Selegiline potentiates the effects of EGb 761 in response to ischemic brain injury.

We evaluated whether combined treatment with selegiline, a selective MAO-B inhibitor, and EGb 761, a standard extract of Ginkgo biloba, has synergistic effects against ischemic reperfusion injury (IRI) in gerbils. Interestingly, we observed that pretreatment with EGb 761 significantly attenuated selegiline-induced hyperactivity. This finding paralleled striatal fos-related antigen immunoreactivity (FRA-IR) in mice. Four minutes of bilateral carotid artery occlusion caused substantial cell loss in the CA1 of the hippocampus 5 days post-ischemic insult. Pretreatment with EGb 761, with or without selegiline, significantly attenuated this neuronal loss. Combined treatment with EGb 761 plus selegiline was more efficacious in preventing this loss. Synaptosomal formations of protein carbonyl, lipid peroxidation (malondialdehyde (MDA) + 4-hydroxyalkenal (4-HDA)), and reactive oxygen species (ROS) in the hippocampus remained elevated 5 days post-ischemic insult. The antioxidant effects appeared to be most significant in the group treated with EGb 761 plus selegiline. This combined treatment produced more significant attenuation of IRI-induced alterations in intramitochondrial calcium accumulation, the mitochondrial transmembrane potential, and mitochondrial Mn-superoxide dismutase-like immunoreactivity (Mn-SOD-IR) than either treatment alone. Our results suggest that co-administration of EGb 761 and selegiline produces significant neuroprotective effects via suppression of oxidative stress and mitochondrial dysfunction without affecting neurological function.

Differential effects of magnesium on basal and agonist-induced EDRF relaxation in canine coronary arteries.

In the present study the effects of alterations in extracellular magnesium concentration on spontaneous release of endothelium-derived relaxing factor (EDRF) and acetylcholine-, thrombin-, and calcium ionophore A23187-induced EDRF-dependent relaxation in isolated canine coronary arteries were determined. Increasing the extracellular magnesium concentration from the standard 1.2 to 2.4 mM did not alter the coronary contractile responses, while complete removal of extracellular magnesium resulted in an EDRF-dependent relaxation. The averaged percent relaxation of coronary arteries previously constricted with either prostaglandin F2 alpha, norepinephrine, or barium chloride was -75.9 +/- 4.4 (mean +/- SEM of 27 vessel segments), -43.1 +/- 4.9% (20 segments), and -25.8 +/- 6.9% (15 segments), respectively. Complete removal of extracellular magnesium also resulted in a slight but significant rightward shift of the concentration-response curves of acetylcholine- and thrombin-induced EDRF-dependent relaxation. The maximum relaxation was decreased to 86.5 +/- 1.7% and 69.5 +/- 4.6% of the control acetylcholine and thrombin relaxation, respectively. In contrast, the calcium ionophore A23187-induced EDRF-dependent relaxation was not altered following magnesium withdrawal, nor was the isoproterenol-induced endothelium-independent relaxation. These results suggest that extracellular magnesium exerts a direct inhibitory effect on the spontaneous release of EDRF and is apparently important for the expression of endothelium-dependent relaxation induced by acetylcholine and thrombin, but not calcium ionophore A23187, in canine coronary arteries.

Altered angiotensin-converting enzyme in lung and extrapulmonary tissues of hypoxia-adapted rats.

The effects of exposing rats to hypoxia (10% O2) at normal atmospheric pressure for periods of 14 or 28 days on angiotensin-converting enzyme (ACE) activity and stores of angiotensin I (ANG I) and angiotensin II (ANG II) in lung, kidney, brain, and testis were examined. ACE activity was measured by spectrophotometric assay, and active sites of ACE were estimated by measuring the binding of 125I-351A [N-(1-carbonyl-3-phenyl-propyl)-L-lysyl-L-proline], a highly specific active site-directed inhibitor of ACE, to tissue homogenates and perfused lungs. Hypoxia exposure produced progressive reductions in ACE activity in lung homogenates and in ACE inhibitor binding to perfused lungs. ANG II levels in lungs from hypoxia-adapted animals were significantly less than air controls, suggesting that the reduction in intrapulmonary ACE activity was associated with reduced local generation of ANG II. ACE activity was increased in kidney and unchanged in brain and testis of hypoxia-adapted rats compared with air controls. Thus the effects of chronic hypoxia on catalytically active ACE and ACE active sites in the intact animal were organ specific. Adaptation to chronic hypoxia did not significantly alter plasma renin activity or ANG I or ANG II levels or serum ACE content. The hypoxia-induced alterations in lung and kidney ACE were reversible after return to a normoxic environment.

Effects of leukotriene B4 on permeability, prostacyclin and thromboxane release by normal and oxygen-preexposed isolated, perfused rat lungs.

This study assessed the hemodynamic and permeability effects of exogenous, synthetic leukotriene B4 (LTB4) on normal rat lungs and lungs from rats preexposed to oxygen for 48 h, which were isolated and perfused at constant flow in vitro. Adult, Sprague-Dawley rats were exposed to air or greater than 97% O2 for 48 h. After exposure, their lungs were removed from the thorax, ventilated with normoxic gas, and perfused at 12 ml/min with Krebs-Ringer bicarbonate buffer which contained 5 mM glucose and 3 mg/ml albumin. A total of 5.55 micrograms of synthetic LTB4 was infused in three separate boluses over 15 minutes. Perfusion and airway pressures were monitored, and the lungs release of 6-ketoprostaglandin F1 alpha and thromboxane B2 (TXB2) into the effluent from the pulmonary vasculature was measured by radioimmunoassay. The LTB4 had no measureable effects on pulmonary vascular pressures. LTB4 infusion caused a pronounced increase in permeability, indicated by increased albumin concentrations in alveolar lavage fluid from O2-preexposed lungs. Release of TXB2 from both air- and O2-preexposed lungs was increased after LTB4 infusion, while the change in 6-ketoprostaglandin F1 alpha release was not statistically significant. Both the increase in permeability enhanced TXB2 released after LTB4 infusion were inhibited by 10 microM indomethacin in the perfusate. These data indicate that exogenous LTB4 increases microvascular permeability in O2-exposed lungs in association with increased release of TXB2 into the pulmonary vascular effluent.

Hypoxia-induced oxygen tolerance: maintenance of endothelial metabolic function.

Hypoxia (10%-12% O2) preadaptation for 4-7 days effectively protects rats from oxygen toxicity. The present study was designed to investigate the hypothesis that the lung's microvascular endothelium shares in development of oxygen tolerance and therefore that endothelial metabolic function would be protected from oxygen toxicity by prior adaptation to hypoxia. Since pulmonary oxygen toxicity decreases lung capillary angiotensin converting enzyme (ACE) activity, we assayed converting enzyme active sites in an isolated perfused rat lung preparation as a marker for the development of oxygen toxicity and tolerance. Rats were exposed to air, hypoxia (10% O2 for 4 days), hyperoxia (greater than 95% O2 for 2 days) alone, or hypoxia followed immediately by hyperoxia. Lung vascular ACE content was quantitated by measuring the single pass binding of an iodinated-converting enzyme inhibitor, 125I-MK351A, a derivative of lisinopril. Hypoxia adaptation per se had no effect on ACE content reflected in normal 125I-MK351A binding, whereas hyperoxia exposure caused a significant decrease in lung vascular ACE. Hyperoxia-induced decreases in ACE content were prevented partially by hypoxia adaptation, indicating that ACE content on luminal endothelial surfaces was protected from oxygen toxicity. In isolated perfused lungs 125I-MK351A binding reflects development of oxygen tolerance after hypoxia preadaptation and suggests that lung endothelial metabolic function is protected from oxygen toxicity.

Lung MK 351A uptake after hypoxia adaptation and subsequent hyperoxia exposure.

This study investigated the effects of hypoxia adaptation (10% O2 for 4 days) on rat lung angiotensin-converting enzyme (ACE) content before and after hyperoxia exposure (greater than 95% O2 for 2 days). The rationale for this investigation was that hyperoxia exposure decreases lung ACE, while hypoxia adaptation produces tolerance (improved survival) to oxygen toxicity in rats. Rats were exposed to air, hypoxia, hyperoxia alone, or hypoxia followed immediately by hyperoxia. The lungs were then excised and perfused in vitro at 12 ml/min with buffer. Lung ACE content was quantitated by measuring the single-pass binding of an iodinated ACE inhibitor, 125I-MK 351A, a derivative of lisinopril. We showed previously that 125I-MK 351A binding correlates quantitatively with ACE activity in lung homogenates and isolated, perfused lungs. Lung internal surface area was estimated by measuring the mean alveolar diameter of 5 micron hematoxylin and eosin sections from lungs fixed in inflation (25 cmH2O transpulmonary pressure). Hypoxia adaptation per se had no effect on 125I-MK 351A binding or estimated alveolar surface area, while hyperoxia exposure caused a significant decrease in both 125I-MK 351A binding and alveolar surface area. These hyperoxia-induced decreases were prevented partially by hypoxia adaptation, indicating a protective effect on both ACE content and surface area. 125I-MK 351A binding in isolated perfused lungs changed in parallel with histologically estimated surface area. These results indicate that hypoxia preadaptation minimizes the oxygen-induced decrease in lung microvascular ACE content.

Hypoxia-induced inhibition of converting enzyme activity: role in vascular regulation.

Systemic and pulmonary vascular reactivity to graded doses of angiotensin I (ANG I), angiotensin II (ANG II), and, as a control, phenylephrine were examined in 14- or 28-day hypoxia-exposed and air control rats. Hypoxic rats exhibited pulmonary hypertension that was reversible on return to room air, but systemic arterial pressure was not altered by hypoxia. Systemic pressor responses to ANG I and ANG II were significantly less in the hypoxic rats than in the control rats at 14 and 28 days but returned to control levels in hypoxic animals that were then returned to room air, demonstrating reversibility of the hypoxia-induced changes in vascular reactivity. Pulmonary pressor responses to ANG I were significantly less at 14 days, whereas responses to ANG II were significantly greater at 28 days, in hypoxic rats than in controls. There were no significant differences in systemic and pulmonary pressor responses to phenylephrine between the hypoxic and air control animals. The altered systemic and pulmonary pressor responsiveness to ANG I and ANG II in hypoxic rats is probably related to mechanisms specific to the renin-angiotensin system, such as inhibition of intrapulmonary angiotensin-converting enzyme activity and down regulation of ANG II receptors in the systemic circulation. Further study is needed to elucidate these mechanisms.

Role of vasopressin in the cardiovascular effects of LY171555, a selective dopamine D2 receptor agonist: studies in conscious Brattleboro and Long-Evans rats.

To elucidate the role of central and peripheral arginine vasopressin (AVP) in the cardiovascular action of LY171555 in conscious rats, we have examined the effects of LY171555 on mean arterial pressure, heart rate, plasma AVP and catecholamine levels in conscious, congenitally AVP-deficient Brattleboro (DI) rats and Long-Evans (LE) control rats. Administration of LY171555 (1 mg/kg i.v.) increased heart rate without altering mean arterial pressure in DI rats but increased both mean arterial pressure and heart rate in LE rats. After pretreatment with domperidone, LY171555 induced both a pressure response and a tachycardia in DI rats. Domperidone pretreatment enhanced the pressor action of LY171555 and attenuated the LY171555-induced tachycardia in LE rats. After pretreatment with phenoxybenzamine, LY171555 induced a depressor and bradycardic response that could be blocked by pretreatment with domperidone in DI rats. Pressor and bradycardic responses to LY171555 were attenuated by phenoxybenzamine pretreatment in LE rats. LY171555 administration increased plasma norepinephrine and epinephrine levels in both DI and LE rats but increased plasma AVP only in LE rats. The vasopressor effect of the alpha-1 adrenoceptor agonist phenylephrine was significantly attenuated in DI rats compared with LE rats, whereas the pressor action of angiotensin II was similar in both groups. These results suggest that the pressor action of LY171555 in conscious LE rats is mediated by an increase in plasma AVP and by activation of sympathetic outflow through the central D2 dopaminergic system but not through the central vasopressinergic system.(ABSTRACT TRUNCATED AT 250 WORDS)

Magnesium deficiency produces endothelium-dependent vasorelaxation in canine coronary arteries.

In the present study, effects of magnesium deficiency on coronary vascular reactivity changes were studied in isolated canine coronary arteries before and after endothelium disruption. Lowering of extracellular magnesium produced a potent vasorelaxant response in coronary arteries with intact endothelium, whereas in arteries with disrupted endothelium a potent vasoconstrictory response was observed. The magnitude of magnesium deficiency-induced vasorelaxation is also dependent on the extracellular calcium concentrations, such that lowering of extracellular calcium from 1.75 to 0.22 mM completely abolished the magnesium deficiency-induced vasorelaxant effects. Similar inhibition, as well as reversal, of magnesium deficiency-induced vasorelaxation was also observed with dichlorobenzamil, a known inhibitor of the Na+/Ca++ exchange mechanism, but not with nifedipine, a known inhibitor of the voltage-operated calcium channel. The magnesium deficiency-induced vasoconstriction, on the other hand, is endothelium-independent. Although a similar dependency on extracellular calcium concentration was observed in this vasoconstrictory response, nifedipine, but not dichlorobenzamil, was found to be effective in inhibiting the vasoconstrictory effect. Hemoglobin, a known inhibitor of the endothelium-derived relaxing factor, also completely inhibited the magnesium deficiency-induced vasorelaxation but had no effect on the latter vasoconstrictory effect. These results suggest that circulating magnesium ion may have a dual effect in the modulation of coronary vascular reactivity. In coronary vessels with intact endothelium, lowering of extracellular magnesium may potentiate calcium-induced basal release of endothelium-derived relaxing factor and result in vasodilation, whereas in pathological vessels with injured endothelial cells, a similar lowering of magnesium may increase coronary vascular tone by accentuating the calcium-induced coronary constrictory response in vascular smooth muscle cells and result in vasoconstriction.