Heterogeneous distribution of adrenergic receptors in coronary arteries and their influence on coronary arterial tone.

In-vitro experiments, using canine large (o.d. 1-2 mm) and small (o.d. 0.4-0.8mm) coronary arteries, were undertaken to determine the functional state of alpha- and beta-adrenergic receptors and their role in regulation of coronary arterial tone. Both large and small coronary arteries of the dog possess alpha and beta adrenergic receptors. Alpha-adrenergic receptors are more numerous on the circumflex vessel than elsewhere. Functionally, beta-adrenergic receptors appear to be dominant under physiological conditions in most types of coronary arteries. In canine coronary arteries, beta-adrenergic receptors have a greater affinity for noradrenaline than adrenaline. Alpha-adrenergic receptors have a greater affinity for adrenaline than noradrenaline. It is concluded that after beta-adrenergic blockade, in both large and small coronary arteries, alpha-adrenergic receptors play an important role in regulation of coronary arterial tone.

Withdrawal of magnesium enhances coronary arterial spasms produced by vasoactive agents.

1 The influence of external magnesium ions ([Mg2+]o) on the sensitivity (i.e. EC50) and contractility (maximum response) of isolated large and small coronary arteries of the dog, obtained from different regions of the myocardium, to vasoactive agents was studied. 2 Removal of [Mg2+]o from the physiological salt solution enhanced, while elevation in [Mg2+]o to 4.8 mM, lowered the contractile sensitivity to three different agents, 5-hydroxytryptamine (5-HT), angiotensin II and KCl. 3 Contractility, of both large and small coronary arteries, to 5-HT and angiotensin II was potentiated and depressed, respectively, by withdrawal and elevation of [Mg2+]o; maximum responses to KCl were not altered by 0 or 4.8 mM [Mg2+]o. 4 Cumulative concentration-contractile effect curves to CaCl2 were shifted leftward on removal of [Mg2+]o; elevation of [Mg2+]o to 4.8 mM shifted the CaCl2 concentration-effect curves to the right. Maximal contractile responses to CaCl2 were enhanced by removal of, and reduced by elevation of, [Mg2+]o. 5 The calcium channel blocking agent, verapamil (10(-6)M), inhibited completely contractile responses to KCl; contractile responses elicited by angiotensin II and 5-HT were attenuated by verapamil. 6 A variety of pharmacological antagonists (phentolamine, propranolol, methysergide, atropine, diphenhydramine), as well as use of a prostaglandin cyclo-oxygenase inhibitor, did not modify the altered contractile responses evoked by angiotensin II or KCl in different concentrations of Mg2+. 7 These results suggest: (1) [Mg2+]o may exert considerably greater influence on receptor-operated rather than membrane-potential sensitive channels involved in Ca2+ transport in coronary arterial smooth muscle; (2) Mg2+ interferes with the affinity (binding) of certain agonists (5-HT and angiotensin II) for their respective receptors in coronary vascular muscle; and (3) a functional pool of Ca2+ which is resistant to Ca2+-depletion, but accessible to activation by 5-HT and angiotensin II is present in canine coronary arterial smooth muscle.

Ca2+ coupling in vascular smooth muscle: Mg2+ and buffer effects on contractility and membrane Ca2+ movements.

An examination of the literature, over the past two decades, reveals that (1) in studies of different types of vascular smooth muscles, Mg2+ is often either left out of physiological salt solutions or reduced in concentration compared with that in blood; and (2) when excitation--contraction coupling processes have been examined in isolated vascular tissues and cells, a number of artificial (synthetic) amine and organic zwitterion buffers have often been substituted for the naturally occurring bicarbonate and phosphate anions found in the blood and in cells. The influence of extracellular magnesium ions ([Mg2+]0) on tone, contractility, reactivity, and divalent cation movements in vascular smooth muscles, and how they may relate to certain vascular disease states, is reviewed. Data are presented and reviewed which indicate that many of the most commonly used artificial buffers (e.g., Tris, HEPES, MOPS, Bicine, PIPES, imidazole) can exert adverse effects on contractility and reactivity of certain arterial and venous smooth muscles. The data reviewed herein suggest that [Mg2+]0 and membrane Mg are important in the regulation of vascular tone, vascular reactivity, and in control of Ca uptake, content, and distribution in smooth muscle cells. [HCO3-]0 and (or) PO4(2-) anions may be important for normal maintenance of excitability and reactivity and in the control of Ca uptake, content, and distribution in smooth muscle cells.

Propranolol induces contractions of canine small and large coronary arteries.

Using isolated canine small (right coronary branch, left coronary branch; o. d. 0.4-0.8 mm) and large (left coronary, circumflex; o. d. 1-2 mm) coronary arteries, the beta-adrenergic antagonist dl-propranolol (5 X 10(-7) to 5 X 10(-5) m/l) was found to produce concentration-dependent contractions. Interestingly, most of these contractile events take place with concentrations of propranolol (0.1-1 microgram/ml) found in the blood of patients who are taking this drug for various therapeutic reasons. These propranolol-induced contractions were enhanced in Krebs-Ringer solution containing slightly elevated (weak contractile) concentrations of potassium (15 mmol/l). Experiments with specific pharmacologic antagonists indicated that propranolol-induced contractions on canine coronary arteries can not be mediated by release (or inhibition) of catecholamines, histamine, serotonin or acetylcholine. Propranolol contractions could be released by low concentrations of potassium ions (4 mmol/l), suggesting that the beta receptor antagonist might inactive coronary arterial membrane Na+, K+-ATPase. Other experiments demonstrated that propranolol can enhance coronary arterial membrane permeability to calcium ions; these observations suggest that propranolol might sensitize coronary vascular smooth muscle cells to calcium ions. Removal of calcium ions from the Krebs-Ringer solution or addition of the calcium entry blocker, verapamil, prevented completely the propranolol-induced contractions. Catecholamines (i.e., epinephrine, norepinephrine, isoproterenol), which normally induce relaxation on these isolated coronary arteries, always induced contraction after use of dl-propranolol. Overall, these experiments suggest that the so-called "beta-blocker poisoning" sometimes noted with propranolol in patients might be brought about by four actions of this drug acting in concert: 1. direct coronary arterial vasospasm; 2. an unmasking of normally silent alpha-adrenergic receptors, thus allowing circulating and released catecholamines to induce potent coronary constriction; 3. attenuation of membrane Na+, K+-ATPase activity, and 4. an enhancement of coronary vascular smooth muscle membrane permeability to calcium ions.

Interactions of magnesium and verapamil on tone and contractility of vascular smooth muscle.

Previous studies on isolated blood vessels indicate that (1) withdrawal of magnesium ([Mg2+]0) induces calcium-dependent contractile responses, and (2) Mg2+]0 and verapamil (VE) inhibit calcium influx across the cell membrane. The present study, using isolated rat aortic strips and portal veins, was designed to assess the interactions of [Mg2+]0 and VE on increases in active tension and contractility induced by withdrawal of [Mg2+]0.[Mg2+]0 was found to: (1) enhance VE-induced inhibition of portal vein amplitude, and (2) antagonize VE-induced enhanced frequency of spontaneous phasic contractions in portal veins. Both [Mg2+]0 and VE could prevent and reverse the increases in active tension developed in aortic smooth muscle upon removal of [Mg2+]0.[Mg2+]0 markedly potentiated the inhibitory effect of verapamil on calcium-induced contractions of K+-depolarized aorta but not in K+-depolarized portal vein. This synergistic effect in aortal could be due to the influence of Mg and VE at the membrane and/or the influence of Mg at intracellular sites resulting in decreased levels of functional (activator) cytoplasmic Ca2+. On the other hand, the lack of synergism in portal vein could indicate that both of these agents might be inactivating the same membrane channels involved in calcium influx.

Sequential changes in serum glucose, triglycerides and cholesterol in aging of normal and alloxan-diabetic rats.

16-week-old Wistar, alloxan diabetic rats exhibited progressive elevations in levels of serum glucose, total triglycerides, cholesterol and creatinine over a period of 8 weeks; hyperglycemia preceded hyperlipidemia and hypercreatininemia and hypertriglyceridemia preceded hypercholesteremia. Age-matched control rats failed to develop any signs of hyperglycemia or hypercreatininemia, but did develop both hypercholesterolemia and hypertriglyceridemia at 24 weeks of age. This suggests that the progressive cardiovascular derangements (e.g., atherosclerosis, hypertension) noted in experimental diabetes mellitus and in the normal aging (and maturation) process may be brought about by distinctly different biochemical processes.

Hypomagnesemia and vasoconstriction: possible relationship to etiology of sudden death ischemic heart disease and hypertensive vascular diseases.

In-vitro experiments are presented which indicate that the concentration of extracellular magnesium ions ([Mg2+]o) can exert profound influences on the contractility and reactivity of arteries, arterioles and veins from a number of regional vasculatures in several mammalian species, including man. Hypomagnesemia can potentiate the contractile activity of a variety of neurohumoral substances and induce vasospasm. Hypermagnesemia can do the reverse, i.e., induce hyporeactivity, relaxation and vasodilatation. Data are also presented to indicate that [Mg2+]o can control the entry, distribution and exit of calcium ions (Ca2+) from vascular smooth muscle cells. Arterial and venous smooth muscles excised from rats with alloxan-diabetes mellitus or spontaneous hypertension (SHR) appear to exhibit vascular membranes which have modifications in their Mg-Ca exchange sites. Data are reviewed which suggest that certain vascular diseases (e.g., sudden-death ischemic heart disease, hypertension, eclampsia, diabetes mellitus) are associated with a Mg-deficiency. Overall, it is suggested that [Mg2+]o and membrane [Mg] may play critical roles in regulating vascular tone and homeostasis.

Autonomic regulation of cardiovascular function in neonates.

The degree of participation of the vascular bed supplied by the superior mesenteric artery in autonomic regulation of cardiovascular function during postnatal maturation was evaluated in piglets, ranging in age from birth to two months. Animals were anaesthetized with 0.25-0.5% halothane in a 50% mixture of N2O and O2, paralysed with decamethonium bromide and artificially ventilated to maintain normal arterial blood gases and pH. Mesenteric arterial flow was recorded continuously with an electromagnetic flow transducer; simultaneously aortic pressure, heart rate, and femoral, renal and carotid flows were monitored. Resistance was calculated as the ratio of mean aortic pressure to mean flow. The central neural regulation of the cardiovascular system was altered: (1) by electrical stimulation of medullary vasoactive sites, (2) by changing the input from visceral or somatic afferents and (3) by subjecting the animals to stress, such as haemorrhage or hypoxia. Postnatal development of adrenergic mechanisms was studied with the aid of adrenergic agonist both in vivo and in vitro (helical strips cut from superior mesenteric artery were placed in an organ bath for isometric tension recordings). Recordings of spontaneous efferent splanchnic discharge permitted a more direct examination of autonomic regulation of the mesenteric vascular bed. The results indicate that the vascular bed supplied by the superior mesenteric artery and innervated by the efferent splanchnic nerve is actively involved in the cardiovascular responses to alterations in the cardiovascular regulatory system. Furthermore, this vascular bed may have an inappropriate response to both feeding and stress such that pathological changes in the gastrointestinal tract may result.

Vascular responsiveness and serum biochemical parameters in alloxan diabetes mellitus.

Responses of isolated aorta and portal vein (PV) to norepinephrine (NE), angiotensin II (AII), KCl, and CaCl2 were investigated in alloxan diabetic rats. Based on serum biochemical parameters (i.e., glucose, cholesterol, triglycerides, and creatinine) (alloxan, 150 mg/kg), diabetic rates were divided into three groups: 1) mildly diabetic at 1 week (only elevated glucose levels), 2) moderately diabetic at 4 wk (elevated glucose, triglycerides, and creatinine), and 3) severely diabetic at 8 wk (all serum biochemical parameters elevated). The sensitivity (i.e., ED50) of aortic smooth muscle from diabetic rats when compared to saline controls was 1) unchanged in mild diabetes; 2) decreased to KCl, AII, and CaCl2 in moderate diabetes; and 3) decreased to KCl, NE, and CaCl2 in severe diabetes. Ability of aortic smooth muscle to develop maximal contractions (i.e., contractility) to all these agonists was markedly diminished in severe diabetes. Spontaneous phasic contractions of PV from diabetic rats exhibited progressively greater tension as the disease advanced. Unlike aortas, contractility of PV to vasoactive agents was not affected at any stage of diabetes. PV sensitivity to AII in moderate diabetes and to Ca in severe diabetes was decreased when compared to saline controls. These differences in reactivity and contractility of aorta and PV in progressive stages of experimental diabetes could be due to alterations in calcium handling and its metabolism in arterial and venous smooth muscle cells in the diabetic state.

Ca2+ uptake and distribution in alloxan-diabetic rat arterial and venous smooth muscle.

This report demonstrates that in experimental diabetes mellitus (DM) calcium uptake and its distribution is altered in rat aortic but not in portal venous smooth muscle. Results are interpreted as consequences of increased calcium binding by aortic smooth muscle in experimental DM, which could account for the hyporeactivity of alloxan diabetic rat aorta reported previously.

Adverse effects of artificial buffers on contractile responses of arterial and venous smooth muscle.

1 In vitro studies were undertaken on rat aortic strips and portal vein segments in order to determine whether or not several commonly used artificial buffers, i.e., tris(hydroxymethyl) aminomethane (Tris), N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid (HEPES), morpholine propanesulphonic acid (MOPS), N,N bis(2-hydroxyethyl) glycine (BICINE) and 1,4-piperazinediethanesulphonic acid (PIPES), can exert direct actions on vascular smooth muscle. 2 All artificial buffers used in 5 mM concentrations were found to inhibit development of spontaneous mechanical activity. 3 Tris, HEPES, MOPS, BICINE and PIPES markedly attenuated contractions induced by adrenaline, angiotensin and KCl. The fast phase components of the agonist-induced contractions were either obliterated or reduced in the presence of the artificial buffers. The sustained slow phase components were greatly reduced and retarded by the artificial buffers. 4 The relative order of artificial buffer potency (i.e., from 100% to 14% inhibition) seems to depend upon the agonist and type of smooth muscle. 5 All of these inhibitory effects were reversible, since normal contractile responses and spontaneous mechanical activity could be obtained by simply reincubating the smooth muscles in Krebs-Ringer bicarbonate buffer. 6 A variety of pharmacological antagonists failed to mimic or affect the inhibitory effects of Tris, HEPES, MOPS, PIPES and BICINE. 7 These data show that five of the most commonly used artificial buffers, to study muscles in vitro, exert adverse effects on contractility of arterial and venous smooth muscle.

Magnesium deficiency produces spasms of coronary arteries: relationship to etiology of sudden death ischemic heart disease.

Isolated coronary arteries from dogs were incubated in Krebs-Ringer bicarbonate isolation and exposed to normal, high, and low concentrations of magnesium in the medium. Sudden withdrawal of magnesium from the medium increased whereas high concentrations of magnesium decreased the basal tension of the arteries. The absence of magnesium in the medium significantly potentiated the contractile responses of both small and large coronary arteries to norepinephrine, acetylcholine, serotonin, angiotensin, and potassium. These data support the hypothesis that magnesium deficiency, associated with sudden death ischemic heart disease, produces coronary arterial spasm.

Pentobarbital sodium inhibits calcium uptake in vascular smooth muscle.

This report demonstrates that the commonly used anesthetic agent, pentobarbital sodium, in concentrations of 1 . 10(-4) to 2 . 10(-3) M inhibits calcium (Ca2+) uptake in both rat aortic and portal venous smooth muscle. The data indicate that total exchangeable Ca2+ in portal vein is reduced by about 15% in 1 . 10(-4) M pentobarbital sodium, while the intracellular exchangeable Ca2+ is reduced by 24%. On the other hand, in aortic smooth muscle, while 5--20 . 10(-4) M pentobarbital sodium reduces total exchangeable Ca2+ by about 15%, intracellular Ca2+ is reduced by 22% in 5 . 10(-4) M pentobarbital sodium and by 38% in 2 . 10(-3) M pentobarbital sodium. The present studies thus reveal that concentrations of pentobarbital sodium known to be present during induction of surgical anesthesia can exert significant inhibitory effects on exchangeability and transmembrane movement of Ca2+ in at least two different types of blood vessels.

Ethanol reduces Ca2+ concentrations in arterial and venous smooth muscle.

The present results, using isolated rat aortic strips and portal vein segments, demonstrate that ethanol (170--430 mM) significantly inhibits calcium uptake in these 2 different types of vascular smooth muscle.

Tris(hydroxymethyl)aminomethane inhibits calcium uptake in vascular smooth muscle.

This report demonstrates that the commonly used buffering agent Tris(hydroxymethyl)aminomethane (Tris) in concentrations of 5 and 30 mM inhibits calcium (Ca2+) uptake in both rat aortic and portal venous smooth muscle. The data indicates that total exchangeable Ca2+ in portal vein is reduced by about 35% in 5 or 30 mM Tris, while the intracellular exchangeable Ca2+ is not significantly altered. On the other hand, in aortic smooth muscle, while 30 mM Tris reduces total exchangeable Ca2+ by about 20%, intracellular Ca2+ is reduced by 44% in 5 mM Tris and by 55% in 30 mM Tris. The present studies, thus, reveal that Tris exerts significant inhibitory effects on exchangeability and transmembrane movement of Ca2+ in at least two different types of smooth muscle.

Influence of tris on contracile responses of isolated rat aorta and portal vein.

The influence of tris(hydroxymethyl)-aminomethane (Tris) on the spontaneous mechanical activity (SMA) and agonist-induced contractile responses of rat portal vein and aorta was investigated. Tris, in a concentration-dependent manner (5, 10, and 30 mM), significantly increased frequency and attenuated amplitude of SMA in portal vein. Tris, in all concentrations, abolished spontaneous activity of aorta. Tris (5 mM) attenuated epinephrine-, angiotensin-, and potassium-induced contractile responses in portal vein and in aorta: the vein responses exhibited the greater sensitivity to Tris inhibition. In the presence of Tris, agonist dose-response curves were shifted to the right, concomitant with a reduction in maximum tension in portal vein. In contrast, in aorta, only a rightward shift of the dose-response curves was observed, withour any change in maximum tensions in the presence of Tri. Tris (5 mM) depressed contractions induced by calcium in potassium-depolarized aortic and venous smooth muscle. These results suggest that Tris may interfere with the binding, translocation, and utilization of calcium ions at or beyond the membrane in venous smooth muscle and at the membrane in aortic smooth muscle.

Morphine-induced supersensitivity to acetylcholine in skeletal muscle and its mechanism of action.

The effect of in vitro morphine pretreatment (250 micron) on the sensitivity of skeletal muscle to acetylcholine (ACh) was investigated using the isolated rectus abdominis muscle of frog. Morphine (15 to 120 min) significantly increased the sensitivity of the tissue to ACh. This increase in the sensitivity was found to be greatest after 120 min. Morphine significantly increased the sensitivity of the tissue to carbachol only after 120 min. In the presence of physostigmine, morphine did not cause further augmentation of responses to ACh. Potassium chloride-induced contractile responses were not influenced by morphine. In the presence of morphine, maximum contractile responses to ACh were significantly decreased. Highly calcium (1.62 mM) in the medium decreased the magnitude of the morphine-induced supersentivity to ACh at 120 min and antagonised the decrease in maximum contractile responses to ACh. These results suggest that morphine is causing supersensitivity by both its anticholinesterase activity and its inhibitory action on presynaptic nerve terminals.