Caffeine-induced contracture in oesophageal striated muscle of normotensive and hypertensive rats

To elucidate whether properties of the sarcoplasmic reticulum are altered, not only in vascular smooth muscle, but also in visceral striated muscle of spontaneously hypertensive rats (SHR), caffeine-induced contractures in oesophageal striated muscle of Wistar Kyoto rats (WKY) and stroke-prone SHR (SHRSP) were compared. In both preparations, 30 mM caffeine induced a contracture with two components. The second component, which was diminished by extracellular Ca(2+) removal or Ni(2+) but not by verapamil, was much smaller in SHRSP. Both components and differences between WKY and SHRSP coincided with changes in intracellular Ca(2+). Although membrane potential was identical between these preparations, caffeine induced slight depolarization only in WKY preparations. Similar depolarization was observed with 10 mM K(+), which induced no contraction. It is suggested that the first and the second components of caffeine-induced contracture were induced by Ca(2+) released from sarcoplasmic reticulum and by Ca(2+) that entered through channels activated by sarcoplasmic reticulum Ca(2+) depletion, respectively. In SHRSP preparations, Ca(2+) from the latter pathway was clearly decreased, although this change is thought not to be related to the initiation of hypertension. These results suggest that Ca(2+) handling properties of cell membrane and sarcoplasmic reticulum are generally altered in muscles of SHRSP.     

Reduced effect of caffeine on twitch contraction of oesophageal striated muscle from stroke-prone spontaneously hypertensive rats

1. There are known differences in the sensitivity to caffeine between skeletal muscle (soleus) of normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). The present study was performed in order to examine differences in the effects of caffeine on twitch contraction between visceral striated muscle using the outer layer of the oesophagus from WKY rats and stroke-prone SHR (SHRSP). 2. Caffeine, at concentrations ranging from 0.3 to 10 mmol/L, exhibited potentiating effects on twitch contraction in preparations from both WKY rats and SHRSP. The potentiating effect of caffeine was markedly less prominent in preparations from SHRSP compared with preparations from WKY rats. 3. The rate of contraction and relaxation, the time to peak tension and 80% relaxation time were not significantly altered by caffeine at concentrations lower than 3 mmol/L in preparations from either strain. 4. With 10 mmol/L caffeine, the rate of relaxation was markedly reduced and the 80% relaxation time was prolonged, with no significant changes in the rate of contraction, in preparations from WKY rats. These changes were significantly smaller in preparations from SHRSP. 5. The duration of the action potential was greater in preparations from SHRSP than in preparations from WKY rats, although the membrane potential and the amplitude of the action potential were not significantly different between preparations from WKY rats and SHRSP. 6. Caffeine, at 10 mmol/L, prolonged the duration of the action potential in preparations from both strains. The effect of caffeine was not different between preparations from WKY rats and SHRSP. 7. The results of the present study suggest that caffeine augments release of Ca2+ from the sarcoplasmic reticulum (SR) at low concentrations and attenuates Ca2+ re-uptake at 10 mmol/L. Decreased reactivity of SR to caffeine may be a cause of the lesser potentiation of twitch contraction by caffeine in preparations from SHRSP.     

Effects of cyclopiazonic acid and thapsigargin on electromechanical activities and intracellular Ca2+ in smooth muscle of carotid artery of hypertensive rats.

1. The effects of cyclopiazonic acid (CPA) and thapsigargin (TG), both of which are known to inhibit sarcoplasmic reticular Ca(2+)-ATPase, on the mechanical activities, intracellular Ca2+ level and electrical activities of smooth muscle of the carotid artery of stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar Kyoto rats (WKY) were compared. 2. Both CPA and TG induced elevation of tension of the smooth muscle, which was composed of a phasic and a tonic component. The level of tension attained, especially the tonic component, was greater in the preparation from SHRSP. 3. The elevation of tension was associated with an increased intracellular Ca2+ level. Both the elevation of tension and the increase in intracellular Ca2+ were diminished by the removal of extracellular Ca2+ or by the application of verapamil. 4. The resting membrane potential of the preparations from SHRSP were depolarized to a greater extent than those from WKY.CPA depolarized the smooth muscle from both SHRSP and WKY, and the final level was also more depolarized in the preparation from SHRSP. 5. These results indicate that the elevation of tension induced by these drugs is mainly due to increased Ca2+ influx through voltage-dependent Ca2+ channels, and the difference in the action between the preparation from SHRSP and that from WKY can be explained mainly by the changes in the channels. 6. Thus, differences in the action of these drugs on the tension of smooth muscle between preparations from WKY and SHRSP can mainly be explained by the difference in the membrane potential which is related to the difference in voltage-dependent Ca2+ influx.     

Abnormal response to ryanodine in oesophageal striated muscle of spontaneously hypertensive rats

The effects of ryanodine on twitch contraction and basal tension of oesophageal striated muscle were compared between preparations from stroke-prone spontaneously hypertensive rats (SHRSP) and normotensive Wistar Kyoto rats (WKY). Ryanodine (3 x 10(-7) M) augmented the twitch contraction in WKY preparations, butt attenuated it in SHRSP preparations. Rates of contraction and relaxation of twitch contraction, normalized to developed tension, were slightly decreased by ryanodine in both preparations. The effect of ryanodine was not different between WKY and SHRSP preparations. Ryanodine elevated the basal tension in WKY preparations but not in SHRSP preparations. Ryanodine elevated the intracellular Ca(2+) level in both preparations, but the response was significantly less in SHRSP preparations. Resting and action potentials were not significantly different between WKY and SHRSP preparations, while the duration of the action potential was significantly longer in SHRSP preparations. Ryanodine did not alter the resting and action potentials of either preparation. These results suggest that the Ca(2+) handling properties, including the ryanodine receptor, of the sarcoplasmic reticulum are genetically altered in oesophageal striated muscle of SHRSP.     

Augmented sphingosylphosphorylcholine-induced Ca2+-sensitization of mesenteric artery contraction in spontaneously hypertensive rat

Sphingosylphosphorylcholine (SPC) is a vasoconstricting lysosphingolipid, and the RhoA/Rho-kinase pathway plays an important role in SPC-induced contraction. Since RhoA/Rho-kinase-mediated signaling is involved in the generation and/or maintenance of hypertension, we compared the effect of SPC on the contractility of endothelium-denuded small mesenteric arteries in spontaneously hypertensive rats (SHR) and Wistar Kyoto rats (WKY). Fura-2 Ca²⁺ signals, contractile responses, and phosphorylation of 20-kDa myosin light chains (MLC₂₀) were measured. Ten μM SPC induced a gradual and sustained vasoconstriction, which was greater in arteries of the SHR (82.5±4.3%, n=9) than in those of the WKY (26.7±4.5%, n=10). In Ca²⁺-free media, SPC gradually increased vascular tone in the SHR, but caused little vasoconstriction in the WKY. In the SHR and WKY, SPC evoked a greater vasoconstriction than did high K⁺depolarization at a given Ca²⁺ ratio, and the Ca²⁺ ratio–tension curve induced by SPC was significantly shifted to the left compared with that induced by high K⁺ depolarization. However, the magnitude of shift to the left was greater in the SHR than in the WKY. The Rho-kinase inhibitor Y-27632 significantly inhibited SPC-induced contractions, but neither the protein kinase C inhibitor calphostin-C nor PD98059, which inhibits activation of some mitogen-activated protein kinases, had any effect on the SHR or the WKY. SPC significantly increased the phosphorylation of MLC₂₀ in both the SHR and the WKY, and Y-27632 inhibited the SPC-induced increase in MLC₂₀ phosphorylation in the SHR. Our results suggest that SPC induces greater vascular tone in the SHR than in the WKY. Furthermore, our results indicate that activation of the Rho-kinase pathway plays an important role in the SPC-induced Ca²⁺ sensitization in the SHR.     

Ca2+ buffering function of sarcoplasmic reticulum in rat tail arteries: comparison in normotensive and spontaneously hypertensive rats

The superficial buffer barrier function of the sarcoplasmic reticulum (SR) during rest and that during stimulation with Bay k 8644, an agonist of L-type Ca2+ channels, were compared in endothelium-denuded strips of tail arteries from 13-week-old normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR), by measuring the effects of cyclopiazonic acid (CPA) and thapsigargin that inhibit SR Ca2+-ATPase and the effect of ryanodine that depletes SR Ca2+. The addition of 10 microM CPA induced a transient contraction that was not significantly different between WKY and SHR. The CPA-induced contraction was strongly inhibited by 100 nM nifedipine and was abolished by Ca2+-free solution in both strains. Thapsigargin (100 nM) or ryanodine (10 microM) induced similar, small transient contractions in the two strains. The addition of Bay k 8644 (1-100 nM) almost failed to induce a contraction in both WKY and SHR. When the strips were preincubated with 10 microM CPA, 100 nM thapsigargin or 10 microM ryanodine, Bay k 8644 induced similar concentration-dependent contractions in the two strains. The amount of Ca2+ stored in the SR, as estimated from the 20 mM caffeine-induced contraction, was not significantly different between WKY and SHR. Our results suggest that the SR of rat tail arteries can buffer a large amount of Ca2+ that enters the cell during the rest and the Bay k 8644 stimulation, and these functions are not altered in SHR.     

Tachycardia in spontaneously hypertensive and normotensive rats after fusaric acid and bupicamide

1. The effects of the dopamine-β-hydroxylase inhibitors bupicamide, fusaric acid, FLA-63 and U-14,624 on blood pressure and heart rate of spontaneously hypertensive rats were examined. 2. Bupicamide and fusaric acid caused marked tachycardia whereas FLA-63 and U-14,624 caused modest bradycardia; all drugs decreased blood pressure. 3. In normotensive rats, fusaric acid caused the same degree of tachycardia as in spontaneously hypertensive rats, but blood pressure was only slightly reduced. 4. Tachycardia after fusaric acid was not due to increased sympathetic activity or decreased parasympathetic activity but required intact catecholamine stores. 5. It is concluded that fusaric acid causes tachycardia by releasing catecholamines indirectly and that a metabolite of fusaric acid is also involved.     

Effects of dihydropyridine-type Ca2+ antagonists on the renal arterial tree in spontaneously hypertensive rats.

The effects of hypertension and of treatment with dihydropyridine-type Ca2+ antagonists and the vasodilator hydralazine on renal arterial tree were investigated in spontaneously hypertensive rats (SHR) with quantitative microanatomical techniques. Pharmacological treatment decreased to a similar extent systolic blood pressure values in SHR. Increased thickness of the tunica media of intrarenal arteries accompanied and luminal narrowing were observed in control SHR. Lercanidipine, manidipine, and nicardipine significantly countered wall thickening and luminal narrowing. Hydralazine countered luminal narrowing only. Dihydropyridines exerted renal vasocilatory activity primarily on resistance arteries, being lercanidipine the only compound active on small sized arteries.     

硝苯地平和环匹阿尼酸对易卒中型肾性高血压大鼠血管平滑肌收缩功能的影响

观察了易卒中型肾性高血压大鼠(SPRHR)主动脉环血管平滑肌在术?后血压升高不同阶段收缩反应性. 与假手术组相比，随着术后血压的升高，低浓度KCl 10～20 μmol·L^-1使SPRHR主动脉环的收缩反应明显增强；苯肾上腺素（Phe）在无钙Krebs液中所致的收缩反应强度降低，而复钙后的收缩张力明显增强；SPRHR主动脉环在无钙Krebs液中温育1 h后，重新复钙后可致收缩反应且为硝苯地平（Nif）所抑制；肌浆网Ca²⁺泵抑制剂环匹阿尼酸（CPA）在SPRHR主动脉环上所致的持续收缩反应也明显强于对照并为Nif所阻断；连续给予Nif，卡托普利180±11 mg·kg^-1·d^-1 8 wk后，SPRHR的血压均明显下降. 在术后1, 4 wk, Nif对部分SPRHR主动脉环在无钙液中复钙所致的收缩反应并无抑制作用且可进一步引起收缩反应. 结果表明，肾性高血压大鼠血管平滑肌收缩功能的高反应性与高血压状态下Ca²⁺转运的失调有关，不同高血压时期的血管平滑肌细胞膜上的Ca²⁺通道特性可能存在差异性.     

硝苯地平和环匹阿尼酸对易卒中型肾性高血压大鼠血管平滑肌收缩功能的影响

观察了易卒中型肾性高血压大鼠（ＳＰＲＨＲ）主动脉环血管平滑肌在术后血压升高不同阶段收缩反应性．与假手术组相比，随着术后血压的升高，低浓度ＫＣｌ１０～２０μｍｏｌ·Ｌ－１使ＳＰＲＨＲ主动脉环的收缩反应明显增强；苯肾上腺素（Ｐｈｅ）在无钙Ｋｒｅｂｓ液中所致的收缩反应强度降低，而复钙后的收缩张力明显增强；ＳＰＲＨＲ主动脉环在无钙Ｋｒｅｂｓ液中温育１ｈ后，重新复钙后可致收缩反应且为硝苯地平（Ｎｉｆ）所抑制；肌浆网Ｃａ２＋泵抑制剂环匹阿尼酸（ＣＰＡ）在ＳＰＲＨＲ主动脉环上所致的持续收缩反应也明显强于对照并为Ｎｉｆ所阻断；连续给予Ｎｉｆ，卡托普利１８０±１１ｍｇ·ｋｇ－１·ｄ－１８ｗｋ后，ＳＰＲＨＲ的血压均明显下降．在术后１，４ｗｋ，Ｎｉｆ对部分ＳＰＲＨＲ主动脉环在无钙液中复钙所致的收缩反应并无抑制作用且可进一步引起收缩反应．结果表明，肾性高血压大鼠血管平滑肌收缩功能的高反应性与高血压状态下Ｃａ２＋转运的失调有关，不同高血压时期的血管平滑肌细胞膜上的Ｃａ２＋通道特性可能存在差异性．     

Potent vasoconstrictor actions of cyclopiazonic acid and thapsigargin on femoral arteries from spontaneously hypertensive rats

1. The Ca²⁺ buffering function of sarcoplasmic reticulum (SR) in the resting state of arteries from spontaneously hypertensive rats (SHR) was examined. Differences in the effects of cyclopiazonic acid (CPA) and thapsigargin, agents which inhibit the Ca²⁺-ATPase of SR, on tension and cellular Ca²⁺ level were assessed in endothelium-denuded strips of femoral arteries from 13-week-old SHR and normotensive Wistar-Kyoto rats (WKY).
2. In resting strips preloaded with fura-PE3, the addition of CPA (10 μM) or thapsigargin (100 nM) caused an elevation of cytosolic Ca²⁺ level ([Ca²⁺]_i) and a contraction. These responses were significantly greater in SHR than in WKY.
3. The addition of verapamil (3 μM) to the resting strips caused a decrease in resting [Ca²⁺]_i, which was significantly greater in SHR than in WKY. In SHR, but not in WKY, this decrease was accompanied by a relaxation from the resting tone, suggesting the maintenance of myogenic tone in the SHR artery.
4. Verapamil (3 μM) abolished differences between SHR and WKY. The effects of verapamil were much greater on the contraction than on the [Ca²⁺]_i.
5. The resting Ca²⁺ influx in arteries measured after a 5 min incubation of the artery with ⁴⁵Ca was not increased by CPA or thapsigargin in either SHR or WKY. The net Ca²⁺ entry measured after a 30 min incubation of the artery with ⁴⁵Ca was decreased by CPA or thapsigargin in both SHR and WKY. The resting Ca²⁺ influx was significantly higher in SHR than in WKY, and was decreased by nifedipine (100 nM) in the SHR artery, but was unchanged in the WKY artery.
6. The resting ⁴⁵Ca efflux from the artery was increased during the addition of CPA (10 μM). This increase was less in SHR than in WKY. The resting ⁴⁵Ca efflux was the same in SHR and WKY.
7. These results suggest that (1) the Ca²⁺ influx via L-type voltage-dependent Ca²⁺ channels (VDCCs) was increased in the resting state of the SHR femoral artery, (2) the greater part of the increased Ca²⁺ influx was buffered by Ca²⁺ uptake into the SR and some Ca²⁺ reached the myofilaments resulting in the maintenance of the myogenic tone, and (3) therefore the functional elimination of SR by CPA or thapsigargin caused a large elevation of [Ca²⁺]_i and a potent contraction in this artery. During this process, the contraction was mainly due to the basal Ca²⁺ influx via L-type VDCCs. The present study also showed the existence of a relatively large compartment of [Ca²⁺]_i which does not contribute to the contraction during the addition of CPA or thapsigargin.

     

Force and intracellular Ca2+ during cyclic nucleotide-mediated relaxation of rat anococcygeus muscle and the effects of cyclopiazonic acid.

1. Simultaneous recordings of tension and [Ca2+]i were made in rat anococcygeus muscle strips to investigate possible mechanisms involved during cyclic nucleotide-mediated relaxation. Relaxation of pre-contracted muscles was induced by sodium nitroprusside (SNP) or forskolin and the effects of cyclopiazonic acid (CPA) on these responses were examined. 2. In muscles pre-contracted with 0.2 microM phenylephrine addition of SNP (10 microM) caused a rapid and near complete relaxation of force. This was accompanied by a decrease in [Ca2+]i, however, this was not of a comparable magnitude to the decrease in force. The level of [Ca2+]i in muscles relaxed with SNP was shown to be associated with substantially higher force levels in the absence of SNP. Forskolin (10 microM) caused a slower, essentially complete relaxation which was associated with a proportional decrease in [Ca2+]i. 3. In muscles pretreated with SNP or forskolin subsequent responses to phenylephrine were attenuated with both force and [Ca2+]i rising slowly to attain eventually levels similar to those observed when the relaxant was applied to pre-contracted muscles. 4. Exposure of the muscles to the sarcoplasmic reticulum Ca(2+)-ATPase inhibitor, CPA (10 microM), resulted in a sustained increase in [Ca2+]i which, in most cases, was not associated with any force development. The relaxation and decrease in [Ca2+]i in response to both SNP and forskolin were attenuated and substantially slowed in the presence of CPA. Overall the extent of this attenuation was greater for SNP. For both SNP and forskolin, CPA attenuated the decrease in [Ca2+]i to a greater extent than the decrease in force. In some cases, SNP-mediated relaxation in the presence of CPA was observed with almost no detectable change in [Ca2+]i. 5. The results suggest that, in the rat anococcygeus muscle under normal circumstances, a lowering of [Ca2+]i can fully account for the relaxation induced by forskolin but not for that induced by SNP, where mechanisms independent of changes in [Ca2+]i appear to contribute. Whilst Ca2+ sequestration into the sarcoplasmic reticulum plays a role in the relaxation mediated by both SNP and forskolin other Ca2+ lowering mechanisms may also be involved, especially in the response to forskolin.     

Age related changes in Ca2+ channels in spontaneously hypertensive rats.

1. The binding of the Ca2+ channel antagonist [3H]PN200 110 to 1,4-dihydropyridine binding sites in cardiac, brain and vascular smooth muscle preparations from WKY and SHR was studied as a function of age and blood pressure. 2. Binding site density in the heart from prehypertensive SHR (6 weeks) was significantly lower than that of WKY. 3. Between 6 and 12 weeks binding site density in SHR increased and between 12 and 24 weeks there was no difference between WKY and SHR. No changes in affinity occurred. 4. In brain a significant fall in binding site density occurred between 6 and 12 weeks and there was (with limited data) an increase in binding site density in tail artery membranes. 5. A good correlation (r = 0.82) exists between blood pressure and cardiac binding site density between 6 and 12 weeks of age in SHR.     

Interaction of cyclopiazonic acid with rat skeletal muscle sarcoplasmic reticulum vesicles. Effect on Ca2+ binding and Ca2+ permeability

The interaction of cyclopiazonic acid with rat skeletal muscle sarcoplasmic reticulum (SR) vesicles was investigated in order to study the mechanism of cyclopiazonic acid inhibition of the Ca2+-ATPase (Goeger et al., Biochem Pharmacol 37: 978-981, 1988). Cyclopiazonic acid at 25 microM prevented the binding of Ca2+ to the high affinity binding site of mixed (light and heavy) SR vesicles and inhibited, in a dose-dependent manner, the Ca2+-dependent phosphorylation of SR vesicles by ATP. Binding of Ca2+ to the high affinity site of the CA2+-ATPase is necessary for both Ca2+ transport and for phosphorylation of the Ca2+-ATPase. We conclude that inhibition of Ca2+ binding to the high affinity site may be responsible, at least in part, for the activity of cyclopiazonic acid. The mechanism of inhibition remains unclear. The inhibition was not reduced after dialysis and was only partially reversed by gel filtration of SR vesicles treated with cyclopiazonic acid. Neither 1 mM glutathione nor dithiothreitol pretreatment had any effect on the inhibition of the Ca2+-ATPase. In addition to its inhibition of Ca2+ uptake and the Ca2+-ATPase, cyclopiazonic acid had significant effects on Ca2+ efflux from both passively and actively loaded SR vesicles. Cyclopiazonic acid impeded the efflux of Ca2+ from passively loaded SR vesicles (in the presence of ruthenium red) when compared to either untreated vesicles or those treated with mersalyl acid, a mercurial which also inhibits the Ca2+-ATPase and is known to induce Ca2+ release by both ruthenium red-sensitive and -insensitive pathways. Treatment of actively loaded vesicles with cyclopiazonic acid resulted in a decreased rate of Ca2+ efflux when compared to SR vesicles in which the Ca2+-ATPase activity was inhibited by ATP depletion with hexokinase and glucose. The results are consistent with the hypothesis that, in mixed SR vesicles, cyclopiazonic acid inhibits both the Ca2+ pump and Ca2+ efflux.     

Vascular reactivity in spontaneously hypertensive normotensive and hypotensive rats

1 Three strains of rats spontaneously hypertensive (HRS), normotensive (NS) and hypotensive (HOS), were selected by repeated inbreeding.2 In order to explain the different blood pressure levels observed, we have studied their vascular reactivity to noradrenaline and angiotensin II. Experiments were performed in anaesthetized ganglion-blocked, vagotomized rats.3 The reactivity to noradrenaline was significantly higher in HRS than in NS and HOS rats.4 The reactivity to angiotensin II appeared identical in the three strains used.     

Ca2+ influx in spontaneously hypertensive rats is sensitive to calcium antagonists

Upon perfusion with a solution containing Ca²⁺, after Ca²⁺-free perfusion, aortic rings from spontaneously hypertensive rats (SHR) responded with a contraction. The contraction was usually transient. When verapamil or nifedipine were added prior to and during Ca²⁺ repletion, the force developed was reduced to 25% of that of uninhibited contraction. Aortas from normotensive rats did not show comparable properties. These findings demonstrate strate that calcium influx in aortic smooth muscle of SHR occurs mainly via voltage-dependent calcium channels.     

Prostanoid-mediated vascular contraction in normotensive and hypertensive rats.

We investigated the role of prostanoids in the constrictor effect of calcium ionophore A23187, endothelin-1 and vasopressin in rings of thoracic aorta obtained from normotensive rats and rats with aortic coarctation-induced hypertension. Isometric tension was measured in aortic rings bathed in buffer with and without indomethacin (10 microM), CGS13080 (10 microM) or SQ29548 (1 microM) to inhibit cyclooxygenase and thromboxane synthase and to block TxA2-PGH2 receptors, respectively. Increases in tension elicited by A23187 and vasopressin in aortic rings from hypertensive rats exceeded responses in rings from normotensive rats. A23187-induced contractions were virtually abolished by indomethacin and SQ29548, and slightly attenuated by CGS13080. These agents also attenuated the contractions elicited by endothelin but not by vasopressin. According to these data, a prostanoid(s) agonist for TxA2-PGH2 receptors contributes to the constrictor effect of A23187 in aortic rings of hypertensive rats, and of endothelin in aortic rings of normotensive and hypertensive rats. Moreover, the expression of prostanoid-mediated contractions as it pertains to the aortic response to A23187 is greatly increased in hypertensive rats.     

Effects of (+)-propranolol on intracellular mechanisms of contraction in striated muscle of the rabbit

Summary In functionally skinned muscle fibers from the rabbit, we studied the effect of propranolol on calcium activation of the contractile proteins and, in separate experiments, on calcium uptake and release from the sarcoplasmic reticulum (SR) while measuring physiological tension. Pieces from isolated papillary muscle (PM), soleus (SL) (slowtwitch skeletal muscle), and adductor magnus (AM) (fasttwitch skeletal muscle) were homogenized (sarcolemma disrupted). A fiber bundle from PM and single fibers from SL and AM were dissected from the homogenate and mounted on a photodiode tension transducer. To study Ca²⁺-activated tension development of the contractile proteins, we used high EGTA (7 mmol/l) to control the free calcium concentration. To study SR function, we used five different solutions to load the calcium into the SR and to release it from the SR with 25 mmol/l caffeine, thus producing a tension transient.In general, propranolol has similar mechanisms of action in the three muscle types. Propranolol (0.1–1.0 mmol/l) increased the submaximal calcium-activated tension development in all muscles but with PM=SL > AM, and this increase was correlated with increases in calcium binding to isolated troponin C. Propranolol increased the maximal calcium-activated tension development in PM and SL, but decreased that in AM. Propranolol at concentrations of 0.3–1.0 mmol/l decreased calcium uptake by the SR but did not change caclium release in any of the three muscles. In PM, however, propranolol at a concentration of 0.1 mmol/l increased calcium uptake by the SR. We conclude that propranolol induces decreases in muscle contraction mainly by decreasing calcium uptake by the SR.     

Effects of cyclopiazonic acid and ryanodine on cytosolic calcium and contraction in vascular smooth muscle.

1. In smooth muscle, both Ca2+ release from the sarcoplasmic reticulum (SR) and Ca2+ influx across the plasma membrane are responsible for the increase in the cytosolic Ca2+ level ([Ca2+]i). To understand further the role of SR on smooth muscle contraction, the effects of an inhibitor of the SR Ca2+ pump, cyclopiazonic acid (CPA 10 microM), an inhibitor of the Ca(2+) -induced Ca2+ release, ryanodine, (10 microM), and an activator of the Ca(2+) -induced Ca2+ release, caffeine (20 mM), on [Ca2+]i and contractile force were examined in the ferret portal vein loaded with a photoprotein, aequorin. 2. CPA induced a small increase in the aequorin signal reaching a maximum at 7 min. Several minutes after the increase in the aequorin signal, muscle tension increased reaching a maximum at 21.5 min. In contrast, ryanodine changed neither the aequorin signal nor contraction. In the presence of ryanodine, caffeine induced a sustained increase in the aequorin signal and transient contraction. After washing ryanodine and caffeine, the aequorin signal and muscle tone returned to their respective control levels. After treatment with ryanodine and caffeine, the second addition of caffeine was almost ineffective whereas CPA still increased the aequorin signal and muscle tension. 3. In the presence of external Ca2+, noradrenaline (NA, 10 microM) induced a transient increase followed by a sustained increase in the aequorin signal and sustained contraction. In contrast, KCl (70 mM) induced sustained increases in the aequorin signal and sustained contraction. In Ca(2+) -free solution, NA induced a small transient increase in the aequorin signal and a small transient contraction. These changes were inhibited in the presence of CPA or on pretreatment of the muscle with ryanodine and caffeine. These results suggest that CPA or ryanodine and caffeine depleted Ca2+ in SR. High K+ was ineffective in the absence of external Ca2+. 4. In the presence of external Ca2+ and CPA, NA and high K+ induced larger aequorin signals than in the absence of CPA, whereas the magnitude and shape of the contractions did not change. In contrast, pretreatment with ryanodine and caffeine did not have such an effect. In the muscle pretreated with ryanodine and caffeine, CPA changed the responses to high K+ and NA in a similar manner to that in the muscle without the pretreatment with ryanodine and caffeine. 5. Dissociation of contraction from [Ca2+]i as measured with aequorin suggests that NA and high K+ increase Ca2+ in two compartments: a compartment containing contractile elements (contractile compartment) and another compartment unrelated to contractile elements (non-contractile compartment). Because CPA augmented the stimulant-induced increase in aequorin signal without changing contraction, the non-contractile compartment may be located near the SR and the CPA-sensitive SR Ca2+ pump may regulate the Ca2+ level in this compartment. However, because CPA changed neither the magnitude nor shape of the contractions in the presence of external Ca2+, the SR Ca2+ pump may have little effect on regulation of Ca2+ level in the contractile compartment. Furthermore, the release of Ca2+ from SR seems to have little effect on the increase in the contractile Ca2+ because ryanodine and caffeine changed neither the aequorin signals nor contractions induced by NA and high K+ in the presence of external Ca2+ in the ferret portal vein.     

Cardiovascular responses to morphiceptin in spontaneously hypertensive and normotensive rats

In the urethane anesthetized spontaneously hypertensive rats (SHR) intraventricular injections of morphiceptin produced dose-related increase in heart rate and blood pressure. In contrary, intraventricular administration of morphiceptin in Wistar rats induced a fall in blood pressure and heart rate. Yohimbine antagonized pressor responses to morphiceptin in SHR. Naloxone counteracted both the stimulatory effects of morphiceptin in SHR as well as hypotensive responses in Wistar rats. Bilateral vagotomy blocked depressant action of morphiceptin in normotensive but failed to alter the pressor effects in SHR rats. After systemic injections of morphiceptin a fall in heart rate and blood pressure was obtained in both strains.