Opioid activities of human β-casomorphins

Summary Opioid activities of human -casomorphin-4,-5,-7 and -8 and, for comparison, of the corresponding bovine -casomorphins were studied in the guinea-pig ileum preparation. Binding parameters, i.e. K _d -values and binding site concentrations, for the interaction of human and bovine -casomorphins with opioid receptors in rat brain homogenates were determined in inhibition experiments, using [³H]-(d-Ala², MePhe⁴, Gly-ol⁵)enkephalin, [³H]-(d-Ala², d-Leu⁵)enkephalin and [³H]ethylketazocin as -, - and -opioid receptor ligands. Analysis of binding data was performed using a non-linear curve fitting program. All -casomorphins examined displayed opioid activity. The affinity was highest for -receptors, less so for -receptors and lowest for -receptors. It is suggested that human -casomorphins might play a role as food hormones.     

A photoetched cell relocation matrix for long-term, quantitative observations of selected individual neurons in culture

A photoetched matrix of indium tin oxide (ITO) on glass has been developed and tested as a tool to assist in the relocation and identification of individual neuronal cells in culture. The matrix is formed by 10-15 micron wide and 300 A thick ITO lines which subdivide a 1-cm2 area into 625 smaller squares. Each of the smaller squares measures 400 micron on a side and contains a photoetched two-letter "address". The address code allows precise relocation of specific regions of a culture as well as verification of the identities of individual neurons selected for repeated observation. Marks at 50 micron intervals along the sides of the address squares permit quantitative analysis of morphological changes, cell migration, reaggregation, etc. The ITO is transparent and does not interfere with visualization of even fine details of cells with high power microscopy.     

Effects of acute and chronic treatment with imipramine on 5-hydroxytryptamine nerve cell groups and on bulbospinal 5-hydroxytryptamine/substance P/thyrotropin releasing hormone immunoreactive neurons in the rat. A morphometric and microdensitometric analysis

Summary Groups of male rats were treated for a period of 14 days with imipramine (10mol/kg) given twice daily. Separate groups of rats received a single dose treatment using the same dose and experimental design as for the repeated treatment. Employing the avidin-biotin immunoperoxidase technique for immunohistochemistry 5-hydroxytryptamine (5-HT)-, substance P (SP)- and thyrotropin releasing hormone (TRH)-like immunoreactivities (IRs) were visualized in consecutive coronal sections of the brain stem and of the spinal cord. The IRs were studied by means of morphometric and microdensitometric procedures using automatic image analysis on profiles representing nerve terminal networks of the ventral horn of the cervical and lumbar enlargements of the spinal cord as well as their coexistence (5-HT/SP and 5-HT/TRH). With the same technique 5-HT IR was measured in the 5-HT nerve cell groups of the medulla oblongata (B 1, B 2, B 3) and of the nucleus raphe dorsalis (B 7) of the midbrain. In addition 5-HT and 5-hydroxyindolacetic acid (5-HIAA) levels were measured in the ventral and dorsal horns of the cervical and lumbar enlargements of the spinal cord using high performance liquid chromatography (HPLC). In the same parts of the spinal cord SP IR was studied by means of radioimmunoassay (RIA).The microdensitometric studies showed that chronic, but not acute, imipramine treatment selectively increased SP IR in the 5-HT/SP/TRH costoring nerve terminals of the medial part of the ventral horn in both the cervical and the lumbar enlargements. Furthermore, quantitative analysis of the entity of coexistence in the 5-HT nerve terminal networks of these areas showed that all the 5-HT nerve terminals contained SP and TRH IRs and that this phenomenon remained after acute and chronic imipramine treatment. The microdensitometric studies on the 5-HT nerve cell groups of the medulla oblongata and of the nucleus raphe dorsalis demonstrated that chronic, but not acute, imipramine treatment selectively increased 5-HT IR in the nerve cell bodies of the lateral part of group B 3 as evaluated from the median grey values. Acute, but not chronic, imipramine treatment significantly increased the field area of 5-HT IR of nerve cell bodies in group B 7, reflecting an increase in the mean profile area of the 5-HT IR nerve cell body profiles. Instead, the mean profile area of 5-HT IR cell bodies of group B 1 was acutely reduced by imipramine.The biochemical studies demonstrated that chronic imipramine treatment selectively reduced 5-HT utilization in the ventral horn of the spinal cord and selectively increased SP IR in the dorsal horn of the lumbar enlargement.In view of these observations it is suggested that chronic imipramine treatment specifically increases SP IR in the 5-HT/SP/TRH costoring nerve terminals of the ventral horn probably related to reduced SP release and reduced 5-HT utilization in these terminals. The results obtained in group B 7 may be explained by a regulation by the³H-imipramine raphe binding sites of fast axonal transport, an influence which may have therapeutic consequences. This mechanism may also be responsible for the increase in 5-HT IR seen upon chronic imipramine treatment in the lateral part of the 5-HT nerve cell body group B 3. Such an effect may lead to a metabolic down-regulation of group B 7, having a possible role for the antidepressant activity of imipramine. The reduction of the mean profile area of 5-HT IR cell bodies of group B 1 seen in the acute treatment can possibly be caused by, noradrenaline (NA) uptake inhibition in inhibitory NA terminals innervating the B 1 group. These results also illustrate the heterogeneities in the responses of the 5-HT nerve cell groups to antidepressant treatment. The ability of chronic imipramine treatment to increase SP IR in the dorsal horn of the lumbar enlargement may reflect the existence of a monoamine-SP interaction in the substantia gelatinosa due to the NA and/or 5-HT uptake blocking activity of imipramine. The existence of such an interaction may help to explain the antinociceptive effect of chronic imipramine treatment.Part of the paper was presented at the 17th International Congress of the International Society of Psychoneuroendocrinology, Bergen, June 29–July 4, 1986.     

Cluster analysis of antibiotic susceptibility patterns of clinical isolates as a tool in nosocomial infection surveillance

Hospital infections represent a major epidemiological problem. The first step in the detection of nosocomial infections consists in assessing the probability that two or more isolates from different patients are similar or different. Many methods are available for typing purposes. Among these, antibiotic susceptibility patterns do not need extra cost or extra work and are available on line every moment they are needed. A mathematical technique of elaboration is proposed for disk zone sizes, in order to assess the probability of two or more clinical isolates to be the same strain. Antibiograms performed according to Kirby-Bauer are evaluated detecting zone sizes by a computer controlled device and then submitted to cluster analysis. Similarity of strains is reported in a dendrogram, in which strains are successively fused. Strains that share a common susceptibility pattern are considered a cluster . At last, epidemiological maps are constructed for each group of strains, in which all the isolates are reported, ordered for patients, plotted on the day the specimen was collected, drawn in a different shape according to the source of specimen, and shadowed by the pattern of its cluster. This method of reporting data directly allows to detect cross infections among patients and can be used as a first typing step before other more expensive procedures.Corresponding author.     

Psychotropic drug utilization patterns in a metropolitan population

Summary Psychotropic drug intake by a random sample of citizens of the city of Munich aged 30–69 years has been assessed. A 1-week prevalence of 9.3% for all psychotropic drug users was found, benzodiazepines accounting for approximately two-thirds (6.6%) of the users. Two-thirds of drug users were women. Drug use in both sexes increased with age. The doses of benzodiazepines prescribed in most cases were less than 10 mg diazepam equivalent per day. Intake of benzodiazepines in combination with analgesics or alcohol (40 g/day) did not appear to represent a major problem. Multiple logistic regression analysis showed that the number of chronic diseases was the strongest predictor of benzodiazepine intake in men, whereas stress and age determined intake in women. Long-term use seemed to be relatively rare at 11% of all benzodiazepine users, so it was not considered to be a severe public health problem.     

Pharmacodynamic modeling of the EEG effects of ketamine and its enantiomers in man

The pharmacodynamics of a racemic mixture of ketamine R,S (±)-ketamine and of each enantiomer, S(+)-ketamine and R(–)-ketamine, were studied in five volunteers. The median frequency of the electroencephalogram (EEG) power spectrum, a continuous noninvasive measure of the degree of central nervous system (CNS) depression (pharmacodynamics), was related to measured serum concentrations of drug (pharmacokinetics). The concentration-effect relationship was described by an inhibitory sigmoid E_max pharmacodynamic model, yielding estimates of both maximal effect (E_max) and sensitivity (IC₅₀) to the racemic and enantiomeric forms of ketamine. R(–)-ketamine was not as effective as R,S(±)-ketamine or S(+)-ketamine in causing EEG slowing. The maximal decrease (mean±SD) of the median frequency (E_max)for R(–)-ketamine was 4.4±0.5 Hz and was significantly different fromR,S (±)-ketamine (7.6 ±1.7 Hz) and S(+)-ketamine (8.3±1.9Hz). The ketamine serum concentration that caused one-half of the maximal median frequency decrease (IC₅₀) was 1.8±0.5g/mL for R(–)-ketamine; 2.0±0.5 g/mL for R,S(±)-ketamine; and 0.8±0.4 g/mL for S(+)-ketamine. Because the maximal effect (E_max) of the R(–)-ketamine was different from that of S(+)-ketamine and R,S(±)-ketamine, it was not possible to directly compare the potency (i.e., IC₅₀) of these compounds. Accordingly, a classical agonist/partial-agonist interaction model was examined, using the separate enantiomer results to predict racemate results. Although the model did not predict racemate results well, its failure was not so great as to provide clear evidence of synergism (or excess antagonism) of the enantiomers.This work was supported in part by a Starter Grant from the American Society of Anesthesiologists, the Biomedical Research Support Grant NIH 2S07RR5353-20, 1981, (P.F.W.); and NIH and NIA Research Grants NS-17956 and AG03104 (D.R.S., A.J.T., L.B.S). The research fellowship of Dr. Schüttler was made possible by a NATO Foundation Grant (300-402-511-3), awarded by the German Academic Exchange Service. This study is part of Dr. Schüttler's Habilitation Thesis for the Faculty of Medicine at the University of Bonn, West Germany. Dr. Verotta is a fellow of the program of advanced training established by EEC and Regione Lombardia on leave of absence from Mario Negri Institute of Pharmacological Research, Milan, Italy.     

Radiographic quantification of alveolar bone level changes

The "random burst" theory has recently been proposed as an explanation of the pattern of periodontal disease progression. The theory predicts that the progression of bone loss at individual sites is not dependent upon previous bone loss and age. A longitudinal radiographic study was designed to test this hypothesis, and to describe the changes in bone level over 2 years in a group of 180 subjects (18-68 years of age) who were not under systematic periodontal treatment. The results indicated that 94% of the sites did not show significant changes in the alveolar bone level during the observation period. The mean annual bone loss for the total population was 0.11 mm. By regressing longitudinal bone loss upon age, it was shown that the rate of bone loss increased rapidly between 33 and 56 years of age while a different pattern was shown for the age intervals 18-32 and 57-68 years. Also, the rate of bone loss increased with increasing initial bone loss. This was less evident in the oldest age group. It was concluded that the progression of bone loss in the present material is consistent with a "burst" theory. However, the progression did not occur randomly with regard to previous loss of alveolar bone and time.     

Symptom patterns and causes of somatization in men: I. Differentiation of two discrete disorders

Two distinct patterns of somatization were identified in 807 Swedish adopted men, using comprehensive lifetime psychiatric and sick-leave records. "Diversiform" somatizers had a high frequency of brief sickness occasions for a wide diversity of complaints, particularly pain in the head, joints, and abdomen. "Asthenic" somatizers had a lower frequency and diversity of complaints. They recuperate more slowly, however, and were more often disabled by fatigue, weakness, and minor illnesses such as upper respiratory infections. Both types of somatizers had associated psychosocial maladjustment, but they had discrete clinical patterns, with infrequent overlap. Diversiform somatizers had a higher risk of alcohol abuse, psychiatric hospitalization, and substandard income than either asthenic somatizers or non-somatizers. Asthenic somatizers had a higher risk of divorce than either diversiform somatizers or non-somatizers. Men with prominent somatization had an excess of psychiatric treatment for alcoholism or anxiety disorders, but, unlike female somatizers, no excess of criminality. These clinical differences suggest that the psychiatric processes associated with somatization may be qualitatively different in men and women. The method used here is generally applicable in genetic epidemiology to identify natural clinical subtypes within a heterogeneous phenotype.     

Correlation between the negative inotropic potency and binding parameters of 1,4-dihydropyridine and phenylalkylamine calcium channel blockers in cat heart

Summary Partially purified plasma membranes were prepared from cat ventricle. The purification factors for the calcium channel ligands (+)-³H-PN 200-110, ³H-nimodipine (1,4-dihydropyridines) and (–)-³-H-desmethoxyverapamil (a phenylalkylamine) were 3.1-, 3.4- and 2.9-fold, respectively, whilst -adrenoceptors labelled with (–)-³H-dihydroalprenolol were purified 3.0-fold.(+)-³H-PN 200-110 bound to 930±140 fmol/mg of membrane protein with a dissociation constant of 70 pmol/l at 25°C. Under the same conditions ³H-nimodipine bound to 490±24 fmol/mg of sites with a K _D of 120 pmol/l. (–)-³-H-desmethoxyverapamil bound to 530±55 fmol/mg of sites with a K _D of 2.47 nmol/l.Twelve 1,4-dihydropyridines were evaluated for binding inhibition constants (K _i) with (+)-³H-PN 200-110 and 13 phenylalkylamines with (–)-³-H-desmethoxyverapamil in radioligand binding assays. Of the twelve 1,4-dihydropyridines evaluated (±)-nitrendipine was the most potent with a K _i-value of 280 pmol/l, nifedipine had a K _i-value of 500 pmol/l and the weakest drug tested, (±)-Bay b4328, had a K _i-value of 14.3 nmol/l. The EC₅₀-values of the same 1,4-dihydropyridines to inhibit the electrically driven cat papillary muscle were 77- to 3,450-fold higher and little correlation existed between K _i and EC₅₀-values.Thirteen phenylakylamines were tested for their potency to inhibit (–)-³-H-desmethoxyverapamil binding. The most potent phenylalkylamine with respect to negative inotropy was (±)-D 595 with an EC₅₀-value of 794 nmol/l, the least potent substance was (±)-Sz 45 with an EC₅₀-value of 39.8 mol/l. The binding inhibition constants for the phenylalkylamines were 13-to 322-fold lower than the values for negative inotropy, but a significant positive correlation between the K _i and EC₅₀-values (n=12, r=0.84) was observed. The absolute differences may reflect the state-dependent binding of phenylalkylamines to the channel.QSAR analysis revealed nearly identical correlations between physicochemical substituent properties on the one hand and binding affinities or functional potency on the other hand. In both cases the electronic properties (F-constant) of ring substituents mainly determine the variance in potency.     

The effect of the β-carboline FG 7142 on the behaviour of male rats in a living cage: An ethological analysis of social and nonsocial behaviour

The effects of FG 7142, a -carboline benzodiazepine receptor partial inverse agonist, on the social behavior of pair-housed rats were investigated. Four 6-min dyadic social encounters in a living cage were observed in a paradigm in which one member of a pair of rats was injected. The four injection groups (n=8) were vehicle control, and FG 7142 at 2.5, 5.0 and 10.0 mg/kg, respectively. All injections were administered 2 min before the start of the first observation trial. Compared to the effects of vehicle alone, FG 7142 decreased aggressive behaviour but did not change the level of total social interaction. Thus there were compensating increases in approaching and avoiding behaviours following the administration of FG 7142. Locomotion declined marginally and immobility increased in FG 7142-injected rats. FG 7142 decreased the incidence of self-grooming. The evidence is consistent with a relatively selective reduction in intraspecies aggression in male rats after the injection of the -carboline inverse agonist.