The neurogenic constrictor response of isolated rat saphenous artery is reduced after 2-week tail suspension

Under real or simulated microgravity conditions the control of arterial vascular tone is greatly disturbed. The low arterial vessel reactivity to sympathetic influences may be the cause of an increase in flow in hind limb skeletal muscles in tail-suspended (TS) rats. Our previous experiments with constant pressure perfusion of rat hind limb demonstrated the reduced vasoconstrictor responses to sympathetic nerve stimulation in TS rats. Responses to exogenous noradrenaline depended on the perfusion conditions. It is known that the vessels of various branching orders noticeably differ in nerve density and in sensitivity to vasoconstrictor agonists. So under neurogenic or exogenous noradrenaline influences the vascular resistance may be increased at different levels of vascular bed, thus making the data analysis seriously complicated. This uncertainty may be overcome by investigation of a single vessel isolated from hind limb vascular bed. The saphenous artery, a resistance artery with dense innervation, is a very convenient object for this purpose. Thus, this study was aimed at comparing the effects of 2-week tail suspension upon the constrictor responses of isolated saphenous artery to neurogenic and exogenous noradrenaline stimuli in rats.     

Slow and incomplete sympathetic reinnervation of rat tail artery restores the amplitude of nerve-evoked contractions provided a perivascular plexus is present

We have investigated the recovery of sympathetic control following reinnervation of denervated rat tail arteries by relating the reappearance of noradrenergic terminals to the amplitude of nerve-evoked contractions of isometrically mounted artery segments in vitro. We have also assessed reactivity to vasoconstrictor agonists. Freezing the collector nerves near the base of the tail in adult rats denervated the artery from ～40 mm along the tail. Restoration of the perivascular plexus declined along the length of the tail, remaining incomplete for >6 mo. After 4 mo, nerve-evoked contractions were prolonged but of comparable amplitude to control at ～60 mm along the tail; they were smaller at ～110 mm. At ～60 mm, facilitation of contractions to short trains of stimuli by the norepinephrine transporter blocker, desmethylimipramine, and by the α2-adrenoceptor antagonist, idazoxan, was reduced in reinnervated arteries. Blockade of nerve-evoked contractions by the α1-adrenoceptor antagonist, prazosin, was less and by idazoxan greater than control after 8 wk but similar to control after 16 wk. Sensitivity of reinnervated arteries to the α1-adrenoceptor agonist, phenylephrine, was raised in the absence but not in the presence of desmethylimipramine. Sensitivity to the α2-adrenoceptor agonist, clonidine, was maintained in 16-wk reinnervated arteries when it had declined in controls. Thus regenerating sympathetic axons have a limited capacity to reinnervate the rat tail artery, but nerve-evoked contractions match control once a relatively sparse perivascular plexus is reestablished. Functional recovery involves prolongation of contractions and deficits in both clearance of released norepinephrine and autoinhibition of norepinephrine release.     

Restoring effect of noradrenalne on diminished neurogenic vasoreactivity

The effect of 0.01-1.0 microM noradrenali on response to electrical field stimulation (EFS) of the juvenile rat tail artery segment was studied. Noradrenali was shown to potentiate the EFS-evoked constriction decreased in the course of experiments or in the acidic solution (pH 6.6) and this potentiation was proportional to the extent of the preceding decrease of the constriction. The more decreased was the EFS-evoked constriction the higher was the noradrenali concentration which produced the maximal potentiation and the wider was the potentiative noradrenali concentration range. The potentiative effect of noradrenali was not prevented by the NO synthase inhibitor NG-nitro-L-arginine. The results suggest that noradrenali can restore the diminished neurogenic reactivity of blood vessels, and this effect is not connected with the change in the NO production.     

Two-component responses to sympathetic nerve stimulation in the rat tail artery

Membrane potential and tension were recorded simultaneously from the smooth muscle of the rat tail artery. A single stimulus to the perivascular nerves caused a tension transient. The tension transient had two components, one due to a muscle action potential and one due to alpha-adrenoceptor activation. During trains of stimuli most of the tension was due to alpha-receptor activation, even when every stimulus caused a smooth muscle action potential.     

Three different vasoactive responses of rat tail artery to nicotine

The vasoactive effects of nicotine on isolated rat tail artery tissues were studied. Nicotine transiently contracted rat tail artery tissues (EC50, 55.6 +/- 2 microM) in an extracellular Ca2+ dependent and endothelium-independent fashion. The blockade of alpha1-adrenoceptors, but not alpha2-adrenoceptors or P2X purinoceptors, inhibited the nicotine-induced contraction by 38 +/- 7% (p < 0.05). Nicotine (1 mM) depolarized membrane by 13 +/- 3 mV, but did not affect L-type Ca2+ channel currents, of the isolated rat tail artery smooth muscle cells. The phenylephrine-precontracted tail artery tissues were relaxed by nicotine (EC50, 0.90 +/- 0.31 mM), which was significantly inhibited after the blockade of nicotinic receptors. Simultaneous removal of phenylephrine and nicotine, after a complete relaxation of the phenylephrine-precontracted tail artery strips was achieved by nicotine at accumulated concentrations (> or =10 mM), triggered a Ca2+-dependent rebound long-lasting vasoconstriction (n = 20). This rebound contraction was abolished in the absence of calcium or in the presence of tetracaine in the bath solution. Pretreatment of vascular tissues with a nicotinic receptor antagonist did not affect the nicotine-induced vasoconstriction or nicotine withdrawal induced rebound contraction. The elucidation of the triphasic vascular effects of nicotine and the underlying mechanisms is important for a better understanding of the complex vascular actions of nicotine.     

Amiloride acts as an alpha-adrenergic antagonist in the isolated rat tail artery

In the final concentration of 100 microM, amiloride increased substantially the overflow of endogenous noradrenaline and decreased that of 3,4-dihydroxyphenylethylene glycol from the rat tail artery into Krebs solution supplemented with 10 microM veratridine. The overflow of the amine into a 120 mM-K version of Krebs solution was unaffected by amiloride, while that of the glycol was reduced. Abolition of the contractile response to 10 microM veratridine by 2 microM phentolamine indicated that the response was due to release of endogenous noradrenaline. Addition of amiloride in the final concentrations of 10 and 100 microM caused relaxation of strips contracted by the alkaloid. The dose-response relations for exogenous noradrenaline measured in the absence or presence of 50 microM amiloride indicated that the drug acted as a reversible competitive alpha-adrenergic antagonist. The phentolamine-resistant component of the contractile response to the 120 mM-K solution was unaffected by 100 microM amiloride. Although the exact site of action of amiloride remains to be determined, it can be concluded that amiloride inhibits adrenergic transmission at a postsynaptic site at a step preceding elevation of myoplasmic Ca2+.     

Tetraethylammonium-evoked oscillatory contractions of rat tail artery: a K-K model

Spontaneously rhythmic contraction of peripheral blood vessels actively modulates the peripheral circulation and blood pressure. However, the underlying mechanisms for the complex rhythmic contraction patterns of various vascular tissues are not yet fully understood. In the present study, the tetraethylammonium (TEA)-induced spontaneously oscillatory contractions of isolated rat tail artery tissues were examined. It was found that TEA evoked arterial oscillatory contractions in a concentration-dependent, but endothelium-independent manner. The voltage-dependent K+ (Kv) channel specific blocker, 4-aminopyridine (4-AP), induced a sustained, but not oscillated, vascular contraction. The presence of 4-AP had no effect on the TEA-induced oscillatory contractions. The blockade of KCa channels with charybdotoxin or apamin did not affect the basal force of vascular tissues. Neither the TEA-induced oscillatory contraction was affected by these blockers. The opening of KATP channels by levcromakalim or their blockade by glybenclamide ceased or increased, respectively, the oscillation of TEA-induced contractions. The absence of Ca2+ or the presence of nifedipine in the bath solution completely abolished the effects of TEA. The inhibition of Ca2+-ATPase in the sarcoplasmic reticulum with micromolar concentrations of thapsigargin or cyclopiazonic acid either abolished or enhanced, respectively, the TEA-induced oscillatory contractions. Ryanodine did not affect the TEA-induced oscillatory contraction. In conclusion, the TEA-induced oscillatory contraction may be initiated by the blockade of the TEA-sensitive delayed rectifier K+ channels and maintained by the TEA-insensitive but ATP-sensitive K+ channels. This K-K model presents a novel mechanism for the depolarization-induced rhythmic contractions of small arteries.     

Norepinephrine stimulates the direct breakdown of phosphatidyl inositol in rat tail artery.

When segments of rat tail artery were labeled with [3H]inositol and then stimulated with norepinephrine (NE), the inositol phosphates produced were primarily IP and IP2, together with a small but significant amount of Ins(1,4,5)P3 and a very small amount of Ins(1,3,4,5)P4. It has been unclear in many studies whether or not the relatively large levels of IP and IP2 produced in [3H]inositol-labeled tissue represent indirect products of phosphatidyl inositol(4,5)bis phosphate breakdown (through Ins(1,4,5)P3) or direct products of phosphatidyl inositol 4 monophosphate and phosphatidyl inositol breakdown. In order to answer this question tail artery segments were prelabeled with [3H]inositol and then permeabilized with beta escin and stimulated with norepinephrine and GTP gamma S, so that increases in IP, IP2, and Ins(1,4,5)P3 were still observed. If these permeable segments were stimulated with agonist in the presence of compounds known to inhibit Ins(1,4,5)P3 5-phosphatase, such as glucose 6P, (2,3)diphosphoglycerate, or Ins(1,4,5)P3, the levels of labeled Ins(1,4,5)P3 and labeled IP2 were increased, while the level of stimulated labeled IP was unchanged. This indicated that some of the IP2 and IP formed in these cells was produced from PIP2 but that some of these compounds might be formed from PIP or PI. When the isomers of inositol monophosphate, Ins 1P and Ins 4P, were separated by HPLC, it was shown that after prelabeled tail artery was stimulated by norepinephrine for periods of 1-2 min, the predominant isomer formed was Ins 4P, indicating either PIP2 or PIP as the source. However, after 5-20 min stimulation, both Ins 1P and Ins 4P were formed in equal amounts, suggesting that during sustained stimulation of smooth muscle PI itself was broken down directly. Therefore it appears that within 1-2 min of norepinephrine addition to vascular smooth muscle the bulk of the IP and IP2 produced are derived from PIP2 via IP3, while after 20 min of norepinephrine treatment much of the IP comes directly from PI. This suggests that the regulation of PLC in this tissue is more complicated than has been previously believed.     

Some effects of the ionophore X-537A on the isolated rat tail artery

The effects of micromolar concentrations of the ionophore X-537A (RO 2-2985) were studied using isolated preparations of the rat tail artery. The ionophore causes complete release of catecholamines from adrenergic nerves, which is the sole cause of the transient contractile response. The amines are released by a nonexocytotic process which seems to be related to the ability of X-537A to act as an efficient transmembrane carrier of Na+, k+, and H+. The ionophore also causes an almost complete and irreversible loss of the cocaine-sensitive component of metaraminol uptake by the tissue. X-537A dissipates the transmembrane concentration gradients of Na and K in the smooth muscle component of the preparation. This effect is unrelated to the release of endogenous catecholamines, and it can also be observed after the Na pump has been inhibited with ouabain. It is fully reversible, though not readily, and it can be induced repeatedly. In catecholamine-depleted strips, X-537A dissipates the transmembrane Na+ and K+ gradients without causing any change in tension. Stimulation of the rate of O2 consumption by X-537A in catecholamine-depleted tissue is reversible, and it is unaffected by ouabain and (or) removal of external Ca2+.     

Male-female differences in the relative contribution of endothelial vasodilators released by rat tail artery

Several different vasodilator substances can be released by vascular endothelium in response to mechanical stimuli and vasoactive agents. The purpose of this study was to determine whether there is a male-female difference in the relative contributions of nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF) to endothelium-dependent vasodilation. Perfusion pressure was measured in isolated tail arteries from male and female rats. Vasodilators released by mechanical shear stress were assessed by constricting the artery with methoxamine; acetylcholine was applied to induce receptor-mediated vasodilation. We used an inhibitor of NO synthase, N(G)-monomethyl-L-arginine acetate (L-NMMA), and elevated levels of K(+) (27 mM) to reveal the relative contributions of NO and EDHF, respectively. Indomethacin was present in all experiments to block prostanoid production. The results indicate that NO was the primary vasodilator released by male tail arteries in response to both mechanical stress and acetylcholine (the L-NMMA-sensitive component of the combined L-NMMA/K(+) effect was 83 +/- 8% and 101 +/- 4%, respectively). However female tail arteries appeared to utilize both NO and EDHF for vascular relaxation (e.g., L-NMMA sensitivity: 58 +/- 9%; K+-sensitivity: 42 +/- 9% in mechanical stress experiments). These findings suggest endothelial regulation differs between males and females.     

Differences in inositol phosphate production in rat tail artery and thoracic aorta

Pharmacomechanical coupling of vascular smooth muscle is believed to be mediated by inositol trisphosphate (IP3). Numerous studies have demonstrated an increase in inositol phosphates following tissue stimulation using either intact aortic strips or cultured cells from aorta. However, little information is available concerning inositol phosphates in vascular tissue other than in the large conduit vessel, the aorta. This present study was designed to examine the role of inositol phosphate metabolism following adrenergic stimulation of the muscular rat tail artery as compared to the aorta. Segments of thoracic aorta and tail artery from male Sprague Dawley rats were labeled with [3H]inositol and stimulated with norepinephrine. The norepinephrine concentration that resulted in a half-maximal stimulation of inositol phosphates was approximately 10(-6) M in both the aorta and tail artery. Although the sensitivity of the two vessels to norepinephrine stimulation were similar, the stimulated levels of IP, IP2, and IP3 were from 1 to 2 orders of magnitude greater in the tail artery than in aorta. IP production in aorta and tail artery was a linear function of time (from 0 to 30 min). Significant levels of IP3 (the 1,4,5-IP3 isomer as determined by HPLC) could only be detected in the tail artery and appeared to be produced optimally after 5 min of stimulation. The several order of magnitude increase in adrenergic stimulated inositol phosphate production in the tail artery was not due to either an increased magnitude of [3H]inositol incorporated into PI, PIP, and PIP2 or to a greater percentage of smooth muscle cells per unit tissue of the rat tail artery. We believe the results of this study demonstrate that the increased inositol phosphate metabolism in the vascular smooth muscle cells of the tail artery is an intrinsic property of the cell. Moreover, due to the significant levels of all inositol phosphates produced in the tail artery, this muscular artery may be a better model, as compared to the aorta, for future studies investigating pharmacomechanical coupling of vascular smooth muscle.     

Barium and strontium as calcium substitutes for contractile responses in the rat tail artery

The ability of Ba2+ and Sr2+ to substitute for Ca2+ in contractile responses of the rat tail artery has been examined. Both Ba2+ and Sr2+ caused comparable contractions in Ca-depleted NA-stimulated, or K+-depolarized strips. Ba2+ and Sr2+ substitute poorly for Ca2+ at noradrenaline-sensitive membrane sites. At high concentrations, the three divalent cations stabilize the membrane in the order: Ca2+ greater than Sr2+ greater than Ba2+. The relaxation rates following high-K+ contractions were similar for all three divalent cations, suggesting a common mechanism for sequestration/extrusion.     

Effects of age and streptozotocin-induced diabetes on biogenic amines in rat tail artery

The changes in amine concentrations in different segments of the rat tail artery have been investigated at different ages and after different durations of streptozotocin-induced diabetes. There was a significant positive slope to the relationship between amine concentrations and age in proximal portion of the normal tail artery, and a significant additional increase in amine concentrations following induction of diabetes. The peak of the latter response occurred between 10 and 20 weeks following the induction of diabetes. There was also a significant dependence on the length of the post-ganglionic neurones, which may be related to the failure of axonal transport of some of the essential enzymes or transporters for these biogenic amines. This model of altered catecholamine metabolism and handling requires further investigation so that alterations in the mechanisms involved in processing of these amines in diabetic autonomic neuropathy may be elucidated. (Mol Cell Biochem 261: 77–82, 2004)     

Effects of age and streptozotocin-induced diabetes on biogenic amines in rat tail artery

The changes in amine concentrations in different segments of the rat tail artery have been investigated at different ages and after different durations of streptozotocin-induced diabetes. There was a significant positive slope to the relationship between amine concentrations and age in proximal portion of the normal tail artery, and a significant additional increase in amine concentrations following induction of diabetes. The peak of the latter response occurred between 10 and 20 weeks following the induction of diabetes. There was also a significant dependence on the length of the post-ganglionic neurones, which may be related to the failure of axonal transport of some of the essential enzymes or transporters for these biogenic amines. This model of altered catecholamine metabolism and handling requires further investigation so that alterations in the mechanisms involved in processing of these amines in diabetic autonomic neuropathy may be elucidated.     

Modulation of [H]noradrenaline release by endothelin-1 in the rat tail artery

The effect of endothelin-1 (ET-1) on the increase in perfusion pressure and the release of noradrenaline produced by electrical field stimulation were examined in isolated perfused/superfused rat tail arteries. ET-1 (1–30 nM) increased, in an identical concentration-dependent manner, the basal perfusion pressure and the stimulation-evoked tritum overflow, whereas the basal outflow of noradrenaline was not changed by the peptide. These results show that, besides its postjunctional vasoconstrictor effect, ET-1 exerts in the rat tail artery a prejunctional action which might be involved in the modulation of stimulation-evoked noradrenaline release from postganglionic nerves.     

Heat vasodilatation of the rat tail.

The vasomotor response of the tail of the albino rat to total-body heating and cooling was studied by skin-temperature recording and plethysmography with the tail at 25 degrees C air temperature. Tail vasodilation started at core temperatures lightly above 37 degrees C and increased to a core temperature up to about 39 degrees C. During cooling of warm rats, tail vasoconstriction started at significantly higher levels of core temperature than the values at which vasodilation appeared when the rat was warmed.     

Effects of dexamethasone and L-canavanine on the intracellular calcium-contraction relation of the rat tail artery during septic shock

The intracellular mechanism by which sepsis lowers vascular reactivity and the subsequent reversal by dexamethasone or nitric oxide synthase (NOS) inhibitors remain unclear. We measured the sensitivity of contraction of the rat tail artery to intracellular Ca2+ in a model of polymicrobial septic shock. At 22 h after cecal ligation and puncture (CLP), rats were treated with an anti-inflammatory glucocorticoid (dexamethasone, 1 mg/kg ip), an inducible NOS inhibitor (L-canavanine, 100 mg/kg ip), or saline. At 24 h after CLP, endothelium-denuded, perfused segments of tail artery were loaded with the intracellular Ca2+-sensitive dye fura 2 in vitro. Intracellular Ca2+ concentration and perfusion pressure were measured simultaneously. The rightward shift of the perfusion pressure-intracellular Ca2+ mobilization curve after norepinephrine stimulation subsequent to CLP indicates decreased intracellular Ca2+ sensitivity of contraction. The relation was restored by dexamethasone (which also restored in vivo blood pressure and flow), but not by L-canavanine (which restored perfusion pressure by further mobilization of intracellular Ca2+). We conclude that CLP lowers vasomotion by lowering intracellular Ca2+ sensitivity, which can be restored with glucocorticoid treatment. The involvement of inducible NOS does not solely account for the sepsis-induced reduction in Ca2+ sensitivity of contraction.