Sympathetic,muscarinic vasodilation in cranial vessels of the cat

Summary Pressor responses evoked by stimulation of the preganglionic sympathetic trunk of the feline superior cervical ganglion have been recorded in vivo from the vascular bed perfused by one external carotid and the vertebral artery. When vasoconstrictor activity is blocked and potential vasodilator activity enhanced by close, intracarotid injection of guanethidine and prostaglandin F₂ respectively, stimulation evokes a weak pressor response followed, on cessation of stimulation, by a prolonged vasodilation lasting for 6–8 min. The magnitude and duration of the poststimulation vasodilation was reduced significantly by atropine. Due to the prolonged nature of the vasodilation, it is unlikely that a sympathetic cholinergic vasodilation in the classical sense is involved.     

Cyclic nucleotide-dependent relaxation pathways in vascular smooth muscle

Vascular smooth muscle tone is controlled by a balance between the cellular signaling pathways that mediate the generation of force (vasoconstriction) and release of force (vasodilation). The initiation of force is associated with increases in intracellular calcium concentrations, activation of myosin light-chain kinase, increases in the phosphorylation of the regulatory myosin light chains, and actin-myosin crossbridge cycling. There are, however, several signaling pathways modulating Ca²⁺ mobilization and Ca²⁺ sensitivity of the contractile machinery that secondarily regulate the contractile response of vascular smooth muscle to receptor agonists. Among these regulatory mechanisms involved in the physiological regulation of vascular tone are the cyclic nucleotides (cAMP and cGMP), which are considered the main messengers that mediate vasodilation under physiological conditions. At least four distinct mechanisms are currently thought to be involved in the vasodilator effect of cyclic nucleotides and their dependent protein kinases: (1) the decrease in cytosolic calcium concentration ([Ca²⁺]c), (2) the hyperpolarization of the smooth muscle cell membrane potential, (3) the reduction in the sensitivity of the contractile machinery by decreasing the [Ca²⁺]c sensitivity of myosin light-chain phosphorylation, and (4) the reduction in the sensitivity of the contractile machinery by uncoupling contraction from myosin light-chain phosphorylation. This review focuses on each of these mechanisms involved in cyclic nucleotide-dependent relaxation of vascular smooth muscle under physiological conditions.     

Sympathetic, muscarinic vasodilation in cranial vessels of the cat

Pressor responses evoked by stimulation of the preganglionic sympathetic trunk of the feline superior cervical ganglion have been recorded in vivo from the vascular bed perfused by one external carotid and the vertebral artery. When vasoconstrictor activity is blocked and potential vasodilator activity enhanced by close, intracarotid injection of guanethidine and prostaglandin F2 alpha respectively, stimulation evokes a weak pressor response followed, on cessation of stimulation, by a prolonged vasodilation lasting for 6-8 min. The magnitude and duration of the poststimulation vasodilation was reduced significantly by atropine. Due to the prolonged nature of the vasodilation, it is unlikely that a sympathetic cholinergic vasodilation in the classical sense is involved.     

P2Y<Subscript>2</Subscript> receptor modulates shear stress-induced cell alignment and actin stress fibers in human umbilical vein endothelial cells

Endothelial cells release ATP in response to fluid shear stress, which activates purinergic (P2) receptor-mediated signaling molecules including endothelial nitric oxide (eNOS), a regulator of vascular tone. While P2 receptor-mediated signaling in the vasculature is well studied, the role of P2Y₂ receptors in shear stress-associated endothelial cell alignment, cytoskeletal alterations, and wound repair remains ill defined. To address these aspects, human umbilical vein endothelial cell (HUVEC) monolayers were cultured on gelatin-coated dishes and subjected to a shear stress of 1 Pa. HUVECs exposed to either P2Y₂ receptor antagonists or siRNA showed impaired fluid shear stress-induced cell alignment, and actin stress fiber formation as early as 6 h. Similarly, when compared to cells expressing the P2Y₂ Arg-Gly-Asp (RGD) wild-type receptors, HUVECs transiently expressing the P2Y₂ Arg-Gly-Glu (RGE) mutant receptors showed reduced cell alignment and actin stress fiber formation in response to shear stress as well as to P2Y₂ receptor agonists in static cultures. Additionally, we observed reduced shear stress-induced phosphorylation of focal adhesion kinase (Y397), and cofilin-1 (S3) with receptor knockdown as well as in cells expressing the P2Y₂ RGE mutant receptors. Consistent with the role of P2Y₂ receptors in vasodilation, receptor knockdown and overexpression of P2Y₂ RGE mutant receptors reduced shear stress-induced phosphorylation of AKT (S473), and eNOS (S1177). Furthermore, in a scratched wound assay, shear stress-induced cell migration was reduced by both pharmacological inhibition and receptor knockdown. Together, our results suggest a novel role for P2Y₂ receptor in shear stress-induced cytoskeletal alterations in HUVECs.     

Occurrence of inhibitory histamine H2-receptors in isolated pulmonary blood vessels of dogs and rats

Summary Isolated, helically cut strips of pulmonary arteries and veins of dogs, and pulmonary arteries of rats, precontracted with norepinephrine or 5-HT exhibited potent concentration-dependent relaxations to impromidine and dimaprit (histamine H₂-agonists). The occurrence of inhibitory histamine H₂-receptors in the pulmonary vasculature could play a modulatory role in hypoxic pulmonary vasoconstriction.Supported by research grants NHLBI-18002, NHLBI-18015 and DA-02339 from the USPHS. We are grateful to Dr D.D.A. Owen of Smith, Kline & French Ltd, England for the generous supply of dimaprit and impromidine. Requests for reprints should be addressed to B.M. Altura.     

Altered brain cyclic AMP-responses in rats reared in enriched or impoverished environments

Summary The accumulation of radioactive cyclic AMP elicited by various neurohormones has been examined in adenine-labeled telencephalon slices from rats raised in enriched or impoverished environments. Basal levels of cyclic AMP and responses of the brain slice cyclic AMP-generating systems to norepinephrine, isoproterenol and adenosine did not differe between the two group of rats, while responses to prostaglandin E₁ were significantly greater with the impoverished group and responses to histamine appeared to be greater with the enriched group.     

Corticotropin releasing factor increases the adrenocortical responsiveness to adrenocorticotropin

Summary In the course of studying the plasma adrenocorticotropic hormone (ACTH) and corticosterone responses to synthetic corticotropin releasing factor (CRF), we noted some disparity in the responses. A higher dose (20 g compared with 5 g per rat i.a.) produced an equal plasma ACTH but greater plasma corticosterone response in adult male rats. Thus, we examined the possibility that CRF increases adrenocortical responsiveness to ACTH. CRF significantly (p<0.0005) increased the plasma corticosterone response to ACTH in rats pretreated with dexamethasone. Thus, synthetic CRF increases corticosterone secretion in rats not only by stimulating ACTH secretion, but also by increasing the adrenocortical responsiveness to ACTH.Acknowledgments. This research was supported by grant MT-5183 from the Medical Research Council of Canada and by the Department of Medicine, University of Kentucky.     

The effects of thermogenic agents on hindlimb oxygen consumption in the dog: ICI D7114 and noradrenaline

The thermogenic action of beta-adrenoceptor agonists may be due, in part, to increased metabolism in skeletal muscle. Previous results suggest that vasoconstriction is also necessary, and that the effect can be blocked by vasodilators. Both noradrenaline and the beta-3 agonist, ICI D7114, were studied using two dog hindlimb protocols. During constant perfusion conditions, ICI D7114 caused a significant increase in hindlimb oxygen consumption although it is a vasodilator. Noradrenaline resulted in a smaller rise in oxygen consumption, and produced a marked vasoconstriction. Both noradrenaline and ICI D7114 resulted in decreased oxygen consumption when the blood flow was allowed to vary in response to the drug treatment. The results suggest that changes in tissue oxygen consumption caused by beta-agonists are not related to vasomotion.     

Increase in Na+, K+ -ATPase enzyme units in liver and kidneys from essential fatty acid deficient rats.

(3H)-Ouabain binding to liver and kidney preparations was utilized to estimate the number of Na+, K+-ATPase enzyme units in livers and kidneys from rats fed 2% corn oil supplemented or fat-free diets. The specific (3H)-ouabain binding in liver and kidney preparations from fatty acid deficient rats was increased approximately 40%, but the affinity of the binding sites for ouabain (Kd-value) remained unchanged. The increased concentration of Na+, K+-ATPase enzyme units observed in the essential fatty acid deficient rats may contribute to the reduced body fat accumulation and elevated heat production observed in these animals.     

Effects of thyroid state on brain stem responses to iontophoretic noradrenaline

Summary Responses to iontophoretic NA were increased in hyperthyroid rats and decreased in hypothyroid animals. No effect was observed on acetylcholine and glycine responses tested in the same way.This work was aided by grants from Consejo de Desarrollo Cientifico y Humanistico U.C.V. and CONICIT proyecto 51.26.SI-086.     

Alteration of mitochondrial bioenergetics due to intravenous injection of a perfluorocarbon emulsion

Wistar albino rats were intravenously injected with 1 ml of an oxyphoretic emulsion of perfluorobutylfurane and killed 3, 7 or 30 days later. Mitochondria isolated from the liver and kidneys of treated rats showed a small decrease in the transmembrane electrical potential and a substantial depression of the rates of both ATP synthesis and ADP-stimulated respiration. These alterations in mitochondrial oxidative phosphorylation appear to be induced by perfluorocarbon and/or tensioactive molecules interacting with hydrophobic cell structures.     

Über den Vitamin-B12-Gehalt von Organen verschieden ernährter Ratten

Summary Adult, male rats bred for over 10 generations on a soy meal-corn-diet had vitamin B₁₂ values of liver and kidney about 10 times lower than the controls. If the deficient ration was supplemented with 5µg/kg of B₁₂, these values were still about 1/5 of the controls. 3 weeks on the deficient diet lowered the B₁₂ levels in the livers and kidneys of previously undepleted rats to about 1/2, and a similar diet containing 0.1% of iodized casein lowered these levels to about 1/3 of the normal values but did not lower the B₁₂-concentration of organs of already deficient rats.Rats bred on the deficient diet and receiving for 1 month a supplement of 30µg/kg of vitamin B₁₂ or the stock diet with a similar B₁₂-content, had normal B₁₂-levels in livers and kidneys.     

Influence du cadmium sur le métabolisme du calcium. Action sur le cytochrome P450

Summary Oral administration of cadmium to female rats for 6 weeks does not reduce the microsomal cytochrome P₄₅₀ levels in the liver and kidneys, nor the cytochrome P₄₅₀ content in the renal mitochondria.     

Nonsympathetic dilator innervation of cat cerebral arteries.

The results presented provide strong support for the presence of vasodilator innervation in the cat cerebral arteries. The dilator innervation is neither adrenergic nor cholinergic and does not originate in the superior cervical ganglia. The nature of the vasodilator transmitter is unidentified. Such innervation, however, may be involved in the regulation of cerebral blood flow, especially in view of the capability of some cat cerebral vessels to develop intrinsic muscle tone.     

Positive and negative inotropic effects of carbachol on the embryonic chick atrium

The effect of carbachol on twitch tension of atrial preparations from chick embryos of different incubation ages (3–14 days) was studied. At every age carbachol evoked negative (at low concentrations) and positive (at higher concentrations) inotropic responses. Maximal response values for both effects increased with age; in 3- and 5-day atria the positive inotropic response prevailed. The muscarinic antagonist pirenzepine inhibited the positive (on 5-day atria) and negative (on strips of 14-day atria) inotropic effects of carbachol with pA₂ values of 6.8 and 8.0, respectively, suggesting that muscarinic receptors mediating these effects belong to different receptor subtypes.     

Dynamic insulin secretion from perifused rat pancreatic islets

Insulin secretion from isolated pancreatic islets of 8- to 12-day-old rats was investigated in a dynamic in vitro (perifusion) system. The aims of the study were (i) to describe a carefully controlled in vitro method to study the mechanism of insulin secretion and to analyse the effects and dynamic interactions of bioactive compounds on isolated rat pancreatic islets, (ii) to validate the method by comparing fundamental data on the functions of the islets obtained with this method to those collected with other techniques; and (iii) to find novel features of the control of insulin secretion. The method was carefully designed to maintain the functional capacity of the explanted cells. A functional standardization system was elaborated consisting of (i) analysis of the changes in the basal hormone secretion of the cells; (ii) evaluating responses to a standard, specific stimuli (50 mM glucose for 3 min); (iii) determining the alteration of the momentary size of the hormone pool with responses to KCl; and (iv) direct determination of the total intracellular hormone content from the extract of the column. The technique provides accurate quantitative data on the dynamic responses to biologically active compounds that act directly on the pancreatic islets. The islets maintained their full responsiveness for up to 7 days, and responses as close as in 1-min intervals could be distinguished. A linear dose-response relationship was found on the glucose-induced insulin release in case of 3-min stimulation with 4 and 500 mM of glucose (lin-log graph). Utilizing this method, we showed that no desensitization to glucose-induced insulin release can be observed if the responsiveness of the cells is properly maintained and the parameters of the stimulation are carefully designed. Exposure of the explanted islets to 10 μM acetylcholine or 30 mM arginine (Arg) induced a transitory elevation of insulin release similar in shape to that experienced after glucose stimulation. Norepinephrine (NE), dopamine (DA) and somatostatin (SS) did not induce any detectable alteration on the basal insulin secretion of the islets. However, 100 nM SS given together with 50 mM glucose, 30 mM Arg or 10 μM acetylcholine significantly reduced the insulin-releasing effect of these substances (by 75.5, 71.5 and 72.5%, respectively). At the same time, SS did not alter the insulin response of the islets to 100 mM elevation of K⁺ concentration. SS also inhibited glucose-induced insulin release in a dose-dependent way (ED₅₀ = 22 nM). A similar dose-dependent inhibitory effect on glucose-induced insulin release was found with NE (ED₅₀ = 89 nM) and DA (ED₅₀ = 2.2 μM). γ-Aminobutyric acid (GABA) did not influence insulin release under similar circumstances. Received 16 January 1998; received after revision 6 May 1998; accepted 8 May 1998     

Reduction of bovine pulmonary hypertension by normoxia,verapamil and hexaprenaline

Summary In calves with hypoxic pulmonary hypertension, resistance was reduced by 40±3% with normoxia, 19±4% with verapamil, and 60±1% with hexoprenaline. It is possible that the increased resistance during normoxia is due partly to vasoconstriction rather than solely to vascular thickening, and that the vasoconstriction is due to an abnormality in calcium metabolism by the hypertensive vasculature.Acknowledgments. We thank Rosann Glas, Mary Munroe, Diann Smith and Linda Panepinto for technical assistance. This work was supported by the Colorado Heart Association and by the National Institutes of Health Grants HL-05973 and HL-29237. Verapamil (Isoptin) was supplied by Knoll Pharmaceutical Co. Hexoprenaline (ST 1512 HCl, Ipradol) was supplied by Österreichische Stickstoffwerke Linz.     

Altered brain cyclic AMP-responses in rats reared in enriched or impoverished environments.

The accumulation of radioactive cyclic AMP elicited by various neurohormones has been examined in adenine-labeled telencephalon slices from rats raised in enriched or impoverished environments. Basal levels of cyclic AMP and responses of the brain slice cyclic AMP-generating systems to norepinephrine, isoproterenol and adenosine did not differ between the two group of rats, while responses to prostaglandin E1 were significantly greater with the impoverished group and responses to histamine appeared to be greater with the enriched group.     

Changes with hypertension and maturation in the response of stomach fundus to acetylcholine

The purpose of these experiments was to compare the contractile response to ACh of stomach fundal strips from hypertensive (SHR) and normotensive (WKY) rats during the development of hypertension. The results indicate that the reactivity to ACh is the same in fundal strips from young SHR and WKY rats; however, with maturation strips from WKY rats undergo a reduction in responsiveness which does not occur in the SHR. Therefore, strips from older SHR rats are more reactive to ACh than are those from age matched WKY rats.     

Phosphocreatine and ATP concentrations increase during flow-stimulated metabolism in a non-contracting muscle

The gracilis muscle was excised from cold-acclimated rats, placed in vitro, and simultaneously perfused via its artery by high pO₂ medium and superfused by low pO₂ medium. With a doubling of the perfusion rate (from 50 to 100 μl/min) phosphocreatine and ATP increased by 39% and 44%, respectively.