Plasma lipoproteins in rats with experimental biliary obstruction. I. A chemical study.

Acute biliary obstruction in the rat is associated with striking alterations of the plasma level and the physico-chemical properties of plasma lipoproteins. 1. The level of very low density lipproteins (VLDL) in plasma increases from 2 to 3 fold. The chemical composition of VLDL is characterized by a high content of phospholipids and cholesterol and by a diminution of the relative content of triacylglycerols and protein. On cellulose acetate electrophoresis, VLDL show a beta-mobility. 2. The plasma concentration of low density lipoproteins (1.019--1.063 g/ml, LDL2) increases several fold above the control level. Phospholipids and unesterified cholesterol are the major components of this fraction which contains only a minute amount of cholesteryl ester (4%) and triacylglycerols (10%). LDL2 contain a component which migrates to the cathode in 1% agar gel electrophoresis. Separation of LDL2 by gel filtration on 2% agarose column results in the identification of three subfractions. Subfraction I contains a large proportion of cholesterol and triacylgylcerols, subfraction II is rich in unesterified cholesterol and phospholipids whereas subfraction III has a chemical composition fairly similar to that of the control LDL2. 3. The level of high density lipoproteins (HDL) also increases after bile duct ligation. The chemical composition of HDL2 (1.063-1.125 g/ml) is characterized by a high content of unesterified cholesterol and phospholipids and by a remarkable reduction in the content of cholesteryl esters and protein.


Response of renal calcium-binding protein. Independence of kidney vitamin D hydroxylation.

Dietary calcium and dietary phosphorus restriction were studied in chicks fed either cholecalciferol or 1alpha-hydroxycholecalciferol. Intestinal calcium absorption and calcium-binding protein of 1alpha-hydroxycholecalciferol-treated chicks remained unchanged under dietary calcium restriction, but increased under dietary phosphorus restriction. Kidney calcium-binding protein was not altered by dietary caclium restriction in chidks treated with either cholecalciferol or 1alpha-hydroxycholecalciferol, but increased under dietary phosphorus restriction independent of the vitamin D source. In contrast to the intestine, calcium-binding activity of the kidney was found to be poorly related to the calcium-binding protein concentration. It is suggested that kidney calcium-binding protein is regulated by a mechanism different from that of intestinal calcium-binding protein, and that its concentration in renal tissue is related to renal caclium excretion or plasma calcium level.


The involvement of receptro sulphydryl groups in the binding of steroids to the cytoplasmic glucocorticoid receptor from rat thymus.

The glucocorticoid receptor protein present in the high-speed supernant fraction of rat thymus tissue is extremely unstable, having a half-life of about 2 h at 4 degrees C. It was found that the decline in steroid-binding capacity could be slowed, though not arrested completely, by the addition of sulphydryl-protecting agents such as 2-mercaptoethanol or dithiothreitol, and by EDTA. The inactivation was also partly reversed by these agents. 0.5 mM N-ethylmaleimideor p-chloromercuriphenylsulphonic acid inactivated the recptor at 4 degrees C, but the presence of bound steroid protected the receptor against this inactivation. Bound steroid did not protect the receptor against the action of higher concentrations of these reagents. Treatment of intact thymus cells with 2,4-dinitrophenol resulted in a reduction in the steroid-binding capacity of the supernatant fraction derived from these cells. This effect of 2,4-dinitrophenol could not be reversed by the presence of dithiothreitol in the extraction buffer. It is concluded that the inactivation of the receptor in vitro is at least partly due to the oxidation of one or more sulphydryl groups necessary for steroid binding; the process of oxidation does not account for the reduction in steroid binding observed in intact thymus cells under conditions of energy deprivation.


The effects of pregnancy and lactation on the activities in rat liver of some enzymes associated with glucose metabolism.

1. The ratio of the combined activities of hexokinase (EC 2.7.1.1) and glucokinase (EC 2.7.1.2) to the activity of glucose-6-phosphatase (EC 3.1.3.9) changed in favour of the glycolytic enzymes during pregnancy and at peak lactation. 2. There were no important changes in the ratio of the activity of phosphofructokinase (EC 2.7.1.11) to that of fructose diphosphatase (EC 3.1.3.11). 3. The ratio of the activity of pyruvate kinase (EC 2.7.1.40) to the combined activities of phosphoenolpyruvate carboxylase (EE 4.1.1.32) and pyruvate carboxylase (EC 6.4.1.1) changed in favour of the glycolytic enzyme during pregnancy and at peak lactation, but changed in favour of the gluconeogenic enzymes immediately after parturition. 4. These changes are considered in relation to the changes in food intake and hormonal status that occur during pregnancy and lactation.


Implications of the reorganizational behavior of condensed phosphates for mitochondrial phosphorylation.

The composition of phosphorus-32 labeled polyphosphoric acid (32PPA) prepared from, (i) H3(32)PO4 and (82 percent w/w) P2O5 and, (ii) by incorporation of radioactivity from H3(32)PO4 into unlabeled polyphosphoric acid (PPA) was examined and found to be the same. The identity of these condensed phosphorate mixtures, as established, argues strongly in favor of a rapid and complete equilibration and provides direct evidence in support of Van Wazer's reorganization theory of condensed phosphates. No evidence for metaphosphates or label exchange in an aqueous environment was found. 32PPA was found to exist in metastable equilibrium with 32PPA as a crystalline solid of different composition. Agreement of hydrolytic rate data from the mixtures of condensed phosphates with literature values for individual species suggests that the hydrolysis of the pyro- and tripoly-phosphate species proceeds independently in the presence of other homologs. The rate of hydrolysis of the condensed phosphates (at pH5.0 and 65 degrees C) was found to be proportional to chain length, increasing about four-fold between pyro- and hexapoly-phosphate. The same correlation is also manifest in the adenosine 5' polyphosphates. A slight enhancement in the rate of hydrolysis of adenosine diphosphate, increasingly more pronounced in the case of adenosine tri- and tetra-phosphate, was also noted. This effect is attributed to the presence of the adenosine moiety. The most interesting aspect of the rate data is to be found in the similarity of the rates of hydrolysis of the adenosine 5' polyphosphates, relative to their inorganic polyphosphate counterparts. The significance of these findings is discussed and the viewpoint is presented that the properties of condensed phosphates are compatible with the requisite characteristics that must be predicated of any in situ phosphoryl precursor in mitochondrial phosphorylation.


The effect of catecholamines and prostaglandins upon human and rat erythrocytes.

Both human and rat erythrocytes respond to low doses (10(-11)--10(-9) M) of L-isoproterenol and L-epinephrine with an increased degree of hypotonic hemolysis and a decreased rate of filtration through standardized paper filters. The receptors in both cell types have many of the characteristics of beta-receptors for catecholamines. However, hormone-receptor interaction in the human cell does not lead to an increase in intracellular cyclic AMP concentration, but in the rat cell, hormone-receptor interaction does lead to a significant increase in cyclic AMP content. Thus, catecholamine-beta-receptor interaction, at least in the human red cell, leads to a change in red cell properties which are not mediated by adenylate cyclase activation. Likewise, prostaglandin E2, at 10(-12)--10(-10) M, causes are increased degree of hypotonic hemolysis and a decreased rate of filtration through standardized paper filters, but it also does not increase the cycliC AMP content of the human erythrocyte but does increase that of the rat erythrocyte. Nevertheless, exogenous cyclic AMP, when added at a concentration of 10(-8) M to washed human erythrocytes, increases the degree of hypotonic hemolysis. Conversely, prostaglandin E1, at 10(-12)--10(-10) M, causes a decreased degree of hypotonic hemolysis and an increased rate of filtration through a standard filter. Both prostaglandin E2 and the catecholamines decrease the size of a rapidly exchangeable calcium pool, and prostaglandin E1 increases it.


Enzymatic and non-enzymatic assay of superoxide dismutase.

Superoxide dismutase from breef brain and rat liver was assayed in an enzymatic system, using xanthine oxidase, and a non-enzymatic system, based on aerobic reduction of nitro-blue tetrazolium in presence of phenazine methosulphate. The non-enzymatic assay is rapid and simple and permits simulatneous analysis of many samples.  Similar results are  found by the two methods of assay of superoxide dismutase.


Desoxyribonucleic acid and pyrimidine synthesis in the rat during intra-uterine growth retardation: responsiveness of several organs.

The response of DNA synthesis, thymidine incorporation and the activities of uridine kinase, carbamyl phosphate synthetase and aspartate transcarbamylase were studied in the rat, following intra-uterine growth retardation. A good correlation was found between the falls of DNA production and thymidine incorporation and the decrease of the activity of the three enzymes during the fetal period. After birth, cell proliferation remained depressed but no further changes occurred in the enzymatic activities.


Neurophysiological observations in diseased newborns.

The early statements that the EEG alone could correctly be used for the assessment of gestational age (cf., among others, Dreyfus-Brisac, 1964; Parmelee et al., 1968; and Nolte et al., 1969), even in pathological babies, need some restrictive qualifications. Likewise, the more recent statements that certain neurophysiological parameters, including nerve conduction velocity and basic EEG patterns are independent not only of gestational age and body weight, but also of various abnormal factors in the pre- and perinatal periods, appear also too rigid, at least as regards the EEG and the other parameters used in this study. As a matter of fact, some authors who had adhered rather strictly to these concepts have later on presented evidence suggesting some deviations in bioelectric brain maturation in some cases, as for example, in small-for-date newborns of toxemic mothers (Schulte et al., 1972). When more exact quantification by coding techniques was applied (Parmelee et al., 1968), the conclusion was also reached that extrauterine and intrauterine development of bioelectric brain activity developed according to the conceptional age, regardless of variable extrauterine experience, although it was recognized that other categories of behavioral and neurophysiological activities, for example, Moro reflex, states stability, crying, sucking, scores, etc., are more labile to environmental and pathological factors (Prechtl et al., 1972; Michaelis et al., 1973).


Biorhythms in toxicology and pharmacology. I. Generalities, ultradian and circadian biorhythms.

According to their periods, rhythms can be classified in ultradian (period inferior to one day), circadian (period equal to one day) and infradian (period superior to one day). Different biorhythms of endogenous origin are modified mostly relatively to their amplitude by definite doses of the effects of pharmacological or toxic agents. Circadian variations of the effects of drugs, of the toxicities of gases as well as circadian modifications of the reactions to different stresses and agressions have been demonstrated amply in animals (mostly nocturnal rodents) and sometimes in man. In a general way reactions are more rapid and more marked in the active period where enzymatic reactions show a peak activity corresponding to a maximal energy output than during the rest and sleep periods.


Stimulation by organic solvents and detergents of conversion of L-sorbose to L-sorbosone by Gluconobacter melanogenus IFO 3293.

Treatment of Gluconobacter melanogenus IFO 3293 cells with benzene, carbon tetrachloride, cyclohexane, deoxycholate, toluene, or xylene stimulated their conversion of L-sorbose to L-sorbosone two- to threefold. The degree of stimulation depended upon the length of exposure time to the agent and the age of the G. melanogenus cells. A rapid decrease in viability of the cells and degradation of cell RNA was noted after treatment with the effective agents. The G. melanogenus cells were unable to absorb L-sorbose actively after toluene treatment.


[Effect of triftazin on the transmission of excitation in the brain of the rabbit under the prolonged administration of the preparation].

Experiments were conducted on awake rabbits to which trifluoperazine was given daily for 2 weeks in a dose of 1 mg/kg a day; a study was made of the reaction of the cortical neurones in response to the electrostimulation of its adjacent areas. In measuring the dispersions of the poststimular histograms it was found that under the effect of trifluoperazine there occurred a reduction of the value and the duration of the responses of the cortical neurons after the infliction of stimulation. The effect obtained after the administration of trifluoperazine was regarded from the point of view of aggravation of the signal conduction in the neuronal reticula.


[Effect of catecholaminergic substances on the proconvulsive properties of the caudate nucleus].

In the freely moving rats stimulants of catecholaminergic tramission (DOPA, apomorphine, d,1-amphetamine, and also their combination with disulfiram) reduced the proconvulsive properties of the caudate nucleus. Under the effect of these substances there occurred a shortening of the cortical electroencephalographic response to a single stimulation of the nucleus in the animals given subconvulsive doses of pentaxylenetetrazol and a reduction of the extent of the spike-wave rhythm induced by the repeated caudate stimuli. On the contrary, inhibitors of catecholaminergic transmission (chlorpromazine, haloperidol, alpha-methyltyrosine and disulfiram) intensified the proconvulsive effect of the caudate nucleus.


[Effect of sodium oxybutyrate on regional blood circulation and on neural regulation of vascular tonus].

The effect of sodium hydroxybutyrate on the blood flow in the aorta, carotid, mesenteric and femoral arteries were studied on cats and dogs. The circulation was assessed by the electromagnetic and resistographic methods, in the anesthetized and nonanesthetized animals. The tonic activity was recorded in the sympathetic nerves and the EEG. Sodium hydroxybutrate was shown to decrease the sympathetic activity, resulting in the increase of the regional circulation and induced the EEG synchronization. The latter effect was more pronounced in the arotid arteries. It can be assumed that sodium hydroxybutyrate affects the nervous control of the blood vessels.


An electrophysiological analysis of the effects of reserpine on adrenergic neuromuscular transmission.

1 An electrophysiological study has been made of the effects of depleting synaptic vesicles (i.e. small vesicles less than 60 nm diameter) of their transmitter with reserpine on the quantity of transmitter released by nerve impulses, using the amplitude of the synaptic potential as a measure of transmitter release. 2 Pretreatment of adrenergic nerve terminals with reserpine sufficient to deplete the terminals of 70% of their noradrenaline (NA) did not change the total number of synaptic vesicles in the terminals, but did reduce the number with a large granular core as well as the quantity of NA released by a single nerve impulse by 80%. 3 Pretreatment of adrenergic nerve terminals with reserpine and iproniazid, to decrease vesicular NA but enhance cytoplasmic NA, had the same effect on synaptic vesicles and on the NA released by a single nerve impulse as did reserpine alone. 4 During a short train of impulses at high frequencies in reserpine pretreated terminals, the quantity of NA released by successive impulses increased until a steady-state release was reached comparable to that in untreated preparations. This facilitated release could be quantitatively predicted in terms of the addition of the individual potentiations introduced by each impulse in the train. 5 These results are consistent with the idea that each quantum of transmitter is stored in a synaptic vesicle, and that these may be released by nerve impulses directly from the terminal by a process of exocytosis.


An electrophysiological analysis of the effect of Ca ions on neuromuscular transmission in the mouse vas deferens.

1 A study has been made of the effects of changing the external calcium concentration [Ca]0 and the external magnesium concentration [Mg]0 on the synaptic potential due to noradrenaline release. 2 When [Ca]0 was varied in the range 0.7 to 1.8 mM, the synaptic potential increased as about the second power of [Ca]0. 3 Increasing [Mg]0 depressed the synaptic potential; however, variation of [Ca]0 in the presence of high [Mg]0 did not significantly change the power relationship between the synaptic potential and [Ca]0. 4 The facilitated increase in the synaptic potential during short trains of impulses at different frequencies was quantitatively predicted on the assumption that each impulse leaves residual Ca ions bound to release receptors in the nerve terminal.


Community care: an analysis of assumptions.

The implementation of a policy of "community care' is seen to involve a number of assumptions, some of which are rarely examined. These can be roughly categorized as involving the nature of mental illness, the nature of community, the course and treatment of mental illness, the proper scope of psychiatry, the burden on the community and the efficacy of social work. Data bearing on these assumptions are reviewed, and the conclusion is offered that they are far from being uncontentious. It is suggested that the movement toward community care has many of the attributes of a moral enterprise which, unless substantiated by benefits to the patient or his family, may be the latest diversion of the psychiatric conscience from the care and treatment of the chronic mentally ill.


Extra-osseous localization of 99Tcm-Sn pyrophosphate.

99Tcm-Sn pyrophosphate bone scans of 250 patients referred for skeletal metastatic survey were analysed to determine the frequency of abnormal extra-osseous localization and the various pathological causes. Twenty-six patients demonstrated abnormal extra-osseous concentration. There were three false positives. Sixty-five per cent of the extra-osseous lesions concentrating pyrophosphate were malignant (carcinoma of lung and breast, metastatic hepatic carcinoma, chondrosarcoma) and the remainder were benign lesions, e.g. sarcoidosis, soft-tissue calcification, post-surgical and irradiation sites. An incidental finding was the unusual frequency of accumulation of pyrophosphate in various joints, especially the knees and shoulders in asymptomatic patients above 70 years of age.


The nitrofurans as sperm immobilising agents: their tissue toxicity and their clinical application.

Nitrofurantoin sodium and nitrofurazone in certain critical concentrations have the ability to cause sperm immobilisation. Intraoperative vas irrigation with these solutions in 24 patients have produced sterility in the immediate postoperative period.


The sympathetic nervous system in alcoholic neuropathy. A clinical and pathological study.

Tests of autonomic function were performed on 12 subjects with alcoholic neuropathy. Abnormal sweat patterns occurred in 8/8 (100 per cent) and an abnormal Valsalva ratio in 2/9 (22 per cent). Postural hypotension and denervation hypersensitivity were absent in all patients examined. A quantitative assessment of baroreceptor function was made. The resting heart period, heart period range and mean gain of alcoholics were within the control range. Quantitative histological studies were performed on the greater splanchnic nerves removed at autopsy in 4 alcoholic subjects. The myelinated fibre density fell within the control range. The absence of significant disturbance of blood pressure control correlates well with the absence of pathology in the greater splanchnic nerve.


Recurrent brachial plexus neuropathy.

The clinical, electrophysiological and pathological changes in 3 patients with recurrent attacks of non-traumatic brachial plexus neuropathy have been described. Two had recurrent attacks and a dominant family history of similar attacks, together with evidence of lesser degrees of nerve involvement outside the brachial plexus. In one patient the attacks were moderately painful, while in the other there was little or no pain. Only one showed undue slowing of motor nerve conduction during ischaemia, but in both cases the sural nerves had the changes of tomaculous neuropathy, with many sausage-shaped swellings of the myelin sheaths, and extensive segmental demyelination and remyelination. The third patient had two attacks of acute brachial plexus neuropathy which were both extremely painful. The clinical features were compatible with a diagnosis of neuralgic amuotrophy. In the second attack, there was vagus nerve involvement and the sural nerve showed evidence of healed extensive segmental demyelination. The various syndromes presenting with acute non-traumatic brachial plexus neuropathy are reviewed, and a tentative nonsological classification advanced. Most patients fall into the category of acute, painful paralysis with amyotrophy, with no family history and no evidence of lesions outside the brachial plexus. It is suggested that the term "neuralgic amyotrophy" be restricted to this group. Patients with features outside this clinical picture probably suffer from other disease entities presenting with brachial plexus neuropathy. The familial cases constitute one or more aetioliogical subgroups, differing from neuralgic amyotrophy in the frequency of recurrences, the relative freedom from pain in the attacks, the frequency of nerve lesions outside the brachial plexus, and of hypotelorism. Individual attacks of acute brachial plexus neuropathy, however, may be identical in patients with the different diseases, and further pathological and biochemical studies are awaited to aid in nosology.


Unmyelinated fibres and Schwann cells of sural nerve in neuropathy.

Electron micrographs of 45 sural nerves from patients with acquired (22) or heredodegenerative neuropathy (23) were analysed with respect to the number of unmyelinated nerve fibres, 37 nerves with respect to the number of Schwann cell sub-units and of structures connected with Schwann cells. Findings were compared with those in 6 nerves from control subjects and referred to the total number rather than to the number per mm2 to eliminate error due to increase in the transverse endoneurial area, present in more than half the diseased nerves. Ninety-one per cent of the diseased nerves showed one or several abnormalities in unmyelinated fibres of their Schwann cells. The best indicator of fibre loss was an increase in the number of Schwann cell sub-units devoid of axons, found in more than half the nerves. This was the only abnormality related with decrease in number of myelinated fibres. The increase in number of empty Schwann cell sub-units was due both to loss of unmyelinated nerve fibres and to proliferation of Schwann cells. Proliferation was indicated by the higher incidence of Schwann cell nuclei in cross-sections of diseased nerves than in controls. The earliest sign of involvement was an increase in number of profiles and of small isolated Schwann cell projections, observed in 33 of 37 diseased nerves, as the only abnormality in 7 nerves. The number of unmyelinated nerve fibres by itself was of little value to indicate loss of fibres, since regeneration often replaced or more than replaced degenerated fibres. Regeneration was indicated by an increase in number or incidence of small unmyelinated fibres, present in nearly half of 45 diseased nerves; and by an increased in the total number, present in a third of the nerves. An increase in the number of collagen pockets and of fibres undergoing degeneration (loss of organelles) and a decrease in the number of unmyelinated fibres per Schwann cell sub-units was present in only a quarter to a third of diseased nerves and was not related to other criteria of loss of fibres or of regneration.


Effects of activity in non-myelinated afferent fibres on the spinocervical tract.

Microelectrode recordings were made from axons of the spinocervical tract (SCT) in unanaesthetized decerebrate-spinal cats. Pure volleys in non-myelinated (C) cutaneous nerve fibres were obtained by DC-polarization block of conduction in the myelinated (A) fibres, and SCT cells were classified according to whether or not they were excited by C fibres. The effects of conditioning C responses in the SCT with A and C fibre input were examined, as were the effects of conditioning A fibre responses in the tract with C fibre inputs. The effects of stimulating descending systems in the cervical spinal cord on the C fibre-evoked responses of SCT cells were determined. SCT units with a slowly-adapting pressure-sensitive component in their receptive fields responded to cutaneous C fibres in addition to A fibres. Units which responded only to hair movement when the receptive field was stimulated mechanically were not excited by C fibres. Responses in SCT neurones produced by C fibres were profoundly inhibited by conditioning volleys in A fibres of both ipsilateral and contralateral cutaneous nerves and by activity in several descending systems. Conditioning with C fibres failed to affect the responses of SCT cells to either A or C fibres. It is concluded that cutaneous C fibres excite some SCT cells but have no other effect on transmission through this system. It is suggested that there are common inhibitory interneurones in the paths from descending systems and cutaneous A and C fibres to the excitatory input to SCT cells.


Depressive effects of morphine upon lamina V cells activities in the dorsal horn of the spinal cat.

The effects of morphine upon the transmission of nociceptive messages at the spinal level have been investigated in spinal cats by studying its effects on the activities of lamina V dorsal horn interneurons. Morphine (2 mg/kg i.v.) induced a direct depressive action at the spinal level, since it strongly reduced both spontaneous and evoked activities of lamina V cells. The spontaneous firing rate and the responses elicited by natural nociceptive stimulation were decreased by 50%. The responses of these units evoked by supramaximal electrical stimulation were reduced to 67% of their initial value; in this case, the depressive effect was much more prominent on the late component of the long duration responses. The observed depressive effects are specific since they are immediately reversed by administration of opiate antagonists (nalorphine or naloxone).


Intracellular postsynaptic potentials of cat lateral geniculate cells and the effects of degeneration of the optic terminals.

(1) The synaptic signal transmission was studied in the lateral geniculate nucleus (LGN) during the late phase of degeneration, 50-80 h after light coagulation of the optic disc and the retinal vessels of one eye. Postsynaptic potentials and action potentials were intracellularly recorded from normal LGN principal cells (N-cells) and cells with degenerating optic terminals (D-cells). EPSPs and postsynaptic action potentials were elicited by electrical stimulation of the optic tract. Two neuronal classes (I and II) were characterized according to their response latency to optic tract stimuli. (2) Spontaneous small EPSPs and slow changes of the membrane potential were found in D-cells and the EPSP-IPSP pattern normally present in N-cells was absent. (3) The responses to optic tract stimuli applied at low stimulus frequencies (less than 5 stimuli/sec) were identical in D-cells and N-cells. (4) At higher stimulus frequencies (greater than 50 stimuli/sec), the EPSP amplitude was, on average, significantly smaller in D-cells than in N-cells. (5) At frequencies above 50 stimuli/sec, the fluctuation of the EPSP amplitudes and the latency scattering of postsynaptic action potentials were larger in D-cells than in N-cells. (6) The slope of the rising phase of the EPSPs increased slightly in N-cells and decreased in D-cells when the stimulus frequency was raised from 5/sec to 100/sec (class I cells). (7) The decay time constant of the EPSP, the latency of the onset and the peak amplitude of EPSPs did not change during degeneration. (8) The hypothesis put forward to account for the results is that the amount of transmitter released by each action potential in degenerating terminals is reduced, but its variability is increased. This is probably caused by a reduction in both the transmitter reserve and the transmitter resynthesis, and also by a larger fluctuation of the size of transmitter quanta.


[Role of ACTH in regulation of aldosterone secretion].

During the nycthemeral cycle, ACTH modulated the aldosterone production and increases plasmatic level at the end of night. The chronic stimulant effect of angiotensin II is a permissive factor of ACTH influence on aldosterone formation.


[Specific insulin binding to chloroplasts isolated from Acetabularia mediterranea].

Kinetics of insulin fixation to chloroplasts, exponential dissociation of bound hormone, competition between radioactive and native insulins for binding and for release indicate that chloroplasts possess insulin receptors.


[Influence of darkness on insulin binding in the unicellular algae Acetabularia mediterranea].

Culture of Acetabularia in darkness increases the number of membrane receptor sites to insulin. This increase has not been observed with anucleated parts of algae. It may be concluded that induction of insulin receptor sites depends on the presence of the cell nucleus.


[Spike activity recorded at the level of optic tract in a primate, Papio papio].

Spike activity has been recorded, in two young baboons, at the part of the optic tract (O.T.) localized between the chiasma and the lateral geniculate (L.G.) during REM and also slow wave sleep, preceding or not a REM period. These spikes, mono or biphasic, were of 60-200 ms duration and of 30-140 muV amplitude. They occurred generally in bursts (up to 50 spikes); 4 to 10 spikes bursts were the more frequent. These O.T. spikes were assimilated to the L.G. spikes.


[Cytoimmunological and histochemical study of cat adenohypophyseal cells, made fluorescent by the Falck and Hillarp technic].

In the Cat, after Falck and Hillarp method, all the fluorescent cells of the PI and the anterior lobe of adenohypophysis can be revealed with specific anti-sera to ACTH(1-24), ACTH(17-39), bovine beta-MSH and porcine beta-LPH. With the lead hematoxyline staining method, two types of cells are recognizable in the anterior lobe, in which the non hormonal constituents of the granules must be different.


[Mouse pancreas involvement in murine hepatitis virus infection].

After intraperitoneal inoculation with mouse hepatitis virus (MHv) into mice, the excretory cells of the pancreas showed a marked vacuolar degeneration concomitantly with hepatitis. MHV-specific antigen was evidenced in the cytoplasm of infected cells. Electron microscopy revealed a number of virions within the matrix, spaces of the endoplasmic reticulum and areas of focal cytoplasmic necrosis. The virus titer of the pancreas was equal or superior to that of the liver at least in early stage of infection.


Filaments and granules in mitochondrial vacuoles in chondrocytes.

As chondrocytes degenerate mitochondria frequently become vacuolated. Vacuoles seem to form by an invagination of mitochondria into their own matrix followed by cytoplasmic invasion. The bounding membrane of vacuoles and the electron dense granules which are intermittently dispersed around its periphery are positive when stained with phosphotungstic acid and with silver nitrate methenamine after periodic acid oxidation, indicating the presence of glycoprotein; they are also positive after staining with colloidal iron and bismuth nitrate, indicating the presence of acid proteoglycans. Within vacuoles and apparently arising from their glycoprotein proteoglycan boundary are small filaments composed of granular and filamentous protions. Vacuoles and their contents are exocytosed thus accounting for the presence of granules and filaments in pericellular lacunae. In pericellular lacunae and in the extracellular matrix, filaments interconnect with granules or with collagen fibrils, or both. Filaments decrease in number as the distance from the cell increases, while granules decrease in size but remain numerous. Filaments and granules decrease in the extracellular matrix as collagen fibrils aggregate and as the matric calcifies. Thus, they may play a role in aggregation of collagen fibrils. Filaments and granules also decrease with increasing age. Filaments and granules do not seem to be derived from Golgi vacuoles. They are present in costal, tracheal, and epiphyseal cartilage in three different species of animals.


An electron microscopic study on the formation of the periostracum in Helisoma (Mollusca).

The ultrastructure of the cells involved in the formation of the periostracum in Helisoma duryi duryi is described. Two distinct types of periostracal cells can be recognized: light and dark. The Golgi complexes of the light cell are involved in the biogenesis of periostracal units which are extruded at the base of the periostracal groove. Here they align into a single row so as to form the membrane-like lamellar covering of the periostracum. While the secretions of the dark periostracal cells contribute to the initial component of the dark layer of the periostracum, the mantle edge gland cells are responsible for its thickening. The adult periostracum is about 2--3 mum thick and it consists of a single dark layer covered by a membrane-like lamellar structure.


Comparison of the effect of various methods of dissociation on the protein composition of rat liver ribosomal subunits.

Rat liver ribosomes were dissociated into subunits using EDTA, sodium pyrophosphate, high concentrations of KC1, as well as by incubation with puromycin in presence of 0.5 M KC1. The subunits obtained were analyzed using the density gradient centrifugation technique and their ribosomal proteins were separated by means of two-dimensional polyacrylamide gel electrophoresis. The ribosomal protein patterns of the two subunits isolated using each of the dissociating method were compared to the protein patterns of monosomes prepared by puromycin treatment alone. Our results revealed that the use of chelating agents to dissociate the ribosomes resulted in the loss of some ribosomal proteins from the small subunit. On the other hand, the use of KC1 in high concentrations to dissociate the ribosomes did not appear to cause any major loss of proteins from the ribosomes except for some acidic proteins.


Inhibition of phosphatidylethanolamine biosynthesis in baby hamster kidney-21 cells infected with Semliki Forest virus.

Semliki Forest virus inhibits phosphatidylethanolamine biosynthesis in baby hamster kidney-21 cells 6 h after infection. Viral infection reduced the incorporation of [1,2-14C]-ethanolamine into intact cells by approximately 50%. A similar reduction in the activity of the ethanolaminephosphotransferase (EC 2.7.8.1) was also observed. The apparent Km for CDPethanolamine was 60 muM for the microsomal enzymes from infected or mock-infected cells. In addition, exogenous diglyceride only stimulated by 1.5-fold the ethanolaminephosphotransferase from virus- or mock-infected cells, whereas the same diglyceride preparations stimulated the cholinephosphotransferase (EC 2.7.8.2) from baby hamster kidney cells by sixfold. Generation of endogenous diglyceride by pretreatment of the microsomes with phospholipase C (EC 3.1.4.3) stimulated the activity of the cholinephosphotransferase but not the ethanolaminephosphotranferase. Semliki Forest virus does not inhibit all microsomal enzymes, since the activities of NADH- K3Fe(CN)6 reductase and NADH dehydrogenase (EC 1.6.99.3) were not affected. The ethanolaminephosphotransferase from virus- and mock-infected cells showed similar profiles of activity as a function of temperature; this result and other studies suggest that that membranous environment of the ethanolaminephosphotransferase was not significantly modified by the virus.


Chemodectoma of the larynx.

The ninettenth reported case of chemodectoma of the larynx is presented and its management discussed. Some reflections on the laryngeal paraganglia are made and the past literature is reviewed.


Chemodectomas of the head and neck.

Chemodectomas of the head and neck are reviewed. Their clinical picture, frequent multicentric origin, microscopie and macroscopic picture are discussed as well as their roentgenologic appearance. The role of angiography in their diagnosis is emphasized.


Infectious pancreatic necrosis (IPN) virus carriers and antibody production in a population of rainbow trout (Salmo gairdneri).

A total of three hundred and fifty-seven 2-year-old rainbow trout from a hatchery were examined for IPN virus over a period of 5 months. The overall virus isolation rate was 23.2%. The kidneys yielded virus most frequently while the spleen and pancreas-caeca yielded virus less frequently, and the gonads and feces yielded virus infrequently. During this period there was a decline in carrier rate as well as a decline in the numbers of infectious virions in the organs. These decreases were accompanied by elevated levels of neutralizing antibody in the sera. Fish with higher antibody levels were found to lose their IPN carrier status more frequently than those with lower antibody levels.


Cytoplasmic inclusions in canine cells infected with infectious canine laryngotracheitis (ICL) adenovirus.

Spherical dark inclusions were observed in both the cytoplasm and nuclei of infectious canine laryngotracheitis (ICL) adenovirus infected MDCK (Madin-Darby Canine Kidney) cells. The distribution of these inclusions in the cells appeared to indicate that they were formed in both the cytoplasm and in the nucleus at about the same time and there did not appear to be movement of these inclusions between the cytoplasm and the nucleus during the early stages of infection. Morphological appearance, 3H-leucine autoradiography, and immunoferritin labelling showed that the cytoplasmic inclusions were similar in nature to the dark nuclear inclusions, and contained the adenovirus hexon antigen, but not the penton base and fiber antigens.


Multiple neoplasms in two siblings with a variant form of Fanconi's anemia.

Two siblings with a variant form of Fanconi's anemia developed multiple neoplasms after prolonged survival and treatment with androgens. One of the siblings developed two separate oral squamous cell carcinomata, and the other developed acute leukemia and hepatoma. Androgens may have had a carcinogenic role in the appearance of the hepatic neoplasm. There is an increased incidence of neoplasm associated with Fanconi's anemia. This may be related to frequent spontaneous chromosomal abberations and/or to increased cellular susceptibility to viral transformation.


Small cell epidermoid carcinoma of the esophagus. An oat-cell-like carcinoma.

A case of small cell anaplastic epidermoid carcinoma of the esophagus is reported. The morphology of this rare esophageal neoplasm simulates that of oat cell carcinoma. Four similar cases have been previously reported. The finding of epidermoid differentiation in two of these tumors, including the present one, raises considerable doubt that any are truly oat cell carcinomas of the esophagus. Although it is theoretically possible that an oat cell carcinoma may arise in the esophagus, there is as yet no conclusive evidence for the existence of such an entity.


Hypercalcemia in small cell (oat cell) carcinoma of the lung. Coincident parathyroid adenoma in one case.

Hypercalcemia is very uncommon in small cell (oat cell) carcinoma of the lung. Two cases of this neoplasm associated with symptomatic hypercalcemia are described. Despite normal skeletal roentgenograms, metastatic bone disease was demonstrated by abnormal bone scans and bone biopsies in both patients. The combination of conventional antihypercalcemia therapy, cytotoxic cancer chemotherapy, and synthetic salmon calcitonin corrected the hypercalcemia despite progression of the small cell carcinoma. One patient with elevated serum immunoreactive parathyroid hormone (PTH) had a parathyroid adenoma at autopsy. This association emphasizes that in cases of bronchogenic small cell carcinoma with hypercalcemia, conincidental primary hyperparathyroidism should be considered.


Extramammary Paget's disease of the external ear canal in association with ceruminous gland carcinoma. A case report.

A case of extramammary Paget's disease of the external ear canal in association with ceruminous gland carcinoma is presented. Extramammary Paget's disease is not uncommonly seen in regions where apocrine glands are prevalent, with or without underlying apocrine gland carcinoma. This is the first case, to our knowledge, of this association in the external ear canal.


Mucinous adenocarcinomas of nose and paranasal sinuses.

This study concerns the Clinicopathologic findings for 18 patients with mucinous adenocarcinomas of nose and/or paranasal sinuses. Males in the 5th decade of life predominated in the series. Nasal obstruction, a growing mass in a sinus, or epistaxis were the most frequent complaints. Ten patients had tumors in the maxillary antrum, and the nasal cavity was the site in 5 patients. Histopathologically, the tumors were moderately to well differentiated, with a few poorly differentiated types. Tumor with the solid pattern of growth were anaplastic; these patients had poorer prognoses. For most patients, treatment consisted of radical surgery alone or in combination with radiotherapy. Of 13 patients for whom survival could be adequately evaluated, 7 died from the tumors, 5 are alive and free of disease more than 4 years, and 1 is living with recurrent tumor 14 months after diagnosis.


Wilms' tumor--histologic variation and prognosis.

A histologic analysis of 75 consecutive cases of Wilms' tumor in the Manchester (England) hospital region has been carried out. A classification based on the tubular status is described. A comparison of the histologic findings with the clinical staging and survival was undertaken, and it was found that the histologic classification was significantly correlated with both stage and survival.


The value of isolation procedures for cytomegalovirus infections in children with leukemia.

Standard contagious isolation procedures were used for 83 patients with acute lymphocytic leukemia and serologic and/or cultural evidence of cytomegalovirus infection. The infection rate, as determined serologically, for 9 months before and 13 months during isolation procedures was not decreased. Since the techniques employed were not helpful in preventing infection with cytomegalovirus in the immunosuppressed host, they have been discontinued at this hospital.


Actinomycin D as the primary agent for gestational trophoblastic disease.

Thirty-one patients with nonmetastatic trophoblastic disease (NMTD) and 39 patients with metastatic trophoblastic disease (MTD) of gestational origin were treated primarily with actinomycin D. Complete and sustained remission was achieved with actinomycin D alone in 94% of patients with NMTD and 67% with MTD. In the nonmetastatic group, 93% of patients without choriocarcinoma (non-CCA) achieved remission with actinomycin D, as compared to 100% (only 3 patients) with CCA. in the metastatic group, 76% of patients without CCA achieved remission with actinomycin D, as compared to 56% where CCA was present. Fourteen of the 15 patients who failed to respond completely to actinomycin D alone subsequently responded to methotrexate (10 patients), triple therapy (3 patients), methotrexate plus triple therapy (1 patient), and methotrexate plus vinblastine (1 patient). One patient died with widespread metastases despite intensive chemotherapy with actinomycin D and triple therapy. No serious toxic side effects were encountered even in treated patients with pre-existing laboratory evidence of impaired hepatic function.


Combination chemotherapy in oat cell carcinoma of the lung.

Twenty-eight patients with oat cell carcinoma of the lung were treated with cyclophosphamide, vincristine, procarbazine, and prednisone with and without irradiation. Median survival was significantly increased in this treatment group (10 months) as compared to a retrospective radiation alone series (5 months). Intensive chemotherapy with this four-drug regimen represents an initial step in effective management of this disorder.


Preoperative irradiation of cancer of the lung: final report of a therapeutic trial. A collaborative study.

Between May, 1963 and December, 1966, 17 medical centers cooperated in two separate but integrated therapeutic trials of primary lung cancer. One study was of patients with lesions considered operable at the time of diagnosis, and the other of patients with initially inoperable cancer but who were considered potentially operable after radiotherapy. Patients operable at the time of diagnosis were randomly assigned to receive either immediate surgery (278 patients) or preoperative radiotherapy followed by surgery (290 patients). All but one were followed until death or 5 years survival. Survival to each anniversary after randomization was almost identical for the two groups. At 5 years the survival rate was 14% after preoperative radiotherapy and 16% after immediate surgery. On the basis of the small standard error of the difference between these survival rates, a large advantage or a large disadvantage for preoperative radiotherapy is unlikely. Recurrence of cancer either locally or as distant metastasis was also similar in the two groups. Postoperative mortality was estimated to be 11% in the immediate surgery group, but cannot be estimated in a comparable fashion for the irradiated group. Certain postoperative complications were more frequent in the irradiated group, but survival during the first was not affected. Out of 425 patients initially considered to be inoperable, 152 were considered resectable after radiotherapy. These patients were randomly assigned to have either a thoracotomy and resection of their cancer if possible (78 patients) or no surgery (74 patients). Survival to each anniversary after randomization was very similar. After 5 years the survival rate was 8% for the group assigned to surgery and 6% for the group assigned to no surgery. The difference has a standard error of 4%.


Fractionation of nuclei and analysis of nuclear proteins of rat liver and Morris hepatoma 7777.

The contributions of nuclear populations to the total profile of nuclear proteins in a tissue were examined in normal rat liver and Morris hepatoma 7777. Comparison by sodium dodecyl sulfate polyacrylamide gel electrophoresis of phenol-soluble nuclear proteins from tumor and control liver revealed additional proteins of molecular weight 60,000, 100,00, and 135,000 and the loss of proteins of about 45,000 and 55,000 in the tumor. Subfractionation of liver nuclei on a 30 to 50% sucrose gradient yielded three nuclear classes with nearly identical complements of the phenol-soluble proteins. Similar fractionation performed on the hepatoma nuclei also produced three nuclear populations. In the hepatoma nuclei, several differences in the phenol-soluble proteins were found between the minor, slowly sedimenting nuclear fraction, and the two major fractions, while the two latter fractions were very similar in their protein composition. Histones derived from both tissues were also compared electrophoretically, indicating a decrease in the concentration of histone H1(0)in all nuclear classes derived from the tumor.


Interaction of normal and tumor transfer RNA methyltransferases with ethionine-induced methyl-deficient rat liver transfer RNA.

The tRNA methyltransferases from normal rat liver and Novikoff hepatoma have been compared with respect to their base specificity, capacity to methylate, and reaction kinetics, using mixed Escherichia coli B transfer RNA (tRNA) and ethionine-induced partially methyl-deficient rat liver tRNA. The pattern of base methylation of the two substrates is different with the use of enzymes from either source. In particular, N1-methylguanine methylation is much greater in the methyl-deficient rat liver tRNA. The enzymes from the two sources also show differences in specificity of base methylation in either substrate, particularly in the percentage of N2-methylguanine synthesized. The Novikoff hepatoma enzymes have a greater capacity for methylation with either type of tRNA than do rat liver enzymes. The methyl-deficient rat liver tRNA is a poorer substrate for the enzymes from both sources than is E. coli B tRNA in terms of rate of methylation as well as total acceptance of methyl groups. The affinity constants are somewhat higher for the methyl-deficient rat liver tRNA than for E. coli B tRNA. The Novikoff hepatoma enzymes, in general, have larger affinity constants than the rat liver enzymes. Maximal velocities for the various base-specific enzymes are lower with the methyl-deficient rat liver tRNA, with the exception of the 1-methylguanine specific enzymes. These enzymes from either rat liver or Novikoff hepatoma exhibit approximately a 2.5-fold greater maximal velocity with methyl-deficient rat liver tRNA.


The inhibition of ribosomal RNA synthesis and maturation in Novikoff hepatoma cells by 5-fluorouridine.

The inhibition of ribosomal RNA (rRNA) maturation by 5-fluorouridine (FUrd) in Novikoff hepatoma cells appears to depend upon the incorporation of the analog into the 45 S rRNA precursor. Precursor synthesized in the presence of FUrd is not processed into mature rRNA, but precursor synthesized in the absence of the analog is processed normally after the addition of the drug. The effect of FUrd on rRNA maturation is concentration dependent. At a concentration of 1 X 10(-4)M, the analog completely inhibits the formation of mature 18 S and 28 S rRNA; while at a concentration of 1 X 10(-7)M, the analog has no significant effect on rRNA maturation. These results suggest that some minimum degree of analog substitution is necessary to inhibit the maturation process. In addition to its inhibition of maturation, FUrd also inhibits the transcription of 45 S rRNA precursor. However, this effect of the drug is less complete and more time dependent that the effect on maturation. The inhibition of rRNA maturation by FUrd persists after removal of the analog from the culture medium. Cells that had been exposed to FUrd for 2 hr were unable to process 45 S rRNA precursor 20 hr after removal of the drug from the medium.


Virus-like particles in cystic mammary adenoma of a snow leopard.

Virus-like particles were observed in the giant cells of a mammary adenoma of a snow leopard kept in captivity. Particles that measured 115 to 125 nm in diameter budded from the lamella of endoplasmic reticulum and were studded on their inner surfaces with dense granules (approximately 12 nm) that gave them their unique ultrastructural morphology. Such particles were not observed extracellularly. Type B or type C particles were not seen in the tumor tissue.


Diminished binding of thyroid-stimulating hormone in a transplantable rat thyroid tumor as a possible cause of hormone unresponsiveness.

The adenylate cyclase activity and the binding of 125I-labeled thyroid-stimulating hormone (TSH) of normal and tumor rat thyroid plasma membranes were compared. No significant difference in the basal and fluoride-sensitive adenylate cyclase activity between normal and tumor plasma membranes was observed. Thyroid plasma membranes responded to TSH, whereas the enzyme from the tumor plasma membranes was TSH insensitive. Thyroid plasma membranes boud 125I-TSH. Tumor plasma membranes bound 125I-TSH poorly. At the highest concentration of unlabeled TSH used, 80% of the 125I-TSH that was bound to thyroid plasma membranes was displaced, whereas only 10% of the 125I-TSH bound to tumor plasma membranes was displaced. Therefore, it seems likely that the failure of this tumor to respond to TSH is due to an alteration in the functional unit of membrane adenylate cyclase at the level of the receptor subunit.


The tissue culture and morphology of human breast tumor cell line BOT-2.

A continuous human breast tumor cell line (BOT-2) was derived from an infiltrating duct carcinoma. The tumor cell line was grown as a monolayer in flasks, but the cells could be readily adapted to growth in roller cultures. These studies indicate that BOT-2 cells have a 16- to 18-hr doubling time and a modal chromosomal number of 63. The original BOT-2 cell culture has been in continuous cultivation for almost 2 years and has been passed 137 times. The BOT-2 cell line has been differentiated from HeLa cells by isoenzyme studies and chromosomal analysis.


Some biochemical characteristics of rat liver and Morris hepatoma nuclei and nuclear membranes.

Nuclei prepared from host liver and from Morris hepatomas 7777 and 7800 have been compared with respect to some of their biochemical characteristics. Several criteria were used to ensure that liver and hepatoma nuclei were of equal purity. These criteria include equal specific activity ratios (homogenate nuclei) for several marker enzymes. Phospholipids, proteins, and sialic acid content were compared in liver and hepatoma sucrose nuclei and in membrane and chromatin fractions obtained from liver or hepatoma nuclei. As determined by sodium dodecyl sulfate polyacrylamide electrophoresis, the only qualitative difference in protein that could detected was in 2 of the 4 nuclear fractions. There was an extra band in each of the 2 hepatoma fractions. Sialic acid was increased in hepatoma nuclei. In addition, a fraction containing most of the inner nuclear membrane from liver nuclei had no sialic acid, whereas the equivalent hepatoma fraction did have sialic acid. Total phospholipids were increased in hepatoma nuclei. This increased phospholipid concentration in hepatoma nuclei as compared to liver nuclei was apparent with sucrose nuclei, citric acid nuclei, membrane-denuded nuclei, chromatin, and nuclear fractions. Determination of the percentages of individual phospholipids making up the total phospholipids extracted revealed that the only significant change in the phospholipid composition of hepatoma nuclei was an increase in sphingomyelin. A large amount of this sphingomyelin was found to be associated with chromatin. The possible significance of chromatin-associated phospholipids is discussed.


Adhesion of malignant and nonmalignant cells to cultured embryonic substrates.

To study adhesion, which is probably the initial step in malignant invasion, we associated tissue culture fragments with living substrates in vitro. Malignant HeLa, hepatoma, and PY cells, as well as nonmalignant BHK cells, were transplanted into cultured chick blastoderms and organ fragments from chick embryos. Adhesion was evaluated by time-lapse cinematography, by flushing with Tyrode's solution, and by histological examination after fixation. It was shown that the adhesion of these tissue culture fragments depends on the nature of the substrate. Substrates of connective tissue, mesenchyme, and the basal side of epithelia proved to be adhesive. In contrast, the apical side of intact epithelia was nonadhesive. Perforated epithelia allowed adhesion at the site of the perforation. In the presence of dilysine, HeLa cells adhere to the apical side of epithelia and to the dorsal side of the upper layer of the blastoderm. We concluded that the apical side of intact epithelia constitutes an inappropriate substrate for adhesion of a large variety of cells, in vitro as well as in vivo. Alteration of this characteristic in the presence of dilysine indicates that long-range electrostatic repulsion might be responsible for the nonadhesive character of the epithelia.


Pathological review of breast lesions analyzed for estrogen receptor protein.

This report provides a detailed pathological review of 333 specimens analyzed for estrogen receptor protein (ERP) and correlates a series of morphological features with ERP results. Included were 147 primary breast carcinomas, 78 metastases, 27 fibroadenomas, and 81 nonneoplastic tissues, all from women. ERP in cytosols was assayed by incubation with [3H]estradiol in the presence and absence of "cold" estradiol followed by dextran-charcoal treatment. Results were summarized as positive (greater 60% inhibition by nontritiated estradiol, greater than 10 fmoles/mg protein), negative (less than 60% inhibition by nontritiated estradiol, less than 10 fmoles/mg protein), or intermediate borderline combinations. ERP in primary tumors ranged from 0.2 to 358 fmoles/mg protein (54.4% positive, 35.4% negative, 10.2% borderline). New findings are: (a) a high frequency of positive ERP in invasive lobular carcinoma (12 of 13, 92.3%) compared to typical ductal tumors (64 of 117, 54.7%); and (b) low frequency of positive ERP(5 of 21, 23.8%) in tumors with a prominent local lymphocyte reaction. Three ERP-positive noncarcinomatous specimens were fibroadenomas of high epithelial cellularity from patients under 30 years. No statistically significant relationship existed between ERP and any other morphological features that were examined.


Acute myocardial infarction in a young female with familial hyperbetalipoproteinemia.

A young woman, 21 years of age, suffered an acute transmural inferior wall myocardial infarction. EKG changes and serum enzymes accompanied a typical history. Coronary angiography perfomed 2 months after the acute episode showed complete occlusion of the right coronary artery and some mild disease of the circumflex and left anterior descending branches. Family history of type II hyperlipoproteinemia was present and she was known to have cholesterol since she was 12 years of age.


The phosphorylation of nuclear proteins in the regenerating and premalignant rat liver and its significance for cell proliferation.

In liver regeneration or neoplastic transformation, phosphorylation of nuclear proteins is stimulated. In the regenerating liver all main histone fractions are involved in this process. The type of histone phosphorylated seems to be dependent on the position of the partially synchronized cells within the generation cycle. At a time when most cells are exhibiting maximum HnRNA-synthesis, histone F2a2 belongs to those fractions with highly stimulated phosphate incorporation. Phosphorylation of this fraction alone is stimulated by cyclic AMP in parallel to a stimulation of HnRNA-synthesis. The preneoplastic liver is characterized by oscillating phosphorylation and dephosphorylation reactions of nearly all histone fractions during the first days of N-nitroso-diethylamine administration. After 2 months of carcinogen feeding a 50-150% stimulation of the phosphorylation of Fl subfractions is observed. The phosphate content of the other histones, however, has returned to the original level. A series of further proteins, isolated together with the histones, show very similar phosphorylation characteristics. These proteins are mostly of non-histone origin. It is suggested that some of them are responsible for the transport of RNA with messenger properties within the cell.


Nuclear secretory particles associated with the calyx cells of the ichneumonid parasitoid Campoletis sonorensis (Cameron).

The present study is an ultrastructural investigation of the calyx region of the ichneumonid endoparasitoid Campoletis sonorensis. It appears that synthesis of electron-dense secretory particles occurs within nuclei of calyx cells. The particles consist of an ovocylindrical electron-dense inner core and a surrounding unit membrane. After their formation the particles pass from the nucleus by budding through both membranes of the nuclear envelope. The particles, along with fully developed parasitoid eggs concentrate within the lateral oviduct lumen. Feulgen histochemical studies suggest the presence of DNA within the calyxfluid. The possible function of the particles is discussed.


Fine structure of the subradular organ of Lepidochitona cinereus (L), (Mollusca, Polyplacophora).

Electron microscopy of the subradular organ of the chiton Lepidochitona cinereus (L) reveals at least three cell types, microvillous, ciliated and mucus-secreting, situated in a single epithelium. The base of the epithelium is abundantly innervated and supplied with muscle cells. The fine structure is consistent with a chemosensory function for the subradular organ.


Amphibian sympathetic ganglia in tissue culture.

1. A culture medium has been developed for amphibian sympathetic nervous tissue but it is suggested that the ionic values should be adjusted to correspond to the concentrations of salts in the plasma of particular species. 2. The morphology, monoamine fluorescence, growth and differentiation of sympathetic ganglia of the frog, Limnodynastes dumerili, have been studied in culture. 3. Two types of neuron could be distinguished largely according to size, namely small, 18 X 20 mum and large, 38 X 42 mum. The possibility that these represent one type at different stages in development or represent functionally distinct neurons is discussed. 4. The sympathetic neurons are extremely sensitive to nerve growth factor (NGF) which caused an increase in the size of the cell bodies, the number of nerve fibres regenerating, the rate of axonal growth and synthesis of catecholamines. 5. Various other cell types appearing in the cultures have been described, including chromaffin, satellite, Schwann, multipolar and epithelial cells as well as fibroblasts, melanocytes and macrophages. The epithelial cells show slow contractions and changes in shape.


Synaptic morphology of the carotid body of the domestic fowl.

Efferent and reciprocal synapses have been demonstrated in the carotid body of the domestic fowl (Gallus gallus domesticus). Synapses were also found with purely afferent morphology, but were probably components of reciprocal synapses. The general morphology of the endings suggested the presence of two types of axon, afferent axons making reciprocal and perhaps afferent synapses with Type I cells, and efferent axons making efferent synapses with Type I cells. A few axo-dendritic synapses were also found. The dense-cored vesicles associated with the afferent components of reciprocal synapses and with the possible true afferent synapses varied in diameter and core but could belong to one population of pre-synaptic vesicles. These observations are consistent wtih a new theory for the carotid body receptor mechanism. This proposes a spontaneously discharging afferent axon inhibited by an inhibitory transmitter substance released by the Type I cell via the "afferent" component of its reciprocal synapse, the "efferent" component inhibiting this release. Besides this chemoreceptor modulation of its afferent axon, the Type I cell may also have a general secretory function.


The ultrastructure, catecholamine, and prolactin contents of the rostral pars distalis of the fish Mugil platanus after reserpine or 6-hydroxydopamine administration.

The rostral pars distalis (RPD) of the teleost Mugil plantanus from animals pretreated with reserpine or 6-hydroxydopamine (6-HODA) were assayed for dopamine (DA) or noradrenaline (NA) or for prolactin hormone. Such determinations were coupled with electron microscopy. It was found that reserpine and 6-HODA produced a significant decrease in the content of DA, NA, and prolactin. Electron microscope studies revealed that prolactin cells became activated as judged by ultrastructural criteria. After 6-HODA treatment type "B" neurosecretory fibers entering the RPD became selectively destroyed. These observations lead us to suggest that prolactin secretion is under inhibitory control by type "B" neurosecretory fibers of adrenergic nature.


Nerve-ending specializations in the central ganglia of Planorbis corneus.

The neuropile in the central ganglia of Planorbis corneus was studied by electron microscopy, using different fixatives. Several types of specialization were observed in the membranes of adjacent axonal processes, varying from only slight membrane thickenings to structures resembling desmosomes. Specialized structures similar to the 'spinule complexes' in mammalian brain are widely distributed in certain areas of neuropile. They consist of a small projection which invaginates a neighbouring neurone; the membranes are closely apposed and there is associated electron-dense material. The problem of identifying chemical synapses in this tissue is discussed. Many axonal processes are varicose, and it is suggested that varicosities may also be sites of transmitter release.


Ultrastructural and pharmacological studies on the afferent synapse of lateral-line sensory cells of the African clawed toad, Xenopus laevis.

Lateral-line sensory cells of Xenopus show considerable similarity to central nervous system synapses in their responses to the zinc iodide-osmium tetroxide, and bismuth-iodide cytochemical staining techniques. Both reserpine and FLA-63 cause a reduction in the diameter and electron density of the synaptic bars, and a dramatic loss of synaptic vesicles surrounding the bars. Aminooxy acetic acid and 3-mercaptopropionic acid cause no significant changes in the sensory-cell afferent synaptic apparatus. The results are discussed in the light of the known effects of the drugs. It is concluded that a monoamine rather than an amino acid may act as the afferent transmitter.


Ganglion cells and paraganglionic cells in the developing superior cervical ganglion of normal and p-chlorophenylalanine treated rats.

Ganglion cells and paraganglionic (PG) cells in the developing rat superior cervical ganglion were studied following postnatal treatment with p-chlorophenylalanine (pCPA) for 5 to 8 days. Litter mates, injected with saline solution, served as controls. Ganglion cells of control animals were differentiated ultrastructurally according to L. Eränkö (1972a) into late sympathicoblasts and young sympathetic nerve cells. In both maturation stages treatment with pCPA caused marked swelling of mitochondria, concomitant with minor changes of other cell organelles. Parallel to the ultrastructural alterations, fluorescence microscopy and cytophotometry revealed a slight diminution of diffuse fluorescence intensity in sympathetic neurons as the expression of a mainly extragranular amine depletion. In distinction from ganglion blocking agents the alterations are regarded as a general toxic effect of pCPA upon maturing sympathetic neurons, which secondarily influences catecholamine storage sites. Following treatment with pCPA, in PG-cells an alteration of mitochondria was scarcely to recognize. Specific granules were distinctly decreased in number, in some cases to an almost complete degree. Concordant to ultrastructural observations a marked diminution of fluorescence intensity was demonstrable in SIF-cells. In addition in these elements the fluorescence spectrum shifted towards the green field. Fluorescence cytophotometric evaluations confirmed the optical impression. Provided, that PG-cells, demonstrated with electron microscopy, are identical with SIF-cells in fluorescence microscopy, the results are discussed on the basis of a specific decrease of primary catecholamines due to an enzyme inhibition involved in catecholamine synthesis.


Evidence for the existence of two types of cAMP binding sites in aggregating cells of Dictyostelium discoideum.

Inhibition by dithiothreitol of the cell-bound phosphodiesterase has allowed the measurement of cAMP binding to aggregation-competent Dictyostelium discoidium amebae. Two classes of binding sites were demonstrated: Type 1, of low affinity (Kd approximately 160 nM) and high capacity (Ro approximately 1 X 10(5) sites per cell); and Type 2, of high affinity (Kd approximately 9 nM) but low capacity (Ro approximately 1.5 X 10(4) sites per cell). Both sites are developmentally regulated and are expressed during aggregation. The specificities of both sites are consistent with the specificity observed in vivo for the chemotactic response.


Effect of proteases on activation of resting chick embryo fibroblasts and on cell surface proteins.

The relationship between activation of resting chick embryo fibroblasts by proteases and proteolytic alteration of the cell surface has been investigated. Five different proteases were examined: trypsin, collagenase, plasmin, alpha-chymotrypsin, and thrombin. All of these proteases, when added to the culture medium at concentrations of 0.08-2.2 mug/ml, stimulated deoxyglucose uptake and induced cell division. The absolute levels of stimulation depended on the specific protease. Activation ranged from a doubling in cell number in 24 hr for trypsin and thrombin down to a 47% increase in cell number for alpha-chymotrypsin. Except in the case of thrombin, the stimulatory effects of these proteases correlated with breakdown of Z, a protein which is the major chick surface protein as revealed by lactoperoxidase-catalyzed iodination and which disappears upon transformation. In the case of thrombin, stimulatory concentrations brought about no detectable loss of surface components. Thus loss of Z is not a necessary condition for activation of chick fibroblasts; it may be a sufficient condition for activation of part of the cell population.


Synthesis, surface deposition, and secretion of immunoglobulins by Abelson virus-transformed lymphosarcoma cell lines.

Three Abelson virus-transformed lymphoma cell lines were established in tissue culture and the immunoglobulin biosynthesis by these cell lines was studied. Two of the cell lines (ABLS-1 and ABLS-5) were found to synthesize monomeric IgM molecules which were deposited in the cell membrane, probably to serve as an antigen receptor. The third cell line (ABLS-8) was found to synthesize membrane-associated IgM as well as cellular IgG molecules. In addition, these cell lines were found to synthesize a protein of 35,000 molecular weight which is also membrane-associated and which has the capability to bind the immunoglobulin (MAID). It is speculated that this protein might play a role in adapting the receptor immunoglobulin molecule to the hydrophobic environment of the cell membrane. The kinetics of amino acid incorporation into immunoglobulins by these cell lines show that they produce immunoglobulins at a rate which is two orders of magnitude smaller than plasmacytoma cells (MOPC 104E). These results suggest that Abelson virus transforms thymus-independent lymphocytes in various stages of maturation and these lymphocytes might be of B cell origin. The T lymphoma (P1798) used as a control cell line was found occasionally to produce minute amounts of immunoglobulin.


The activation of phospholipase C from Clostridium Welchii by quinine: an absolute requirement for calcium ions.

Quinine activates the hydrolysis of phosphatidyl choline suspensions by phospholipase C (E.C. 3.1.4.3) obtained from Clostridium welchii. Low levels of calcium are an absolute requirement for this activation: Mg2+, Ba2+, Sr2+, and Zn2+ are ineffective. The induction period, or lag phase for this enzyme is dependent upon both calcium concentration and substrate interfacial surface area. At low concentrations (less then 50 muM) calcium ions affect the induction period but not the maximal rate of hydrolysis, whereas guinine predominantly affects the rate of hydrolysis by alterations in the surface charge carried by the substrate.


Phosphorus-31 studies on lecithin in ethanol solutions.

31P relaxation times of lecithin in ethanol solutions have been measured in dependence on temperature and water concentration. Trial calculations have been carried out on the assumption of a 2-site exchange model. The results suggest first, the relaxation behaviour is determined by various motional and exchange processes; second, at 29 MHz the dipole-dipole interaction between 31P and protons provides the dominant contribution; third, in general we are not concerned with the case of "extreme narrowing". Morover, there are no negligible intermolecular contributions to relaxation.


[Comparative study on the ultrastructure of the primary mammary bud of male and female embryos at the stage of sexual differentiation].

The observation of embryonic mammary glands in male and female mice under the electron microscope has shown on one hand that the cellular ultrastructure of the buds is characterized by embryonic tissue only slightly differentiated, and on the other that, under the influence of testicular hormone, the detachment of the anlage from the epidermis in fourteen-day old embryos results in an important modification in the relationship between the ectoderm and the mesenchyme at the level of the neck of the gland rudiment. The basal membrane is destroyed and numerous centers of necrosis can be seen starting from the dense bodies in the ectodermic cells of the ruptured zone.


[Lipid overload induced in myocardial cells by vegetable oils. Observation using electron microscopy].

Comparative studies were made to estimate the lipid overloading of myocardiac cells in rats fed by groups, for 3 and 6 days, one of the following dietary oils: peanut oil, maize oil, sunflower oi, soyabean oil, rapeseed-primor oil, rapeseed-canbra oil and rapeseed oil.


[Ultrastructure of the rat neurohypophysis following electric stimulation of the supraoptic nuclei].

By means of electrical stimulation of the supraoptic nuclei, in a stereotaxic apparatus, partial or complete degranulation of the neurohypophysis is obtained, with often a continued presence of large vesicles; the number of microvesicles is always increased.


[A strain of amaril virus isolated from Aedes africanus in the Ivory Coast].

The yellow fever virus has been isolated from a pool of Aedes africanus caught on men in a region situated between forest and savannah near Touba, in the Ivory Coast. This isolation, occurring at an interepidemic period, proves for the first time in West Africa, the part played by this mosquito in the transmission of the yellow fever virus.


Simplified Scatchard-plot assay for estrogen receptor in human breast tumor.

In this two-point Scatchard-plot assay, with which a test of competitive inhibition is combined, the sample is mechanically homogenized in a buffer containing dithiothreitol, ultracentrifuged to obtain a fat-free cytosol, the protein content of which is then adjusted, and free and bound labeled estradiol are separated with dextran-coated charcoal after overnight incubation. We tested the method for precision and reliability by assaying such cytosols from pregnant rabbit uteri before and after dilution with kidney cytosol, and by assaying several other target and nontarget animal and human tissues. The Scatchard plot data were more reliable than tests for percent inhibition of binding by a competitor (diethylstilbestrol). For a tumor tissue to be judged positive it must bind at least 8 fmol of estradiol per milligram of protein and have a Kd of 0.1 to 5 X 10(-10) mol/liter. Some non-target tissues showed less than 70% inhibition by 10(4)-fold concentrations (over labeled estradiol) of inhibitor. Of 19 breast-tumor specimens, seven were found to be positive.


Quantitation of estrogen receptors in human breast cancer by agar gel electrophoresis.

Samples from human primary and metastatic mammary carcinomas were examined for the presence of specific estrogen-binding proteins. Cytosols were incubated with [3H]estradiol-17beta with and without an excess of unlabelled estradiol or estrogen antagonist. After incubation, the mixtures were treated with dextran-charcoal to remove non-specifically bound steroid. Low-temperature agar gel electrophoresis was used to separate receptor- and non-receptor-bound steroid. Statistically significant differences were found between the tissue protein contents of primary mammary carcinomas and skin metastases. Estrogen receptor and tissue protein contents of the tumors seem to be related to the age of the patients. Objective remissions with endocrine therapy were obtained in patients with carcinomas which possessed a substantial concentration of estrogen receptors.


Application of a simple urinary 11-deoxy-17-oxosteroid assay to the study of adrenocortical function in a selected group of hirsute women.

Using a relatively simple 11-deoxy-17-oxosteroid assay a group of hirsute patients have been studied before and following the administration of ACTH. These patients were selected on the basis that each showed an exaggerated response of urinary testosterone excretion to ACTH stimulation. Fifteen of eighteen patients were found to show an exaggerated urinary 11-deoxy-17-oxosteroid response to ACTH stimulation in addition to the excessive urinary testosterone response, suggesting that dysfunction of the adrenal cortex might be a factor contributing to the hirsutism of these fifteen patients.


A simple method for the determination of oestrogen receptor concentrations in breast tumours and other tissues.

A simple method for the determination of oestrogen receptor activity in breast tumours and other tissues is described. By competition between radioactive and non-radioactive oestradiol-17beta for binding to a tissue extract prepared with centrifugation at low speed, the presence or absence of receptors is decided from the unmanipulated data. The concentration of receptors (P0) and dissociation constant of binding (Kd) are calculated by Scatchard analysis. In two relatively homogeneous tissues, assay precision was of the order of 16% for receptor concentration and 21-33% for the dissociation constant of binding: in heterogeneous breast tumors, much poorer precision can be expected. Sensitivity determined empirically is approximately 0.1 fmol/mg tissue. The method has been applied to the determination of receptor concentrations in human breast cancers, benign breast tumours, non-malignant breast, gynaecomastic breast, and in various tissues of the rat. Receptor activity was detected in 70% of the human breast cancers examined, and in lower amounts, in 21% of the benign human breast tissues. Activity was also detected in rat uterus and mammary gland, and in low amounts in 82% of the rat mammary tumours analysed. Dissociation constant of binding was generally of the order of 0.5 x 10(-10) M. The advantages of the method are discussed.


Plasma lipoprotein abnormalities in a case of primary high-density lipoprotein (HDL) deficiency.

A 53-year-old patient with primary HDL-deficiency is reported. About 2% of the normal concentration of alpha1 HDL was present in his plasma. The alpha1-high-density-lipoproteins separated into two fast-moving components in polyacrylamide gel electrophoresis. The Apo HDL contained both the main apolipoproteins, Apo A-I and Apo A-II, but in disproportionally reduced amounts, the concentration of Apo A-I being reduced about 360-fold, and that of Apo A-II about 14-fold. Concomitantly, the amount of the Apo C polypeptides in the HDL-fractions was decreased to about 5.5% and the activity of the enzyme lecithin cholesterol acyltransferase (EC 2.3.1.4.3) in plasma was found to be only 40% of normal. Apoprotein D was present in the LDL in association with Apo B, forming an abnormal, fast-moving LDL-complex. Apo A-I and Apo A-II were both of normal size as determined by SDS-PAGE, and reduction with thiols resulted in the shift of the M.W. of Apo A-II from 17,000 daltons to about 8,500 daltons. Both proteins were found in the same position as their normal counterparts in analytical isoelectric focusing. The most likely explanation for the multiple lipoprotein abnormalities seems to be that a defect in the regulation or structure of Apo A-I is the basis of the HDL-deficeincy.


Complement-induced mixed aggregation of T and B lymphocytes and myeloid cells and the effect of adjuvant.

Complement induces mixed aggregation of murine spleen cells bearing C3 receptors. In the present studies, we show with immunofluorescent techniques that these aggregates are formed by T and B lymphocytes and myeloid cells, B cells constituting the predominant component in normal mice. There are striking changes in the quantity and quality of the clumps after in vitro treatment with adjuvant, mostly due to an increase in macrophages and possibly in T cells. The possible meaning of these phenomena in T-cell-B-cell co-operation and in the mode of action of adjuvants is discussed.


Thymus transplantation. Reconstitution of cellular immunity in a four-year-old patient with T-cell deficiency.

The case is reported of a 4-year-old girl affected with recurrent infections; anaemia, thrombocytopenia, haemorrhages and hepatosplenomegaly. Immunological investigations revealed a defect in cellular immunity related to the thymus-dependent system, hypergammaglobulinaemia (especially of class IgE), and very high titres of antibodies against Epstein-Barr virus (EBV). After foetal thymus transplantation, correction of the immunological defect and significant clinical improvement were noted, as well as a decrease of IgE and EBV antibody titres.


Dermatoglyphics of congenital abnormalities without chromosomal aberrations. A review of the clinical applications.

This review outlines the dermatoglyphics of congenital abnormalities without chromosomal aberrations. When combined with other clinical features of a particular disease, dermatoglyphics can serve to strengthen a diagnostic impression and may be useful as a screening device to select individuals for additional diagnostic studies.


[Raised cAMP content of striated muscle in experimental chronic renal failure (author's transl)].

3', 5'-Adenosine monophosphate (cAMP) levels were determined in skeletal and heart muscle tissue of rats in chronic renal failure. Compared to normal animals no alteration in cAMP concentration was observed in heart muscle, whereas the cAMP levels in skeletal muscle were increased by 34%. This cAMP rise may be caused by the elevated plasma catecholamines, glucagon or parathyroid hormone levels in uraemia. The results suggest that the increased cAMP levels in skeletal muscle of rats in chronic renal failure contribute to the raised cAMP levels in the plasma.


[Plasma-aldosterone and renin activity in Bartter's syndrome (author's transl)].

The temporal changes of aldosterone concentration and renin activity in plasma under the influence of stimulating and inhibiting factors were tested in two patients with Bartter's syndrome. There was a normal response to potassium and sodium administration, spironolactone, angiotensin II, ACTH and dexamethasone. Changes in plasma-aldosterone concentration, as measured at frequent intervals during angiotensin II infusion, suggest differential regulation of aldosterone synthesis by angiotensin II. In one patient the symptoms improved with administration of glucocorticoid derivatives.


Nuclear binding capacity appears to limit the hepatic response to L-triiodothyronine (T3).

In vivo saturation studies of thyroidectomized, propyl thiouracil, and L-triiodothyronine treated animals indicate that the hepatic nuclear binding capacity is not significantly influenced by the thyroidal status of the tissues. The constancy of the nuclear binding capacity for L-triiodothyronine facilitated an analysis of the relationship between nuclear occupancy and hepatic response to an intravenous injection of a triiodothyronine pulse. The activity of the mitochondrial enzyme alpha-glycerophosphate dehydrogenase was used as an index of tissue response to the administered triiodothyronine. Results were compatible with the following model. Saturation of nuclear sites is rapidly followed by a maximal rate of enzyme synthesis. When the sites are saturated, the activity of alpha-glycerophosphate dehydrogenase appeared to be independent of the dose administered. Higher doses of triiodothyronine achieve greater effects simply by occupying nuclear sites for a longer period. The apparent ability of nuclear sites to constrain tissue response to hormone administration provides additional supporting data for the physiological relevance of these sites in the initiation of hormonal action.


Androgen nuclear exchange activity in rat testis.

Androgen nuclear exchange activity has been measured in the rat testis. Exchange activity was increased 2-10-fold by pretreatment of the tissue with testosterone and persisted for at least 21 days after hypophysectomy in mature animals. Competition for exchange activity using 3H-testosterone or 3H-dihydrotestosterone was significant for a 500-fold excess of testosterone, dihydrotestosterone, progesterone, and cyproterone acetate. The activity was found in tubules and in isolated germ cells from mature rat testes. These results suggest that the molecular mechanism of androgen action in the testis may be quite complex and may involve components both in the more mature germ cells as well.


Prolactin binding activity on the crop sacs of juvenile, mature, parent and prolactin-injected pigeons.

Specific prolactin (PRL) binding activity of lactoperoxidase catalyzed 125I-labeled ovine-PRL was determined in a membrane-rich particulate fraction of pigeon crop sacs. Levels of TSH, LH or FSH as high as 1000 ng each were unable to displace the 125I-o-PRL bound to 600 mug of crop sac microsomal protein, whereas competitive displacement was achieved with as little as 0.5 ng unlabeled PRL. Ovine GH exhibited some cross reactivity when incubated in amounts greater than 500 ng, but this could be accounted for by its stated PRL contamination. Specific PRL binding activities were determined in juvenile and mature pigeons with unstimulated crop sacs, and parent pigeons with 'crop milk' and mature birds injected with PRL for 4 days. Crop sacs from juvenile birds contained approximately twice as much binding activity as crop sacs from mature pigeons. Parent and PRL injected pigeons, each with proliferated crop sac epithelium, exhibited 4-5 times as much specific PRL binding as the non-proliferated crops from juvenile or mature birds. These results show that the pigeon crop sac contains specific binding sites for PRL, and that the crop sac response to PRL is associated with an increase in PRL binding activity.


Cyclic AMP in the rat ovary: effect of endogenous LH secretion.

Cyclic AMP levels increased in ovaries from normal cycling rats on the afternoon of proestrus at the same time as the plasma LH peak occurred. Plasma progesterone was also elevated at this time. These results complement the extensive in vitro data indicating that cyclic AMP mediates the action of LH on the ovary.


Hormone receptors, 5. Binding of glucagon and insulin to human circulating mononuclear cells in diabetes mellitus.

Preparations of human peripheral mononuclear leukocytes, containing over 85% lymphocytes, specifically bound glucagon and insulin. Binding of physiological concentrations of both hormones was seen to be substantially diminished when cells were derived from either adult-type diabetics or individuals with a unilateral genetic history of diabetes.


Some structural requirements for LH-RH actions.

LH-RH and analogues of LH-RH, [2-4] LH-RH, [Lys8] LH-RH, [Ala4] LH-RH and [3-10] LH-RH were tested in vitro for their ability to stimulate LH and FSH release, respectively. Furthermore, the influence of these peptides on binding of LH-RH to the anterior pituitary plasma membrane fractions of female rats was studied to obtain some information on the structural requirements for LH-RH actions. Significant increase of LH release above the control level was recorded for LH-RH, [Lys8] LH-RH and [Ala4] LH-RH. LH release in the presence of the tripeptide [2-4] LH-RH and the octapeptide [3-10] LH-RH was statistically indistinguishable from that of basal levels. Significant stimulation of FSH release was noted only in the presence of LH-RH and [Lys8] LH-RH. Changes in the binding characteristics of LH-RH to the pituitary plasma membrane in the presence of the LH-RH analogues suggest a proportionality between releasing ability and receptor affinity. Glu and His residues of the LH molecule were found to be critical for binding to the receptor. Residual binding ability of TRH to LH-RH receptor is consistent with current molecular models of LH-RH and TRH, respectively. Furthermore, the data presented suggest that the amino acid in position 8 of the LH-RH molecule is of importance for the three dimensional structure of LH-RH.


Sex steroids influence LH-RH-receptor interaction.

Intact male rats (I) and rats 30 days after orchidectomy (OR) were treated in vivo with a single dose of 1.5 or 0.5 mg testosterone propionate (TP), 22.5 or 7.5 mug estradiol benzoate (EB) and a mixture of TP plus EB, respectively. The release of LH and FSH from anterior pituitaries of steroid hormone pretreated animals induced by the administration of LH-RH was studied in vitro. LH-RH stimulated FSH release from pituitaries of I and OR was inhibited by higher doses of EB or a mixture of EB plus TP. On the other hand, EB and combination of EB plus TP stimulated LH release in I, whereas EB and TP inhibited LH-RH induced release of LH in OR. At the lower dose levels no inhibition of LH-RH stimulated FSH release from pituitaries of steroid pretreated I or OR animals occured. LH-RH induced LH release was only observed at thise dose level in I pretreated with TP and OR pretreated with EB. On the other hand, pretreatment of ovariectomized rats (OV) with sex steroids (5 mug 17 beta-estradiol, 3 mg progesterone, and mixture of 5 mug 17 beta-estradiol and 3 mg progesterone, resp., each injected through 6 days) resulted in restoration of LH-RH induced FSH release lost 30 days after ovariectomy. Furthermore, LH-RH induced release of LH and FSH from male rat pituitaries at different times after orc  dectomy exhibited a pattern from that recorded for ovariectomized animals. [125I] LH-RH binding assays with anterior pituitary plasma membrane fractions isolated from I revealed two binding sites with apparent affinity constants of 4.3 x 10(-9) M and 2.2 x 10(-8) M, respectively. The low affinity binding site is lost upon orchidectomy. Scatchard plot analyses of binding data obtained for LH-RH reaction with plasma membrane fractions isolated from anterior pituitaries of OR pretreated with sex steroids showed that the second binding site could be regenerated by treatment with the low dose of TP. From the present experiments it is concluded that: 1. LH-RH induced gonadotropin release is affected by sex steroids at the pituitary plasma membrane level. 2. One hypothalamic hormone may well specifically control LH and FSH release. 3. Mechanisms controlling LH-RH induced gonadotropin release are different in the male and female.


3,3',5'-Triiodothyronine (reverse T3) and 3,3',5-triiodothyronine (T3) in fetal and adult sheep: studies of metabolic clearance rates, production rates, serum binding, and thyroidal content relative to thyroxine.

To examine the mechanism(s) responsible for high serum concentration of 3,3',5'-triiodothyronine (reverse T3, rT3) and low serum concentration of 3,3',5-triiodothyronine (T3) in the fetus, we studied metabolic clearance rates (MCR) and production rates (PR) of rT3, T3, and thyroxine (T4) in adult nonpregnant sheep and sheep fetuses in utero. The mean fetal MCR-rT3 was significantly lower than that in adult sheep, and the mean fetal PR-rT3 significantly higher. The mean fetal MCR-T3 was higher than, and the mean fetal PR-T3 similar to that in adult sheep. The mean fetal MCR-T4 and PR-T4 were both significantly higher than the corresponding values in adult sheep. The ratios of mean PR-rT3 to PR-T4 (rT3/T4) were similar in fetal and adult sheep. However, the ratio of mean PR-T3 to PR-T4 (T3/T4) in the fetal sheep was much lower than that in the adult sheep. The low fetal MCR-rT3 was not attributable to high serum binding of rT3. On the basis of the thyroidal content and kinetics of iodothyronines, it was estimated that whereas thyroidal secretion may account for nearly all of serum T3 (or PR-T3) in the fetus and about 50% of serum T3 in adults, it accounts for only about 3% of the serum rT3 (or PR-rT3) in both fetal and adult sheep. The results suggest a) that elevated serum rT3 in the fetus is due to its decreased clearance and increased production by mono-deiodination of T4, and b) that low serum T3 in the fetus is due to its increased clearance and decreased production by mono-deiodination of T4. In addition, on the basis of discordant changes in the production of T3 and rT3 from T4, it appears that there may exist two separate, apparently specific, iodothyronine deiodinating activities--one cleaving the iodine atom at the 5'-position and the other acting in the iodine atom at the 5-position of the T4 molecule; 5'-iodothyronine deiodinating activity is apparently reduced in the fetus.


Thyroid-stimulating hormone regulation of ornithine decarboxylase activity in the thyroid.

TSH (1.0 U im) caused a 22-fold increase in thyroidal ornithine decarboxylase activity (ODC) 6 hours after administration in intact rats. Hypophysectomized rats treated with 1 U TSH showed a 5-fold increase in thyroid ODC activity. This stimulation appeared to be specific for TSH since hormones known to induce ODC activity in other target tissues, such as ACTH or LH, showed no significant stimulation. DIBUTYRYL CYCLIC AMP and aminopylline caused a 12-fold increase in ODC activity 5 hours after administration. Prostaglandins have also been implicated in the TSH-induced stimulation of cyclic AMP. Indomethacin (1.0 mg/100 g body wt, ip), an inhibitor of prostaglandin synthesis, was administered 3 hours before TSH with a resulting 30% diminution (P less than .001) in ODC activity compared with the administration of TSH alone. To rule out the possibility that the increase in ODC activity with TSH might be due to increased thyroid hormone secretion, ODC activity was evaluated 6 hours after triiodothyronine administration (60 mug/100 g body wt), and no significant increase in thyroid ODC activity was found. Stimulation of ODC activity was 90% inhibited by the intraperitoneal administration of actinomycin D (80 mug/100 g body wt) or cycloheximide (400 mug/100 g body wt) given simultaneously with TSH. These results indicated that TSH specifically stimulated thyroid ODC activity, which may be important for the growth-promoting action of the hormone on the thyroid gland. This action may be mediated by cAMP and prostaglandins and may require new protein synthesis.


The prosence of estrogen receptor in kidneys from normal and androgen-insensitive tfm/y mice.

In vitro and in vivo binding of [3H]estradiol to cytosol proteins and nuclei was studied in kidneys from normal and androgen-insensitive tfm/y mice. The use of tfm/y mice permitted study of the estrogen-receptor complex in the absence of androgen receptor. A high affinity, [3H]estradiol-labeled, 8S molecule was demonstrated in low salt sucrose gradients. This macromolecule sedimented more slowly in gradients containing KC1 (0.5 M). Binding of [3H]estradiol was inhibited by 100-fold excess estrone, estradiol, or diethylstillbestrol but was not affected by testosterone, androstenedione, or progesterone. Studies of equilibrium-binding kinetics for estradiol indicated a Kd of 1.4 X 10(-9) M and 4.4 X 10(-14) mol of binding sites/mg cytosol protein. Furthermore, the binder was an acidic, heat-labile protein (pI = 4.8) which could be precipitated from cytosol with 33% ammonium sulfate, and needed sulfhydryl groups for activity. The demonstration of an estradiol-binding protein in vitro was correlated with specific [3H]estradiol uptake by tfm/y kidney nuclei in vivo. We concluded that the mouse kidney contains an estradiol-binding protein, distinct from that for androgens, which has many of the characteristics of a steroid receptor. The presence of an estrogen receptor in both normal and tfm/y mice indicates that a genetic defect in one receptor does not influence the properties of another. We concluded that androgen and estrogen receptors are under independent genetic control.


Androgen induction of a specific uterine protein.

Studies were conducted to determine the ability of androgens in vitro to elicit the induction of a specific uterine protein (IP) normally attributed to estrogens. Both 5alpha-dihydrotestosterone (5alpha-DHT) and testosterone were effective in stimulating IP synthesis in rat uterine tissue in a concentration dependent manner (0.1 muM to 50 muM). 5alpha-DHT was more effective than testosterone and reached approximately 85% of the estradiol stimulated IP response at 10 muM and 50 muM; whereas testosterone was only able to achieve about a 70% IP response at 50 muM. This androgen stimulated IP synthesis was stereospecific since cis-testosterone and 5beta-DHT, inactive androgen isomers, were unable to evoke a detectable IP response at any concentration studied. Antiandrogens were unable to inhibit the 5alpha-DHT stimulated IP synthesis but antiestrogens were able to greatly inhibit the 5alpha-DHT and testosterone stimulated IP responses in a concentration dependent manner. The anti-estrogens, themselves, were very weak inducers of the IP response. The nuclear accumulation of the estrogen receptor by various androgens and inactive androgen isomers was also determined. Approximately 100% nuclear accumulation of receptor was attained with 1 muM 5alpha-DHT, whereas 50 muM testosterone was needed for 100% uptake. 5beta-DHT was unable to translocate the receptor at the lower concentrations tested, but caused a significant nuclear accumulation of 50 muM. Cis-testosterone was unable to cause the nuclear accumulation of the estrogen receptor at all concentrations studied. These studies suggest that some of the estrogen receptors accumulated in the nuclei by androgens, inactive androgen isomers, or antiestrogens may not be capable of eliciting a biological response.