Inhibition of aldosterone production in adrenal cell suspensions by serum from patients with manic-depressive psychosis.

A method of preparing a suspension of cells of the zona glomerulosa from rat adrenal capsules treated with crude collagenase is described. The cells responded to ACTH, angiotensin II and serotonin by increased production of aldosterone. Pooled human sera or individual human sera (from healthy normal or non-psychiatric in-patients) to a final concentration of 30% had no effect on ACTH-stimulated production of aldosterone. Many serum samples from five patients with manic-depressive psychosis, however, caused a reduction in aldosterone production; 65% of those samples taken during depression, 44% of the samples taken during manic episodes and 23% of the samples taken when the mood was normal. Sera from manic-depressive patients also reduced the production of aldosterone caused by angiotensin II or serotonin. This effect of serum from manic-depressives in vitro may be related to the abnormalities of aldosterone control in such patients.

The inhibition of human prostatic aromatase activity by imidazole drugs including ketoconazole and 4-hydroxyandrostenedione.

Ketoconazole, an orally active imidazole drug and bifonazole, clotrimazole, econazole, isoconazole, miconazole and tioconazole are known as inhibitors of cytochrome P450 dependent steroidogenic enzymes including human placental aromatase. The aim of the present study was to investigate the effectiveness of these imidazole drugs to inhibit human prostatic aromatase activity compared with the known inhibitor of aromatase 4-hydroxyandrostenedione (4-OHA). The imidazole drugs and 4-OHA inhibited prostatic aromatase activity in a dose-dependent manner. The order of decreasing inhibitory potency determined from IC50 values (mumol/L) was: 4-OHA (1.57) greater than bifonazole (1.6) greater than tioconazole (1.69) greater than clotrimazole (1.73) greater than econazole (1.87) greater than miconazole (2.0) greater than isoconazole (2.2) greater than ketoconazole (4.7). The IC50 values for the inhibition of prostatic homogenate aromatase activity are 3-9-fold higher than that for the inhibition of human placental aromatase activity, previously reported, except that of ketoconazole which was 1.5-fold lower than that for the inhibition of placental microsomal aromatase.

A method for deriving normal ranges from laboratory specimens applied to uric acid in males.

Plasma specimens from male blood donors gave a normal range for uric acid concentration of 3.3 to 7.5 mg/100 ml. Specimens from male outpatients, which were received in the laboratory for the analysis of urea but not of uric acid and proved to have normal concentrations of urea, showed a positively skewed distribution of values for uric acid from which a ;normal range' of 3.2 to 7.6 mg/100 ml was derived. We suggest that specimens from outpatients selected in the way described here may prove to be a convenient source of data for determining the normal range of other substances.

The effect of ibuprofen on the excretion of steroid metabolites.

An inexpensive gas chromatographic method is described that allows simultaneous measurement in urine of androsterone (A), aetiocholanolone (E), 11-hydroxyandrosterone (11-OA), 11-hydroxyaetiocholanolone (11-OE), pregnanediol (PD), pregnanetriol (PT), tetrahydrocortisone (THE) and tetrahydrocortisol (THF). Dehydroepiandrosterone was also resolved by the column. Ibuprofen was administered to five healthy normal males at a dose used therapeutically in rheumatoid arthritis (RA). The above urinary steroids were measured weekly during a control period, during a four week period of drug treatment and for four weeks after drug treatment had ceased. The excretion of A fell to a mean of 63% of the control value (p less than 0.02) and returned to the control value within two weeks. 11-OA, which showed a greater variability than A, fell to the same extent (p less than 0.1). No other steroid measured showed a change that could be related to the drug. This relatively limited effect of ibuprofen on steroid metabolism makes it a suitable drug for maintaining patients with RA during studies of their steroid metabolism.

Effects of analytical error on the clinical discrimination of the urinary 11-hydroxycorticosteroid assay.

A model consisting of overlapping Gaussian distributions of disease and reference values has been used to calculate the effects of analytical imprecision on the proportions of patients wrongly classified by a test. Using published data the model has been applied to the urinary excretion of 11-hydroxycorticosteroids in the diagnosis of Cushing's syndrome. At the lowest levels of imprecision encountered in hospital laboratories, doubling the imprecision increased false negatives (missed diagnoses) from 3.6% to 4.1% and false positives from 9.4% to 11.2%. Although improved imprecision may thus produce only marginal improvements of misclassification, there is no level of imprecision below which further improvement does not improve misclassification. It is suggested that the concept of an 'acceptable level of imprecision' should be replaced by the concept of costing improvements of imprecision, equating the benefits in terms of patient classification with cost in terms of additional resources.

Suppression of plasma androgens by the antiandrogen flutamide in prostatic cancer patients treated with Zoladex, a GnRH analogue.

Chronic treatment with the GnRH (gonadotrophin hormone releasing hormone) agonist Zoladex causes suppression of testicular androgens. Use of antiandrogens has been advocated to block the effects of the initial surge of androgens, and to block any presumed effects of adrenal androgens. We have measured plasma concentrations of androgens and possible precursors before and during treatment in the following prostate cancer patients: 10 who received Zoladex alone (Z), nine who received Zoladex + the anti-androgen flutamide (Z + F) and five who were orchidectomized (O). Testosterone fell in the Z + F group to 0.84 +/- 0.21 nmol/l (mean +/- SD) significantly lower (Wilcoxon P less than 0.05) than after Z (1.58 +/- 1.84 nmol/l) alone. Progesterone and 17 alpha-hydryxyprogesterone did not change significantly in any group. Androstenedione and dehydroepiandrosterone sulphate (DHAS) showed significant falls in Z + F (from 3.44 +/- 0.34 to 1.92 +/- 0.18 mumol/l and from 3.88 +/- 0.64 to 1.92 +/- 0.36 mumol/l respectively) but not in other groups. These results are consistent with our demonstration of an inhibitory effect of flutamide, hydroxyflutamide and other antiandrogens on human adrenal microsomal 17 alpha-hydroxylase and 17,20-lyase activities in vitro.

Inhibition of human adrenal steroidogenic enzymes in vitro by imidazole drugs including ketoconazole.

The effect of several imidazole containing drugs including keto on human adrenal 17 alpha-hydroxylase, 17,20-lyase, 21-hydroxylase, 11 beta-hydroxylase and 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta-HSD-I) activities was studied in vitro. The order of decreasing inhibitory potency as determined from ID50 values for both 17 alpha-hydroxylase (ID50 values ranged from 1.13-4.17 mumol/l) and 17,20-lyase (0.57-1.95 mumol/l) activities was: bifon greater than clot greater than keto greater than micon greater than econ greater than isocon greater than tiocon. Using [3H]progesterone (5.50-12.25 mumol/l) as the substrate for the 21-hydroxylase activity the order of decreasing inhibitory potency was: clot greater than bifon greater than isocon greater than micon greater than tiocon greater than econ greater than tiocon greater than keto. For the 11 beta-hydroxylation of [3H]deoxycortisol (1.48-2.34 mumol/l) the order of decreasing inhibitory potency was keto greater than bifon greater than clot greater than micon greater than econ greater than isocon greater than tiocon. The cytochrome P-450 dependent enzyme most sensitive to inhibition was 17,20-lyase and the least sensitive was 21-hydroxylase whereas the imidazole drugs were without effect on the cytochrome P-450 independent 3 beta-HSD-I activity. In agreement with previous results a common structural feature of the imidazole drugs having an inhibitory effect was the presence of aromatic rings on the N-1 substituent of the imidazole ring.

Inhibition of rat testicular 17 alpha-hydroxylase and 17,20-lyase activities by anti-androgens (flutamide, hydroxyflutamide, RU23908, cyproterone acetate) in vitro.

Flutamide, hydroxyflutamide, RU23908 and cyproterone acetate (CPA) inhibited rat testicular microsomal 17 alpha-hydroxylase and 17,20-lyase activities in vitro. The Km of [3H] progesterone for 17 alpha-hydroxylase was 45 +/- 0.62 nmol/l (+/- SEM, n = 12) and the Km of [3H] 17 alpha-hydroxyprogesterone for 17,20-lyase was 192 +/- 0.42 nmol/l (+/- SEM, n = 12). The Ki values for 17 alpha-hydroxylase, determined from Lineweaver-Burk plots were 102 +/- 3.2 mumol/l (+/- SEM, n = 6), 363 +/- 3.8 mumol/l (+/- SEM, n = 6), 118 +/- 1.4 mumol/l (+/- SEM, n = 6) and 123 +/- 2.1 mumol/l (+/- SEM, n = 6) for flutamide, hydroxyflutamide, RU23908 and CPA respectively. Flutamide and CPA were mixed-type inhibitors, whereas hydroxyflutamide and RU23908 were competitive inhibitors of 17 alpha-hydroxylase activity. Ki values for 17,20-lyase were 33 +/- 3.1 mumol/l (+/- SEM, n = 6), 112 +/- 3.1 mumol/l (+/- SEM, n = 6), 69 +/- 4.4 mumol/l (+/- SEM, n = 6) and 71 +/- 3.2 mumol/l (+/- SEM, n = 6) for flutamide, hydroxyflutamide, RU23908 and CPA, respectively. Inhibition was found to be competitive in each case. Although the characteristic action of anti-androgens is at the receptor level, these results demonstrate that anti-androgens may also have inhibitory effects on androgen biosynthesis which could prove to be of clinical significance.

Inhibition of testicular 17 alpha-hydroxylase and 17,20-lyase but not 3 beta-hydroxysteroid dehydrogenase-isomerase or 17 beta-hydroxysteroid oxidoreductase by ketoconazole and other imidazole drugs.

Ketoconazole, an orally active antifungal drug, is known to inhibit testicular androgen production both in vitro and in vivo. The aim of the present study was to examine the effect of ketoconazole and 13 other imidazole drugs on rat testicular microsomal 17 alpha-hydroxylase, 17,20-lyase, 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta-HSD-I) and 17 beta-hydroxysteroid oxidoreductase (17 beta-HSOR). The order of decreasing inhibitory effect (determined from Ki values) on 17 alpha-hydroxylase (substrate [3H]progesterone; Km = 89 +/- 0.65 nmol/l; SEM, n = 8) was bifonazole (Ki = 86 +/- 3.3 nmol/l; SEM, n = 4) greater than ketoconazole (160 +/- 4.92) greater than clotrimazole (170 +/- 5.81) greater than miconazole (599 +/- 7.22) greater than econazole (688 +/- 6.98) greater than tioconazole (901 +/- 1.71) greater than isoconazole (1090 +/- 6.96) and on 17,20-lyase (substrate, [3H]17 alpha-hydroxyprogesterone; Km = 250 +/- 0.75 nmol/l; SEM, n = 8) was bifonazole (56.5 +/- 3.4) greater than clotrimazole (81.5 +/- 3.1) greater than ketoconazole (84 +/- 3.5) greater than miconazole (243 +/- 6.3) greater than econazole (325 +/- 5.1) greater than tioconazole (505 +/- 5.2) greater than isoconazole (610 +/- 6.34). However, these imidazole drugs did not inhibit the 3 beta-HSD-I or 17 beta-HSOR activities. A common structural feature of the imidazole drugs having an inhibitory effect was the presence of one or more aromatic rings on the imidazole side chain. In contrast, the imidazole drugs having the imidazole ring fused to a benezene ring, i.e. benzimidazoles (astemizole, mebendazole, thiabendazole) and those having an aliphatic side chain on the N-1 of the imidazole ring (carbimazole, metronidazole, nimorazole, tinidazole) did not inhibit 17 alpha-hydroxylase, 3 beta-HSD-I or 17 beta-HSOR enzyme activities. However some did inhibit 17,20-lyase activity but only at high concentrations. The results of the present study suggest that some imidazole drugs may be useful in clinical situations requiring the suppression of androgen production, for example in the treatment of hormone-dependent prostatic cancer.

Structure-activity relationships of the inhibition of human placental aromatase by imidazole drugs including ketoconazole.

The aim of the present study was to investigate the effectiveness of several imidazole drugs to inhibit human placental aromatase compared with the known inhibitors of aromatase, 4-hydroxyandrostenedione (4-OHA) and aminoglutethimide (AG). Inhibition was similar with both androstenedione and testosterone as substrates. The order of decreasing inhibitory effect (determined from ID50 values) was: 4-OHA greater than tioconazole greater than econazole greater than bifonazole greater than clotrimazole greater than micomazole greater than isoconazole greater than ketoconazole greater than AG greater than nimorazole. The imidazole drugs and AG were reversible inhibitors of aromatase activity, in contrast 4-OHA was an irreversible inhibitor. Astemizole inhibited less than 40% whereas metronidazole, carbimazole, mebendazole, tinidazole and thiabendazole inhibited less than 20% of aromatase activity at 100 mumol/l. The imidazole drugs and AG were without effect on 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta-HSD-I) and 17 beta-hydroxysteroid oxidoreductase activity. In contrast 4-OHA was found to be a potent, reversible inhibitor of 3 beta-HSD-I with an ID50 value of 2.15 mumol/l. A common structural feature of the imidazole drugs having an inhibitory effect was the presence of one or more aromatic rings on the N-1 substituent. In contrast, the imidazole drugs having the imidazole ring fused to a benzene ring, i.e. benzimidazoles (astemizole, mebendazole, thiabendazole) and those having an aliphatic side chain on the N-1 of the imidazole ring (carbimazole, metronidazole, nimorazole, tinidazole) were only weak inhibitors of aromatase.

The effect of ketoconazole related imidazole drugs and antiandrogens on [3H] R 1881 binding to the prostatic androgen receptor and [3H]5 alpha-dihydrotestosterone and [3H]cortisol binding to plasma proteins.

Ketoconazole an orally active imidazole drug and bifonazole, clotrimazole, econazole, isoconazole, miconazole and tioconazole are known inhibitors of cytochrome P-450 dependent steroidogenic enzymes. The aim of the present study was to determine whether these imidazole drugs also have an effect on [3H]R1881 binding to the human prostatic androgen receptor, [3H]5 alpha-dihydrotestosterone (5 alpha-DHT) binding to plasma sex hormone binding globulin (SHBG) and [3H]cortisol binding to plasma corticosteroid binding globulin (CBG). In comparison the effect of both steroidal (cyproterone acetate; CPA) and non-steroidal (anandron, flutamide, hydroxyflutamide, ICI 176344) antiandrogens on these steroid binding proteins was also determined. The results of the present study show that the imidazole drugs were without effect on [3H]R1881 binding to the androgen receptor and on [3H]cortisol binding to CBG up to 100 mumol/l. However, they were weak competitors of [3H]5 alpha-DHT binding to SHBG inhibiting 20-53% of binding at 100 mumol/l. In comparison the antiandrogens were strong competitors of [3H]R1881 binding to the androgen receptor, the order of decreasing potency, determined from ID50 (mumol/l) values were CPA (0.073) greater than ICI 176344 (0.4) greater than anandron (0.63) greater than hydroxyflutamide (1) greater than flutamide (greater than 100). The non-steroidal antiandrogens were without effect on [3H]cortisol binding to CBG whereas CPA caused 36% inhibition of binding at 100 mumol/l. Of the antiandrogens studied CPA was the strongest competitor of [3H]5 alpha-DHT binding to SHBG with an ID50 of 23 mumol/l, in contrast the non-steroidal antiandrogens were weak competitors causing less than 40% inhibition at 100 mumol/l. It is concluded that the primary mode of action of the imidazole drugs is through the inhibition of cytochrome P-450 dependent steroidogenic enzymes with little or no effect on steroid binding proteins. In comparison, the antiandrogens were strong competitors of [3H] binding to the androgen receptor but relatively weaker competitors of [3H] steroids binding to plasma binding proteins.

Arginine vasopressin in manic-depressive psychosis.

Urinary excretions of arginine vasopressin (AVP), sodium, potassium, osmoles and creatinine were measured in three in-patients with bipolar manic-depressive psychosis on at least eight 24-hour periods in each affective phase. Mood and body weight were recorded twice daily. The excretion by each patient of sodium, water and osmoles was greater in mania than during depression. Comparison of electrolytes and osmoles suggested that the increase was due to increased intake of salt and water rather than of total diet. There was a fall of mean AVP excretion during mania, the magnitude of the fall being related to the increase of water throughput. Compared with controls, AVP excretion was high and variable. It did not show the normal relationship to urine osmolality. Days with very high AVP were not associated with any characteristic feature of the other measurements; nor were they confined to any one point in the manic-depressive cycle. AVP does not appear to play a major role in the salt and water changes characteristic of manic-depressive psychosis and we have no evidence of its having any direct relationship to mood changes. We suggest that the observed abnormalities of AVP excretion are another manifestation of the central defect of this disease.

The effect of temperature and anoxia of kidney on the subsequent oxidative phosphorylation of mitochondria.

1. Kidneys were kept anoxic at 4 degrees , 20 degrees and 38 degrees . Mitochondria were then isolated and their oxidative phosphorylation and respiration were determined. 2. Under all conditions the rate of phosphate esterification was affected to a greater extent, or earlier, than oxygen consumption. 3. Glutamate and succinate were used as substrates. The depression of P/O ratio was greater for glutamate at 4 degrees , and for succinate at 20 degrees . 4. Anoxia abolished the inhibiting effect of fluoride on respiration. 5. Phosphate esterification, after anoxia, was higher in the presence of fluoride than its absence, whereas in control preparations they were the same. 6. The decrease in P/O ratio did not appear to be due to activation of adenosine triphosphatase, as activities of both Mg(2+)-and dinitrophenol-activated adenosine triphosphatases were decreased after anoxia.