activation increases Ca2+sensitivity of permeabilized lymphatic muscle via myosin light chain 20 phosphorylation-dependent and -independent mechanisms

The contractile activity of muscle cells lining the walls of collecting lymphatics is responsible for generating and regulating flow within the lymphatic system. Activation of PKC signaling contributes to the regulation of smooth muscle contraction by enhancing sensitivity of the contractile apparatus to Ca(2+). It is currently unknown whether PKC signaling contributes to the regulation of lymphatic muscle contraction. We hypothesized that the activation of PKC signaling would increase the sensitivity of the lymphatic myofilament to Ca(2+). To test this hypothesis, we determined the effects of PKC activation with phorbol esters [PMA or phorbol dibutyrate (PDBu)] on the contractile behavior of α-toxin-permeabilized rat mesenteric and cervical lymphatics or the thoracic duct. The addition of PMA or PDBu induced a significant increase in the contractile force of submaximally activated α-toxin-permeabilized lymphatic muscle independent of a change in intracellular Ca(2+) concentration, and the Ca(2+)-force relationship of lymphatic muscle was significantly left shifted, indicating greater myofilament Ca(2+) sensitivity. Phorbol esters increased the maximal rate of force development, whereas the rate of relaxation was reduced. Western blot and immunohistochemistry data indicated that the initial rapid increase in tension development after stimulation by PDBu was associated with myosin light chain (MLC)20 phosphorylation; however, the later, steady-state Ca(2+) sensitization of permeabilized lymphatic muscle was not associated with increased phosphorylation of MLC20 at Ser(19), 17-kDa C-kinase-potentiated protein phosphatase-1 inhibitor at Thr(38), or caldesmon at Ser(789). Thus, these data indicate that PKC-dependent Ca(2+) sensitization of lymphatic muscle may involve MLC20 phosphorylation-dependent and -independent mechanism(s).

Dysfunction of coronary smooth muscle Ca2+ regulation in the progression of metabolic syndrome and coronary artery disease in Ossabaw miniature swine

Coronary smooth muscle (CSM) cell Ca2+ regulation was studied in the Ossabaw swine model of metabolic syndrome (MetS) and coronary artery disease (CAD). MetS was induced by excess calorie atherogenic diet for 9 or 12 months and compared to lean controls on normal diet. Isometric tension studies on isolated coronary arteries revealed that 9 months of MetS increased tension development to KCl depolarization vs. lean controls, whereas 12 months reduced tension vs. control. MetS increases CAD and CSM Ca2+ dysregulation. CSM were isolated enzymatically and digitally imaged with the fluorescent Ca2+ indicator fura‐2. There was no difference in basal Ca2+ levels between groups. We released sarcoplasmic reticulum (SR) Ca2+ stores with caffeine. The peak Ca2+ release was increased in 9 month and decreased in 12 month MetS vs. lean. This Ca2+ response largely represents the caffeine‐sensitive SR Ca2+ store capacity. After the SR store depletion, a sustained Ca2+ signal above basal levels remained in the 9 month MetS group, reflecting store‐operated Ca2+ entry and decreased Ca2+ extrusion ability. Intravascular ultrasound and micro‐computed tomography imaging showed greater atherosclerosis and extracellular calcification in 12 vs. 9 month MetS. Collectively, these data suggest that CSM undergo dedifferentiation from a contractile to an osteogenic phenotype with progressively increasing CAD. (NIH HL062552, HL092245)

Sphingosine-1-phosphate induced increase in tension development of aortic valve leaflets is mediated via sphingosine-1-phosphate 2 receptor

Sphingosine-1-phosphate (S1P) has emerged as potent bioactive lipid with multiple functions in cardiovascular pathophysiology. However, nothing is known about the potential role of S1P in heart valves and about the signaling pathways involved. Accordingly cusp tissue has been shown to have contractile properties, but little is known as to which receptors mediate these effects. Here we investigated the effect of S1P on tension development of isolated strips of porcine aortic valve leaflets. Exposure to 90 mM KCl evoked a mean increase in force of contraction of 0.5±0.03 mN under control conditions without significant difference between the experimental groups. Increasing concentrations of S1P significantly enhanced force of contraction (112.6±2.3% at 1µM, 118.1±3.9% at 2µM, and 124.5±4.0% at 5µM) compared with control conditions. The S1P effect was completely blunted in the presence of S1P2 receptor inhibitor JTE-013. This is in accordance with previous results of our group showing that S1P induces contraction of valvular interstitial cells (VICs) from porcine aortic valves by signaling via S1P2, RhoA, and RhoA-associated protein kinase. In this way, S1P may contribute to regulation of tissue tension in aortic valves.

Food restriction impairs myocardial inotropic response to calcium and β-adrenergic stimulation in spontaneously hypertensive rats

Although long-term food restriction (FR) has been shown to induce cardiac remodeling and dysfunction, there are few data on the effects of FR on pressure-overloaded hearts. The aim of this study was to examine the effects of FR on cardiac muscle performance during inotropic stimulation in the myocardium of spontaneously hypertensive rats (SHRs). Male 60-day-old SHRs were subjected to FR for 90 days. Food-restricted animals received 50% of the ad libitum amount of food consumed by the control group. Myocardial function was studied in isolated left ventricular papillary muscle under isometric contraction in basal condition (1.25 mmol/L extracellular Ca(2+) concentration) and after 3 inotropic maneuvers: (1) at postrest contraction of 30 seconds, (2) at extracellular Ca(2+) concentration of 5.2 mmol/L, and (3) after beta-adrenergic stimulation with 10(-6) mol/L isoproterenol. At basal condition, time from peak tension to 50% relaxation was greater in the food-restricted group (P < .05). Inotropic stimulation with postrest contraction and isoproterenol promoted a significant lower increase of developed tension, maximum rate of tension development, and maximum rate of tension decline in the food-restricted compared to the control group. The elevation of extracellular Ca(2+) concentration induced a lower increase of developed tension, maximum rate of tension development, and time from peak tension to 50% relaxation in the food-restricted than in the control group. In conclusion, long-term FR promotes impairment of myocardial inotropic response to calcium and beta-adrenergic stimulation in SHRs.

Abstract 387: Prolactin-Releasing Peptide, the Novel Endogenous Ligand of the Orphan Receptor GPR10, Regulates Cardiac Contractility

The hypothalamic prolactin-releasing peptide (PrRP), which is the endogenous ligand of the orphan receptor GPR10, plays an important role in the regulation of the central stress response, sleep regulation and feeding behavior. Although specific binding sites have been found in the rat myocardium, the functional importance of PrRP in the regulation of cardiovascular system is still unknown. The objective of the present study was to characterize the direct cardiac effects of PrRP as well as the underlying signaling mechanisms. In the isolated perfused, paced rat heart preparation (male 7-week-old, Sprague-Dawley rats, n=138), intracoronary infusion of PrRP-31 (1–30 nmol/L) for 15 min induced a dose-dependent, slowly developing positive inotropic effect, with a maximal increase in developed tension at the dose of 10 nmol/L (13.6%±3.1, P&lt; 0.001). Next we tested if various counter-regulatory mechanisms could modify the effect of PrRP. The phosphodiesterase inhibitor IBMX (10 μmol/L) failed to alter the inotropic response to PrRP (10.7±1.9%, P=N.S.) excluding the involvement of cAMP-dependent mechanisms. In contrast, the specific protein kinase C-alfa (PKC-alpha) inhibitor Ro32– 0432 (100 nmol/L) significantly increased the inotropic response to PrRP (21.6±3.4%, P&lt;0.05) as well as the phosphorylation of phospholamban at Ser-16 (1.5±0.09 vs. 1.0±0.18; P&lt;0.001) as measured by Western blotting. Moreover, the protein phosphatase (PP1/PP2A) inhibitor calyculin A (CyA) augmented the effect of PrRP on contractility (20.7 ±1.7%, P&lt;0.001) similarly to PKC-alpha inhibition, whereas the PP2A inhibitor okadaic acid (10 nmol/L) had no effect (7.8±1.0 %, P=N.S.). Furthermore, long-term (120 min) administration of PrRP resulted in a greater increase in contractility (20.3±3.1 %) as compared to short-term infusion (15 min; P&lt;0.05). Our results provide the first evidence for the functional significance of PrRP in the heart. The positive inotropic effect of PrRP is mediated via cAMP-independent mechanisms, which are suppressed by enhanced activity of PKC-alpha and PP1. We suggest that PrRP may play an important role in the long-term regulation of myocardial contractility.

The enhanced endothelin-1-induced contraction in cultured coronary arteries from mature female pigs is not antagonized by 17β-estradiol

We have previously reported that organ cultured coronary arteries from market-age pigs (6–9 months of age) exhibit an enhanced contraction to the atherosclerotic-associated peptide, endothelin-1 (ET-1). The objective of this study was to investigate the interaction of 17β-estradiol with ET-1 in organ cultured coronary arteries from older female pigs (3–4 years old). A cumulative concentration–response relationship (1 × 10 − 9 M to 3 × 10 − 7 M) was generated to ET-1, and the isometric tension measured in fresh and organ cultured (4 days at 37 °C) arterial rings that were each pre-incubated for 50 min in different concentrations (1 × 10 − 9 M to 1 × 10 − 5 M) of 17β-estradiol. Compared to freshly used arteries, culturing induced a 2-fold increase in tension development to ET-1 (3 × 10 − 7 M). Although 17β-estradiol previously relaxed pre-constricted (with a 60 mM KCl solution) arteries, it did not affect the constrictive response to ET-1. Also, using an ET-1 ELISA we found that 17β-estradiol did not effect ET-1 production in intact arteries. Our results indicate that 17β-estradiol does not attenuate the production and constrictive properties of ET-1 in coronary arteries demonstrating a dedifferentiated cell phenotype.

Abstract 578: Urotensin-II Upregulation in Human Heart Failure: Direct Effects on Contractility and Sarcoplasmic Reticulum Calcium Load

While studies have reported that Urotensin II (UII) is upregulated in failing (F) hearts, the pathophysiologic implications of increased UII on cardiac function is incompletely understood. We investigated the direct effects of exogenous UII on in vitro contractile performance and sarcoplasmic reticulum (SR) Ca 2+ load in F and non failing (NF) human myocardium. Methods: Isometric force was measured in F (n=11) and NF (n=6) right ventricular trabeculae (0.5 Hz, 37°C) before and after bath application of UII (UII; 0.1, 1.0, 10, 100, 1000 nM). Single and paired rapid cooling contractures (RCCs) were employed to define SR Ca 2+ load and the balance between SERCA and the NCX. Results: UII induced increases in developed tension (DT) in NF trabeculae and dose-dependent decreases in DT in F trabeculae. UII did not alter resting tension in either group. RCC amplitude increased during graded increases in UII in NF trabeculae, suggesting increases in SR calcium load, but remained stable in F trabeculae. In NF trabeculae, UII did not alter the RCC2/RCC1 ratio at any concentration, but in F trabeculae the RCC2/RCC1 ratio increased at 0.1 nM UII then exhibited dose-dependent decreases at higher concentrations. Conclusion: In contrast to the positive inotropic response elicited in NF hearts, UII is a negative inotrope in ventricular trabeculae obtained from F human hearts. Based on the RCC data, UII has little effect on SR Ca 2+ load in F myocardium suggesting that negative inotropic responses reflect decreases in myofilament Ca 2+ sensitivity. The paired RCC data suggests that U-II also affects the functional competition between SERCA and NCX differently in F and NF human hearts.

Effects of Remifentanil on the Contractility of Failing Human Heart Muscle

The purpose of this study was to determine the direct effects of remifentanil on inotropy of human myocardial tissue removed from failing explanted hearts of cardiac transplant recipients. In vitro, prospective study with repeated measures. Research laboratory in a tertiary referral center. The authors studied the effects of remifentanil on isometric myocardial contractile parameters in failing atrial and ventricular myocardial muscles. The authors also used 1 micromol/L of isoproterenol to test adrenergic responsiveness of failing myocardial tissues in the presence of supratherapeutic remifentanil concentrations. Muscles were studied at 37 degrees C and 1 Hz, with concentrations of remifentanil covering a wide range of therapeutic as well as supratherapeutic concentrations (ie, between 0.001 and 100 microg/mL). After the dose-response curve was obtained, the author measured the inotropic response to 1 micromol/L of isoproterenol in the presence of the cumulative dose of remifentanil. Over the wide range of concentrations, remifentanil had no significant effect on contractile function. In the presence of supratherapeutic concentration of remifentanil, isoproterenol produced a 97% +/- 58% increase in developed tension in failing atrial muscles and a 91% +/- 40% increase in failing ventricular muscles, which did not significantly differ from control muscles. The authors showed that remifentanil had no significant negative inotropic effects in failing human heart muscle and that myocardial contractility remained fully responsive to beta-adrenergic stimulation at all remifentanil concentrations studied.

Communication: Effect of diperoxovandate on isolated rat heart

Diperoxovanadate (DPV), a product of vanadate is gaining importance as a biologically active vanadium compound. The aim of the present study was to evaluate the chronotropic and inotropic activity of DPV using isolated rat heart and to determine the concentration at which it is toxic to the heart. The study was carried out using modified Langendorff\'s setup. DPV was injected at varying concentrations (from 10-9 to 10-4) either as bolus (0.1 ml) or the heart was perfused continuously with varying concentrations of DPV for 10 min. Low concentration of DPV did not produce any significant effect on chronotropy and on developed tension. However, as the dose of DPV was increased, tension developed and heart rate was enhanced to significant extent (P < 0.05) and both were found to be maximum at a dose of 10-7M. Further increase in DPV dose did not show either an increase in force or rate of contraction of heart but instead produced a relative decrease in both of these parameters when compared with the 10-7 M dose. When heart was perfused with a dose of 10-7 M DPV continuously for 10 min there was a significant increase in heart rate and developed tension (P < 0.01). It was also found that at a dose of 10-5M, DPV showed not only further increase in developed tension but also produced marked disturbances in the rhythm indicating cardiac toxicity. This was further confirmed by lactate dehydrogenase (LDH) activity determination. Keywords: diperoxovanadate, reperfusion, isolated rat heart, hydrogen peroxide, lactate dehydrogenase Ethiopian Pharmaceutical Journal, vol. 22 (2004): 47-52

Interplay between nitric oxide and VIP in CCK-8-induced phasic contractile activity in the rabbit sphincter of Oddi

The sphincter of Oddi (SO) plays an important role in delivery of bile into the duodenum. To establish whether vasoactive intestinal polypeptide (VIP) and nitric oxide (NO) were involved in phasic contractile activity of the rabbit SO stimulated by cholecystokinin-octapeptide (CCK-8). Isolated SO muscle rings were cleaned of fat and mounted horizontally on two small L-shaped hooks one of which was connected to a force transducer for the measurement of isometric tension. The experiments were carried out in a thermostatically controlled (37+/-0.2 degrees) organ bath (5 mL) containing Krebs solution. The organ fluid was gassed with 95% O(2) and 50 mL/L CO(2) to keep the pH at 7.40+/-0.05. Contractile responses to CCK-8 (1 micromol/L) were evaluated in the presence and absence of N(G)-nitro-L-arginine (LNNA), an inhibitor of NO synthase (100 micromol/L), and (p-chloro-D-Phe(6)-Leu(17))-VIP (VIPa, 30 micromol/L), a VIP receptor antagonist. CCK-8 stimulated the phasic activity of the SO. NO synthase inhibition increased the frequency and amplitude of contractions with a slight increase in developed tension. Pre-incubation with VIPa also attenuated this CCK-8 effect. The combined application of LNNA and VIPa abolished the phasic activity of the muscle rings with a marked increase in tension in response to CCK-8. VIP and NO together contribute to an increase in phasic activity of SO.

Progestins used in hormonal replacement therapy display different effects in coronary arteries from New Zealand white rabbits

Objectives: The aim of this study was in an animal model to assess the vascular effects of different progestins commonly used in hormonal replacement treatment. Methods: Fifty-six non-atherosclerotic, ovariectomized New Zealand white rabbits were randomized into seven groups: (1) medroxyprogesterone acetate (MPA), (2) norethisterone acetate (NETA), (3) conjugated equine estrogens (CEE), (4) 17-β-estradiol (E2), (5) MPA+CEE, (6) NETA+E2, (7) or placebo ( n=8) and given hormonal treatment through the diet for 4 weeks. Ring segments from the left proximal coronary artery and from the distal part of the left anterior descending coronary artery were microdissected and mounted for isometric tension recordings in a myograph. The vasoconstrictory responses induced by potassium, endothelin-1, calcium and N w-nitro- l-arginine methyl ester, and the vasodilatory response induced by acetylcholine and sodiumnitroprusside were investigated. The maximum contraction/relaxation ( E max) and the concentration required to induce half the maximum response (EC 50) were determined. EC 50 values were expressed as the negative logarithm to the molar concentration, pD 2=− log EC 50 . Results: Treatment with MPA alone caused when compared to treatment with NETA an increase in tension development in the distal coronary artery after the addition of potassium (6.36±0.36 versus 4.31±0.42 P<0.005) (single dose response, mN/mm, mean±S.E.M.) and endothelin-1 (9.41±0.82 versus 6.43±0.73 P<0.05) ( E max, mN/mm, mean±S.E.M.). Treatment with MPA compared to placebo caused an endothelin-1 induced increase of E max in the distal coronary artery (9.21±0.87 versus 6.51± 0.65 P<0.05) and a calcium induced increase of pD 2 in both coronary arteries (2.98±0.19 versus 2.42±0.12 P<0.05, proximal coronary artery) (3.26±0.09 versus 2.9±0.1 P<0.05, distal coronary artery) (pD 2, mean±S.E.M.). Treatment with NETA compared to placebo in the proximal coronary artery, after the addition of sodiumnitroprusside caused a decrease of pD 2 (5.33±0.19 versus 5.94±0.13 P<0.05). Treatment with E2 compared to treatment with CEE in the proximal coronary artery caused a decrease of pD 2 after the addition of sodiumnitroprusside (5.00±0.16 versus 5.77±0.28 P<0.05). No significant differences were found between MPA+CEE and NETA+E2. Conclusion: Treatment with MPA alone seems to enhance the contractile response to potassium and endothelin-1 in the distal coronary artery compared to NETA, indicating that different progestins used in hormonal replacement treatment may display different effects on contractile functions of coronary arteries.

Moxonidine, an antihypertensive agent, is permissive to alpha1-adrenergic receptor pathway in the rat-tail artery.

To investigate whether alpha1-adrenergic receptors were involved in the contractile response of tail arteries to moxonidine, isolated ring segments of tail arteries from male adult Sprague-Dawley rats were studied. Moxonidine (EC50 = 1.3 microM) and the alpha1-agonist phenylephrine (EC50 = 2.5 microM) increased tension development in the rat-tail artery similarly. The response to moxonidine (1 microM) could be blocked by both alpha1-adrenoceptor blockers prazosin (IC50 = 1 nM), and urapidil (IC50 = 14 nM), and also by alpha2-adrenoceptor blockers, yohimbine (IC50 = 49 nM) and efaroxan (IC50 = 49 nM). Combination drug treatment (urapidil and yohimbine, or yohimbine and prazosin) was more effective in blocking the contractile response to moxonidine, than treatment with prazosin or urapidil alone. Comparison of pA2 values for prazosin in the presence of moxonidine (9.35) or phenylephrine (10.2) confirm that alpha1-adrenergic receptors are involved in the contractile response of rat-tail artery to moxonidine.

In vivo monitoring of intracellular free calcium changes during uterine activation by prostaglandin f(2alpha) and oxytocin.

It has been well established that oxytocin (OXT) increases intracellular free calcium ([Ca(2+)](i)) by targeting both intracellular and extracellular stores, but the mechanisms involved in the increase through activation with prostaglandin F(2alpha) (PGF(2alpha)) are still incompletely understood. This study was designed to elucidate the source(s) of increased [Ca(2+)](i) in response to PGF(2alpha) (10(-6) M) or OXT (10(-8) M) administration in the near-term rat myometrium. The animals were divided into an in vitro group (n= 8), where the developed tension of uterine strips was assessed, and an in vivo group (n= 5), where a lobe of the uterus with intact innervation and circulation was loaded with the fluorescent indicator Indo-1 AM to assess [Ca(2+)](i). PGF(2alpha) and OXT induced a 30.1% and 35.9%, respectively, increase in developed tension in the potassium chloride-depolarized myometrial strips. Nifedipine reduced the PGF(2alpha) and OXT increased tension by 65.8% and 49.4%, respectively. In vivo, both PGF(2alpha) and OXT increased [Ca(2+)](i) in the potassium chloride-depolarized uterine muscle by 35.7% and 44.6%, respectively, increases similar to the rises in tension in vitro. Nifedipine reduced these effects of PGF(2alpha) and OXT by 45.3% and 39.6%. These findings indicate that in near-term myometrium the source of increased [Ca(2+)](i) after administration of PGF(2alpha), similar to OXT, is both extracellular and intracellular.

Decreased expression of phospholamban is not associated with lower beta-adrenergic activation in rat atria.

The aim of the study was to find out whether low phospholamban level in atria as compared with ventricles is associated with differences in sarcoplasmic reticular Ca2+-uptake and contractile performance. Relationship between phospholamban and beta-adrenergic stimulation in rat left atria and papillary muscles were examined by means of contractile measurements, sarcoplasmic reticular oxalate-supported Ca2+-uptake, and Western blotting of phosphorylated phospholamban. Phosphoprotein determination after beta-adrenergic stimulation demonstrated that the levels of Ser16 and Thr17 phosphorylated phospholamban in atria remained at about one-third of that in ventricles. However, comparison of sarcoplasmic reticular Ca2+-uptake in control and isoproterenol perfused preparations demonstrated that the effect of beta-adrenergic stimulation on sarcoplasmic reticular Ca2+-uptake was stronger in atrial preparations. Moreover, atria responded to isoproterenol with much larger increases in developed tension, contractility and relaxation rates than papillary muscles. Thus, despite lower level of phospholamban, the beta-adrenergic activation of sarcoplasmic reticular Ca2+-uptake and contractile indices are higher in atria.

Impact of NO on ET-1- and AM-induced inotropic responses: potentiation by combined administration.

We characterize herein the impact of myocardial nitric oxide (NO) synthesis on the inotropic response to two cardioactive peptides, endothelin-1 (ET-1) and adrenomedullin (AM). In the isolated perfused rat heart preparation, intracoronary infusion of AM (0.03 and 1 nmol/l) and ET-1 (0.08 and 1 nmol/l) for 30 min induced a dose-dependent, gradual increase in developed tension, the maximal responses being equal. Inhibition of myocardial NO synthase (NOS) by N(omega)-nitro-L-arginine methyl ester (L-NAME; 300 micromol/l) enhanced the inotropic response to ET-1 at a concentration of 1 nmol/l; meanwhile, the effect of AM was not augmented significantly. The inotropic response to simultaneous administration of low, equipotent doses of AM (0.03 nmol/l) and ET-1 (0.08 nmol/l) was significantly smaller than that of either peptide alone. This depressed response was more than overcome by concomitant administration of L-NAME. In conclusion, this study reveals that the maximal inotropic response to ET-1 can be augmented by inhibition of myocardial NOS, whereas it has only a minor impact on the effect of AM. The inotropic response to combined administration of low doses of AM and ET-1 is substantially suppressed by endogenous NO, whereas the individual effects of the peptides at these doses are not the subject of secondary modulation by NO.

Oxygen free radicals and excitation-contraction coupling.

Oxygen free radicals (OFR) contribute to contractile failure, rigor, and calcium (Ca2+) overload in ischemic/reperfused myocardium. Using both multicellular and isolated single-cell preparations, our laboratory has identified two fundamental mechanisms contributing to the deleterious effects of OFR: (i) impaired myocardial metabolism, and (ii) altered myocardial calcium handling. Impaired metabolism leads to activation of metabolically sensitive K+ currents, which shorten the action potential, thereby decreasing the duration of systole. Ultimately, high-energy phosphate depletion secondary to metabolic failure results in rigor. Altered myocardial Ca2+ handling is evidenced by a decrease in Ca2+ entry via L-type Ca2+ channels [another cause of decreased action potential duration (APD)], a reduction in sarcoplasmic reticulum (SR) Ca2+ content, slowed Ca2+ uptake in diastole, and increased sodium-calcium exchange (NaCaX) activity. The increase in NaCaX activity may contribute to the early increase in developed tension frequently observed in multicellular preparations exposed to free radicals, as well as the SR depletion occurring early on in voltage-clamped isolated cell preparations. Increased NaCaX activity is likely to be a critical factor underlying the late Ca2+ overload that occurs in the setting of increased intracellular Na+, and which leads to irreversible injury. The extent to which free radical-mediated metabolic inhibition participates in the dysfunction of the L-type Ca2+ channel is uncertain. The altered activity of the SR Ca2+ pump and NaCaX are more likely caused by direct actions of OFR on these proteins.