[Metabolism of sulfate and phosphate conjugates of 19-nortestosterone enantiomers].

The sulfate and phosphate conjugates of 19-nortestosterone (17beta-hydroxy-estr-4-ene-3-one) and ent-19-nortestosterone (ent-17beta-hydroxy-estr-4-ene-3-one) have been synthesized. The sodium salts of the conjugates were i.v. infused to 2 women. On days 1--4 following application, the 19-nortestosterone and 17-ketosteroid excretion in urine as well as the 19-nortestosterone conjugation have been estimated. The 19-nortestosterone enantiomers were excreted in nearly the same amounts. In comparison to testosterone, the excretion of unchanged steroid (20--50%) is significantly higher. The sulfate conjugates of the enantiomers are not hydrolyzed in vivo, whereas the phosphate conjugates are completely splitted. Hydrolysis of ent-nortestosterone phosphate takes place obviously more slowly. Urinary 17-ketosteroids are significantly increased only following infusion of 19-nortestosterone phosphate.


Structural changes in the medial and central amygdala of the male rat, following neonatal castration and androgen treatment.

In contrast to the lateral amygdalar region, in the medial and central parts of the amygdala a significant female-like increase of neuronal nuclear volumes was found in adult male rats orchidectomized on the 1st day of life. These permanent structural changes of the medial and central amygdala could be prevented by androgen administration on the 3rd day of life.


[Endocrinologic and pharmacologic-toxicologic findings with ethinyl-estradiol sulfonate. 8th communication. Complement and lysozyme in the serum of beagles after 6 months application of ethinylestradiol sulfonate].

In course of studies for the chronic toxicity of estrogens after oral application two parameters of the unspecific resistance - complement (CH50U and CB50U) and lysozyme - in sera of 30 beagle-dogs were studied. The doses of the daily (over 6 months) applicated hormones were 1,0 mg/kg ethinylöstradiol and 1,0, 0,1 and 0,01 mg/kg ethinylöstradiolsulfonate (depotform). In the majority of all measurements no significant variations of serum complement and -lysozyme could be detected, however after application of high doses of the hormone ethinylöstradiol a little reduction of complement and significant elevations of lysozyme were observed.


Thyroid testis inter-relationship in Indian palm squirrel, Funambulus pennanti Wroughton.

The present work attempts to find out a relationship between testicular functions and thyroid activity in a seasonal breeder, Indian palm squirrel, Funambulus pennanti Wroughton. In addition to radioiodine studies used for assessing thyroid status during different months in the year, biochemical estimations and histological studies have also been undertaken with a view to determine thyroid and testicular functions. Testicular recrudescence in Indian palm squirrel starts in January and peak activity is reached during April and May. June marks the beginning of testicular regression and the maximal regression is reached in October. Fluctuations in thyroid function are clearly manifested by variations observed in radioiodine uptake during different phases of reproductive cycle. Cytoarchitecture of thyroid also changes somewhat during different periods of the year and these changes are in accordance with the results obtained from radioiodine uptake studies. The present investigation suggests that the thyroid has a definite and significant role to play during the period of testicular recrudescence and regression in seasonal breeder Funambulus pennanti and it seems to help in tapering off sexual arousal and regression rather than sudden switching on and off of the reproductive cycle. In addition, it seems to facilitate peak testicular activity. However, mechanisms whereby thyroid influences various testicular functions seem to involve a complex series of interactions.


The response of thyroid C-cell system in rat to long-term hypercalcemia.

Long-term hypercalcemia induced in rats by administration of vitamine D3 and CaCl2 for 60 days resulted in strong hyperplasia and hypertrophy of C-cells. The extent of hyperplasia varied greatly in individual animals. Histochemical reactions, especially the masked metachromasy with toluidine blue, demonstrated cell groups in which no reaction was observed beside those exhibiting a very strong reaction. Impregnation with silver according to Cajal showed a diminished number of argyrophilic granules in the C-cells, which had undergone hyperplasia. The reactions for non-specific esterases and cholinesterases were similar both in the experimental animals and in the controls. An enlargement of the C-cell nuclei in the experimental group was also pronounced. The total serum calcium level was only slightly increased in this group. It is concluded that the results of staining and the enlargement in nuclear volume of C-cells reflect the increased activity of these cells. Hyperplasia of the C-cells may represent a type of adaptation of the endocrine system in order to maintain calcium homeostasis.


Decreased antidiuretic response in diabetes insipidus during prolonged treatment with carbamazepine presumably due to enzym-induction.

Carbamazepine (CA)-induced antidiuresis and fluorescence in urinary cortisol assay was compared during short-term and prolonged treatment in 3 patients with pituitary diabetes insipidus. The fluorescence induced by excreted CA-metabolite(s) (probably CA-10-11 epoxide and its derivatives) was used for the rough clinical estimation of the rate of metabolic elimination of the drug. A marked decrease was observed both in the antidiuretic response induced by CA and excretion of CA metabolite(s) during prolonged treatment - with a fixed dose of CA. These findings suggest that diminishing antidiuretic effectiveness might be due to auto-induction of CA. Auto-induction of CA may explain the delay in improvement of renal concentrating operation in patients with DI during long-term treatment with fixed doses of CA. Another clinical consequence of this mechanism may be the prevention of the development of water intoxication in neuro-psychiatric patients undergoing chronic treatment with CA.


Relationship between plasma renin activity and urinary catecholamines in various types of hypertension.

The urinary excretion of free noradrenaline (NA), adrenaline (A), dopamine (DA), the DA/NA ratio in the urine, plasma renin activity (PRA) and their mutual relationship were investigated in 71 patients suffering from different types of arterial hypertension. In spite of the fact that the mean values of excreted catecholamines, with the exception of pheochromocytoma, lie within the range of values found in healthy controls, certain differences were found in spectrum of excreted catecholamines. In patients with labile, malignant and renovascular hypertension and in pheochromcytoma the higher mean excretion of NA and the low DA/NA ratio was accompanied by the higher PRA in comparison with fixed benign essential hypertension. On the other hand, in hypertension with low PRA (essential hypertension with suppressed renin and Conn's syndrome) a low excretion of NA and high DA/NA ratio was found. There was a significant, if not even very close negative correlation between the PRA and DA/NA ratios both in recumbent and upright position. The rise of PRA on standing up was followed by an increased excretion of NA while the excretion of DA did not change or decreased. Hence the DA/NA ratio when standing up showed a decreasing tendency as compared with values when lying down. Application of the beta-blocker Inderal decreased the PRA and the blood pressure not only in juvenile hypertensive patients with hyperkinetic circulation but also in the early phases of renovascular hypertension. It thus appears that endogenous catecholamines, first of all the ratio between the renin-inhibiting DA and the renin-stimulating NA, participate as one of several factors in the regulation of secretion and of the plasma levels of renin not only in juvenile hypertensive patients with hyperkinetic circulation but also in other types of hypertension.


Diabetes mellitus and osteoporosis.

In 428 non selected diabetics, the authors determined the metacarpus index of Barnett-Nordin. Upon comparing the data with those of the control group it was shown that opposite to most references one has not to reckon in diabetes with the development of porosis and also that the decrease of the calcium content concomitant with age, respectively, does not significantly differ from that of the control group. Spontaneous vertebral compression occurred more rarely in diabetics than would have been expected on the basis of the literary data. The relation was examined between the type of diabetes, the time of its persisting and osteoporosis. The difference in the observations--compared to the previous ones - is, by the authors, assigned to the fact that nowadays no such lasting and severe metabolic disturbances develop in diabetics that would lead to rare-faction of the bony system and also that hormonal factors can become effective against the development of osteoporosis.


Clinical and metabolic studies in children of diabetic-mothers.

Children of diabetic mothers are a high-risk group. The perinatal mortality and the malformation frequency of the newborns as well as the psychosomatic, somatic and speech development during childhood until the age of 15 were statistically significantly influenced by quality of metabolic compensation, toxaemia and urinary tract infections during pregnancy. To evaluate the diabetogenic risk in children of diabetic mothers metabolic follow-up studies were performed. A statistically significant insulin-high-response without disturbed carbohydrate tolerance was seen in 25% after intravenous glucose infusion and in 28% after oral glucose load. 17% exhibited a paradoxical growth hormone reaction, while the intravenous glucose tolerance and the insulin secretion was in the normal range. Both symptoms could be early diabetic signs in children of diabetic mothers. The oGTT had pathological results in 18% and the glucose infusion test in 8%.


Liver phospholipid distribution in hypophysectomized catfish, Clarias batrachus and Heteropneustes fossilis.

The percentage of certain phospholipid fractions such as phosphatylcholine, phosphatidylserine and phosphatidylinositol was more in the liver of Clarias batrachus than that of the corresponding fractions in Heteropneustes fossilis. Hypophysectomy in both the species resulted in an increase in phosphatidylcholine, but reduction in other fractions. The phosphatidylinositol however, remained unchanged in C. batrachus, but it disappeared in H. fossilis as a result of hypophysectomy.


Insulin resistant diabetes mellitus without the presence of insulin antibodies. A case report.

A case of a 26-year old woman suffering from an insulin resistant diabetes mellitus for 14 years is described. Acanthosis nigricans was diagnosed in the patient's second year and the syndrome of Stein-Leventhal at the age of 15. Diabetes could not be properly controlled either with the daily dosis of insulin as high as 1140 U or with peroral tolbutamide. Fasting serum IRI concentrations were elevated, the secretoric response to the stimulation by glucose or tolbutamide was substantial and protracted. The hypoglycemic response to the i.v. application of commercial insulin was insignificant. Serum growth hormone levels were normal. No presence of insulin antibodies in the serum was detected.


Opposite effects of spironolactone and amiloride on potassium excretion in a patient with ascites due to liver cirrhosis.

Potassium excretion was enhanced by spironolactone and it was inhibited by amiloride in a patient with ascites. One drug's effect was counteracted by the other when they were used in combination. Individual variation in the sites of action of antikaluretic drugs might be a possible explanation for the unpredictable changes in potassium excretion induced by these agents.


Prevalence of epilepsy in children of Multnomah County, Oregon.

The prevalence of epilepsy among children in the third grade in Multnomah County, Oregon, was investigated by use of a questionnaire similar to that used by Rose et al. (1973). Of the 7,039 potential respondents, 5,317 (75.5%) responded to the questionnaire. A stratified random sample of 161 children was drawn from the respondents, and of these 129 (80%) underwent physical and EEG examinations. This replication of the study by Rose et al. (1973) yielded prevalence estimates which were somewhat lower than those reported by Rose et al. for Washington County, Maryland. Our findings suggest that the questionnaire method of estimating the prevalence of epilepsy has promise; further investigation of the method is in progress.


EEG and clinical studies of the development of alumina cream epileptic focus in split-brain cats.

Alumina cream epileptic focus was established in the right sensorimotor cortex in 20 split-brain cats (partial or complete). EEG and behavioral observations were made in a period ranging from 24 to 836 days. Four types of EEG changes after alumina cream injection were differentiated. These types could be related to the direct effects of brain damage and to development of epilepsy. Spikes and sharp waves and paroxysmal discharges (focal and multifocal) were observed in about 60% of the cats. Clinical seizures developed in about the same percentage of the animals. These values are below those reported for cats with intact interhemispheric commissures. Diphenylhydantoin (DPH) was given orally in a daily dose of up to 15 mg/kg body weight in 9 animals with developed epileptic EEG activity. Five of them had epileptic seizures. DPH was introduced not earlier than 1.5 months after intracortical alumina cream injection. The plasma level of DPH varied between 7-20 mug/ml. This dose produced chronic symptoms of intoxication. Neither EEG changes nor clinical seizures were entirely controlled by this drug. Additional doses of Relanium (diazepam), and phenobarbital were necessary to stop generalized seizures or status epilepticus.


Effects of drugs modifying brain levels of catecholamines on photically induced epilepsy in Papio papio.

The behavioral and electrographic effects of DL-amphetamine, disulfiram, FLA-63, and propranolol on the photically induced epilepsy of the Senegalese baboon were evaluated. Amphetamine produced somatic hypokinesia with enhancement of eye movements, diminution of spontaneous paroxysmal activity, and little change in photosensitivity. The acute effects of disulfiram and FLA-63 were poor, but the latter was lethal in the days following administration. Propranolol showed no consistent effects. Participation of catecholamine processes in this type of experimental reflex epilepsy are discussed.


A teachers' questionnaire description of epileptic children.

A teachers' questionnaire was completed for 120 children from normal schools, half of whom suffered from epilepsy. The rate of perceived disturbance among the 30 epileptic children who, for a variety of reasons, had been admitted to a neuropsychiatric hospital was as high as that of a matched group of psychiatric inpatients. The other epileptic children were seen to exhibit significantly more signs of disturbance than their normal school controls with whom they had been matched for behavior. The two epileptic groups shared certain behavioral characteristics. These did not fit into typical "neurotic" or "antisocial" patterns and may relate to cerebral dysfunction.


Phenytoin-induced depression of salivary IgA and gingival hyperplasia.

IgA concentrations were determined in saliva from epileptic patients taking phenytoin and in saliva from healthy controls, by single radial immunodiffusion technique. Mean salivary IgA in epileptic children was 7.23 mg/ml; in healthy children, 16.44 mg/ml. Corresponding values for adults were 13.53 and 19.48 mg/ml, respectively. In 14 out of 84 samples, salivary IgA levels were too low for quantitative analysis. Salivary IgA levels were normal in untreated patients and fell during treatment with phenytoin. Gingival inflammation was commonly accompanied by an increase of salivary IgG and serum-derived IgA, whereas compensatory increase of IgM was infrequent. Phenytoin-induced deficiency of salivary IgA can result in increased susceptibility to gingival inflammation, which is considered one of the predisposing factors for subsequent development of gingival hyperplasia.


Differences between patients with temporal lobe seizures and those with other forms of epileptic attacks.

Seventy-eight patients with temporal lobe type seizures were precisely matched for age, sex, and Wechsler I.Q. levels with patients suffering from other seizure types. Statistically significant differences were established between the groups in regard to treatment response, etiologies, and behavioral variables. Further analysis of the data showed that the behavioral problems of the temporal lobe patients were present only in that group that suffered from more than one seizure type. That temporal lobe epilepsy group having only one seizure type differed from the controls with one seizure type in regard to etiological factors; they actually showed better performance on certain subscales of the WAIS, as well as better employment records. The clinical impression of the temporal lobe epilepsy patient usually presenting more difficulties than other patients with epilepsy results from the fact that the temporal lobe patients have, in the overwhelming majority of instances, more than one seizure type.


Insect growth regulators and insect control: a critical appraisal.

Insect growth regulators (IGRs) of the juvenile hormone type alter physiological processes essential to insect development and appear to act specifically on insects. Three natural juvenile hormones have been found in insects but not in other organisms. Future use of antagonists or inhibitors of hormone synthesis may be technically possible as an advantageous extension of pest control by IGRs. A documented survey of the properties, metabolism, toxicology, and uses of the most commercially advanced chemical, methoprene, shows it to be environmentally acceptable and toxicologically innocuous. Derivation of its current use patterns is discussed and limitations on these are noted. Residue levels and their measurement in the ppb region have allowed exemption from the requirement of tolerances in the EPA registered use of methoprene for mosquito control. Tolerances for foods accompany its fully approved use for control of manure breeding flies through a cattle feed supplement. The human health effects of using this chemical appear to be purely beneficial, but further advances through new IGR chemicals appear unlikely without major changes in regulatory and legislative policy.


Public health advantages of biological insect controls.

Biological control is not new, it is simply newly appreciated. This renewed appreciation stems from the widespread insecticide treadmill which is largely a product of insecticide disruption of the balance of insect communities. Biological control is a natural phenomenon; the regulation of plant and animal numbers by natural enemies. In this broad sense, biological control is vital to public health because it keeps the myriad insect species from out-competing us. It also has direct public health advantages as where natural enemies are manipulated to control disease vectoring insects. Insecticide distruption of biological control by insecticides and the resulting pesticide treadmill have serious public health implications. One is the increased pesticide load in the environment. The other is the acceleration of pesticide resistance in disease vectoring insects. The treadmill and its associated hazards will not abate so long as chemical control dominates our pest management strategy.


Integrated pest management.

Integrated pest management, in which the conventional pesticides are augmented by one or more nonchemical control practices, has been receiving renewed interest. What is new in this revitalization of an old technique is the careful and more knowledgeable application of a variety of control techniques.


Health-related aspects of integrated pest management.

Basic concepts and philosophy of integrated pest management are presented in order to dispel several misconceptions and to provide the necessary background information for discussion of its relationship to the health-related effects of pesticide use. Implications for human health of current pesticide practices in Central America are examined to illustrate major problems associated with the injudicious use of insecticides, i.e., human pesticide poisonings, development of insect resistance, and persistence in the environment. Mitigation of these problems would ideally be achieved through the efforts and cooperation of a multidisciplinary team of scientists and technical people in the medical and agricultural sciences. The dilemma associated with the development of integrated pest control systems in developing countries is discussed. The FAO/UNEP Global Programme was reviewed.


Organotin compounds and their use for insect and mite control.

A brief overview of biological activity of organotin compounds is given, with emphasis on activity against plant-feeding arthropods. Plictran miticide, containing cyhexatin (tricyclohexylhydroxystannane), the first product to be fully commercialized for control of plant-feeding mites on a broad range of agricultural crops, is used as a model to indicate the chemistry and potential performance, phytotoxicity, residues, limitations on usage, environmental acceptability, and use patterns for products containing similar organotin compounds.


Biochemical and physiological effects of chlordimeform.

Chlordimeform is a relatively new acaricide/insecticide, whose mode of action we have investigated. It appears to interfere with amine-mediated control of nervous and endocrine systems in a variety of ways. Specifically, chlordimeform causes a build-up of the amines 5-hydroxytryptamine and to a lesser extent norepinephrine in the rat brain in vivo, antagonizes the in vivo action of reserpine in the rat (reserpine depletes amine stores in the CNS), inhibits monoamine oxidase from rat liver in vitro, and causes hypotension in rabbits. In the American cockroach it directly stimulates the heart in situ, acts synergistically with tryptamine in vivo, inhibits amine-N-acetyltransferase from cockroach head in vitro, causes accumulation of indolamines in cockroaches in vivo, and blocks the stimulation of adenylate cyclase by octopamine in the cockroach CNS in situ. It also inhibits tryptamine metabolism in whole mites in vitro.


The husbandry and veterinary care of wild horses in captivity.

A short account of the natural history of the wild Equidae is given as a background to their management in captivity. In general the approach to their husbandry and veterinary care is similar to that for the domestic horse. A mixture of etorphine and acepromazine has been found to be the most effective for handling and conventional methods of anaesthesia can be safely superimposed on these if required. The data collected at Whipsnade on the haematology and biochemistry of these species is summarised in tabular form and an analysis is included of the post-mortem examinations of 105 equine carcases from the Society's collection.


Percutaneous needle muscle biopsy in the horse.

The use of the technique of percutaneous needle biopsy in obtaining skeletal muscle samples in the horse is described. The biochemical, ultrastructural and histochemical investigations that can be carried out on this biopsy specimen are outlined. Analyses performed on the specimen may be used to obtain information on racing potential and state of fitness. These studies on normal horses will provide information for future investigations into the structural and biochemical alterations in muscle disorders in the equine.


Congenital deformities in two Clydesdale foals.

The deformities observed in 2 Clydesdale foals are described. Both had abnormal joint positions in the forelimbs and discrepancies in the symmetry of the vertebral column. The changes were only mild in one case but extreme in the other where it was accompanied by torticollis, scoliosis and vertebral fusion. A comparison is made with deformities described in the contracted foal syndrome and some of the developmental implications discussed.


An investigation into the genetics of 'wobbler disease' in thoroughbred horses in Britain.

Sixty-seven TB horses in Britain suffering from clinical wobbler disease were divided into 5 groups, based on the presence or absence of pathological lesions and their nature. These groups were compared genetically with a control group of 67 TBs known not to exhibit signs of wobbling. Though analysed for both simple and complex modes of inheritance no evidence of a genetic basis was found in any group. The high breed incidence in TBs may be real or apparent. Considerably more male than female wobblers are reported. Other forms of possibly inherited ataxia in equidae are reviewed and reference is made to pathologically comparable conditions occurring in man and in the large breeds of dogs.


Successful repair of a diaphragmatic hernia in a foal.

The diagnosis and successful surgical treatment of a diaphragmatic hernia in a 12 weeks old standardbred foal are described. The clinical syndrome in horses generally is contrasted with that seen in the foal described in this paper. Some of the factors contributing to the clinical signs are discussed.


The use of Dopram as a respiratory stimulant following Immobilon in the pony.

The effects of the analeptic agent, Dopram (doxapram hydrochloride) were investigated in 2 ponies during Immobilon - induced neuroleptanalgesia. Although Dopram was demonstrated to exert a degree of respiratory stimulation, this was concluded to provide no overall advantage. The etorphine-induced hypoxic hypoxia was only partially reversed, and there was additional cardiovascular stimulation, in contrast to the previously reported tendency for arterial blood pressure to return towards conscious control values during the course of action of Immobilon.


An interventricular septal defect, pulmonary stenosis and bicuspid pulmonary valve in a Welsh pony foal.

A 6 month Welsh pony filly with an interventricular septal defect in conjunction with a pulmonary stenosis due to a bicuspid pulmonary valve is described. The animal had poor exercise tolerance, a loud pansystolic murmur and a precordial thrill. Blood pressure and oxygen tension values obtained during cardiac catheterization suggested the diagnosis which was confirmed at autopsy.


Succinate accumulation in man during exercise.

It has been demonstrated in several diving vertebrates that succinate, a component of the Krebs cycle, accumulates in blood during breath-hold dives. The production of succinate is thought to result from amino acid catabolism. Our purpose was to determine whether succinate accumulation occurs in man during muscular activity requiring anaerobic energy contribution. Experiments using an endurance athlete included apneic work on an underwater ergometer and treadmill running to exhaustion. During 1 min breath-hold "dives" in cold water while exercising at a work rate equivalent to 62% of VO2max, venous succinate increased from 42 mumoles/l (M X 10(-6)) at rest to 125 M X 10(-6). The treadmill run elicited VO2max and increased succinate from a similar resting value to 93 M X 10(-6). Increases in alanine, lactate, and pyruvate were observed for both types of exercise. The findings confirm that succinate accumulation also occurs in man. It was suggested that amino acid catabolism may provide a source of anaerobic energy production in addition to glycolysis. However, the importance of the proposed energy pathway remains to be quantified.


Effects of an anabolic steroid and sprint training on selected histochemical and morphological observations in rat skeletal muscle types.

The effects on selected histochemical and morphological parameters of anabolic steroid administration and of high-intensity sprint running, separately, and in combination, were studied in young adult male rats. Dianabol (methandrostenolone) 1 mg/day for 8 weeks had no significant effects on phosphorylase or glycogen staining intensities and on fiber area in skeletal muscles of either trained or sedentary animals. The program of sprint training resulted in significantly decreased intensities of phosphorylase in all ten regions of the gastrocnemius, plantaris, and soleus muscles that were studied. Glycogen localization was significantly increased with training in five regions of the gastrocnemius and plantaris muscles which contained predominantly fast-switch fibers. No changes in fiber area occurred with the training program. We conclude from these results that (a) normal androgen levels in young, healthy male animals are sufficiently high so that the intake of large doses of anabolic steroid does not result in the stimulation of glycogen metabolism or hypertrophy of skeletal muscle; (b) the changes induced by high-intensity, short-duration sprint training suggest that the existing glycolytic capacity of muscle is adequate to supply the muscles energy needs even during the stress of very strenous exercise, and that more fast-twitch fibers were recruited by the exercise regimen than slow-twitch fibers.


Metabolic changes during exercise testing of patients with ischaemic heart disease.

Nineteen patients who had recovered from a mild to moderately severe myocardial infraction 6--18 months previously were studied in order to investigate the relationship between the severity of myocardial insufficiency and metabolic changes developing during and after exercise. The patients were compared with six controls by means of a graded exercise test on a treadmill. Electrocardiographic records were made and blood pressure measured during and after the exercise and venous blood samples were taken for measurement of lactate, pyruvate, glycerol and free fatty acids. In five patients with moderate or advanced ischaemic electrocardiograph changes the blood lactate and pyruvate mean concentrations continued to rise up to a further 50% after they stopped exercising. The other patients, including some who developed symptoms suggestive of myocardial ischaemia, and all the controls, failed to show these marked increases in blood concentrations after the exercise. We conclude that the development of myocardial insufficiency during exercise is associated with marked increases in lactate and pyruvate concentrations in the peripheral blood particularly after the exercise has ceased.


Effect of physical fitness on vanillylmandelic acid excretion during immersion.

The effects of 4--6 h head-out immersion on excretion of vanillylmandelic acid (VMA), blood pressure and plasma volume were estimated in 8 endurance-trained (TR) and 8 untrained (UT) subjects. In the trained only a slight increase of VMA excretion occurred (4 h value: +2.7 +/- 10.9 ng/ml GFR), but there was a highly significant increase in the UT (+29.0 +/- 17.2 ng/ml GFR). VMA values during control experiments in supine position tended to decrease in both groups. Systolic and diastolic blood pressure fell by 20 mm Hg after beginning of immersion; in the UT plasma volume was reduced while it remained constant in TR. The results indicate that orthostatic intolerance (o.i.) after immersion is not effected by decreased sympathetic innervation of vessels; in contrast it seems to be partly compensated for by an elevated sympathetic activity at least in the UT. As a main cause for the post-immersion o.i. one might suggest a decrease in renin activity.


Age-changes of the force-frequency-relationship and the duration of action potential of isolated papillary muscles of guinea pig.

The force-frequency-relationship and the duration of the action potential recorded intracellularly were compared on isolated isometrically arranged papillary muscles of young (3--4 months old) and senile (36--40 months old) guinea pigs. The muscles were in Krebs-Henseleit-solution at 37 degrees C electrically stimulated with frequencies between 30 and 420 imp./min. The following results were obtained: 1. The isolated papillary muscles of the two groups are almost similar with respect to length, weight and cross-sectional are. 2. In the range of stimulation rate from 30--420 imp./min the duration of action potential in the senile group was 120--135% of that in the young group. 3. The shape of slow repolarization of action potential was steeper in the senile group. 4. The time to peak tension and the relaxation time were longer in the senile group. 5. The maximally developed tension was the same in both groups, but in the senile group it was reached at a stimulation rate of 120 imp./min whereas in the young group, the optimal frequency was 290 imp./min. 6. The greatest value for the maximal rate of tension rise was 2.4 times smaller in the senile group. The results are discussed in relation to the age dependent intensity of membrane currents and to a possible altered activity of sarcoplasmic reticulum.


Effect of training on the resting heart rate of rats.

Adult male rats were progressively trained 5 days/weeks on a motor-driven treadmill. The training period lasted 12 weeks and consisted of 60 min/day of wind-sprints and endurance work. No significant difference in resting heart rates was observed between the control and exercise groups during week 1 (394 +/- 7 vs. 388 +/- 5). However, at week 12 the exercise group had a lower resting heart rate (359 +/- 6 vs. 331 +/- 4). Heart rates observed following saline, propranolol, atropine, and propranolol plus atropine injections were lower in the exercise group in all cases. The difference in heart rates between the control and exercise groups was 19 beats/min following propranolol plus atropine which was less than the 28 beats/min difference observed under control conditions. With atropine and then with propranolol the differences were 33 and 27 beats/min. These heart rate differences were observed without the presence of cardiac hypertrophy as assesssed from ventricle weights. Our data indicate that the bradycardia resulting from exercise training is due primarily to changes other than neural influences on the heart.


Prolonged exercise in prepubertal boys. I. Cardiovascular and metabolic adjustment.

The circulatory and metabolic changes during prolonged exercise (60 min) were studied in a group of 10 healthy prepubertal boys. The boys worked on the treadmill and on the bicycle ergometer at two levels of load, namely at oxygen uptakes of 36--39%, and 60% of the respective VO2 max for both types of exercise. The identical relative work loads represent higher metabolic rate at the treadmill exercise. The rectal temperature was higher after exercises on treadmill than on bicycle. The heart rate increase during prolonged exercise was lower in boys than in adults in both types of exercise. Better cardiovascular adjustment is suggested. The blood lactate increase was negligible in the initial phase of exercise with subsequent removal from the blood during extended exercise, which was more pronounced at the treadmill exercises. The increasing blood free glycerol concentration indicates similar fat share on energy release in children as in adults. No biological handicap could be found for prolonged exercise in children. The preference of short-time exercises in children lies probably in the psychological sphere.


Prolonged exercise in prepubertal boys. II. Changes in plasma volume and in some blood constituents.

A group of 10 prepubertal boys was studied during prolonged exercise (60 min) on bicycle ergometer and on treadmill at two levels of work load (appr. 40% and 60% VO2 max). The hematocrit, serum proteins, Cl- and K+ were followed, and from the blood hematocrit changes the plasma volume changes were calculated. At the exercises of lower intensity of both types a slight hemodilution was found (appr. +5% increase in plasma volume), at higher intensity practically no changes could be demonstrated. These findings are supported by the values of serum protein concentration, where no increase was found, and by the fact that at the lower work loads a rather decreasing trend was found for this blood constituent. These findings are at variance with those in adults under similar conditions. The authors suggest that different changes of plasma volume during exercise in boys than in adults could be related to the disparate lactate production and fate in these age groups.


Purification and properties of L-4-hydroxymandelate oxidase from Pseudomonas convexa.

An inducible membrane-bound L-4-hydroxymandelate oxidase (decarboxylating) from Pseudomonas convexa has been solubilized and partially purified. It catalyzes the conversion of L-4-hydroxymandelic acid to 4-hydroxybenzaldehyde in a single step with the stoichiometric consumption of O2 and liberation of CO2. The enzyme is optimally active at pH 6.6 and at 55 degrees C. It requires FAD and Mn2+ for its activity. The membrane-bound enzyme is more stable than the solubilized and purified enzyme. After solubilization it gradually loses its activity when kept at 5 degrees C which can be fully reactivated by freezing and thawing. The Km values for DL-4-hydroxymandelate and FAD are 0.44 mM and 0.038 mM respectively. The enzyme is highly specific for DL-4-hydroxymandelic acid. DL-3,4-Dihydroxymandelic acid competitively inhibited the enzyme reaction. From the Dixon plot the Ki for DL-3,4-dihydroxymandelic acid was calculated to be 1.8 X 10(-4) M. The enzyme is completely inactivated by thiol compounds and not affected by thiol inhibitors. The enzyme is also inhibited by denaturing agents, heavy metal ions and by chelating agents.


Gel isoelectric focusing of human-serum transferrin.

Analysis of human serum transferrin on gel isoelectric focusing resolved this iron-transport protein into two iron-containing components which were identified by the use of radioactively labelled iron and iron-specific stain. These two components were found to be monoferric transferrin (Fe-transferrin) and diferric transferrin (Fe2-transferrin) with isoelectric points of 5.6 and 5.2 respectively. The estimation of the relative proportions of these two components in serum did not correspond to the calculated theoretical ratio based on random binding of iron to the two binding sites of transferrin. However, the analysis of partially resatured apotransferrin gave a ratio corresponding to a random distribution of iron. The significance of these results is discussed.


Initiation of the polypeptide chain by reticulocyte cell-free systems. Survey of different inhibitors of translation.

In order to elucidate the mechanism of action of inhibitors that block the initiation of protein synthesis in mammalian systems, we have studied the following steps: (a) formation of the ternary complex Met-tRNAr-IF-E2-GTP, (b) binding of the initiator Met-tRNAf to the 40-S ribosomal subunit in the presence of initiation factors and dependent or not on the addition of mRNA, (c) formation of the initiation complex with 80-S ribosomes and (d) formation of the first peptide bond. Adrenochrome, aurintricarboxylic acid, polydextran sulphate, pyrochatechol violet and showdomycin block the formation of the ternary complex Met-tRNAf-IF-E2-GTP. Edeine A1, aurintricarboxylic acid and polydextran sulphate block the binding of the mRNA to the 40-S ribosomal subunit. Pactamycin induces the formation of stable smaller initiation complexes which are unable to go through the subsequent steps of initiation. Stimulation of the binding of the initiator Met-tRNAf to the 80-S ribosome in the presence of initiation factors is observed with sparsomycin and antibiotics of the sesquiterpene family (verrucarin A, trichodermin and trichothecin). However, these antibiotics block the reaction of the bound Met-tRNAf with puromycin. Narciclasine has no effect on the binding of the initiator to the ribosome but strongly blocks its reaction with puromycin. We have developed a simple technique to detect the Met-tRNAf-40-S-subunit-poly(A, G, U) initiation complexes by chromatography on Sepharose 6B columns. The requirements for the formation of such complexes measured by this technique and its comparison with the sucrose gradient centrifugation method are described.


Isolation and characterization of the lipopolysaccharide of Chromatium vinosum.

Lipolysaccharide was isolated from Chromatium vinosum by phenol/water extraction. The lipopolysaccharide is found exclusively in the phenol phase and can be cleaved into a sugar moiety and a lipid A fraction by hydrolysis in 10% acetic acid at 100 degrees C for 3-4 h. The sugar moiety contains the neutral sugars 3-O-methyl-D-ribose, D-ribose, L-arabinose, mannosamine and glucose, and smaller quantities of D-rhamnose, D-glycero-D-manno-heptose (tentatively identified), quinovosamine and 2-keto-3-deoxyoctonate. L-glycero-D-manno-heptose was not detected. The 2-keto-3-deoxyoctonate linkage in C. vinosum lipopolysaccharide is more resistant to acid hydrolysis than that of Escherichia coli. The lipid A fraction contains glucosamine, mannose and the fatty acids of the lipopolysaccharide. The major fatty acid is beta-hydroxymyristic acid, with smaller amounts of lauric and palmitic acids as well as 14-carbon mono-unsaturated fatty acid, also being present. The phosphorus content of the C. vinosum lipopolysaccharide was found to be approximately 0.1%. Erythrocytes sensitized with alkali-treated C. vinosum lipopolysaccharide were agglutinated by antisera prepared against heat-killed cells. Untreated or heat-treated lipopolysaccharide did not sensitize erythrocytes. The lethal toxicity to mice of the C. vinosum lipopolysaccharide is about one-tenth as that from Salmonella abortus equi.


Affinity labelling of the estradiol-17 beta dehydrogenase from human placenta with substrate analogs.

Affinity labelling of the estradiol-17 beta dehydrogenase of human placenta has been performed using derivatives of estradiol-17 beta carrying alkylating groups in nine different positions on the steroid nucleus. The active-site-directed character of the inhibition is confirmed by the following observations: the affinity labels are substrates or competitive inhibitors, the enzyme is protected against inactivation and alkylation by the substrate and by the coenzyme, the stoichiometry of the alkylation is two moles of inhibitor per 68 000 g of enzyme (dimer). The alkylation of a histidine residue which is fast and extensive when the alkylation side chain is on the C-3 carbon atom, is dramatically decreased when alkylating side chain is shifted towards rings B and D. These results allow the location of this histidine in the vicinity of ring A and probably on the beta face of the steroid nucleus. The reactivity of a cysteine located on the active site was quite different, showing increasing alkylation when the alkylating substituent of the affinity labels was shifted from C-3 to C-16 of the steroid nucleus. The correlation of this result and that obtained using an alkylating analog of NAD (3-chloroacetyl-pyridine-adenine dinucleotide) indicates that this cysteine is located in the catalytic region of the active site, at the junction of the ring D of the steroid nucleus with the nicotinamide moiety of the coenzyme.


Role of GTP in eukaryotic polypeptide-chain initiation. Purification and properties of a factor from Artemia salina embryos which interacts with initiator transfer RNA and guanine nucleotides.

1. A factor, which makes a ternary complex with GTP and eukaryotic initiator tRNA (Met-tRNAi), has been purified 100-fold from developed cysts of Artemia salina. Some of the properties of the purified factor have been studied. 2. Mg2+ appears to inhibit ternary complex formation. 3. Little or no ternary complex is formed when 5 muM GTP is replaced by an identical concentration of UTP, CTP or ATP. The analog, guanosine 5'-(beta, gamma-imino)triphosphate [GMP-P(NHP)] seems to be a much better substitute for GTP than guanosine 5'-(beta, gamma-methylene)triphosphate [GMP-P(CH2)P]. Since GMP-P(NH)P is as effective as GTP in ternary complex formation, it would appear that GTP plays the role of an allosteric effector in this step of eukaryotic polypeptide chain initiation. 4. GDP inhibits both the rate and extent of ternary complex formation. The inhibition is largely reversed by adding a 5-fold molar excess of GTP over GDP. DGDP is slightly less inhibitory than GDP. UDP and CDP are much less inhibitory than GDP and very little inhibition is obtained with ADP. 5. The preformed ternary complex is rapidly and completely destroyed in the presence of N-ethylmaleimide. The results suggest that free--SH groups of the factor may be essential for maintaining the integrity of the ternary complex.


Effects of thyroid hormone deficiency on the distribution of hepatic metabolites and control of pathways of carbohydrate metabolism in liver and adipose tissue of the rat.

1. Measurements have been made of the hepatic metabolites in normal and thyroidectomized rats and enzymes of the glycolytic route, the pentose phosphate pathway, the tricarboxylic acid cycle and lipogenesis together with the flux of glucose through alternative pathways of glucose metabolism and into lipid. 2. There is a significant fall in the content of ADP, AMP, citrate, long-chain acyl-CoA derivatives and a rise of 3-phosphoglycerate, 2-phosphoglycerate and phosphoenolpyruvate following thyroidectomy. The observed changes in the glycolytic intermediates may be correlated with the increased activity of phosphofructokinase relative to pyruvate kinase and fructose bisphosphatase. 3. The NAD+/NADH quotient of the mitochondrial compartment, calculated from the reactants and Keq of 3-hydroxybutyrate dehydrogenase, becomes significantly more oxidized in hypothyroid animals. The redox state of the cytosolic NAD and NADP couples remains relatively unchanged. 4. The changes in the hepatic content of CoA derivatives and citrate and in the mitochondrial redox state are interpreted as indicating a depressed rate of lipid oxidation. 5. Calculations of the compartmentation of metabolites between the cytosol and mitochondria indicate a very marked decrease in mitochondrial citrate, 2-oxoglutarate and glutamate with smaller changes in aspartate and malate. These changes are interpreted as providing evidence for the importance of modifications in the malate-aspartate shuttle in hypothyroidism; this is further supported by measured changes in the distribution and activities of the component enzymes of the hydrogen shuttles. 6. There is a diminished activity of glucokinase and of enzymes of the glycolytic pathway below phosphofructokinase in livers from hypothyroid rats. The oxidative enzymes of the pentose phosphate pathway, ATP-citrate lyase, 'malic' enzyme and fatty acid synthetase also decrease markedly. There is a striking parallelism between the changes of enzyme profile of liver and adipose tissue in hypothyroidism. 7. Adrenal glands from hypothyroid rats showed a generalized delcine in enzyme activity in parallel with the fall in tissue weight. Cytosolic glycerol-3-phosphate dehydrogenase decreased sharply. 8. The present results are discussed in relation to data in the literature on the increased activities of cyclic AMP and cyclic GMP phosphodiesterases in hypothyroidism, observations which integrate the relationships between anabolic and catabolic pathways of carbohydrate and lipid metabolism with alterations in hormone sensitivity in hypothyroidism.


Globin messenger RNA from anaemic rabbit spleen. Size of its polyadenylate segment.

The size of the polyadenylate segment of globin messenger RNA isolated from spleens of anaemic rabbits was estimated by comparison of its electrophoretic migration in polyacrylamide gels to that of synthetic poly(A) segments of known lengths. Conditions of enzymic degradation of mRNA with pancreatic ribonuclease and T1 ribonuclease were carefully established in order to ensure complete degradation of the heteropolymeric part of mRNA without affecting the polyadenylate sequence. The poly (A) segments of spleen globin mRNA were found to be 25-90 nucleotides long whilst those of peripheral blood reticulocytes from the same animals were only 10-30 residues long. Since spleen contains young erythroid cells and since anucleated blood reticulocytes constitute a statistically older population of the same cell line, these results support the idea that the poly(A) segment of mRNA shortens when the message ages.


Purification of six neurotoxins from the venom of Dendroaspis viridis. Primary structure of two long toxins.

Six neurotoxins were purified from Dendroaspis viridis venom using gel filtration and equilibrium chromatography. The amino acid sequences of two of these neurotoxins (72 and 73 residues, five disulphide bridges) have been determined using almost exclusively automated Edman degradation. These two sequences are very similar: the only differences lies in the presence of one extra glycine at the C-terminal end of one of them. There is a good homology with the sequences of toxins now isolated from other Elapidae and Hydrophidae venoms.


Taipoxin, an extremely potent presynaptic neurotoxin from the venom of the australian snake taipan (Oxyuranus s. scutellatus). Isolation, characterization, quaternary structure and pharmacological properties.

Taipoxin (taipan toxin), purified from the venom of the Australian taipan (Oxyuranus s. scutellatus) by gel filtration on Sephadex G-75 followed by column zone electrophoresis, is the most lethal neurotoxin yet isolated from any snake venom. The LD50 is 2 mug/kg in the mouse. The main physiological effect is a gradual reduction to complete stop of evoked and spontaneous release of acetylcholine from motor nerve terminals. Intoxicated animals die of asphyxia caused by neuromuscular blockage of the respiratory muscles. Taipoxin is a moderately acidic sialo-glycoprotein (pI 5) with a molecular weight of 45 600 as calculated from composition data or 46 800 as determined by meniscus depletion sedimentation equilibrium. Taipoxin is a 1:1:1 ternary complex of subunits designated alpha, beta and gamma which dissociate completely at low pH and high ionic strength or in 6 M guanidine hydrochloride. The dissociation by guanidine at neutral pH is reversible, while the acid-induced dissociation is not . The alpha and beta components consist of 120 amino acid residues cross-linked by seven disulfide bridges, whereas the gamma component has 135 residues and eight disulfides. The very basic (pI Greater than 10) alpha component contains 13 residues of arginine and is the only subunit displaying lethal neurotoxicity (mouse LD50 approximately 300 mug/kg). The neutral beta fraction was separated by ion-exchange chromatography into two iso-component, beta 1 and beta2, which differ slightly in amino acid composition. The very acidic gamma component contains all of the carbohydrate, which includes 4-5 residues of sialic aid. The three subunits are homologous in sequence although the gamma component is eight residues longer on the N-terminus and must also contain extra amino acids elsewhere.


The arrangement of 18-S and 28-S ribosomal ribonucleic acids within the 40-S precursor molecule of Xenopus laevis.

The arrangement of 18-S rRNA and 28-S rRNA within their 40-S common precursor molecule (pre-rRNA) of Xenopus laevis was investigated by electron microscopic analysis of secondary structure of nascent pre-rRNA chains of oocytes, and by 5'-end analysis of 18-S rRNA and 28-S rRNA hybridized to the EcoRI fragment of rDNA cloned as plasmid pCD42. Secondary structure mapping of phenol-extracted RNA from nucleolar cores revealed complete pre-rRNA chains or molecules at various stages of processing and pre-rRNA molecules apparently lacking one end. In this latter group, which was regarded as representing nascent chains, more than 90% of the molecules had no 28-S rRNA REGION. This shows that the 28-S rRNA sequence is transcribed after the 18-S rRNA region and hence must be located nearer to the 3' end of the pre-rRNA molecule. For 5' end-group determination [3H]uridine-labelled 18-S rRNA and 28-S rRNA were hybridized, as fragments of about 200 nucleotides, to the plasmid pCD42 containing coding sequences for four-fifths of the 18-S rRNA sequence, the external transcribed spacer, the non-transcribed spacer and a tenth of the 28-S rRNA sequence. The RNA was recovered from the hybrids and analyzed for uridine 3',5'-bisphosphate (pUp) after alkaline hydrolysis. The pUp content of the hybridized 18-S rRNA fragments was 20-fold higher than in those of 28-S rRNA, THUS DEMONSTRATING THAT THE 5' END OF THE 18-S rRNA is located next to the external spacer region. From these results it is concluded that the 18-S rRNA is located close to the 5' end of the 40-S pre-rRNA molecule.


Acceleration of the reaction between thrombin and antithrombin III by non-stoichiometric amounts of heparin.

The accelerating effect of heparin on the reaction between purified human antithrombin and thrombin has been investigated by measuring the amount of thrombin inactivated during a short incubation of the enzyme with the inhibitor in the presence and absence of non-stoichiometric amounts of heparin. It was demonstrated that one molecule of heparin was able to promote the binding of a large number of antithrombin molecules to thrombin. Thus heparin may affect the rate of the inactivation of thrombin by antithrombin in a catalytic manner.


Molecular forms of Electrophorus acetylcholinesterase. Molecular weight and composition.

Molecular weights for the series of six Electrophorus acetylcholinesterase forms have been determined either by the sedimentation-diffusion equilibrium method or, particularly in the case of the very scarce G' and G inches forms, from their Stokes radius and sedimentation coefficient values. Both methods are in excellent agreement. The results provide good evidence for the model previously proposed, G inches, G' and G containing one, two and four subunits, whereas A, C and D possess, in addition to respectively one, two and three tetrameric sets of such subunits, a structural element, the tail. Although the amino acid composition of 'tailed' and globular forms did not reveal any significant feature of this element, its mass, about 100 000 daltons, could be deduced from a comparison of molecular weights for the two classes of acetylcholinesterase forms. This value is in close agreement with electron microscopic data. The tail is thought to consist of three 30 000-dalton strands.


Affinity chromatography of acetylcholinesterase. The importance of hydrophobic interactions.

An easily prepared affinity column for acetylcholinesterase is described, which may be operated at ionic strength high enough to prevent aggregation of the asymmetric forms of the enzyme. Specific elution by tetraethylammonium or decamethonium was quantitative. The performance of this column is comparable to that of the column described by Dudai and Silman. It is shown that the hexyl 'spacer arm' strongly participates in the enzyme binding and that its replacement with the more hydrophilic spermine chain lowers the affinity. The hexyl chain itself is shown to bind acetylcholinesterase, although with lower affinity and capacity than the complete column. This binding is also partly reversed by inhibitors. Such hydrophobic columns bind the native asymmetric forms of the enzyme more strongly than the lytic globular ones. The aromatic quaternary ligang inhibits Electrophorus but not Torpedo acetylcholinesterase; therefore the column does not retain the Torpedo enzyme. Differences in Km between acetylcholinesterases of the two species also point to differences in their active sites.


Chemical modification of the tryptophan residues of wheat-germ agglutinin. Effect on fluorescence and saccharide-binding properties.

The oxidation of the tryptophan residues of wheat germ agglutinin by N-bromosuccinimide was investigated under non-denaturing and denaturing conditions. All three tryptophan residues present in wheat germ agglutinin subunit (molecular weight 18 000) could be modified in 0.1 M acetic acid/8 M urea, pH 3.9. One of the residues failed, however, to react with N-bromosuccinimide when the modification was in 0.1 M citrate buffer, pH 6.0. Tryptophan fluorescence of the protein was quenched concomitantly with the oxidation of two tryptophan residues even when the modification was carried out in acetic acid urea. After oxidation of two tryptophan residues per subunit of wheat germ agglutinin, only 15% of the original tryptophan fluorescence remained; upon excitation at 280 nm, tyrosine fluorescence centered at 305 nm could be resolved. The results suggest that there are only two emitters in the protein and that the third tryptophan residue is buried in the native protein and can be modified only in acetic acid urea. This tryptophan residue is quenched in the native protein. Saturation of wheat germ agglutinin with tri-N-acetylchitotriose did not protect the tryptophan residues from oxidation by N-bromosuccinimide. Under these conditions, however, the reactivity of the tryptophan residues towards N-bromosuccinimide was reduced and a higher concentration of the reagent was required to achieve the same extent of oxidation as in the absence of the saccharide. Oxidation of one tryptophan residue per subunit in acetic acid urea led to almost complete loss (97%) of hemagglutinating activity, a 3.5-fold decrease in the affinity constant for tri-N-acetylchitotriose and loss of ability of the subunits (SO20,w = 2.0 S) to reassociate to the native dimer (So20,w = 3.5 S) after dialysis against a non-denaturing buffer. No significant changes in the circular dichroism spectrum of wheat germ agglutinin were observed after oxidation of the three tryptophan residues, suggesting that no gross conformational changes occurred. The steric relationships between the fluorescent tryptophan residues of wheat germ agglutinin and saccharides are discussed.


Nanosecond-pulse fluorimetry of wheat-germ agglutinin (lectin).

Nanosecond-pulse fluorimetry of wheat germ agglutinin is analyzed as a function of both excitation and emission wavelengths. When excited at 280 nm, wheat germ agglutinin fluorescence exhibited three lifetimes: one corresponding to the tyrosine residues as a whole and two others corresponding to the tryptophyl emission. The tyrosine contribution to the emission spectrum deduced from this method was in good agreement with that reported previously in steady-state fluorescence experiments [Privat, J.P. and Monsigny, M. (1975) Eur. J. Biochem. 60, 555-567]. The fluorescence decay of each tryptophan residue was not a single exponential function when wheat germ agglutinin was excited at 295 nm. This could be related to the microenvironment of the indole chromophores in the protein. The comparison of the quantum yield and of average lifetime showed that some tryptophan residues were completely quenched. Energy transfer from tyrosines to tryptophan residues previously detected in steady-state fluorescence was also revealed by fluorescence decay measurements. Comparison of both methods showed that an important part of transfers occurred with a very fast rate equal to or greater than 10(10) s-1. Both lifetimes and the ratio of the short and the long-lived component were found dependent on tri-N-acetylchitotriose binding.


6-Methylsalicylic acid synthetase from Penicillium patulum. Some catalytic properties of the enzyme and its relation to fatty acid synthetase.

1. The specificity of 6-methylsalicylic acid synthetase with respect to the priming substrate was studied. If acetyl-CoA was replaced by propionyl-CoA 6-ethylsalicylic acid was synthesized. The rate of this synthesis was about 13% that of 6-methylsalicylic acid synthesis in presence of acetyl-CoA. 2. 6-Methylsalicylic acid synthetase contains an acetyl transferase activity as demonstrated by the transfer of acetyl residues from acetyl-CoA to pantetheine. This transferase also catalyses propionyl transfer but only with an about 13 times slower rate. 3. Analogous to fatty acid synthetase, treatment with iodoacetamide converts 6-methylsalicylic acid synthetase into a malonyl-CoA decarboxylase. Under certain conditons the iodoacetamide-treated enzyme could catalyse 6-methylsalicylic acid formation from malonyl CoA and NADPH without the external addition of acetyl CoA.


The metabolism of O-acyl-N-acylneuraminic acids. Biosynthesis of O-acylated sialic acids in bovine and equine submandibular glands.

1. The enzymic synthesis of 4-O-acetylneuraminic acid, 4-O-acetyl-N-glycolyneuraminic acid, 4-O-glycolyl-N-acetylneuraminic acid, 9-O-acetyl-N-acetylneuraminic acid and 9-O-acetyl-N-glycolyneuraminic acid is shown using radioactive precursors with surviving slices, membrane fractions or particle-free homogenates from bovine and equine submandibular glands. 2. Acetyl-CoA: N-acetylneuraminate-9(or 7)-O-acetyltransferase activity was found in a microsome fraction and in the cytosol of bovine submandibular glands. The properties of the membrane-bound enzyme acting on endogenous, glycoprotein-bound N-acetyl- and N-glycolylneuraminic acids were compared with those of the soluble enzyme, O-acetylating exogenous, non-glycosidically bound N-acetyl- and N-glycolyneuraminic acids. 3. A rapid, radioactive assay for the membrane-bound enzyme activity is described. The enzyme activity shows an optimum at pH 7 and has  a Km for acetyl-CoA of 0.1 mM. The enzyme is inhibited by p-chloromercuribenzoate and iodoacetate. Divalent cations, EDTA and glutathione have no influence on its activity while CoA proved to be a competitive inhibitor with a Ki of 0.56 mM. 4. The soluble enzyme activity, assayed using a radioactive procedure, shows Km values of 0.01 mM, 0.5 mM and 0.39 mM for acetyl-CoA, N-acetylneuraminic acid and N-glycolylneuraminic acid respectively. The general properties are similar to those found for the membrane-bound enzyme, except that membrane-bound activity is stable for longer on storage at 4 degrees C. 5. Acetyl-CoA, acyl-CoA and CoA concentrations of 33 nmol, 65 nmol and 106 nmol/g wet tissue respectively are found in fresh bovine submandibular glands. 6. The occurrence of the CMP-glycosides of N-acetylneuraminic acid, 9-O-acetyl-N-acetyl-neuraminic acid and N-glycolylneuraminic acid in bovine submandibular glands is demonstrated. 7. The results are discussed in relation to the general metabolism of acylneuraminic acids.


A new familial intrauterine growth retardation syndrome the "3-M syndrome".

Two pairs of siblings are described with proportionate dwarfism due to skeletal hypoplasia of prenatal onset. The head size was normal for age and disproportionately large for height. The patients had a characteristic face different from that seen in the Silver-Russell syndrome. The family data are in accordance with autosomal recessive inheritance. In spite of some similarities, the bulk of clinical and genetic evidence suggests that the described intrauterine growth retardation syndrome is different from the Silver-Russell syndrome and presents an apparently "new" entity which has been designated 3-M syndrome.


Latex phagocytosis by polymorphonuclear leukocytes. In vitro and in vivo studies with a simple screening test.

A simple semiquantitative slide test to assess the phagocytic activity of periphera polymorphonuclear leukocytes is described. Leukocytes are isolated from one drop of blood and latex particles are offered for phagocytosis. Cellular capacities and serum opsonins can be assessed separately. In the presence of pooled adult serum no defect of phagocytosis was seen in preterm and full-term newborns nor in pediatric patients with infectious or other diseases. In 4 of 7 premature newborns a definite deficiency of serum opsonins was demonstrated. In in vitro studies complement (plus possibly other heat-labile factors) promoted phagocytosis. Phagocytic rates were depressed by endotoxin and phenylbutazone.


Analysis of etiologic factors in cerebral palsy with severe mental retardation. I. Analysis of gestational, parturitional and neonatal data.

We analyzed the gestational, parturitional, neonatal (GPN) histories of 281 severely mentally retarded patients with cerebral palsy to define the etiology or pathogenesis of cerebral palsy in each patient. No association between type of cerebral palsy and GPN histories was found except for an increase in spastic-athetoid patients in the breech delivery subgroup. Significant findings include: increased incidence of prematurity and postmaturity, small and large for-gestational age (GA) fetal size, a normal birthweight for GA distribution of patients with diabetic mothers, an excess of mothers greater than or equal to 35 and less than or equal to 20 years old, an increased immediately-previous sib interval of 2.59 years suggestive of an "infertility factor", an unremarkable GPN history in one third of the cases, in another one third GPN problems not usually associated with a high risk of CNS damage, and in one third gross complications which were probably responsible for the CP, including: an increased incidence of breech deliveries, twinning, prolonged and precipitous labor and placental complications; no increased association of athetosis and Rh incompatibility or incidence of toxemia was found. Disseminated intravascular coagulation due to prenatal death of a twin may have been the cause of brain damage in several patients.


Serum lactate dehydrogenase isoenzyme pattern in neuroblastoma.

Total serum LDH activity and isoenzyme distribution were studied in children with neuroblastoma at the time of hospital admission. The total LDH was determined in 26 cases, and 20 (77%) of them showed elevation of its activity. On the other hand, in 9 of these 26 cases, the isoenzyme distribution was determined along with the total LDH. All 9 cases, 4 of them with normal total LDH activity, showed an abnormal isoenzyme pattern with a percentage increase in the intermediate fractions (malignant pattern). The results suggest the usefulness of the determination of serum LDH isoenzymes as a screening procedure in children with malignant tumors including neuroblastoma.


A genetic and statistical study of the respiratory distress syndrome.

The hospital records of 197 infants with the respiratory distress syndrome (RDS) were reviewed and the families of 111 of them subsequently contacted to obtain a family history. After correcting for biasis of ascertainment, the incidence of RDS among the full sibs was found to be between 12 and 19% depending on whether the individuals diagnosed as "possible RDS" were counted as affected. Among the low birth weight (LBW, less than or equal to 2.5 kg) and/or preterm (less than or equal to 37 weeks gestation) infants in the sibships, the incidence of RDS was 32-50%. Considering only sibs born after the probands yielded the empiric recurrence risk of 17--27% for all younger sibs and 39--67% for LBW/preterm younger sibs. The risk for maternal half-sibs was of about the same magnitude as that for full sibs, while the risk for paternal half-sibs was minimal. Among the LBW/preterm first cousins of probands, only the infants of maternal aunts showed an RDS incidence clearly higher than that in the general population. We think these data suggest a genetically determined maternal factor predisposing the infants of certain mothers to RDS. Other significant findings include: 1) an excess of males among the probands but a normal sex ratio among the sibs of the probands; 2) a decrease in mean birth weight and mean length of gestation for not only the probands but also their sibs; 3) a decrease in the mean parental ages at the birth of the probands; 4) a relative dearth of first-born and an excess of second-born infants among the probands; 5) an increased incidence of stillbirths in the sibships; 6) an increased number of probands born by cesarean section; and 7) a twin concordance of 75%.


Prenatal diagnosis of osteogenesis imperfecta congenita by means of fetography.

By means of fetography, prenatal diagnosis of osteogenesis imperfecta congenita was performed on two pregnant women in the 34th week of gestation with familial histories of the disease. In 1 of the 2 cases, the fetogram revealed abnormal twisting of the torso, shortening, and severe angulation deformities of the extremities with callus formation and parchmentlike skull despite clear-cut outline of the scalp. On the basis of these findings, the fetus was diagnosed as osteogenesis imperfecta congenita and subsequently confirmed. Thus, the excellent fetal resolving power of fetography permitted successful prenatal diagnosis of the disease far more easily and precisely than plain roentgenography.


Studies of malformation syndromes of man XXIX: the Wiedemann-Beckwith syndrome. Clinical, genetic and pathogenetic studies of 12 cases.

This report describes 12 patients with the Wiedemann-Beckwith syndrome (WBS), including 6 familial cases from 2 families. The clinical manifestations do not allow for a differentiation between familial and sporadic cases. Consistent morphologic features include organomegaly, cytomegaly and nucleomegaly. The pathogenetic process may involve few or many organs and tissues and may represent a nuclear/mitotic dysfunction. Clinically, the manifestations are hyperplasia, hypoplasia, dysplasia, neoplasia and defects in differentiation. Secondary functional disturbances are at times prominent. The differential diagnosis of the WBS includes 1) the Wilm's tumor (WT)-aniridia syndrome: 2) the "tumor-hypertrophy syndrome" which includes WT, adenocortical tumors or hepatoblastoma; 3) the WT-pseudohermaphroditism syndrome; and 4) the "tumor-nevus syndrome" with or without malformations (particularly duplications) of the urinary tract. The latter two conditions are apparently not associated with hemihypertrophy. Familial occurrence suggests that some cases of the WBS may be due to delayed mutation. Carriers of the premutated allele appear to belong to two classes: those with a high risk of producing affected offspring and those who transmit the premutated allele but have no affected offspring.


In vitro studies on the way in which the activation of factor VIII is affected in mixtures of plasma with hemophilia A plasma.

Factor VIII exchange test experiments with hemophilia A plasmas were performed to find out how the results were affected by submitting plasmas and plasma mixtures to different incubation periods at 37 degrees C, heat precipitation, and ether extraction. The experiments led to the following results: 1. In plasma mixtures, frequently higher factor VIII activities are found than can be expected from the single activities of the used plasmas. Activity increases are factor-specific. 2. The component to be activated is in the hemophilic plasma; the activity-increasing agent is in the normal plasma. These results lead to the hypothesis that hemophilia A patients have sufficient quantities of inactive factor VIII, but it stays inactive for lack of the necessary activator. In normal plasma the activator is in balance or surplus to the inactive factor. By adding normal plasma, thus supplying free activator, the inactive factor VIII of hemophilia A plasma can, under suitable conditions, be developed into active factor VIII.


Comparative studies on coagulation activities in normal plasmas and their mixtures.

By single factor exchange test experiments it was proved that mixtures of normal plasmas usually have the coagulation activity that can be concluded from the single activities of the used plasmas. In individual cases, however, normal plasmas influence each other in the mixture to the effect that coagulation is increased or decreased. This has to be considered when plasma pools for standard curves are established.


Intestinal adaptation after jejuno-ileal bypass operation for massive obesity.

Intestinal adaptation was studied in six patients with massive obesity treated by jejuno-ileal bypass operation. Glucose absorption in the jejunum was measured by a perfusion technique. The morphometric and enzymatic measurements were carried out on biopsies from the proximal jejunum and the distal ileum. Results obtained before and six months after the operation were compared. The glucose absorption per unit length of jejunum was unchanged at a glucose concentration of 66 mmol/l in the perfusate but increased significantly at a glucose concentration of 133 mmol/l (p less 0.025). The mean sucrase activity did not change, whereas the lactase activity increased significantly in the jejunum and ileum. The mean villus height increased significantly, while the epithelial cell height and cell width were unchanged both in the jejunum and the ileum, suggesting that the operation resulted in epithelial cell hyperplasia. The glucose absorption in the jejunum was positively correlated with the villus height (r = 0.76), which suggests that the increased glucose absorption was related to an increased number of epithelial cells.


Peptide-bound hydroxylysine and large polypeptides related to collagen synthesis.

About 10% of the urinary hydroxylysine is linked to peptides. There is no significant difference between the excretion of peptide-bound hydroxylysine and hydroxylysine measured in the non-dialysable fraction of the urine. Non-dialysable hydroxylysine is highly glycosylated and contains mainly glucosyl-galactosyl-hydroxylysine. It is concluded that the urinary peptide-bound hydroxylysine is present in the non-dialysable collagen polypeptides. Their unusual carbohydrate composition is discussed.


Effect of lead on hepatic delta-aminolaevulinic acid synthetase activity in the rat: a model for drug sensitivity in intermittent acute porphyria.

The hereditary hepatic porphyrias are disorders of porphyrin and haem synthesis characterized by a marked idiosyncrasy towards a variety of lipid soluble drugs. Most of these agents are inducers of the haemoprotein cytochrome P450, the terminal oxidase in drug metabolism. The primary genetic defect in intermittent acute porphyria is a partial deficiency of uroporphyrinogen I synthetase, which may result in a secondary derepression of delta-aminoaevulinic acid synthetase, the rate-limiting enzyme in the haem pathway. Analogous defects at more distant sites may explain the other hereditary hepatic porphyrias. As drug sensitivity may be related to the defect in haem synthesis, we investigated the effects of experimental partial blocks in haem synthesis produced by lead in rats. Drug effects on delta-aminolaevulinic acid synthetase, cytochrome P450, And drug metabolism were studied. Our findings indicate: a) While partial impairment of haem biosynthesis has only minor effects on delta-aminolaevulinic acid synthetase activity, it greatly enhances the sensitivity of delta-aminolaevulinic acid synthetase to induction by drugs and steroids, which when given alone, have little or no inducing effect on the enzyme. b) The experimental partial block in haem synthesis delays and impairs drug-mediated induction cytochrome P450 and drug metabolism in vitro. The findings may explain why a large number of structurally unrelated compounds with little effect on normal liver can precipitate "aucte porphyria".


Plasma renin activity in diabetic autonomic neuropathy.

Postural changes in plasma renin activity were studied in three groups of age and duration-matched male diabetics (potent, impotent and with postural hypotension) and in non-diabetic control subjects. Those diabetic subjects with postural hypotension due to automatic neuropathy had no increase in plasma renin activity to the erect posture whereas both the potent and impotent groups had similar plasma renin activity responses to the control subjects. There was a significant inverse correlation between the rise in plasma renin activity on standing and the postural drop in blood pressure (r = 0.476, P less than 0.01) but no correlation with other tests of autonomic reflex function such as the Valsalva manoeuvre and blood pressure response to sustained handgrip. The results suggested that the lesion responsible for the postural hypotension is in the efferent sympathetic pathway. However, neuropathy per se did not wholly explain the decreased postural plasma renin activity response. Diabetic nephropathy, with involvement of cells of juxtaglomerular apparatus, may also be implicated.


The role of the beta-adrnergic receptor in the secretion of gastrin: studies in normal subjects and in patients with duodenal ulcers.

Intravenous infusion of isoproterenol, a beta-adrenergic receptor stimulatory agent, increased serum gastrin concentration significantly more in patients with a duodenal ulcer than in healthy subjects. The rise in pulse rate, blood glucose concentration and in serum insulin was the same in both groups of subjects. Gastrin secretion was also increased significantly more in the patients than in the control subjects after a beef-meal. Basal serum gastrin concentrations were higher in the patients than in the control subjects and correlated to the rise in serum gastrin during both tests in the patients with a duodenal ulcer. Isoproterenol and meal stimulated gastrin secretion, expressed as percent of the basal value, were twice as higher in the patients as in the control subjects. The combined administration of isoproterenol and the meal had an additive effect on the rise in serum gastrin. Isoproterenol stimulated gastrin secretion was completely suppressed by propranolol, a beta-adrenergic receptor blocking agent, which had no effect on meal stimulated gastrin secretion. It is concluded that the mechanism of the hypersecretion of gastrin in patients with a duodenal ulcer did not involve a specific abnormality of the beta-adrenergic receptor or the receptor which recognized proteins and their digested products. There is no established role of beta-adrenergic receptor activity in the hypersecretion of gastrin in patients with duodenal ulcers. It is suggested that the beta-adrenergic receptor may have some yet unknown function unrelated to the acute secretory response of gastrin.


Plasma and urinary dopamine: studies during fasting and exercise and in tetraplegic man.

Dopamine, noradrenaline and adrenaline were measured in plasma and in urine, using double-isotope derivative techniques, in 46 normal subjects and in 17 tetraplegic patients with physiologically complete cervical spinal cord transections above the sympathetic outflow. Dopamine was present in plasma in normal subjects in a concentration of 0.33 mug/l +/- 0.06 (SEM). Twenty-four hour urinary excretion of dopamine averaged 248 mug +/- 22. There was a significant correlation between the 24 h urinary excretion of dopamine and of noradrenaline. In the normal subjects plasma dopamine and the urinary excretion of dopamine did not change during three days of fasting while urinary excretion of adrenaline increased twofold. In the normal subjects exercise significantly increased plasma dopamine from 0.25 mug/l to 0.43 mug/l, but significantly decreased the urinary excretion of dopamine. Exercise significantly increased the excretion of noradrenaline. In the tetraplegic patients the plasma dopamine concentration and the urinary excretion of dopamine were lower but not significantly different from the corresponding values in the normal subjects. Plasma noradrenaline and the urinary excretion of noradrenaline and adrenaline were significantly lower in the tetraplegic patients. It is concluded that dopamine is present in human plasma in concentrations similar to that of noradrenaline. Free dopamine in plasma and urine of normal subjects is not dependent on foot intake. Urinary dopamine may be derived from circulating dopamine. Urinary dopamine does not necessarily appear to reflect changes in plasma dopamine. The relationship between plasma dopamine and changes in adrenergic nervous activity deserves further investigation.


Haemodynamic effects of procainamide and phenytoin at apparent therapeutic plasma levels.

The haemodynamic effects of procainamide and phenytoin after intravenous administration to 19 patients with suspected congenital heart lesions were studied. The drugs were injected into the pulmonary artery in a dose of 5 mg per kg bodyweight, at an infusion rate of 50 mg per minute, thus giving plasma concentrations within the suggested therapeutic ranges. The diastolic pressures in the pulmonary artery and the right ventricle were reduced by procainamide and the converse effect was noted after phenytoin. The heart rate was significantly increased by both drugs but with phenytoin this effect was only transitory. The latter drug also caused a small and transient fall in cardiac output and mean arterial pressure. It was concluded that of the two drugs phenytoin seemed to have the most marked myocardial depressant effect.


Studies on hydralazine. II. Elimination rate and steady-state concentration in patients with impaired renal function.

Following a single 50 mg dose of hydralazine (Apresoline) in 13 patients with impaired renal function, a decrease in glomerular filtration rate (GFR) was correlated with an increase in serum half-life (T1/2) of the drug (r=-0.69; p less than 0.01). The T1/2 was 15.8 h in one patient with a GFR of 16 ml-min-1, as compared to a T1/2 of 1.7-3.0 h found previously in 16 healthy volunteers. In 49 patients on long-term antihypertensive treatment with hydralazine, the ratio between the minimum steady-state drug concentration and the daily dose of hydralazine (Cminss : Dose) increased as the GFR decreased. This accumulation of the drug was particularly evident in patients with a GFR less than 30 ml-min-1 (r=-0.63; p less than 0.01; n=19). As renal excretion of unchanged hydralazine is generally regarded as unimportant, the slower elimination rate in chronic renal failure was probably caused by a slower rate of metabolic conversion. It was found, however, that the renal excretion of hydralazine could easily have been underestimated, as only 12.7% of an initial hydralazine concentration of 200 ng-ml-1 in urine could be recovered after storage of the samples at room temperature for 24 h.


Investigation of the pharmacodynamics and pharmacokinetics of 2-(2,4-dimethoxyphenyl)-Imidazo-(4,5-b)-pyridine hydrochloride (AR-L 57 CL) in man.

AR-L 57 CL is an imidazole derivative which has been shown in animal studies to have a pronounced positive inotropic effect. This effect and the pharmacokinetics of AR-L 57 CL have been investigated by non-invasive methods in 8 healthy volunteers. After a single intravenous dose of 200 mg, administered as part of Phase 1 of the clinical studies, AR-L 57 CL plasma concentrations were measured by fluorimetry at intervals for up to one hour. Its inotropic action on the heart was demonstrated by changes in the systolic time intervals: QS2 = duration of electro-mechanical systole; PEP = pre-ejection period; LVET = left ventricular ejection time. The decrease in plasma concentration could be expressed in terms of an open two-compartment pharmacokinetic model. The shorter elimination phase had a t1/2 of 4 min and the longer a t1/2 of 30 min. Immediately after injection, QS2 and PEP (corrected for heart rate) as well as PEP/LVET (independent of heart rate) decreased considerably. They had returned to normal by 22 min after injection. The plasma concentrations of AR-L 57 CL of 2 - 5 mug-equivalents/ml showed a highly significant correlation with the decrease in systolic time intervals. Both systolic and diastolic blood pressure rose briefly after injection. The AV conduction time fell initially and the heart-rate increased briefly. Thus AR-L 57 CL was shown to be a short acting drug with a high degree of positive inotropic action. It did not cause bradycardia or increase atrioventricular transmission time and appeared to be easily controllable.


Influence of sex on the clinical pharmacology of flutiorex, a new anorectic drug.

The effects of flutiorex, a new anorectic agent, and of a placebo on food intake and the activity of the sympathetic and central nervous systems have been compared in a double blind trial in two groups of healthy volunteers, one of five males and the other of five females. Flutiorex 20 mg orally had a significant anorectic effect both in males and females, the observed reduction in food intake being 34.0 and 27.5%, respectively. It caused alpha-adrenergic stimulation (blood pressure and pupil diameter) and central nervous system excitation (critical flicker frequency), both of which were more marked in males than in females. Flutiorex was better tolerated by women than by men. Measurement of the blood level of flutiorex and its de-ethylated metabolite, norflutiorex, showed that both compounds were detectable in four of the five male subjects, but in only one of the five females. Sex-linked differences in the pharmacokinetics of flutiorex may explain the greater intensity of its effects in males.


Pharmacokinetics of amitriptyline infused intravenously in man.

Amitriptyline was given to four male volunteers by constant rate intravenous infusion. Blood samples were collected before, during and at various times after the infusion for estimation of the serum concentrations of amitriptyline. The level of nortriptyline never reached a detectable level. A two compartment open model was shown to be applicable to the data obtained. The meaning of the parameters obtained by a non-linear, least squares curve fitting procedure is discussed and the values are compared to those recently published for nortriptyline. The calculated biological half-life of amitriptyline was about 17 hours, a figure which differs considerably from previously calculated values for volunteers, but is in accordance with some newer results from patients.


Quantitative contributions of IgG, IgM and C3 to erythrophagocytosis and rosette formation by peritoneal macrophages, and anti-opsonin activity of dextran sulfate 500.

In vitro phagocytosis by guinea pig peritoneal macrophages of immune complexes (EA) was shown to be dependent on IgG antibody in a dose-dependent fashion. C3b enhanced phagocytosis of EA at limited IgG antibody concentrations only. When IgM antibody was used for sensitization of sheep red blood cells (SRBC), phagocytosis and rosette formation did not occur in the absence of bound C3. The polyanion, dextran sulfate 500 (DS), was shown to depress both rosette formation and phagocytosis of EAIgG, C1423 and EAIgMC1423, as well as immune adherence of human group 0  erythrocytes and hemolytic activity of C3. This effect of DS was seen only when it was actually present in the incubation medium.


Analysis of antibody populations to oligo-D-alanine. VIII. Sequential studies on properties of antibody subpopulations: rate constant, affinity, and isoelectric focusing spectrum.

Allotypically marked antisera to the hapten oligo-D-alanine were studied over periods up to 10 months in individual mice and in serial transfers of both nonimmune and memory cells to irradiated recipients. Parameters observed were divalent association rate constants, affinity and autoradiography of isoelectric focusing (IEF) spectra. The two allotypic subpopulations of heterozygous mice are shown to be independent with respect to affinity. Association rate constant, affinity, and IEF spectra of the antibody populations remain largely stable in most mice over prolonged periods. There is an occasional isolated dramatic change in a single property of one allotypic subpopulation; this may be in the direction of either lower or higher rate constant or affinity. While different recipients of naive spleen cells from a single donor, when immunized, differ one from the other in the measured antibody properties, the characteristics now established in each recipient persist in a lineage of transferred memory cells up to 4 generations, with infrequent changes in isolated properties of only one allotypic subpopulation. The limited changes observed would be in agreement with a model of moderate somatic variability superimposed on a small spectrum of germ line genes.


Enzymatic modification of lymphocyte receptors for antigen. III. Resistance of receptors to trypsin at the peak of the immune response.

Using sheep erythrocytes (SRBC) as the antigen, two subpopulations of spleen antigen-binding lymphocytes could be distinguished by a marked difference in the susceptibility of their receptors to trypsin. In unimmunized animals, 30% of the antigen-binding cells were trypsin-resistant, whereas at 5 days after immunization, 80-90% were trypsin-resistant, indicating an increase of about 50-fold in trypsin-resistant antigen-binding cells per spleen. In contrast, trypsin-sensitive cells per spleen were only 4-fold higher on day 5 than before immunization. The rise in % trypsin sensitivity preceded the increase in rosettes per spleen, implying that immunization produced a preferential increase in trypsin-resistant antigen binding cells partly by converting sensitive cells to resistant cells. After the 5th day, the trypsin sensitivity of antigen-binding cells slowly returned toward the unimmunized level, but a booster injection of SRBC restored trypsin resistance. Trypsin resistance was not lost in the presence of sodium azide or protein synthesis inhibitors. But a slightly increased trypsin susceptibility was conferred by 2-deoxyglucose, implying that glycolysis or the glycosylation of protein may be involved in maintaining trypsin resistance.


Characteristics of the effector cells mediating cytotoxicity against antibody-coated target cells. II. The mouse nonadherent K cell.

A cell of lymphoid morphology capable of killing antibody-coated chicken erythrocytes was isolated from nonimmune mouse spleen using a combination of carbonyl iron treatment and glass bead column passage. This non-phagocytic effector cell, which is referred to as the nonadherent K (killer) cell, is distinguished from the non-phagocytic myeloid K cell described earlier (Greenberg, A.H., Shen, L. and Roitt, I.M., Clin. Exp. Immunol. 1973. 15:251) by its relatively weak surface adherence properties and low concentration within the mouse spleen. The cell is further characterized by its relatively large size, lack of theta or immunoglobulin determinants, the presence of Mg++-independent complement receptors, affinity for aggregated IgG2 myeloma proteins, inhibition by cytochalasin B and good survival in cell culture. The possible lineage of the cell is discussed.


Close linkage between genes coding for allotypic markers on chicken IgG and IgM.

The allotypic markers Gla and Gld (present on chicken IgG molecules) and Mla and Mlc (present on IgM) have previously been defined. The inheritance of these markers was studied in the F2 progeny of a cross between birds of the inbred lines CB (Gla+,d-,Mla+,c-) and WA (Gla-,d+,Mla-,c+). The results suggest that antigens Gla and Gld, and Mla and Mlc, are genetically controlled by alleles at two loci (Gl and Ml, respectively) and also show that these loci are closely linked.


Precipitin and mitogenic behavior of dimeric and tetrameric concanavalin A.

The results of double diffusion and quantitative precipitin tests show that in contrast to the concanavalin A tetramer, the dimer is incapable of precipitating glycogen. The results support previous work suggesting that dimers composed of two fragmented monomer units possess only one binding site for carbohydrate (McKenzie and Sawyer, J. Biol. Chem. 1973. 248: 549). On the other hand, dimer and tetramer species had similar mitogenic activity as measured by the incorporation of radioactive thymidine by mouse spleen lymphocytes and induced cap formation to similar degrees. Results are discussed in terms of the affinity and valency of the polymeric forms and their stability at the cell surface.


Genetic control of the immune response to the radom linear terpolymer of L-glutamic acid, L-lysine and L-leucine (GLleu) by complementing Ir genes.

Three levels of humoral responsiveness (none, high or marginal) were noted following secondary immunization with random linear terpolymer of L-glutamic acid, L-lysine and L-leucine (GLleu). These immune responses were under dual H-2-linked Ir gene control. Gene complementation was observed in appropriate F1 offspring between two nonresponder strains and in selected intra-I region recombinants derived from crossing over between nonresponder parental haplotypes. One of the Ir-GLleu genes, beta, was localized within the I-A or I-B subregions. The other Ir-GLleu locus, termed alpha, was mapped in the I-C or S subregions. Differences in the strain distribution of immune responsiveness to GLleu and related terpolymer GLphe (which is also under complementing Ir gene control) are primarily attributable to differences among the Ir-GLleu and Ir-GLphe beta alleles. A correlation exists between the presence of the Ir-GLleu alpha(¿allele and the gene coding for specificity Ia.7.


Analysis of cardiac chronotropic responses to some autonomic blocking agents in conscious trained dogs.

The changes of heart rate in response to i.v. administration of methylatropine (0.5 mg/kg) and/or propranolol (2 mg/Kg) or practolol (2.5 mg/Kg) were studied in conscious trained dogs. Cholinergic blockade alone or combined blockade of sympathetic and parasympathetic effector systems resulted in cardiac acceleration. Conversely, beta-adrenoceptor antagonism with either propranolol or practolol reduced heart rate. The data were analysed by means of a new method, whereby the heart (HRN) of the dog is considered to be the product of the intrinsic heart rate (HR0) and 3 further factors: HRN-HR0 times S times V times W (Multiplicative model). 2 of these factors represent the tonic sympathetic (S) and parasympathetic (V) influences, whereas the third (W) represents the sympathetic-parasympathetic interaction. This type of analysis reveals that W was approximately 1, i.e., the sympathetic-parasympathetic interaction did not play any significant role in determining the heart rate of conscious resting dogs (HRN = HRO-S-V-W = HRO-S-V). The change of heart rate due to the action of parasympathetic system (-53% of the intrinsic heart rate) was more important than the change caused by the action of the sympathetic system (26% of the intrinsic heart rate).


The roles of noradrenaline and dopamine in contraversive circling behaviour seen after unilateral electrolytic lesions of the locus coeruleus.

Unilateral electrolytic lesions of the locus coeruleus in rats result in spontaneous ipsiversive rotation, which is then replaced by contraversive rotation. One week after lesioning, when spontaneous turning ceases, apomorphine and d-amphetamine elicit contraversive circling behaviour, which was not affected by noradrenergic receptor blockade but was abolished by dopamine receptor blockade. The drug-induced contraversive circling response was also reproduced by piribedil but not clonidine. Combined unilateral electrolytic locus coeruleus and substantia nigra lesions on the same side resulted in apomorphine- and d-amphetamine-induced ipsilateral rotational behaviour which was indistinguishable from that seen with substantia nigra lesions alone. In rats with unilateral locus coeruleus lesions, the dose of intrastriatally injected apomorphine required to produce circling was less on the lesioned than the non-lesioned side. Direct injection of noradrenaline into one substantia nigra caused contraversive circling. Direct injection of phenoxybenzamine into one substantia nigra followed by apomorphine caused ipsiversive circling. The results suggest that the circling behaviour seen after unilateral locus coeruleus lesions depends on an asymmetry of striatal dopamine receptor activity and are consistent with a proposed coeruleus-nigral noradrenergic pathway, which enhances impulse flow in the dopaminergic nigrostriatal system.


Effects of physostigmine and electrical stimulation on the acetylcholine content of the guinea-pig ileum.

Incubation with physostigmine (7.7 muM) caused an approximately 2 fold increase in the acetylcholine content of the myenteric plexus--longitudinal muscle preparation of the guinea-pig ileum. This effect was due mainly to an increase in 'free' acetylcholine, which was directly assayable in either the homogenate after removal of cell debris or the supernatant fraction (100,000 g for 60 min) after subcellular fractionation. Acetylcholine output during stimulation at 0.017, 0.1 or 1 Hz was maintained for 60 min at a rate 2--4 times greater than the non-stimulated output; there was no change in content. At 10 HZ, output was high at the start of stimulation and then decreased continuously; there was a proportionate loss of mainly 'free' acetylcholine from the tissue. Mn2+, hexamethonium, morphine and noradrenaline, which depressed acetylcholine output during stimulation at 0.1 HZ, had no effect on the acetylcholine content nor did they affect the increase in acetylcholine content during incubation with physostigmine.


Effects of salsolinol on smooth muscle responses to various biogenic amines.

Isolated smooth muscle tissues from rats, guinea pigs and rabbits were used to investigate pharmacological effects of salsolinol (1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline). By itself, salsolinol did not produce any profound or sustained sympathomimetic or 5-hydroxytryptamine-like activity in the concentrations used in these experiments (7.7 and 77  muM). Salsolinol was found to antagonize the contraction of the rabbit aorta induced by noradrenaline but did not affect the adrenaline-induced relaxation of the guinea pig tracheal chain. On both the rat fundus and oestrus rat uterus, salsolinol exhibited significant anti-5-hydroxytryptamine activity but responses of these tissues to acetylcholine and histamine remained unaffected, indicating that the action was not non-specific. These results suggest that in vivo formation of salsolinol after ethanol ingestion could produce significant pharmacological effects.