High Concentrations Of Noradrenaline Research Articles

EARLY ENDOTOXIC SHOCK RESULTS IN ENHANCED VASODILATOR RESPONSES TO NITROGLYCERIN BUT UNALTERED RESPONSES TO NEUROPEPTIDES CALCITONIN GENE-RELATED PEPTIDE AND SUBSTANCE P

To determine the role that vasoactive neuropeptides, calcitonin gene-related peptide, and substance P play in tissue-blood flow regulation during early septic shock, we examined the responsiveness of arteries removed from pigs 3 h after administration of Escherichia coli lipopolysaccharide or saline vehicle. The carotid, cranial mesenteric, and left anterior descending coronary arteries were excised, and rings were cut from each vessel. Constrictor responses were obtained to cumulative doses of norepinephrine or potassium chloride. Rings were reconstricted and challenged with acetylcholine, substance P, calcitonin gene-related peptide, and nitroglycerin. Lipopolysaccharide significantly increased the cranial mesenteric artery's response to high concentrations of norepinephrine and the response to nitroglycerin in all vessels. This enhancement of responses to nitroglycerin suggests augmented smooth-muscle responsiveness to an exogenous source of nitric oxide, possibly associated with early depression of basal endothelial function. Depression of agonist-induced nitric oxide release may mask such enhancement with endothelial-dependent dilators and may enhance the response to adrenergic constrictors in some vascular beds.

Cloning, expression, and localization of a chloride-facilitated, cocaine-sensitive serotonin transporter from Drosophila melanogaster.

We report here on the isolation and characterization of a serotonin (5HT) transporter from Drosophila melanogaster. A 3.1-kb complementary DNA clone (dSERT) was found to encode a protein of 622 amino acid residues with a predicted molecular mass of approximately 69 kDa and a putative transmembrane topology characteristic of cloned members of the mammalian Na+/Cl- neurotransmitter cotransporter gene family. dSERT displays highest overall amino acid sequence identity with the mammalian 5HT (51%), norepinephrine (47%), and dopamine (47%) transporters and shares with all transporters 104 absolutely conserved amino acid residues. Upon transient expression in HeLa cells, dSERT exhibited saturable, high-affinity, and sodium-dependent [3H]5HT uptake with estimated Km and Vmax values of approximately 500 nM and 5.2 x 10(-18) mol per cell per min, respectively. In marked contrast to the human SERT (hSERT), 5HT-mediated transport by dSERT was not absolutely dependent on extracellular Cl-, while the sodium-dependent uptake of 5HT was facilitated by increased extracellular Cl- concentrations. dSERT displays a pharmacological profile and rank order of potency consistent with, but not identical to, mammalian 5HT transporters. Comparison of the affinities of various compounds for the inhibition of 5HT transport by both dSERT and hSERT revealed that antidepressants were 3- to 300-fold less potent on dSERT than on hSERT, while mazindol displayed approximately 30-fold greater potency for dSERT. Both cocaine and RTI-55 inhibited 5HT uptake by dSERT with estimated inhibition constants of approximately 500 nM, while high concentrations (> 10 microM) of dopamine, norepinephrine, octopamine, tyramine, and histamine failed to inhibit transport. In situ hybridization reveals the selective expression of dSERT mRNA to specific cell bodies in the ventral ganglion of the embryonic and larval Drosophila nervous system with a distribution pattern virtually identical to that of 5HT-containing neurons. The dSERT gene was mapped to position 60C on chromosome 2. The availability of the gene encoding the unique ion dependence and pharmacological characteristics of dSERT may allow for identification of those amino acid residues and structural motifs that confer the pharmacologic specificity and genetic regulation of the 5HT transport process.

Cardiorespiratory and stress hormone responses during first dose surfactant administration in neonates with RDS

Our hypothesis was that surfactant instilled into the trachea, followed by body positioning maneuvers utilized to enhance drug distribution, could alter hemodynamic function and stimulate the release of catecholamines. We conducted a prospective randomized study designed to compare the immediate physiologic effects of the first dose of Exosurf Neonatal (5 mL/kg; n = 16) or Survanta (4 mL/kg; n = 18), when surfactant administration was standardized with strict adherence to drug company protocol. Physiologic variables were monitored continuously. Arterial blood gases (ABG) and plasma catecholamine concentrations were measured before, and 5 minutes after, surfactant administration. Both surfactants had an immediate effect on arterial oxygen saturation (SaO2), partial pressure of oxygen in arterial blood (PaO2), and oxygen index (OI). The improvement in oxygenation after surfactant therapy was similar in both groups. There was no significant difference in the mean umbilical arterial blood pressure (ABP) following surfactant therapy in both groups. High concentrations of plasma norepinephrine (reflecting activity of the sympathetic nerves) and epinephrine (a measure of secretion from the adrenal medulla) indicate that preterm infants with respiratory distress syndrome (RDS) prior to treatment mount a substantial stress response. The currently recommended techniques for instillation of surfactant appear not to trigger a significant further surge of plasma catecholamines or to acutely change mean ABP. Alternatively, it may be possible that the lack of response was because catecholamine release was already maximal.

Myocardial work load is a major determinant of norepinephrine-induced left ventricular dysfunction

This study was conducted to determine whether increased myocardial energy demand plays a role in norepinephrine (NE)-induced left ventricular (LV) dysfunction. A range of arterial pressure-heart rate (P-R) products (myocardial energy demand) was produced in both conscious and pentobarbital sodium-anesthetized rabbits with the same dose of NE (10 micrograms priming bolus plus 2.5 micrograms.kg-1 x min-1 for 2.5 h). After NE treatment, LV function was evaluated in vitro and found to be markedly diminished in the rabbits that had an elevated P-R product. In contrast, LV function was not significantly affected when the P-R product was maintained near control levels during NE treatment. In separate experiments, rabbit hearts were isolated and exposed to NE (10,000 or 50,000 pg/ml) for 2.5 h under low P-R product conditions. These hearts exhibited a dose-dependent decrease in LV function that was modest compared with that observed in rabbits that had elevated P-R products during in vivo NE treatment. Our results suggest that high concentrations of NE may cause modest degrees of LV dysfunction independently of increases in myocardial energy demand, but the LV dysfunction is exacerbated when myocardial energy demand is elevated.

Vascular Structural and Functional Alterations Before and After the Development of Hypertension in SHR

The time-course of the development of vascular and cardiac hypertrophy, as well as of arterial dysfunction, in human and experimental hypertension is still unclear. Moreover, the interrelationships between structural and functional vascular alterations are presently under debate. The aim of this study was to assess the arteriolar wall thickness and left ventricular mass as well as the vascular response to norepinephrine and acetylcholine in spontaneously hypertensive rats (SHR), before and after the development of hypertension, as compared to age-matched normotensive Wistar-Kyoto rats (WKY). Seventeen SHR (4 to 12 weeks old) and 17 WKY were included in the study. Blood pressure was measured noninvasively. After killing the animals, relative left ventricular mass (RLVM) was calculated, and mesenteric arcades were dissected and mounted on a micromyograph. Functional and structural characteristics of the vessels were measured: media thickness (MT), media/lumen ratio (M/L), and wall tension in response to norepinephrine and acetylcholine. At 4 weeks of age, no difference in blood pressure and RLVM between SHR and WKY was detected, but MT and M/L of mesenteric small resistance arteries were significantly greater in SHR. An increased response to norepinephrine was observed in terms of wall tension, but not of active media stress at the two higher norepinephrine concentrations. No difference in the dose-response curve to acetylcholine between SHR and WKY was observed. At 8 and 12 weeks of age systolic blood pressure was significantly higher in SHR; RLVM, MT, and M/L were also higher at this stage.(ABSTRACT TRUNCATED AT 250 WORDS)

The potential of intact human platelets for sulfoconjugation of norepinephrine in vitro

The aim of the present study was to investigate the potential of human platelets for sulfoconjugation of plasma norepinephrine. For this purpose the uptake and the efflux of norepinephrine and norepinephrine sulfate were studied in vitro in intact human platelets. Incubation of platelets with 0.5 μmol/1 norepinephrine led to a rapid uptake of norepinephrine within 1 min (22.76±4.14 pmol/2.5 × 10 9 platelets/min), whereby 63.7% was transformed to norepinephrine sulfate. The substrate kinetics showed a different slope of norepinephrine uptake with increasing norepinephrine concentrations in the incubation medium (0.1−0.5 μmol/1). At 0.1 μmol/1 norepinephrine the uptake was more efficient than at higher norepinephrine concentrations. Incubation with norepinephrine sulfate slightly decreased intraplatelet norepinephrine sulfate (p≤0.01). Incubation of human platelets in a catecholamine free medium led to an efflux of norepinephrine and norepinephrine sulfate (21.8%, 11.2% of total intraplatelet content, respectively). The present findings indicate a net uptake of norepinephrine into the platelets. At physiological norepinephrine concentrations the uptake seems to be achieved predominantly by an active transport mechanism. Moreover, data indicate a net efflux of norepinephrine sulfate. Therefore it is concluded that human platelets might contribute to the formation of plasma norepinephrine sulfate by norepinephrine uptake, intraplatelet sulfoconjugation and release of norepinephrine sulfate.

Modulation of adrenergic responses by prostanoids in the isolated mesenteric vascular bed of the rat

1. 1. The influences of PGE 2, PGE 1, iloprost and carbocyclic thromboxane A 2 (cTxA 2) on the response to adrenergic nerve stimulation, exogenous noradrenaline and perfusion with methoxamine, have been compared in the rat mesenteric vascular bed. 2. 2. PGE 2, PGE 1 and cTxA 2 enhanced the vasoconstrictor response elicited by field stimulation, as well as that induced by noradrenaline and methoxamine. 3. 3. Iloprost did not affect the increase in perfusion pressure induced by field stimulation, slightly reduced the vasoconstrictor response to high concentrations of noradrenaline, and significantly attenuated the increase in vascular tone induced by perfusion with methoxamine. 4. 4. These findings suggest that, in the rat mesenteric vascular bed, prejunctional mechanisms are not involved in the interference exerted by different prostanoids on the vascular response to adrenergic activation.

Investigations of the dual contractile/relaxant properties showed by antioquine in rat aorta

1. In the present study we assessed the activity of antioquine, a bisbenzyltetrahydroisoquinoline alkaloid isolated from Pseudoxandra sclerocarpa, by examining its effects on the contractile activity of rat isolated aorta, specific binding of [3H]-(+)-cis-diltiazem, [3H]-nitrendipine and [3H]-prazosin to cerebral cortical membranes and the different molecular forms of cyclic nucleotide phosphodiesterases (PDE) isolated from bovine aorta. 2. Contractions in rat aorta induced by high concentrations of KCl (80 mM) and noradrenaline (1 microM) were inhibited by antioquine in a concentration-dependent manner (0.1 microM- 300 microM). The alkaloid appeared more potent against KCl-induced contractions. This inhibitory effect was observed at both 37 degrees C and 25 degrees C. 3. Paradoxically, at the highest concentration tested (300 microM) antioquine induced a contractile response of similar magnitude in the presence and absence of extracellular calcium, at 37 degrees C. This activity was greatly attenuated at 25 degrees C. Antioquine-induced contractions were not inhibited by prazosin (0.1 microM), nifedipine (1 microM) or diltiazem (100 microM). On the contrary, prazosin and nifedipine slightly increased the contractions in the presence of extracellular calcium. Papaverine (100 microM) partially inhibited the contractile response to antioquine both in the presence and absence of extracellular calcium. 4. At 25 degrees C, in Ca(2+)-free solution, antioquine (300 microM) did not modify the contractile response (phasic and tonic) evoked by noradrenaline, but increased the phasic contraction induced by caffeine. At 37 degrees C, the contraction elicited by antioquine made it impossible to observe the noradrenaline-induced one. 5. Antioquine showed affinity for the [3H]-prazosin binding site and for the [3H]-(+)-cis-diltiazembinding site of the Ca2+-channel receptor complex, but had no effect at the dihydropyridine binding site in rat cerebral cortex.6. Antioquine weakly inhibited some PDE forms isolated from bovine aorta: a CaM-PDE (PDE I)which preferentially hydrolyzes cyclic GMP and is activated by calmodulin, and a rolipram-sensitive cyclic AMP-PDE (PDE IV) which hydrolyzed cyclic AMP. Antioquine did not exert any inhibitory effect on the other forms of PDE, a cyclic GMP selective form (PDE V) and a low Km cyclic AMP-PDEthat is inhibited by cyclic GMP (CGI-PDE, PDE III).7. The present work provides evidence that antioquine has properties both as a calcium entry blocker(possibly through the benzothiazepine recognition site in the calcium channel) and as a contractile agent.Its mechanism of action as a contractile agent is not related to Ca2+-entry and is hypothetically similar to that of calyculin-A or okadaic acid. The possible involvement of a-adrenoceptors in this paradoxical effect cannot be excluded. The rigidity of the molecule provides an interesting model for analyzing this contractile mechanism and the intracellular processes involved.

Role of noradrenergic innervation of brown adipose tissue in thermoregulatory deficits following prenatal alcohol exposure.

The development of thermoregulation in newborns is delayed by prenatal alcohol exposure in an animal model of moderate maternal drinking. Newborn mammals generate heat primarily via nonshivering thermogenesis in brown adipose tissue (BAT), which is activated by the sympathetic nervous system. In this study, the effects of prenatal alcohol exposure on the development of the sympathetic innervation of BAT was investigated by assessing the concentration of norepinephrine (NE) in interscapular BAT. Pregnant dams were given either a liquid diet with 35% of the calories derived from alcohol, a liquid diet without alcohol to control for any effects of the liquid diet administration, or ad libitum food and water. Interscapular brown adipose tissue was excised from 5-, 10-, and 20-day-old male and female offspring. At 5 days of age, alcohol-exposed pups had significantly lower NE concentrations than did pups in either control group. However, 20-day-old alcohol-exposed pups had significantly higher NE concentrations than either control group. These results suggest a delay in the development of the sympathetic activation of BAT thermogenesis, followed by a compensatory overactivation. These findings may have important implications for understanding the mechanisms underlying thermoregulatory deficits seen after prenatal alcohol exposure. In addition, these results suggest that maternal alcohol consumption may increase the risk of sudden infant death syndrome, which has been linked to inappropriate BAT thermogenesis.

Noradrenaline as a presynaptic inhibitory neurotransmitter in ganglia of the guinea-pig gall-bladder.

1. The effects of noradrenaline on guinea-pig gall-bladder ganglia were investigated with intracellular recording techniques. 2. Noradrenaline (0.01-100 microM) decreased the amplitude of the fast excitatory postsynaptic potential (EPSP) that was evoked by stimulation of interganglionic fibre tracts. High concentrations of noradrenaline (10-100 microM) caused an inhibition ranging from 93-100%. The noradrenaline concentration that resulted in half-maximal inhibition (EC50) of the EPSP was 280 nM. 3. Experiments with selective agonists and antagonists indicated that the alpha 2-adrenoreceptor was involved in the inhibition of the EPSP. Clonidine (0.001-100 microM) reduced the EPSP in a concentration-dependent manner with an EC50 of 30 nM. Yohimbine (100-300 nM) caused a rightward shift of the noradrenaline concentration-effect relationship, with a dissociation equilibrium constant of 1.4 nM. 4. Release of endogenous catecholamines by tyramine (100 microM) in the presence of desipramine (1.0 microM), caused a yohimbine-sensitive decrease in the amplitude of the EPSP. Treatment with tyramine did not affect the amplitude of the EPSP in tissue that had undergone prior chemical sympathectomy with 6-hydroxydopamine. 5. Electrical stimulation of the vascular plexus (1-3 s; 10-20 Hz; 10 mA) decreased the amplitude of the EPSP. In some cases suprathreshold responses were reduced to subthreshold EPSPs following stimulation of the vascular plexus. Yohimbine (300 nM) reversibly inhibited the effects of vascular plexus stimulation. 6. Noradrenaline did not modify the responses of gall-bladder neurones to exogenously applied acetylcholine. Also, application of noradrenaline, by superfusion (0.001-100 microM) or by pressure microejection (1.0 mM), had no effect on the resting membrane potential, membrane conductance, or action potential characteristics of gall-bladder neurones. 7. Immunoreactivity for type A monoamine oxidase (MAO-A) was found in the vascular plexus and the ganglionated plexus of the gall-bladder. 8. These results show that noradrenaline has an alpha 2-adrenoreceptor-mediated presynaptic inhibitory effect on fast synaptic transmission in the ganglia of the guinea-pig gall-bladder. It is proposed that vagal terminals may be an important target of this adrenergic inhibitory input to the gall-bladder.

The Alterations of Norepinephrine and Acetylcholine Concentrations in Immature Rat Urinary Bladder Caused by Streptozotocin-Induced Diabetes

The concentrations of norepinephrine (NE) and acetylcholine (ACh) in the body and base of the bladder were investigated two, four and eight weeks after the administration of streptozotocin to four-week-old rats. At four and eight weeks after following dosage, NE concentrations in the base of the bladder of the diabetic rats were significantly higher than those of control rats. At the two-week diabetic condition, ACh concentrations in both the body and base of the bladder diabetic rats were about two-fold higher than in the respective control levels and remained constant for eight weeks. However, the control rats showed dramatic increase in ACh concentration from two to four weeks after dosage. There was no significant difference in the response of bladder muscle strips to phenylephrine, isoproterenol and ACh in the diabetic vs. control rats throughout the experiment. These observations suggest that the alteration of autonomic nervous system may occur at an early stage of diabetes mellitus in immature rats and that the high concentration of NE in the diabetic bladder base may reflect an adaptive overfunction of the adrenergic system probably to maintain continence. Furthermore, the lack of change in ACh concentration with age in diabetic rats may suggest an impaired development of the cholinergic system. Accepted for publication January 21, 1992.

Catecholamines in cerebrospinal fluid are increased by behavioral arousal and myocardial ischemia

To study the central neural mechanisms involved in malignant ventricular arrhythmia, concentrations of norepinephrine in the cerebrospinal fluid were measured during behavioral stimulation and during coronary artery occlusion. Pigs were instrumented via thoracotomy with catheters to measure mean arterial pressure and plasma catecholamines and with silk snares around the left anterior descending coronary artery for occlusion after recovery from surgery. Cannulas were placed in the lateral ventricle of the brain to sample cerebrospinal fluid. Behavioral arousal was induced by lifting the pig in a canvas sling for 5 min. Mean arterial pressure, heart rate, and both plasma and cerebrospinal fluid norepinephrine concentrations increased significantly after lifting stimulation. In a separate experiment, 5 min after coronary artery occlusion, both plasma catecholamines and norepinephrine in cerebrospinal fluid were significantly elevated. Furthermore, pigs in which ventricular fibrillation occurred after occlusion had significantly higher concentrations of norepinephrine in cerebrospinal fluid before coronary artery occlusion.

Histochemical study of the heart of the axolotl (Ambystoma mexicanum).

We have investigated the presence of cells containing monoamines, substance P, and neuron-specific enolase (NSE) in the heart and in the pericardial wall of a urodele amphibian, the axolotl. Fibers containing substance P-like immunoreactivity were present in the heart but not in the pericardial wall. Also present in the heart were small branched cells, which stained metachromatically with toluidine blue. Similar cells were found in the peritoneum and were tentatively identified as mast cells. NSE-immunoreactive fibers were found both in the heart and in the pericardial wall. Small intensely fluorescent (SIF) cells of the pericardial wall contained a high concentration of norepinephrine but no other monoamines, substance P, or NSE. Comparison with data available for the mudpuppy, Necturus maculosus, a closely related amphibian species, suggests that the innervation of the heart in the axolotl is substantially different.

Effects of catecholamine stimulation on myocardial hibernation

We have recently shown that low-flow (10%) ischemia in the isolated piglet heart causes an abrupt fall in mechanical function and metabolic activity (acute hibernation), with nearly complete preservation of high-energy phosphates and glycogen after 2 hours of ischemia. We attempted to determine if norepinephrine, as occurs in vivo, would modify the hibernation process. Piglet hearts were perfused at 37° C with red blood cell-enhanced Krebs-Henseleit solution. Performance of the left ventricle was assessed isovolumetrically. With control coronary flow, norepinephrine (40 ng/ml) caused a ≈50% increase in pressure-rate product and the rate of change of pressure. When coronary flow was reduced to 10%, these measures fell to levels indentical to those of ischemic hearts not exposed to norepinephrine. Changes in myocardial O 2 metabolism paralleled mechanical function. Lactate release was quantitatively similar in both groups. However, myocardial adenosine triphosphate was reduced from 29 ± 1 to 13 ± 2 μmol/gm and glycogen from 300 ± 46 to 77 ± 15 μmol/gm by the presence of norepinephrine. Left ventricular compliance was reduced to 51 ± 5%, compared with 87 ± 8% in the group without norepinephrine ( p < 0.001). In the norepinephrine group, correlation between left ventricular stiffness and adenosine triphosphate was poor ( r = −0.32). Thus hibernating myocardium does not manifest progressive deterioration in the presence of high concentrations of norepinephrine. Diastolic function is less well preserved, however.

A biphasic effect of noradrenaline on renin release from rat juxtaglomerular cells in vitro is mediated by alpha 1- and beta-adrenoceptors.

The direct effect of noradrenaline on renin release from juxtaglomerular (JG) cells in vitro were investigated in a dynamic superfusion system of dispersed rat renal cortical cells. At low concentrations (1-100 nmol/l), noradrenaline stimulated renin release in a dose-dependent manner, while at higher concentrations (0.1-1 mmol/l) it inhibited renin release. The stimulatory effect of 0.1 mumol noradrenaline/l was completely blocked by a beta-adrenoceptor antagonist, propranolol (0.1 mumol/l). When applied at concentrations of 1 mumol/l or 10 mumol/l, noradrenaline had no consistent effect on renin release, although 10 mumol noradrenaline/l had an inhibitory effect in the presence of propranolol (0.1 mumol/l). The inhibitory effect of noradrenaline (0.1 mmol/l) was converted to a stimulatory effect by the addition of an alpha 1-adrenoceptor antagonist (bunazosin, 1 mumol/l), but was not altered by the addition of an alpha 2-adrenoceptor antagonist (yohimbine, 1 mumol/l). These results indicate that low concentrations of noradrenaline directly stimulate renin release from JG cells by the activation of beta-adrenoceptors, while high concentrations of noradrenaline inhibit renin release by the activation of alpha 1-adrenoceptors. Accordingly, a dynamic balance may exist between beta-adrenergic stimulation and alpha 1-adrenergic depression of renin release.

Differences Between Rat and Guinea Pig Aorta in Postreceptor Mechanisms of α1-Adrenoceptors

The relative importance of different postreceptor mechanisms associated with alpha 1-adrenoceptor activation in guinea pig aorta and rat aorta was compared. In both tissues, a concentration-dependent correlation was observed between contractile responses produced by high concentrations of norepinephrine (NE) and inositol-1-phosphate (IP1) accumulation. Blockade of Ca2+ entry through voltage-dependent membrane channels by nifedipine had no inhibitory action on NE-induced contractile responses in guinea pig aorta, but significantly inhibited NE-induced contractile responses in rat aorta. Nifedipine had no major effect on NE-induced IP1 accumulation in either tissue. A medium with no Ca2+ inhibited NE-induced contractile responses and IP1 accumulation in guinea pig aorta, but had a more marked effect on contractile responses and IP1 accumulation in rat aorta. The combination of a high concentration of nifedipine and a medium with no Ca2+ almost completely inhibited NE-induced contractile responses and IP1 accumulation in both tissues. Exposure to EGTA also virtually completely inhibited NE-induced contractile response and IP1 accumulation in both tissues. Significant 45Ca2+ entry was stimulated in rat aorta by NE, and this entry was completely blocked by nifedipine. No significant 45Ca2+ entry was stimulated by NE in guinea pig aorta. We conclude that the most important postreceptor event linked to alpha 1-adrenoceptor stimulation in guinea pig aorta is activation of the phosphatidylinositol pathway, whereas in rat aorta Ca2+ entry through voltage-dependent membrane channels is as important as activation of the phosphatidylinositol pathway. We also conclude that activation of the phosphatidylinositol pathway in both tissues is critically dependent on the presence of a small amount of Ca2+, which may enter from the external medium.

Regional sympathetic denervation in von Recklinghausen's disease with coronary spasm and myocarditis

Two patients with von Recklinghausen's disease presented with coronary artery spasm, transient high concentrations of norepinephrine, catecholamine-induced myocarditis, and regional sympathetic denervation of the heart that was assessed by I–123 metaiodobenzylguanidine scintigram. It is postulated that regional sympathetic denervation of the heart may precipitate both coronary artery spasm and catecholamine-induced myocarditis in susceptible patients with von Recklinghausen's disease.

Clodronate inhibits contraction and prevents the action of L-type calcium channel antagonists in vascular smooth muscle.

Clodronate (dichloromethylenebisphosphonate) decreased vasoconstriction of the isolated perfused rat tail artery mediated by norepinephrine and by Ca2+ in a K(+)-depolarizing solution. The norepinephrine contractile response was divided into two components by sequential manipulation of the composition of the perfusion fluid, where the first component is due to the release of Ca2+ from intracellular stores and the second to the influx of Ca2+ from extracellular fluid. Clodronate (20 microM) decreased only the first component of the response at a norepinephrine concentration of 50 nM, and both components of the response at a higher norepinephrine concentration (100 nM). The L-type Ca2+ channel blocking drugs, nicardipine (10 nM) and verapamil (1 microM), reduced the second component of the norepinephrine-mediated vasoconstriction, but in the presence of clodronate (20 microM) this blocking action was prevented. These results were confirmed by examining the interaction between clodronate and nicardipine on norepinephrine and K(+)-mediated lanthanum (La(3+)-resistant unidirectional 45Ca uptake. Nicardipine (1-10 nM) decreased the norepinephrine (100 nM) and K(+)-induced (60 mM) La(3+)-resistant unidirection 45Ca uptake in a concentration-dependent manner, but in the presence of clodronate (20 microM) this concentration-dependent response was abolished. Thus, clodronate not only reduced agonist-induced Ca2+ release from intracellular stores and Ca2+ influx through L-type Ca2+ channels but also prevented L-type Ca2+ channel antagonists from exerting their effect. These results indicate clodronate has two sites of action during vascular smooth muscle contraction: the first on intracellular mobilization of Ca2+ and the second on L-type Ca2+ channels.

Acetylcholine-induced release of endothelium-derived relaxing factor from lymphatic endothelial cells

Ring segments of dog thoracic ducts precontracted with a high concentration of norepinephrine (NE) relaxed in a concentration-dependent manner in response to acetylcholine (ACh) or sodium nitroprusside (SNP). Pretreatment with atropine inhibited the ACh-induced relaxation in a competitive manner. The Schild plot showed a slope of 1.1 +/- 0.2 and a pA2 value of 10.4 +/- 0.4 (n = 6 ring segments). Removal of endothelium caused a complete inhibition of the ACh-induced relaxations in precontracted dog thoracic ducts. ACh, which failed to relax precontraction of the ring segment when mounted separately, induced relaxation in the same preparation when it was mounted as a "sandwich" with the longitudinal strip. The ACh-induced relaxations in the lymphatic preparations with endothelium were suppressed or abolished by pretreatment with oxyhemoglobin (10(-6) and 10(-5) M), methylene blue (10(-6) and 10(-5) M), and NG-monomethyl-L-arginine (3 x 10(-5) M), but the relaxations were unaffected by aspirin (10(-5) M). Pretreatment with methylene blue (10(-5) M) also caused a significant reduction of the SNP-induced relaxations in the precontracted thoracic ducts. It may be concluded that ACh-induced relaxation in dog thoracic ducts precontracted with NE is mediated by high-affinity muscarinic receptors in the lymphatic endothelial cells. Also, stimulation of the endothelial muscarinic receptors liberates a transferable endothelium-derived relaxing factor, which results in the relaxation of the lymphatic smooth muscles via the accumulation of cellular guanosine 3',5'-cyclic monophosphate.

Comparative effects of adrenaline and noradrenaline on the synthesis and secretion of soluble protein by isolated hepatocytes

1. 1. Adrenaline (A) supplementation of the incubation medium of monolayer cultures of hepatocytes at 0.1, 0.2, 0.3, 1 and 10 ng/ml resulted in consistently enhanced levels of secreted and newly synthesised non-secreted proteins; supplementation with 100 and 1000 ng/ml resulted in lower or unchanged levels. These effects were most consistent 6 hr after a medium change. 2. 2. Noradrenaline supplementation of the medium resulted in increased levels of secreted and non-secreted proteins at low concentrations (<2.4 ng/ml) at 6 hr after a medium change, but significantly decreased levels of both populations at high concentrations of noradrenaline (NA) (2.4 < NA < 1000 ng/ml), at 6–9 hr after a medium change and sustained to 24 hr, the most significant decrease being at 10 ng/ml. 3. 3. All combinations of concentrations of A + NA resulted in non-dose-dependent decreased levels of both the secreted and non-secreted soluble protein fractions. The most significant decreases occurred at concentrations of (1 + 5) and (10 + 10) ng/ml adrenaline + noradrenaline. 4. 4. Medium supplementation with adrenaline, noradrenaline or a combination of the two had no effect on the uptake of [ 3H]leucine by the cells. 5. 5. The results are discussed in relation to receptor status on the hepatocyte membranes.