Search

Purpose: This study was performed to examine the role of the vascular endothelium in modulating arterial reactivity to adrenergic vasoconstriction in subcutaneous arteries from patients with type II diabetes., Methods: Small subcutaneous arteries (inner diameter = 90 to 180 microns) from control subjects (n = 22) and patients with diabetes (n = 18) were dissected from skin biopsies obtained at surgery and mounted on a specialized arteriograph that allowed for continuous measurement of lumen diameter under controlled pressure. The sensitivity to norepinephrine was compared in arteries that were either intact, denuded of endothelium, or intact and exposed to N omega-nitro-L-arginine (L-NNA), an inhibitor of nitric oxide synthesis. Stimulated release of nitric oxide by acetylcholine and smooth muscle cell responses to sodium nitroprusside were also evaluated in diabetic and control arteries., Results: Sensitivity to norepinephrine was augmented in diabetic arteries and the amount of agonist necessary to contract the vessels 50% of maximum (EC50) decreased from 0.35 +/- 0.05 mumol/L in the control arteries to 0.16 +/- 0.06 mumol/L in the diabetic arteries (p < 0.05). Both endothelial removal and blockade of nitric oxide synthesis increased sensitivity to norepinephrine in control arteries (EC50 denuded = 0.14 +/- 0.03 mumol/L and EC50 L-NNA = 0.14 +/- 0.04 mumol/L; p < 0.01) but failed to augment sensitivity in diabetic arteries (EC50 denuded = 0.17 +/- 0.05 mumol/L and EC50 L-NNA = 0.15 +/- 0.04 mumol/L; p > 0.05). Stimulated release of nitric oxide by acetylcholine was increased in the diabetic arteries: EC50 control = 0.04 +/- 0.01 mumol/L versus EC50 diabetic = 0.009 +/- 0.001 mumol/L (p < 0.05). Sensitivity of vascular smooth muscle to sodium nitroprusside was similar in both nondiabetic and diabetic arteries., Conclusions: The endothelium mitigates adrenergic reactivity in control arteries, which is lacking in diabetic arteries and results in enhanced reactivity to norepinephrine; increased sensitivity of diabetic arteries to acetylcholine, however, indicates a possible alteration at the receptor level.

Melatonin research has primarily utilized blood as the source of samples, but there is now increasing interest in measuring levels of the hormone found in saliva. One impediment to this approach is that several melatonin assays involve a column-extraction step that can prove very time-consuming or even impossible when salivary samples are excessively viscous. We have treated 67 samples with dithiothreitol to enhance their passage through the column. Following this treatment, all samples passed freely through the columns. The minimum and maximum values measured were 0.7 - 50.0 pg/ml for the untreated controls and 1.0 - 51.9 pg/ml for the treated samples. The means (+/-SEM) for these groups were 9.5 +/- 1.6 and 9.9 +/- 1.7, respectively, and were not significantly different from one another as assessed by Student's t-test (P = 0.08). In summary, we have found that this technique permits us to obtain values on samples which would otherwise be unusable and that such treatment does not alter the melatonin values yielded by RIA analysis.

Previous data in rat conductance vessels indicated that cellular mevalonate contributes to vascular tone and systemic blood pressure control. Using exogenous mevalonate (M) or lovastatin, a 3-hydroxy-3-methyl-glutaryl CoA (HMG-CoA) reductase inhibitor (L), we characterized the role of mevalonate availability in resistance artery function, both in experimental animals and humans. Rat mesenteric artery resistance vessels (MARV, n = 9) were incubated for 48 h with either L, M, L + M, or vehicle (V) and tested for reactivity to NE, serotonin, acetylcholine, atrial natriuretic peptide, and sodium nitroprusside (SNP). Lovastatin increased sensitivity to NE (P < 0.03) and serotonin (P < 0.003), and significantly impaired the response to all three vasodilators. These effects were reversed by co-incubation with mevalonate. Mevalonate alone had no effect. In separate experiments, intravascular free Ca2+ concentration (ivfCa2+) was determined in fura-2AM loaded MARV. Basal ivfCa2+ was increased after a 48-h exposure to L (52.7 +/- 4.6 nM, L, vs. 29.7 +/- 2.4 nM, V, n = 12, P < 0.003), as were ivfCa2+ levels following stimulation with low (100 nM) NE concentrations. Similar ivfCa2+ concentrations were achieved during maximum contraction with NE (10 mM) in both groups. Human resistance arteries of human adipose tissue were also studied. Lovastatin increased the sensitivity to NE (ED50 = 372 +/- 56 nM, V, and 99 +/- 33 nM, L, P < 0.001) and significantly decreased the relaxation to acetylcholine and SNP of human vessels. We conclude that mevalonate availability directly contribute to resistance vessel function and vascular signal transduction systems in both experimental animals and humans. The study calls for the identification of non-sterol, mevalonate-derived vasoactive metabolites, and suggests that disorders of the mevalonate pathway can alter vascular tone and cause hypertension.

Human saphenous veins were obtained at operation and assayed immediately (n = 10). The veins were cut into rings, suspended in organ chambers, and connected to force transducers for recording of isometric tension. One ring served as control; others were treated with either the alpha 1-adrenoceptor antagonist prazosin (Pz, 3 x 10(-7) M) or the alpha 2-adrenoceptor antagonist rauwolscine (Rw1, 10(-7) M). Cooling from 37 degrees to 24 degrees C had no significant effect on the resting tone of quiescent rings. Electrical stimulation (0.2-16 Hz) caused frequency-dependent contractions in control vessels. The contractions were inhibited by Pz (p < 0.001) and by Rw (p < 0.001). In control rings, cooling potentiated contractions evoked at all frequencies. Similar augmentations were induced by cooling in rings treated with the alpha 1-antagonist Pz. In contrast, rings treated with Rw before being electrically stimulated showed no significant change in contractile force when cooled. The data indicate that in the human saphenous vein, both alpha 1- and alpha 2-adrenoceptors are innervated, contributing to contractile response evoked by neuronal excitation. Cold augments saphenous vein reactivity to endogenously released norepinephrine (NE) by an apparent increase in the responsiveness of alpha 2-adrenoceptors to agonists. This relationship between temperature and adrenoceptor responsiveness is consistent with the hypothesized role of alpha 2-adrenoceptors in cold-induced vasospasm.

Recently, the obese Zucker rat (OZR), an animal model of non-insulin-dependent (type II) diabetes, was shown to respond to converting enzyme inhibition with decreased albuminuria and a marked attenuation of glomerular injury. It was hypothesized that the OZR would possess low plasma renin values and an increased vascular responsiveness to angiotensin II, and therefore, the renin-angiotensin system (PRA, active renin, inactive renin, renal renin content, and plasma angiotensinogen) and vascular reactivity in OZR at 10 and 24 wk of age were investigated. PRA and renin concentration, inactive plasma renin, and renal renin content were all significantly (P < 0.05) reduced in OZR when compared with age-matched lean controls. The ratio of inactive to total renin was significantly increased in the OZR. OZR aortic ring vascular reactivity to KCl, norepinephrine, and angiotensin II was assessed. Despite essentially equal or increased contractile responses to KCl and norepinephrine at both 10 and 24 wk of age, the OZR was not more sensitive to angiotensin II and displayed a significantly reduced contractile response to angiotensin II at 24 wk of age, when compared with lean age-matched controls. It was concluded that the renal protective effect of converting enzyme inhibition in OZR, despite significantly reduced PRA and concentration, inactive plasma renin, and renal renin content, may not be due to a diabetes-induced increased vascular reactivity to angiotensin II.

Purpose: We hypothesized that the clinical benefit of elastic compression stockings (ECS) is at least in part due to an increase in subcutaneous pressure that may promote resorption of extracellular fluids, providing more efficient diffusion of oxygen and nutrients from the microcirculation to the skin and subcutaneous tissues., Methods: To test this hypothesis we designed and standardized a device for measuring subcutaneous pressure in patients. We then measured the supine perimalleolar subcutaneous pressure from a single limb in four groups: group 1 consisted of control subjects (n = 8); group 2 consisted of patients with varicose veins and superficial venous insufficiency without lipodermatosclerosis or edema (n = 5); group 3 consisted of patients with deep venous insufficiency and lipodermatosclerosis but without edema (n = 8); and group 4 consisted of patients with deep venous insufficiency, lipodermatosclerosis, and clinically evident edema (n = 8). Measurements were made at baseline and after application of 20 to 30 mm Hg and 30 to 40 mm Hg ECS., Results: There was no significant difference in the baseline subcutaneous pressure between the three groups without clinical edema (p > 0.05). Baseline perimalleolar pressure was elevated, however, in group 4 patients compared with groups 1, 2, and 3 (p < 0.05). All three groups with chronic venous insufficiency (CVI) (groups 2, 3, 4) demonstrated increases in subcutaneous pressure with application of ECS, which was statistically significant in groups 3 and 4. There was no difference between the increase in perimalleolar subcutaneous pressure induced by 20 to 30 mm Hg or 30 to 40 mm Hg ECS in groups 3 and 4., Conclusions: Patients with CVI and edema have significant elevations in supine resting perimalleolar subcutaneous pressure compared with control subjects and patients with CVI without edema. Twenty to 30 mm Hg and 30 to 40 mm Hg ECS increased measured perimalleolar subcutaneous pressure in patients with CVI with and without clinical edema but not in control patients. These results suggest the mechanism of benefit of ECS in patients with CVI is due at least in part to an increase in subcutaneous pressure that may act to promote more efficient absorption of perimalleolar extracellular fluid.

Purpose: Animal models have been used to assess the function of vascular smooth muscle and endothelium of veins grafted into arterial circulation. The primary model consists of grafting the external jugular vein into the carotid artery of the rabbit. These studies suggest a selective increase in the responsiveness of the grafted veins to serotonin. However, in both human cardiac and peripheral vascular operations, the saphenous, not the jugular, is the vein most frequently used. Thus the propriety of the rabbit model is unknown., Methods: Human saphenous veins and vein grafts were obtained from patients undergoing leg vein bypass graft revisions (n = 8). The reversed vein grafts were placed into arterial circulation for periods ranging from 4 to 26 months before removal (mean 16 months). All vessels were immediately cut into rings and suspended in organ chambers for recording isometric contractions to norepinephrine and serotonin., Results: The maximal contractions elicited by both norepinephrine and serotonin were reduced in human vein grafts in comparison to the results in human saphenous vein (maximal response to norepinephrine 1.42 +/- 0.34 gm [vein graft] vs 4.59 +/- 1.13 gm [saphenous vein], p = 0.031; maximal response to serotonin 2.68 +/- 0.58 gm [vein graft] vs 4.72 +/- 1.11 gm [saphenous vein], p = 0.042). Human vein grafts were less responsive to norepinephrine than was saphenous vein (negative log of concentration that caused 50% of the maximal response -5.91 +/- 0.10 and -6.84 +/- 0.22, respectively; p < 0.009). After precontraction with norepinephrine (to 30% of the maximal response), saphenous vein, but not vein grafts, demonstrated endothelium-dependent relaxation to acetylcholine (maximum relaxation 27.4% +/- 6.8%; p = 0.001)., Conclusions: Human saphenous veins grafted into arterial circulation exhibit loss of endothelium-dependent relaxation to acetylcholine and diminished contractions to agonists (norepinephrine and serotonin). In contrast to rabbit data, serotonin elicits dose-dependent contractions in both human saphenous vein and human vein grafts. Since the vascular wall contractility varies widely across species, the relevance of rabbit vein graft data to human bypass grafts is uncertain.

Human saphenous veins (HSV; n = 17) were obtained at surgery and assayed immediately. The veins were cut into rings, suspended in organ chambers and connected to force transducers for the recording of isometric tension. Tail arteries (RTA) from male Sprague-Dawley rats were similarly prepared. In quiescent rings, cooling from 37 to 24 degrees C had no significant effect. In the presence of alpha-adrenoceptor blockade, potassium chloride (KCl, 10-80 mM) caused concentration-dependent contractions that were inhibited slightly by cooling. Serotonin (5-HT, 10(-8)-10(-5) M) elicited concentration-dependent contractions in both the HSV and RTA with EC50's (-log concentration required to induce contractions 50% of maximal) of 6.31 +/- 0.03 and 6.55 +/- 0.06, respectively. These contractions were subject to blockade with the S2-selective 5-HT antagonist, ketanserin. When either HSVs or RTAs were contracted with 5-HT and cooled, a potent augmentation of the contractions ensued. The data indicate that, while acute moderate cooling does not significantly augment either resting tone or KCl-induced contractions of HSV or RTA, S2-receptor-mediated events are enhanced, yielding potent augmentations of the vascular response to the platelet-derived compound, serotonin. The relationship between temperature and serotonergic receptor responsiveness may be of clinical importance as a mechanism contributing to cold-induced vasospasm.

Experiments were performed to assess the effects of acute moderate cooling on postjunctional alpha 1- and alpha 2-adrenoceptors in isolated rings of tail arteries from male Sprague-Dawley rats. Rings were contracted with norepinephrine (NE; 10(-9) to 10(-4) M) alone or in the presence of prazosin (Pz; 3 x 10(-7) M) or rauwolscine (Rw; 10(-7) M). NE concentration-response curves were inhibited by alpha 1-blockade (Pz) but not significantly affected by alpha 2-blockade (Rw). In all rings, cooling caused an increase in the slope of the dose-response curve and a significant increase in the concentration of agonist required to evoke contractions, as assessed by that concentration of NE required to evoke a contraction equal to 10% of maximal (EC10). Cooling inhibited contractions evoked by the selective alpha 1-adrenergic agonist phenylephrine (PE) as assessed by EC10 but had no significant effect on the weak contractions elicited by the selective alpha 2-adrenergic agonist B-HT 920. Prior elevation of tone with either KCl or prostaglandin F2 alpha enhanced alpha 2-mediated contractions. These contractions were augmented by cooling, whereas those caused by either KCl or prostaglandin F2 alpha alone were not significantly affected. Our results suggest that alpha 2-adrenoceptor-mediated responses in this blood vessel are dependent on the level of preexisting tone and are potentiated by cooling.

Human saphenous veins were obtained at surgery and assayed immediately (n = 10). The veins were cut into rings, suspended in organ chambers, and connected to force transducers for the recording of isometric tension. One ring served as control whereas others were treated with the alpha 1-adrenoceptor antagonist prazosin (3 X 10(-7) mol/L), or the alpha 2-adrenoceptor antagonist rauwolscine (10(-7) mol/L). In quiescent rings cooling from 37 degrees C to 24 degrees C had no significant effect. Norepinephrine (10(-8)-10(-5) mol/L) caused concentration-dependent contractions with an EC20 (-log concentration of norepinephrine required to induce contractions 20% of maximal) = 6.97 +/- 0.10. The contractions were inhibited by prazosin (EC20 = 5.89 +/- 0.17, p less than 0.001) and rauwolscine (ED20 = 5.78 +/- 0.11, p less than 0.001). In control rings cooling potentiated contractions evoked at concentrations of norepinephrine below 10(-6) mol/L and inhibited those at higher concentrations. In rings treated with alpha-antagonists cooling depressed the maximal contractile responses. Contractions to the alpha 1-agonist, phenylephrine (10(-7)-10(-4) mol/L), were inhibited by cooling, whereas those to the alpha 2-specific agonist B-HT 920 (10(-7)-10(-4) mol/L) showed a pattern similar to that seen with norepinephrine. The data indicate that the human saphenous vein possesses both alpha 1- and alpha 2-adrenoceptors postjunctionally, and that both contribute to contractile responses. Cold augments saphenous vein reactivity to norepinephrine by an apparent increase in the responsiveness of alpha 2-adrenoceptors to agonists. The relationship between temperature and adrenoceptor responsiveness may be of pivotal importance in defining the mechanism of cold-induced vasospasm.

1. Previous studies have suggested that elevated ketone levels are associated with increased survival time in rodents exposed to hypoxia. In this study the association between whole blood BHB (beta-hydroxybutyrate) and hypoxic survival time was investigated in hibernating and non-hibernating ground squirrels and in rats. 2. Non-hibernating ground squirrels and rats were exposed to hypoxia (4.5% O2). One hundred per cent of ground squirrels survived 1 hr of hypoxia vs 20% of rats. 3. Ketone levels were significantly higher in ground squirrels than rats during hypoxia, and rats surviving the longest had the highest ketone levels. 4. When hibernation was induced in ground squirrels there was a significant increase in beta-hydroxy-butyrate from 0.45 to 1.6 mM (P = 0.0005). 5. Ground squirrel heart mitochondrial respiratory control ratios and ATP synthesis rates indicated no preferential ketone utilization which might suggest a possible extramitochondrial role of BHB during hypoxia. 6. We conclude that elevated blood BHB levels are associated with increased hypoxic survival and they may have evolved in response to life-threatening hypoxia as experienced during hibernation.

This study was designed to assess differences in potassium-induced relaxation in two rodents, the Sprague-Dawley albino rat and the 13-lined ground squirrel, Citellus tridecimlineatus. Femoral arteries from both species were cut into helical strips for isometric force recording. After norepinephrine-induced contraction in potassium-free solution, the arterial strips relaxed in response to the introduction of potassium (0.25-20 mM) into the bath. Potassium-induced relaxation was greater in rat than ground squirrel arteries. The concentrations required to induce half-maximal relaxation were approximately 2.5 mM for both species. Potassium-induced relaxation varied with the duration of incubation in potassium-free solution, and with the contractile magnitude induced by varying norepinephrine concentrations. Ouabain inhibited potassium-induced relaxation, with the ground squirrel showing greater sensitivity to the cardiac glycoside than did the rat. Acute cooling (from 37 to 17 degrees C) caused a reduction of the contractile response to norepinephrine in rat arteries, whereas those taken from ground squirrels maintained contractions at, or above, those attained at 37 degrees C. In addition, potassium-induced relaxation in ground squirrel vessels was more sensitive to inhibition by cold than it was in those from the rat. The results show that the characteristics of potassium-induced relaxation (ouabain and temperature sensitivity, magnitude of response, etc.) are species related (ground squirrel vs. rat).

Psychological stress results in a rise in body temperature. Here we report that in rats, hyperthermia induced by open-field stress can be blocked by administration of the antipyretic drug sodium salicylate. These data suggest that this rise in body temperature is a true fever, perhaps mediated by prostaglandins.

Experiments were designed to determine the effects of acute cooling (from 37-24 degrees C) on responses to alpha-1 and alpha-2 adrenergic activation in the central ear artery of the rabbit. Rings were suspended in physiological salt solution for recording of isometric force. Cooling quiescent vessels resulted in an increase in force. This myogenic response was sensitive to depletion of either intra- or extracellular calcium, as well as treatment with the calcium antagonists verapamil and diltiazem. Cooling did not significantly alter contractile responses to norepinephrine in ear arteries under control conditions or under alpha-2 adrenergic blockade (rauwolscine). Cooling arteries under alpha-1 adrenergic blockade (prazosin) caused augmentations which were not significantly different in magnitude from the myogenic responses to cooling exhibited by the same rings when unstimulated. The alpha-1 adrenergic agonist phenylephrine caused concentration-dependent contractions which were not altered by cooling. The alpha-2 adrenergic agonist UK 14,304 evoked only weak contractions; cooling rings exposed to UK 14,304 caused further increases in tension but no more so than when quiescent. A similar pattern was seen to hold for contractions elicited by electrical stimulation. Thus, in the central ear artery of the rabbit cooling appears to have very little effect on adrenergically induced contractions but does cause the development of myogenic tone.

The alteration of renin release by alpha- and beta-adrenoceptors located on the juxtaglomerular cells has been shown to be temperature sensitive in nonhibernating mammals. These experiments investigate the effects of temperature on renin secretion by cortical slices of kidneys from the thirteen-lined ground squirrel Spermophilus tridecemlineatus. At 37 degrees C, beta-stimulation (isoproterenol 10(-7) M) increased the release of renin by slices taken from nonhibernating ground squirrels but had no effect on those taken from hibernating squirrels. The alpha-agonist phenylephrine (10(-5) M) had no effect on slices from nonhibernating squirrels but enhanced the release rate in those from hibernating ground squirrels, providing the first evidence of in vitro stimulation of renin release by an alpha-agonist. When incubated at 11 degrees C, kidney slices from both hibernating and nonhibernating animals were unresponsive to both alpha- and beta-agonists until incubation times were doubled. Under these prolonged conditions, phenylephrine again stimulated renin release. These results indicate that both in vitro and in vivo cooling alter the responses of alpha- and beta-adrenoceptors to renin-releasing stimuli.

Experiments were designed to determine the effects of cooling on alpha-1 and alpha-2 adrenergic responses in a cutaneous vein of the rabbit. Rings of saphenous vein were suspended in physiological salt solution for the recording of isometric force. Cooling (from 37-24 degrees C) caused no significant increase in force in quiescent rings. Similarly, the same degree of cooling had no significant effect on the response to exogenous norepinephrine (10(-9)-10(-5) M), whether under control conditions or in the presence of either the alpha-1 adrenergic antagonist prazosin (3 X 10(-7) M) or the alpha-2 adrenergic antagonist rauwolscine (10(-7) M). Contractions evoked by the alpha-1 adrenergic agonist phenylephrine were reduced, but those induced by the alpha-2 adrenergic agonist UK 14,304 (10(-9)-10(-5) M) were unaffected by the same degree of cooling. Cooling augmented the response elicited by electrical field stimulation of the sympathetic nerves, although only under conditions of alpha-1 or combined alpha-1 and alpha-2 adrenergic blockade. Data obtained with the sympathomimetic tyramine suggest that both alpha-1 and alpha-2 adrenoceptors are innervated in this blood vessel. Together, the present data suggest that the effects of acute cooling on the saphenous vein of the rabbit, unlike that of the dog, are not mediated by changes in the affinity of postjunctional alpha-2 adrenoceptors.

Experiments were designed to determine the mechanism of action underlying relaxation of vascular smooth muscle induced by ammonium ions. In particular, the possibility that these ions might be an endothelium-derived relaxing factor was examined. Rings of large canine femoral, mesenteric and coronary arteries and of small arteries from the gracilis muscle were suspended in organ chambers for the recording of isometric force. Membrane potential was recorded with intracellular microelectrodes in smooth muscle cells from the mesenteric artery. Ammonium ions induced relaxation which were independent of the presence of the endothelium. The relaxations were not prevented by adrenergic, serotonergic, muscarinic and histaminic blockers, by scavengers of oxygen-derived radicals or by inhibitors of soluble guanylate cyclase. The relaxations were prevented by a decrease in extracellular calcium concentration and by inhibition of the Na+/K+ pump. The results are compatible with the hypothesis that the relaxation induced by ammonium ions is related to changes in intracellular pH and, at high concentration of these ions, possibly to activation of the Na+/K+ pump. Ammonium ions are neither the endothelium-derived relaxing factor which activates guanylate cyclase nor the factor that induces endothelium-derived hyperpolarization. Inasmuch as relatively low concentrations of the ion induce relaxation of small arteries of skeletal muscle, they could contribute to exercise hyperemia.

Peripheral vascular resistance in the ground squirrel (Spermophilus tridecemlineatus) increases when the animal enters hibernation. The goals of this study were to determine if a change in vascular reactivity contributes to this hemodynamic response, and to compare the effects of temperature on vascular responsiveness in a hibernator (ground squirrel) and a nonhibernating mammal (rat). Helically cut strips of aortae and femoral arteries were mounted in organ chambers (37 degrees C) and isometric contractions were recorded. The arteries were made to contract in response to exogenous norepinephrine (5.9 X 10(-7) M). Cooling the organ chamber (11 degrees C) potentiated contractions to norepinephrine (5-15% increase) in ground squirrel femoral arteries but depressed those (80-100% decrease) in ground squirrel aortae and rat aortae and femoral arteries. Contractions in response to depolarizing concentrations of potassium in ground squirrel femoral arteries were depressed by cooling (11 degrees C), suggesting that the augmented response to norepinephrine at low temperature is specific. Treatment with indomethacin, propanolol, and ouabain did not alter the potentiating effect of temperature on contractions to norepinephrine in ground squirrel femoral arteries. Apparently, the potentiation is not related to prostaglandins generated in the vascular wall, to blockade of beta-adrenergic receptors, nor to inhibition of the electrogenic sodium pump. The observations are consistent with the hypothesis that a change in vascular responsiveness contributes to the regional control of blood flow in hibernation. This adaptive response is specific in that it does not occur in the aorta of the ground squirrel and the response is not present in the vasculature of the rat, a nonhibernating mammal.