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Twenty hypertensive subjects participated in three clinical trials of 13 days each, to examine the effects of Alsepa fish oil [20:5, n-3 eicosapentaenoic acid (EPA) 180 mg, and 22:6 n-3 docosahexaenoic acid (DHA) 120 mg] on n-3 for n-6 polyunsaturated fatty acids (PUFA) exchange on serum phospholipids, blood pressure (BP), triglycerides (TG) and primary hemostasis. After 13 days, plasma phospholipids showed an increase in sigma n-3 (EPA and DHA) from 2.0 to 5.9% (P0.01), and a decrease in sigma n-6 (arachidonic acid and linoleic acid) from 29.8 to 22.6% (P0.01). A concomitantly significant reduction in systolic BP (SBP) (158.7 +/- 23.8 mm Hg to 146.5 +/- 17.0 mm Hg, P = 0.04), and diastolic BP (DBP) (80.8 +/- 8.4 mm Hg to 72.9 +/- 14.9 mm Hg, P = 0.04) as well as a significant decrease in platelet adhesion and aggregation on extra cellular matrix measured as a percentage of surface coverage (11.9 +/- 4.8% to 4.2 +/- 3.2%, P = 0.0001) was observed. In addition, a significant reduction in baseline dependent TG was observed; the higher the baseline level TG, the more pronounced the reduction (average 159.2 +/- 74.6 mg% to 108.0 +/- 46.1 mg%, P = 0.001). No change was observed in total cholesterol, high and low density lipoprotein (HDL, LDL), platelet and fibrinogen. Repeated fasting and refeeding with fish oil facilitated plasma exchange of n-3 for n-6 PUFA, improved BP, clinical metabolic parameters and lowered platelet reactivity in the vessel wall (primary hemostasis). In severe and life-threatening situations, the beneficial effects of fish oil should be considered for rapid exchange of n-3 for n-6 PUFA. In this study we describe a novel approach for rapid fatty acid exchange by fasting/refeeding with fish oil supplementation, as well as improved BP, plasma lipids and primary hemostasis. Further research is required on the therapeutic use of fish oils and the physiological mechanisms involved in fatty acid exchange.

Oxygen-free radical intermediates/scavengers were measured in 43 patients with essential hypertension who, although under antihypertensive therapy (without angiotensin-converting enzyme inhibitors), still had high blood pressure values. Measurements were taken before and 30, 60 and 120 min after sublingual administration of 25 mg captopril. Both systolic and diastolic blood pressures were reduced significantly. Twenty normotensive healthy volunteers were used as controls. The hypertensive patients had lower glutathione peroxide activity (GSHPx), higher superoxide dismutase (SOD) and serum glutathione reductase activity (GSHRx) compared with the controls. After captopril (30, 60 min) the glutathione and GSHPx increased compared with the pretreatment values. SOD and GSHRx remained high compared with the controls, while whole blood glucose-6-phosphate dehydrogenase remained low. Another group of 19 essential hypertensive patients, free of any antihypertensive medication (for at least 3 weeks), had lower GSHPx, SOD and higher GSHRx than the normal control group. Our results show significant differences in the oxygen free radical scavenger system of hypertensives compared with the normal subjects. It may be that captopril has a concomitant scavenging action together with its antihypertensive effect. Our study raises the question whether cell/organ damage will occur in hypertensive patients exposed to oxidative stress during periods of low antioxidative capacity.

An RIA procedure for measuring cortisol in the circulation of Tilapia aurea is described and evaluated. Cortisol, chromatographically purified from the fish plasma, accounted for its total immunoreactivity. The correlation between observed and expected cortisol values in the accuracy test was r = 0.977. The within-assay variance was 10% and the between-assays variance was 18.9%. The sensitivity of the whole RIA procedure was 0.24 ng/ml when the dilution was 1:100 and 1.2 ng/ml when the dilution was 1:500. Female T. aurea collected in winter and kept at 17° had regressed ovaries, refused food, and their plasma cortisol levels at rest, measured after 1 and 2 months in captivity, were 74 ± 3.5 and 65 ± 6.1 ng/ml, respectively (mean ± SEM). Two weeks after their exposure to 28° the fish resumed feeding, the GSI had doubled, and the plasma resting cortisol level had decreased to 40 ± 4.0 ng/ml. The cortisol level remained low in the subsequent 11 days. Fish collected in the breeding season (April) had a resting cortisol level of 35 ± 4.0 ng/ml (mean ± SEM, n = 28). Nine days following ovariectomy no change could be found in plasma cortisol. Cortisol level in ovariectomized fish injected with estradiol-17β did not differ from that of oil-injected controls. In this nonmigratory, freshwater fish there is no relation between either the state of the ovary, or ovarian hormone, and the level of cortisol. The high level of plasma cortisol in fish exposed to low temperature may be attributed either to the slow rate of cortisol clearance or to the high activity of the interrenal tissue in the fasting fish.

An electron microscopic description is presented of the mitochondria-rich‘chloride‘cells of the gill epithelium in the cichlid fish Tilapia aurea kept in freshwater or adapted to seawater and injected with the pesticide p,p′DDT. The‘chloride‘cells may have a light or dark (electron dense) appearance. Dark‘chloride‘cells are more common in seawateradapted than in freshwater T. aurea. Characteristic features of‘chloride‘cells include an abundance of mitochondria and an elaborate tubular reticulum, which communicates directly with the basal and lateral membranes of the cells. The apical portion of the‘chloride‘cells is often larger in seawater-adapted than in freshwater fish. Differences in‘chloride‘cell ultrastructure, which can possibly be associated with a response to different salinities, arc described and discussed. No significant ultrastructural differences are detectable in response to injections of the pesticide.

SUMMARY The effect of ACTH on cortisol concentration in the plasma of the carp has been studied; cortisol was determined using a competitive protein-binding radioassay. Blood was sampled at 12.00 h (the diurnal minimum) by cardiac puncture. The plasma cortisol level of undisturbed carp (blood obtained 1–3 min after netting) was 46 ± 14 (s.e.m.) ng/ml. One hour after injection of ACTH (1·0 i.u./100 g body wt., i.m.) the level of cortisol increased to 415 ± 62 ng/ml and reached a maximum of 656 ± 62 ng/ml after 2 h. The level of cortisol in saline-injected control carp after 2 h was 72 ± 33 ng/ml. A transient rise of cortisol was noted in both groups 30 min after injection (315 ± 51, 315 ± 62 ng/ml). This was attributed to a surge of endogenous ACTH, presumably due to handling. Carp head kidneys were superfused in vitro. They released cortisol spontaneously, but the releasing rate declined exponentially and reached a minimum after 135 min. Addition of ACTH to the medium was followed by a prompt increase in cortisol release. The rate of cortisol release was maximal 30–45 min after addition of ACTH and was identical to the rate at the beginning of the superfusion.

The purpose of this study was to investigate possible effects of organochlorine residues on cortisol balance in a freshwater fish. The combined stress of netting, handling and blood sampling in Sarotherodon aureus normally results in a twofold increase of circulating cortisol, within 30–60 min. Fish treated with o,p'DDD (50 mg kg−1) had higher resting level of cortisol than before treatment (155.8±12.7 v. 86.9± 11.2 ngmh−1; n=16). No further increase in cortisol level occurred in these fish after exposure to stress; this inability to respond was maintained for more than 120 days. A recovery of the response to ACTH by superfused interrenal tissue from the treated fish was observed 255 days after treatment. Successive blood sampling of o,p'DDD-treated fish, injected intracardially with labelled cortisol, have shown that the half-life of the steroid in the circulation is prolonged by 370% compared with controls. The hepatic metabolism of cortisol was studied by incubating liver slices with the labelled hormone. Hepatic tissue from o,p'DDD-treated fish metabolized cortisol slower than livers from controls. The high resting level of cortisol in plasma of fish treated with o,p'DDD may be attributed to the retarded metabolism of the steroid by the liver. The lack of augmented cortisol levels in response to stress is attributed to the interference of the organochlorine with the response of the interrenal tissue to the stress-induced surge of endogenous ACTH.

A superfusion system was developed in which secretion of cortisol from the interrenal tissue of Sarotherodon aureus could be stimulated by ACTH. Head kidneys from four fish were cut into fragments and superfused with Eagle's basal medium containing 4 mm-NaHCO3 and bovine serum albumin (50 mg/100 ml). The superfused medium was collected every 15 min before stimulation and every 5 min thereafter. Cortisol was measured in the medium by radioimmunoassay. The rate of cortisol secretion increased considerably after a 5 min pulse of 0·1 mu. porcine ACTH/ml, reaching a peak of 11·64 ± 2·40 (s.e.m.) ng/min (n = 5) after 20 min. In the subsequent 35 min the secretion rate decreased to a level of 0·49 ± 0·21 ng/min which was within the range of the baseline (0·2–0·7 ng/min). The amount of cortisol secreted by the superfused tissue in response to the corticotrophin, calculated from the area under the peak, was dose-dependent at the range of 0·06–273 mu. ACTH. Cortisol secretion in this system could also be stimulated by a crude pituitary extract from the same fish. Using the dose–response line, the adrenocorticotrophic activity in the pituitary gland of S. aureus was estimated as 0·2 i.u./g or 1·87 mu. per gland, in porcine ACTH equivalents. It was possible to substitute ACTH with dibutyryl cyclic AMP. The response of the interrenal tissue, i.e. the increase in secretion of cortisol, was dose-dependent at the range of 2–20 mmol/l. The response of the superfused interrenal tissue to ACTH could be extended by the addition of the synthetic phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (0·1 mmol/l), to the medium. These results indicated that the stimulation of cortisol secretion from the interrenal tissue of this fish by ACTH is, most probably, mediated by cyclic AMP as a second messenger.

Superfused interrenal tissue of Sarotherodon aureus responded to ACTH with increased release of cortisol. The response to ACTH of interrenal tissue taken from fish treated with the organochlorine insecticide 1,1-dichloro-2-(o-chlorophenyl)-2-(p-chlorophenyl) ethane (o,p-DDD; 50 mg/kg) in vivo was almost totally abolished. Lower doses were less effective. The response was also suppressed by exposure in vitro of the interrenal tissue to the organochlorine; there seemed to be a dose–response to o,p-DDD over the range of 0·023 to 1 mg/l. Also 1,1-dichloro-2,2-bis (p-chlorophenyl) ethylene (p,p′-DDE) was effective in suppressing the response to ACTH at concentrations of 50–150 mg/l; 1,1,1-trichloro-2,2-bis (p-chlorophenyl) ethane (p,p′-DDT) or the polychlorinated biphenyl, Aroclor 1254, were ineffective at 50 mg/l. No suppressive effect of o,p-DDD on output of cortisol could be detected in superfusion when N6,O2′-dibutyryl cyclic AMP (20 mmol/l) was substituted for ACTH. This may indicate that the interference of o,p-DDD with the interrenal response to ACTH results from an interruption in the generation of cyclic AMP.