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The experiments presented here deal with the effects of the inhalation of dichlorvos [dimethyl-(2,2 dichlorvinyl)-phosphate, DDVP] vapor on acetyl-cholinesterase (ACHE) activity in rat bronchial tissue. Exposure to DDVP concentrations of 0.8 and 1.8 μg/l for 3 days reduced ACHE activity in the bronchial tissue (62.8±0.8 and 51.6±1.6% of the control), but did not elicit any changes in blood ACHE activity (101±4.5% of the control each). Higher concentrations (4.3 μg/l) induced a decline in ACHE activity also in the blood (38.2±1.1% of the control). In the histochemical preparations used to demonstrate CHE activity in bronchial tissue (thiolacetic acid method), a staining of the bronchial glands and smooth muscles characteristic of the enzyme activity was strongly reduced after exposure of the animals to even the lowest dose applied (0.2 μg/l). The question of whether localized inhibition of ACHE in the bronchial tissue might cause increases in airway resistance due to activation of a broncho-bronchial reflex is discussed. This efferent cholinergic mechanism has been found to be at least partly responsible for maintenance of bronchospasm and hypersecretion in chronic obstructive deseases of the respiratory system. [ABSTRACT FROM AUTHOR]

Ischemic preconditioning (IP) has been found to intensify ischemia-induced spreading of connexin 43 (Cx43) into the extrajunctional plasma membrane of the rat heart. Since ischemia is known to induce connexin dephosphorylation and plasmalemmal permeabilization, we considered whether IP might delay dephosphorylation of also extrajunctional connexin and thus impede myocyte permeabilization. Regional myocardial ischemia of both 15 and 45 min duration with and without IP, respectively, was induced in anesthetized rats. Sections of the hearts were immunostained for phosphorylated and dephosphorylated Cx43. The ratios of immunofluorescence in gap junctions and extrajunctional membranes, respectively, versus myocyte interiors were determined as relative fluorescence units (RFU). IP, however, was not found to reduce gap junctional dephosphorylated Cx43 (normal perfusion: 1.23 +/- 0.06 RFU; 15 min ischemia without and with IP: 1.80 +/- 0.09 and 2.10 +/- 0.24 RFU; 45 min ischemia without and with IP: 3.12 +/- 0.55 and 2.57 +/- 0.33 RFU, respectively). Extrajunctionally, only dephosphorylated Cx43 was found. Its occurrence was not prevented by IP, it was rather intensified by IP (normal perfusion: 1.02 +/- 0.01 RFU, 15 min ischemia without and with IP: 1.06 +/- 0.03 and 1.15 +/- 0.05 RFU (P < 0.02), and 45 min ischemia without and with IP: 1.30 +/- 0.07 and 1.27 +/- 0.07 RFU, respectively). Although under ischemia, junctional and extrajunctional Cx43 predominated in the dephosphorylated state, only a small fraction of myocytes were found permeabilized to propidium iodide. In conclusion, IP did not prevent ischemia-induced Cx43 dephosphorylation in gap junctions and extrajunctional plasma membranes. It is thus more likely that changes in non-channel functions or other localizations of Cx43 are involved in IP-induced myocardial protection., ((c) 2010 Elsevier Inc. All rights reserved.)

Connexins are known to play an essential role in the ischemic preconditioning (IP) of the heart; their functional role in this process, however, has not been clearly defined. For this reason, anesthetized rats were subjected to regional myocardial ischemia, with or without IP or reperfusion. In frozen sections of hearts, fluorescence immunohistochemical staining for connexin43 (Cx43) was performed. In contrast to undisturbed zones, tissue that had been subjected to ischemia revealed Cx43 immunostaining not only in the gap junctions but also in a conspicuous pattern in the free cellular membranes of the myocytes. In myocardium that was exposed to IP only, the ratio of immunofluorescence intensity in the free cellular membrane to that in the interior of the cell was 1.22 +/- 0.04 (ratio in non-ischemia-exposed area = 1.04 +/- 0.01). When 15 or 45 min of permanent ischemia followed IP, the effect became more evident (ratio = 1.31 +/- 0.03 and 1.46 +/- 0.03, respectively) and proved to be significantly greater than in the corresponding non-IP groups (ratio = 1.16 +/- 0.03 and 1.30 +/- 0.03, respectively, P < 0.01). Reperfusion led to an overall weakening of fluorescence intensities and a disappearance of the observed IP-specific differences. We conclude that IP initiates a redistribution of Cx43 from its natural position in the gap junctions toward the free plasma membrane, thereby improving the cell's chance of survival during the subsequent phase of prolonged ischemia by an unknown, supposedly gap junction-independent, mechanism.

To investigate the localization of the earliest damage in ischemic and ischemic-reperfused myocardium, anesthetized rats were subjected to coronary occlusion for 15, 30, 45, or 90 min. One-half of the animals in each group had no reperfusion, whereas the other half was reperfused for 14 min. With the use of histological methods, preferentially in the periphery of the area at risk, localized zones were detected that lacked the hypoxia-specific increase in NADH fluorescence. The extent of these areas displaying injured tissue was found to be significantly smaller in the ischemic-nonreperfused hearts than in the ischemic-reperfused organs (15-min ischemia: 0.22 +/- 0.12% vs. 43.0 +/- 5.0%; 30-min ischemia: 5.7 +/- 2.7% vs. 64.6 +/- 2.9%; 45-min ischemia: 5.6 +/- 1.2% vs. 66.0 +/- 7.5%; 90-min ischemia: 39.3 +/- 5.5% vs. 86.7 +/- 1.8% of the area at risk). The results point to a localized initiation of the damage close to the surrounding oxygen-supplied tissue during ischemia and an expansion of this injury by intercellular actions into yet-intact areas upon reperfusion.

Myoglobin is known to become nephrotoxic when released in greater amounts from skeletal muscle into the general circulation during shock. The present study deals with the question as to whether a myoglobin-induced increase in vascular tone additionally contributes to the detrimental role of this protein in hypovolemic shock. Anesthetized rats were subjected to 250 mg kg x h(-1) myoglobin infused i.v. during hemorrhagic hypotension of 50 mmHg. Shock survival time was measured, as were blood flow and vascular resistance in kidney, intestine, brain, and heart, using the microsphere method. Rats subjected to only myoglobin or hemorrhage survived a period of >120 min; in contrast, rats, exposed to both myoglobin and hemorrhage died at 68 +/- 9 min. When the animals subjected to only hemorrhage and to myoglobin/ hemorrhage were compared, significantly lower values were found in the latter group with respect to blood flow in the kidney (1.7 +/- 0.1 vs. 0.2 +/- 0.05 ml x min(-1) x g(-1)), small intestine (1.0 +/- 0.1 vs. 0.5 +/- 0.1 ml x min(-1) x g(-1)), cardiac output (112 +/- 5 vs. 62 +/- 10 ml(-1) x min(-1) x kg(-1)), and significantly higher values of total peripheral vascular resistance (0.45 +/- 0.02 vs. 0.81 +/- 0.12 mmHg x min x ml(-1) x kg) at 50 min of hypotension. It is assumed that these effects of myoglobin are induced by its ability to scavenge endogenous nitric oxide, because a modified, non-nitrosylable myoglobin was unable to induce such effects. The results support the view that a pathological release of myoglobin into the general circulation causes increases in vascular resistance of vital organs that may contribute to decompensation of tissue supply when occurring in hypovolemic shock.

Background: Inflammatory leukocyte-endothelium interactions, mediated by selectins, contribute to renal ischemia/reperfusion (I/R) injury. We examined the influence of the soluble P-selectin glycoprotein ligand 1 (sPSGL) on early I/R-induced changes in a rat kidney transplantation model with long cold ischemia., Methods: After 24 hr of cold storage, syngeneic kidneys were grafted into bilaterally nephrectomized rats. Before transplantation, recipients received either 1 mg/kg of sPSGL or vehicle (n=8 per group). Six hours after reperfusion, grafts were removed for light microscopy and immunohistochemistry. Capillary blood flow was measured under a fluorescence microscope by using the concentric-circles method., Results: A greater proportion, 74.7+/-7.2% (sPSGL) vs. 28+/-7.4% (controls), of all dye-labeled outer medullary capillaries appeared in the 12-microm radius (P<0.01), indicating dense blood flow, whereas 7.6+/-2.9% vs. 43.3+/-9.7%, respectively, appeared in the 60-microm radius (P<0.05), indicating rarefied blood flow. In the sPSGL-treated group, the extent of severe tubular damage within the inner stripe of the outer medulla was lower compared with controls (37.5+/-8.3% vs. 78.4+/-3.5%, P<0.01). Outer medullary heat shock protein 72 expression was 14.5+/-1.6% in the sPSGL-treated group compared with 9.6+/-1.4% in controls (P<0.05). The number of infiltrating polymorphonuclear leukocytes was similar in both groups. Treatment with sPSGL had no influence on the serum creatinine level., Conclusions: Our data suggest that impairment of outer medullary blood flow is crucial in I/R injury of kidney grafts with prolonged cold storage. Reduction of capillary blood flow perturbations by sPSGL protects tubular cells from severe structural damage. Blocking early selectin-mediated leukocyte adhesion may have therapeutic implications in improving the prognosis of renal transplants with severe I/R injury.

The influence on myocyte viability of ischemia-induced changes in capillary perfusion was studied in the hearts of anesthetized rats subjected to partial occlusion of the left coronary artery for 45 min. Timed plasma labeling was applied to determine perfusion patterns. Changes in the fluorescence of preloaded potential-sensitive dyes [tetramethylrhodamine methyl ester (TMRM) and bis-oxonol], of trypan blue, and of endogeneous NADH were utilized in characterizing myocyte viability in histological sections of the heart. Within the hypoperfused zone, localized areas appeared vascularly nonlabeled for periods of at least 10 min. Within these areas a reduction in TMRM fluorescence occurred in 82. 5% of the tissue, signaling a reduced resting membrane potential. In the same areas 37.7% of the myocytes revealed an NADH fluorescence lower than that regularly found in anoxic tissues. This correlated with an especially low level of TMRM, with increased fluorescence bis-oxonol and with an accumulation of trypan blue. In conclusion, in localized hypoperfusion-induced zones lacking capillary flow, an inhomogeneous pattern of reductions in myocyte viability develops, which appears to be relevant in ischemia-induced arrhythmias.

The clinical use of alkylphosphocholines (APC) in cancer patients is restricted because of the high gastrointestinal toxicity and the need for oral administration. Therefore we evaluated the clinical pharmacology of erucylphosphocholine (ErPC), the first derivative of the APC family suitable for intravenous administration with strong antineoplastic activity, in vitro and in vivo in rats. The pharmacokinetic parameters after a single intravenous dose of 40 mg/kg were calculated using a two-compartment model: C(max) = 1.6 +/- 0.3 micromol/ml, T(1/2alpha) = 0.18 +/- 0.09 h, T(1/2beta) = 3.3 +/- 0.88 h, clearance = 9.7 +/- 1.2 ml/h, AUC = 2.5 +/- 0.3 micromol/ml per h and Vss = 40.4 +/- 7.9 ml. Biodistribution studies were performed after repeated ErPC administration at different doses. Intravenous injections of 20 mg/kg given at intervals of 48 h for up to 4 weeks were well tolerated. Neither clinical evaluation nor laboratory parameters (haematology and clinical chemistry) revealed toxic side effects. In contrast, higher doses of ErPC (40 mg/kg per 48 h) led to weight loss. After 2 and 4 weeks of therapy with 20 mg/kg per 48 h a high ErPC accumulation was found in the adrenal glands, small intestine and brain. The brain to serum concentration ratios averaged 2.1 after 2 weeks and 4.5 after 4 weeks. Significant leucocytosis and thrombocytosis were observed after 4 weeks of ErPC treatment. The findings suggest that ErPC is a suitable candidate for clinical trials. In particular, owing to the high accumulation in brain tissue, ErPC is a potential agent for chemotherapy against malignant brain tumours.

The effects of myoglobin on renal microcirculation were studied in anesthetized rats subjected to hemorrhagic hypotension. Capillary flow distribution was determined by allowing two dyes to circulate for 3 and 1 min, respectively, freezing the left kidney and quantifying the dye distribution in histological sections by analyzing the distances of regularly spaced test points to the next dye-labeled capillary. Control experiments showed 88% of distances to be < 12 microns in the cortex [medullary outer stripe (OS): 77%, inner stripe (IS): 93%] and no distance to be > 60 microns. Myoglobin induced disturbances in intrarenal perfusion with a significantly higher potency of (Fe2+)- as compared to (Fe3+)-myoglobin. With the reduced species, the fraction of distances > 60 microns increased to 54% in the cortex (OS: 69%; IS: 67%). L-NAME, an inhibitor of nitric oxide synthesis, induced similar defects of perfusion. The cGMP analogue 8-Br-cGMP was able to nearly completely prevent these effects. The results support the view that myoglobin when released during hemorrhagic hypotension impairs renal microcirculation supposedly by scavenging the endogenous relaxing factor nitric oxide.

A method is described which allows to observe the pattern of capillary plasma filling simultaneously with the redox state (NADH-fluorescence) of the myocytes in the hypoperfused myocardium. In anesthetized rats the coronary perfusion pressure was reduced to a defined level, the blood plasma labeled with dye-conjugated albumin and a myocardial biopsy-sampled. Freeze-substitution of histological sections allowed to detect the intravascular plasma label as well as the myocyte NADH-fluorescence on a microscopic level. It was found that areas showing increased cellular NADH-fluorescence did not arise in a distribution congruent with zones of failing capillary plasma flow in the hypoperfused myocardium.