Enzymatic methylation of carboxyl groups of chromaffin granule membrane proteins. Carboxyl groups of membrane and soluble proteins from bovine adrenal medulla chromaffin granules were enzymatically methylated. The methylated peptides were resolved using gel electrophoresis under acidic conditions in the presence of N-cetylpyridinium chloride. There was a selective methylation of two groups of membrane peptides which did not correspond to any of the chromaffin granule soluble proteins. Dopamine beta-hydroxylase, an acidic protein accounting for up to 25% of the membrane proteins, was a poor substrate for protein carboxylmethylase. The methyl esters of membrane proteins were more labile than those of the chromaffin granule soluble proteins. At all pH values tested, membrane protein-methyl esters were hydrolyzed three times more rapidly than the soluble protein-methyl esters.


Characterization of the low density lipoprotein receptor in membranes prepared from human fibroblasts. An ultracentrifugation assay has been developed to measure low density lipoprotein (LDL) receptor activity in membranes prepared from cultured human fibroblasts. The binding site for 125I-labeled LDL in isolated membranes reflected the properties of the LDL receptor previously demonstrated in intact fibroblasts. It exhibited high affinity (Kd approximately 4 microgram of LDL protein/ml), specificity (LDL approximately 400-fold more effective than high density lipoprotein in competing with 125I-LDL for the binding site), dependence on calcium, and susceptibility to destruction by pronase. The number of LDL receptors detected in the in vitro membrane binding assay was similar to the number detected in intact cells. The number of receptors was reduced in membranes from fibroblasts that were grown in the presence of 25-hydroxycholesterol plus cholesterol and in fibroblast membranes from a subject with homozygous familial hypercholesterolemia, two situations in which the number of LDL receptors in intact fibroblasts is known to be reduced. The availability of a membrane binding assay that faithfully reflects the properties of the physiologic LDL receptor of intact cells should permit the characterization of this receptor in organs from intact humans and animals.


Isolation and characterization of ornithine transcarbamylase from normal human liver. We report experiments describing the isolation and characterization of ornithine transcarbamylase from normal human liver. Our preparative procedure employs initial centrifugation and heat steps, intermediate batch-wise adsorption and desorption from ion exchange resins and column chromatographic elution from hydroxylapatite, and final purification by gel filtration chromatography and glycerol density gradient centrifugation. The enzyme, purified 580-fold in this way, is homogeneous as judged by native and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Human ornithine transcarbamylase has a molecular weight of 114,000 and is a trimer of identical 38,000 molecular weight subunits. It focuses at pH 6.8 as a single band on polyacrylamide gel, has a COOH-terminal phenylalanine, an NH2-terminal glycine, an apparent Km for L-ornithine of 0.4 mM and for carbamyl phosphate of 0.16 mM, and a pH optimum of 7.7. The enzyme is quite stable over a temperature range from -50 degrees to +60 degrees C and over the pH range from 5.8 to 8.2. The quaternary structure and amino acid composition of the human enzyme are very similar to those of its bovine homologue.


Changes in hepatic levels of tyrosine aminotransferase messenger RNA during induction by hydrocortisone. Messenger RNA specific for tyrosine aminotransferase was quantitated by microinjection into oocytes of Xenopus laevis. The heterologously translated enzyme was identified by specific immunoprecipitation and found to be identical with authentic aminotransferase by several criteria. The level of functional message present in rat liver increases during hydrocortisone induction, and this increase is directly proportional to the increased rate of synthesis of the enzyme. Kinetic analysis of the changes in tyrosine aminotransferase mRNA levels during induction and withdrawal indicates that the steroid does not affect the stability of the message, which has a half-life of approximately 1.2 h. Hydrocortisone, therefore, acts to increase the rate of synthesis of the specific messenger by stimulating either its transcription or processing to functional mRNA.


Magnesium-induced inner membrane aggregation in heart mitochondria. Mg(2+) at an optimal concentration of 2mM (ph 6.5) induces large increases (up to 30 percent) in the optical density of bovine heart mitochondria incubated under conditions of low ionic strength (< approx. 0.01). The increases are associated with aggregation (sticking together) of the inner membranes and are little affected by changes in the energy status of the mitochondria. Virtually all of a number of other polyvalent cations tested and Ag(+) induce increases in mitochondrial optical density similar to those induced by Mg(2+), their approximate order of concentration effectiveness in respect to Mg(2+) being: La(3+) > Pb(2+) = Cu(2+) > Cd(2+) > Zn(2+) > Ag(+) > Mn(2+) > Ca(2+) > Mg(2+). With the exception of Mg(2+), all of these cations appear to induce swelling of the mitochondria concomitant with inner membrane aggregation. The inhibitors of the adenine nucleotide transport reaction carboxyatratyloside and bongkrekic acid are capable of preventing and reversing Mg(2+)-induced aggregation at the same low concentration required for complete inhibition of phosphorylating respiration, suggesting that they inhibit the aggregation by binding to the adenine nucleotide carrier. The findings are interpreted to indicate (a) that the inner mitochondrial membrane is normally prevented from aggregating by virtue of its net negative outer surface change, (b) that the cations induce the membrane to aggregate by binding at its outer surface, decreasing the net negative charge, and (c) that carboxyatractyloside and bongkrekic acid inhibit the aggregation by binding to the outer surface of the membrane, increasing the net negative charge.


Polymerization of actin. IV. Role of Ca++ and H+ in the assembly of actin and in membrane fusion in the acrosomal reaction of echinoderm sperm. When Pisaster, Asterias, or Thyone sperm are treated with the ionophore A23187 or X537A, an acrosomal reaction similar but not identical to a normal acrosomal reaction is induced in all the sperm. Based upon the response of the sperm, the acrosomal reaction consists of a series of temporally related steps. These include the fusion of the acrosomal vacuole with the cell surface, the polymerization of the actin, the alignment of the actin filaments, an increase in volume, an increase in the limiting membrane, and changes in the shape of the nucleus. In this report, we have concentrated on the first two steps in this sequence. Although fusion of the acrosomal vacuole with the cell surface requires Ca++, we found that the polymerization of actin instead appears to be dependent upon an increase in intracellular pH. This conclusion was reached by applying to sperm A23187, X537A, or nigericin, ionophores which all carry H+ at high affinity, yet vary in their affinity for other cations. When sperm are suspended in isotonic NaCl, isotonic KCl, calcium-free seawater, or seawater, all at pH 8.0, and the ionophore is added, the actin polymerizes explosively and an efflux of H+ from the cell occurs. However, if the pH, of the external medium is maintained at 6.5, the presumed intracellular pH, no effect is observed. And, finally, if egg jelly is added to sperm (the natural stimulus for the acrosomal reaction) at pH 8.0, H+ is also released. On the basis of these observations and those presented in earlier papers in this series, we conclude that a rise in intracellular pH induces the actin to disassociate from its binding proteins. Now it can polymerize.


Polymerization of actin. V. A new organelle, the actomere, that initates the assembly of actin filaments in Thyone sperm. Between the acrosomal vacuole and the nucleus is a cup of amorphous material (profilactin) which is transformed into filaments during the acrosomal reaction. In the center of this cup in untreated Thyone sperm is a dense material which I refer to as the actomere; it is composed of 20-25 filaments embedded in a dense matrix. To visualize the substructure of the actomere, the profilactin around it must be removed. This is achieved either by demembranating the sperm with Triton X-100 and then raising the pH to 8.0, or by adding inophores to intact sperm at pH 8.0. Under these conditions, the actomere remains as a unit while the rest of the profilactin is solubilized or polymerized. When demembranated sperm are incubated under conditions in which the actin should polymerize, filaments grow from the end of the actomere: the actomere thus appears to behave as a nucleating body. This observation is strengthened by experiments in which untreated sperm are incubated in seawater or isotonic NaCl at pH 7.0 and the ionophore X537A is added; in this case, only a partial polymerization of the actin occurs and the acrosomal vacuole does not fuse with the cell surface. The actin filaments that do form, however, are attached to the apical end of the actomere. In fact, the elongating filaments push their way into and frequently through the acrosomal vacuole. Thus, it appears that the sperm organizes the actin filaments by controlling their nucleation. My model is that the cell controls the ammount of unbound actin such that it is slightly above the critical concentration for polymerization. Then, spontaneous nucleation is unfavored and polymerization would proceed from existing nuclei such as the actomer.


The development of lysosomal apparatus. I. Lysosomal enzyme activities in the liver of mice at perinatal stages and those of their mothers. The enzymatic activity of five acid hydrolases: acid phosphatase, arylsulfatase A, deoxyribonuclease, beta-glucuronidase, and cathepsin D, was assayed in fetal (fifteenth and eighteenth days of pregnancy) and neonatal (Days 0, 5, 10, and 15 post-partum) mouse liver. With the exception of cathepsin D, the activity increased around birth to levels varying according to the enzyme. Histochemical observations of other authors appear to justify, at least in part, the present results, which indicate that late days of fetal development and early neonatal life may constitute a transitional stage to full lysosomal enzyme functionality of the adult organ. The livers of the mothers were also assayed for the same enzymes. Each activity showed a peculiar pattern which was, in turn, different from that found in the liver of the litter for the same enzyme, probably as a cause of the metabolic requirement of the gland. The hypothesis that the lysosomes are heterogeneous in their enzyme composition is suggested by the variety of enzymatic patterns found in the liver of the litters and their mothers.


PH oscillations in cell suspensions of Dictyostelium discoideum: their relation to cyclic-amp signals. Cells of Dictyostelium discoideum known to release cyclic AMP (cAMP) rhythmically in the form of pulses, change with the same period of about 8 min the pH of their medium. The pH is used here as an indicator to investigate the effect of externally added cAMP pulses on the oscillations. Both a temporary increase in amplitude and a permanent phase shift can be induced. The phase-response curve indicates that the period can be increased and decreased by rhythmic stimulation with cAMP pulses.


Interpatient microbiological cross-contamination after dental radiographic examination. Pairs of patients were evaluated for microbiological cross-contamination after radiographic examination. In 30 of these pairs of patients there was the possibility of transference of S pyogenes, S aureus, or D pneumoniae. Such transference was observed in 23 (77%) of these 30 pairs of patients. The vectors for such transfer include the hands of the X-ray technician and the radiographic equipment. Further, it was found that each of these organisms would survive for at least 48 hours after being placed on an X-ray tube. Since interpatient microbiological cross-contamination can occur after routine radiographic examination, in some cases, disinfection of the radiographic equipment is indicated.


Tear pH: how predictable?

Tear pH: how predictable? While the tear pH of a patient is now known to be continually shifting throughout the waking day, such variations were found to be contained within fairly narrow limits (a range here of about 0.6 of a pH unit). Further, sufficient repetition of these shifts occurred over 10 days observed that patterns distinctive of the individual patient often emerged.


A novel immunoadsorbent: use for the preparation of monospecific antibodies to the hepatitis B antigen. Controlled pore glass (CPG) adsorbs hepatitis B surface antigen (HBsAg) from whole plasma with a high degree of specificity. The resultant complex is stable at acid pH and in the presence of high concentrations of sodium thiocyanate. The adsorbed HBsAg is qualitatively and quantitatively similar to the soluble material in its ability to bind antibodies to HBsAg (anti-HBs). The HBsAg in 1 ml of strongly reactive plasma is adsorbed by 100 mg of CPG, which can then specifically bind 32,000 passive hemagglutination units of anti-HBs. Bound antibody can be eluted in 77% yield by acid or by chaotropic ions and the CPG-HBsAg complex can be reused in further adsorption-elution cycles. Antibody to HBsAg can be purified 144-fold in a single step by using this technique. The preparation of monospecific subtyping reagents for HBsAg and of immunochemically purified anti-HBs is described.


Sphingomyelinase in pig and human epidermis. The enzyme sphingomyelinase (sphingomyelin phosphorylcholine phosphohydrolase E.C.3.1.4.12) which hydrolyzes sphingomyelin to ceramide (N-acylsphingosine) and phosphorylcholine was identified in the subcellular fractions of pig and human epidermis. The enzyme has an optimum pH of 4.5 to 5 and is activated by Triton X-100 (0.1% w/v). Approximately two-thirds of the enzyme activity in both the pig and human epidermal homogenates was in the soluble subcellular fraction and more than half of the enzyme activity in the subcellular particulate fraction was solubilized by freeze-thawing. The pH optimum suggests that epidermal sphingomyelinase is probably a lysozomal enzyme. The enzymes in both pig and human epidermis exhibited Michaelis-Menten kinetics. The soluble sphingomyelinase in pig epidermis had an apparent Km, 4.5 X 10(-5) M and that in human epidermis an apparent Km 7.7 X 10(-5) M. The pig epidermal sphingomyelinase had no special requirement for either divalent or heavy metal ions and was not inhibited by sulfydryl group-blocking agents but it was moderately inhibited by dithiothreitol. No evidence was found in either pig or human epidermis for the presence of a phospholipase C (E.C.3.1.4.3) which hydrolyzes phosphatidylcholine to diglyceride and phosphorylcholine but there was suggestive evidence of another catabolic pathway for phosphatidylcholine.


Collecting duct hydrogen ion secretion in the rabbit: role of potassium. The purpose of this study was to investigate distal nephron hydrogen ion secretion in the intact animal. The rabbit was chosen as the experimental model because it produces acid urine containing little ammonium. Upon replacement of the usual rabbit diet with milk, plus administration of an acid load (10 mEq/kg), the urine pH fell consistently from very alkaline values (PH greater than 7.4) to 4.8 +/- 0.2. Despite the ability to achieve high urine-to-blood hydrogen ion concentration gradients, the U-B PCO2, an index of collecting duct hydrogen ion secretion, was virtually zero. In these studies, the urine bicarbonate and buffer concentration were comparable to those observed in dog, rat, and man in which a high U-B PCO2 gradient was achieved. The rabbits studied had low plasma potassium concentrations (less than 3 mEq/L). Since potassium deficiency has been implicated in impaired urine acidification, potassium was administered, and it resulted in an increase in collecting duct hydrogen ion secretion as evidenced by a further fall in minimum urine pH during acidemia and a prompt rise in the U-B PCO2 during alkali administration. In summary, rabbits had a very low but not absent rate of collecting duct hydrogen ion secretion. Potassium administration increased the rate of hydrogen ion secretion in a segment of the collecting duct in which hydrogen ion secretion is reflected by an increase U-B PCO2.


The distribution of bumps in the tail of the locust photoreceptor afterpotential. An extended tail or prolonged depolarizing afterpotential (PDA) follows the receptor potential of a locust retinula cell when the stimulating light is in the intensity range that saturates the receptor potential. The amplitude and duration of this afterpotential depend on the intensity and duration of the stimulus. As the afterpotential decays, apparently exponentially, it becomes resolved into bumps, which we call light-induced dark bumps (LID bumps). The intervals between light-induced dark bumps are distributed in a way that is indistinguishable from a random (Poisson) distribution. As previously demonstrated, LID bumps are indistinguishable from bumps directly induced by low intensity light in light-adapted cells, which in turn grade into the slightly larger bumps produced, each by a single photon, in dark-adapted cells. The light-induced dark bumps continue for up to an hour in darkness, slowly becoming like dark-adapted bumps in amplitude and shape. To account for the random occurrence and discrete features of bumps after so long a latency, we propose that intense light generates a significant amount of an intermediate molecule or packet which decays slowly to start the same process that normally generates bumps with a short delay.


Some responses of the electric ray (Torpedo marmorata) to low ambient oxygen tensions. I. Blood samples were taken during prolonged hypoxia experiments in which the inspired water oxygen tension was less than 10 mmHg. The oxygen tension of the post-branchial blood was about 5 mmHg and its pH shows a significant lowering from normoxic levels. 2. The decrease in blood pH is correlated with increases in levels of lactate and pyruvate. The lactate/pyruvate ratio increases during hypoxia. 3. An increase in blood succinate was also found, and strongly suggests the accumulation of multiple anaerobic end-products within the tissues. 4. Recovery of normoxic levels of succinate takes place almost immediately following the restart of ventilation whereas the decrease in lactate concentration is slower. 5. It is concluded that these adaptations may be related to the habitat of the fish at low tide in pools where the Po2 may fall very markedly.


The identification of an octopaminergic neurone and the modulation of a myogenic rhythm in the locust. An octopaminergic neurone in an insect is demonstrated. This cell (DUMETi) is the dorsal unpaired median neurone which projects to the extensor tibiae muscle of the locust metathoracic leg. Its soma was physiologically identified, isolated and shown to contain about 0.1 pmol of octopamine. Octopamine is about four times more concentrated in the axon than in the soma. The concentration in the soma is at least 800 times more than that in the soma of an identified motoneurone (which controls the fast extensor of the tibia). The effects of DUMETi on a myogenic rhythm in the extensor muscle of the locust metathoracic leg can be mimicked by superfusion with low concentrations of octopamine. The myogenic bundle possesses at least two types of aminergic receptor: one which slows the rhythm (and has a high-affinity for octopamine) and a second which accelerates the rhythm (and has a low-affinity for octopamine but a high-affinity for the indolalkylamine, 5-hydroxytryptamine). The roles of the two receptor types in modulating the rhythm are discussed in relation to the function of the rhythm.


Transplantation of allogeneic bone marrow without graft-versus-host disease using total lymphoid irradiation. Bone marrow (BM) and skin allografts from C57BL/Ka (H-2b/b) mice were transplanted to BALB/c (H-2d/d) recipients treated with total lymphoid irradiation (TLI), whole-body irradiation (WBI), or fractionated thymic irradiation TLI prolonged skin allograft survival about five times as long as that in untreated controls, and allowed for permanent engraftment of BM cells in approximately equal to 90% of recipients. None of the BM recipients showed clinical signs of graft-versus-host disease (GVHD) (diarrhea, weight loss, hunched back, etc.). On the other hand, recipients given WBI and allogeneic BM cells developed severe clinical GVHD. The majority of the latter recipients died within 12 days after BM transplantation, and 95% died within 61 days. Although TLI protected BALB/c mice against GVHD induced by BM cells, all recipients given TLI and allogeneic spleen cells developed lethal GVHD. Thymic irradiation alone marginally prolonged skin allograft survival, and did not allow for allogeneic BM engraftment. These results suggest that TLI may be a useful regimen in clinical BM transplantation, since this form of radiotherapy is used extensively in humans and has few severe side effects.


Loss of proliferative capacity in immunohemopoietic stem cells caused by serial transplantation rather than aging. Marrow stem cell lines from old donors and those from young controls gave equally rapid rates of colony growth on spleens of irradiated mice. Old and young stem cell lines competed equally well with chromosomally marked marrow stem cells from a young donor in producing cell types that are stimulated by bleeding; old cells competed 70% as well as young in producing cell types stimulated by phytohemagglutinin (PHA) in vitro. After a single serial transplantation, the rates of colony growth declined 1.5- to 2.5-fold, and the ability to compete declined 2- to 4-fold for bleeding-stimulated and 4- to 10-fold for PHA-stimulated cells. Thus, immediate stem cell proliferative capacities decline much more after one serial transplantation than after a lifetime of normal function.


A comparative study of the electrode systems of three pH and blood gas apparatus. We present a comparative evaluation of the electrode systems of three modern blood gas analysers: IL-413, ABL-1 and AVL-937C. The response curves, accuracy and precision of the pH-, pCO2- and pO2-electrodes were established with tonometered blood and buffer solutions. pH values (range 6.8-7.8) measured on the AVL deviate (-0.03 pH for blood and +0.03 pH for buffer) from those of BMS2 Mk2; whereas on the IL and ABL analysers the pH values deviate by not more than 0.01 pH. The standard deviation was better than 0.005 pH. pCO2 values of blood and buffer (range 14-106 mm Hg) deviate from the calculated tonometer values by quantities ranging from 3 to 10 mm Hg. The average precision (CV)1) of the pCO2 measurement on each analyser was better than 1.8%. pO2 values of blood (range 0-130 mm Hg) did not differ by more than 3 mm Hg from the calculated values. Above 130 mm Hg a linear negative increasing difference was seen. For buffer solutions a linear relationship between pO2 difference and pO2 value was found over the whole range from zero up to 642 mm Hg: a positive difference below and a negative difference above the pO2 of the previous calibration; if the calibration pO2 is higher, the sample pO2 is shifted to a higher value. The average precision of the pO2 measurements was better than 3%. In the (patho)-physiological range the three instruments may provide suitable results for the clinician. Suggestions are made for standardization and improvement of the electrode systems.


Activation of rat pheochromocytoma tyrosine hydroxylase by a cyclic AMP-dependent protein kinase in a cell-free system. Short term exposure of PC-12 cells to dibutyryl cyclic AMP (dB-cAMP) results in an activation of tyrosine hydroxylase. In the cell-free system the PC-12 tyrosine hydroxylase activity is stimulated by addition of c-AMP, Mg+2 and ATP. Exogenous c-AMP dependent protein kinase further stumulates tyrosine hydroxylase activity. The kinetic data suggests that the PC-12 tyrosine hydroxylase in the basal state is in a non-phosphorylated form but under phosphorylating conditions the enzyme is activated and its kinetics properties are altered.


Intestinal transport of weak electrolytes: Determinants of influx at the luminal surface. The determinants of weak electrolyte influx into everted segments of rat small intestine have been studied. Preliminary experiments showed that the observed influxes could be described as unidirectional, diffusional fluxes of the nonionized compound uncomplicated by a parallel ionic component. It is shown that the determinants of weak electrolyte influx in this situation may be described in terms of the resistance of the unstirred layer to movement from the bulk phase to the cell surface, the degree of ionization of the weak electrolyte at the cell surface, and the cellular permeability to the nonionized weak electrolyte. Quantitative considerations indicated that the unstirred layer was totally rate-limiting in the cases of some poorly ionized, or highly permeant compounds, but the unstirred layer was not totally rate limiting for most of the compounds studied. Calculation of cellular permeabilities for the nonionized forms of weak electrolytes required assumptions to be made concerning the pH value in the surface fluid layer. A uniform set of permeability data including both weak acids and weak bases was obtained only when it was assumed that the pH in the surface fluid layer was equal to that in the bulk phase, and it was concluded that these studies do not support the concept of a microclimate of distinctive pH at the epithelial surface as a determinant of weak electrolyte transport.


Double mode of action of black widow spider venom on frog neuromuscular junction. Black widow spider venom (BWSV) contains a toxin, alpha-latrotoxin, which is capable of stimulating vesicle release, resulting eventually in depletion of vesicles and block of neuromuscular transmission at the frog neuromuscular junction. Since it has been shown that alpha-latrotoxin very markedly increases the cation conductance of artificial lipid bilayers, it was postulated that BWSV stimulates release by opening channels permeable to Ca2+ and, in the case of Ca2+-free Ringer's, to Na+ which would release Ca2+ from intracellular stores. To test this hypothesis we chose as a sodium substitute, glucosamine, which is impermeable to the venom-induced channels in the lipid bilayers and to the postsynaptic membrane of the frog neuromuscular junction. Electron microscopical analysis showed that up to 75 min perfusion in Na+ and Ca2+-free medium did not alter the ultrastructure of the nerve terminals. However when BWSV was applied in this medium a significant depletion was noticeable within 15 min and after 60 min the terminals were depleted of vesicles whereas the mitochondria were unchanged in number and structure. If BWSV is applied for 60 min in glucosamine Ringer's containing 1.8 mM Ca2+, most of the nerve terminals still have synaptic vesicles scattered in the cytoplasm or clustered around amorphous structures and the mitochondria are swollen. Application of large doses of BWSV in low Ca2+ Ringer's leads to damage of the mitochondria and to very pronounced swelling of the nerve endings, whereas this is not observed if the dose of venom is applied in Na+-free and Ca2+-free Ringer's. Electrophysiological recording showed that neuromuscular transmission is already blocked after 15 min treatment with BWSV in glucosamine-Ringer's. From these results we conclude that BWSV increases the conductance of the nerve terminal membrane to cations such as Na+ and Ca2+ and stimulates release by a mechanism which may not involve its ionophore property.


N-Halo derivatives V: Comparative antimicrobial activity of soft N-chloramine systems. Comparative antimicrobial activity studies for certain new classes of soft N-chloramines derived from alpha-aminiisobutyric acid and 2-amino-2-methyl-1-propanol were examined using the minimum inhibitory concentration (MIC) and/or the contact germicidal efficiency (CGE) procedures. Several factors significantly aliphatic chain length in a homologous series, (b) the degree of chlorination of thenitrogen atom, (c) the solution pH, (d) the presence of a denaturant, and (e) the nature of a positive charge.


Stability of aqueous solutions of mibolerone. The kinetics and mechanism of degradation of mibolerone were studied in aqueous buffered solutions in the pH range of 1-8 at 67.5 degrees. Mibolerone showed maximum stability between pH 5.5 and 6.4. At pH 1-2, the major degradative pathway was dehydration followed by migration of the 18-methyl group to form 7alpha,17,17-trimethylgona-4,13-dien-3-one. While there was only one degradation product at pH 1-2, the degradation at pH 7-8 was complex. As many as 12 degradation products were detected by GLC. Mass spectral data indicated that the majority of these products were either oxidation products or isomers. At pH 7.6, the apparent first-order rate constants exhibited marked dependency on buffer concentration. Incorporation of a sequestering agent into the solutions eliminated this dependency, suggesting that trace metal impurities from the buffer reagents were catalyzing the degradation. This was confirmed by degradation studies of solutions in water for injection containing 5 ppm of trace metal ions. Sn+2, Cu"2, and Fe+2 accelerated the degradation, with Fe+2 having the most catalytic effect. The temperature dependence of the rate of degradation was studied in 0.05 M phosphate buffer at pH 6.4. The activation energy was 19.6 +/- 1.63 kcal/mole.


Effects of pH on the myofilaments and the sarcoplasmic reticulum of skinned cells from cardiace and skeletal muscles. 1. The effects of decreasing pH from 7.40 to 6.20 on the tension developed by direct activation of the myofilaments and by Ca2+ release from the sarcoplasmic reticulum were studied comparatively in segments of single cells of skeletal muscle (frog semitendinosus) and cardiac muscle (rat ventricle) from which the sarcolemma had been removed by micro-dissection (skinned muscle cells). 2. The concentration of free Ca2+ in the solutions was buffered with ethylene glycol-bis (beta-aminoethylether N,N'-tetraacetic acid (EGTA). The change of the buffer capacity of a given [total EGTA] caused by varying pH and the uncertainty about the value of the equilibrium constant for Ca-EGTA have been taken into account in the interpretation of the results. 3. Decreasing pH from 7.40 to 6.20 produced an increase in the [free Ca2+] required for the myofilaments to develop 50% of the maximum tension by a factor of about 5 in skinned cardiac cells but of only 3 in skeletal muscle fibres. In addition, acidosis depressed the maximum tension developed in the presence of a saturating [free Ca2+] by approximately the same amount in the two tissues. 4. The pH optimum for loading the sarcoplasmic reticulum of skinned fibres from skeletal muscle decreased when the pCa (-log [free Ca2+]) in the loading solution decreased. The optimum was pH 7.40-7.00 for a loading at pCa 7.75, pH 7.00-6.60 at pCa 7.00 and pH 6.60-6.20 at pCa 6.00. 5. The pH optimum for loading the sarcoplasmic reticulum of skinned cardiac cells with a solution at pCa 7.75 was about pH 7.40 as in skeletal muscle fibres. But the cardiac sarcoplasmic reticulum could not be loaded with a [free Ca2+] much higher than pCa 7.75 because a higher [free Ca2+] triggered a Ca2+-induced release of Ca2+ from the sarcoplasmic reticulum. 6. The pH optimum of about 7.40 for the loading of the cardiac sarcoplasmic reticulum was also optimum for the Ca2+-induced release of Ca2+ from it. 7. It was concluded that the effects of acidosis on the cardiac sarcoplasmic reticulum accentuate the depressive action of decreasing pH on the myofilaments. This may explain the pronounced depression of contractility observed during acidosis in cardiac muscle. In contrast, a moderate acidosis causes an effect on skeletal muscle sarcoplasmic reticulum that could compensate for the depressive action on the myofilaments, which is, in addition, less pronounced than in cardiac muscle.


The effects of pH and curare on the time course of end-plate currents at the neuromuscular junction of the frog. 1. The effect of pH changes on synaptic currents has been analysed by external recording of the miniature end-plate currents (m.e.p.c.s) or by recording in voltage-clamped end-plates the current elicited by nerve stimulation (e.p.c.). 2. Changes in pH do not appreciably effect the peak amplitude of the current produced by a single quantum or by short ionophoretic pulses of acetylcholine. 3. The time constant of decay of the m.e.p.c.s is prolonged by about 50% in acid pH and shortened by about the same amount in alkaline pH. This effect is independent of the cholinesterase activity of the end-plate. 4. In curarized preparations the decay of the e.p.c. is shorter than in Mg-blocked end-plate even in the absence of cholinesterase blocking agents. 5. The action of pH on the decays can be explained by a titration of the surface charges of the membrane which effects the voltage dependent reaction that controls the rate of closing of the synaptic channels.


Effects of dithiothreitol on end-plate currents. 1. End-plate currents have been studied in frog cutaneus pectoris nerve-muscle preparations mounted in continuously flowing solution, using the voltage clamp technique. 2. Exposure of the muscle to 1 mM-dithiothreitol reduced the amplitude of end-plate currents by a factor of 2.7 (mean; range 1.6-3.4; twelve fibres). 3. 1 mM-dithiothreitol also caused a 2.7-fold (2.3-3.1) increase in the rate of decay, and a 1.4-fold (1.3-1.6) decrease in the time to peak of end-plate currents. During the onset of action of dithiothreitol, there was little or no indication of departure of end-plate current decay from a simple exponential. 4. Dithiothreitol actions on amplitude and decay of end-plate currents developed with similar time courses and both effects were slower in onset at pH 7.2 than at pH 8.5. 5. The actions of dithiothreitol were reversed by exposure of the muscle to 1 mM-5,5'-dithio-bis-(2-nitrobenzoic acid). 6. Following dithiothreitol treatment, the rates of decay of end-plate currents continued to depend on membrane potential; there was little or no change in the slope of the relation between in (rate of decay) and membrane potential, consistent with little or no change in the dipole moment of a gating molecule for ion channels. 7. Dithiothreitol changed the relation between peak end-plate current and membrane potential, so that peak conductance increased at more negative membrane potentials; this finding could be accounted for in terms of the closure of ion-channel gates becoming faster though remaining voltage-sensitive after exposure to dithiothreitol. 8. It is concluded that dithiothreitol causes changes in the kinetics of gating of ion channels associated with receptors and that these changes accompany changes in the binding of ACh to receptors.


Anion transport of the red cell under non-equilibrium conditions. 1. The exchange of sulphate for chloride across the human red cell membrane was measured in both directions, i.e. by sulphate influx and sulphate efflux. The influence of the concomitant transient pH changes was minimized by phosphate buffer and by choosing experimental conditions of moderate pH sensitivity (pH 6.4 and 7.8). Sulphate self exchange was determined in chloride-free erythrocyte suspensions. 2. The transport of external sulphate into red cells proceeded at a 15-fold greater rate if its initial concentration was raised from 5 to 95 mM. In contrast the velocity constant of sulphate efflux into sodium chloride medium increased only twofold when the intracellular sulphate concentration was increased. To explain this asymmetry it is proposed that external sulphate ions are more able to complete with chloride for the anion transport sites that those present in the cell interior. 3. The transient membrane potential due to the uneven distribution of sulphate and chloride was shown by the rapid introduction of chromate into the cells. When the  erythrocytes contained chloride and the external anion was sulphate, the cells took up chromate 50 times (16.5 degrees C) faster than with equilibrium chloride distribution. 4. Chloride efflux into sodium sulphate media was measured by a chloride-sensitive electrode. Under buffered conditions in neutral and alkaline media two kinetic components were observed as the result of chloride exchange against hydroxyl and sulphate ions. At pH 6.4, chloride efflux was characterized by a single velocity constant identical to that of sulphate movement in the opposite direction. The results show that under appropriate circumstances net chloride efflux measurements can provide comparative data on the anion permeability of the red cell membrane.


5-Chloro-2-phenyl-1-benzo[b]thiophene-3-alkanimines, potential antipsychotic agents. The title compounds (most notably 2a) were synthesized on the basis of the N-methylation hypothesis of schizophrenia. They were evaluated in dopamine and haloperidol receptor assays. The binding characteristics were comparable in some cases to known neuroleptics.


Chemical and electrochemical oxidation of 7-hydroxychlorpromazine. The oxidation of 7-hydroxychlorpromazine, a process associated with several side effects of chlorpromazine therapy, was examined in vitro by electrochemistry and rapid-scanning spectrophotometry. At pH 2, the oxidation results in a quantitative yield of 7,8-dioxochlorpromazine, but several intermediates are observable during the course of the reaction. These include a quinone imine with a half-life of 0.1 s, a monosubstituted benzoquinone with a half-life of approximately 50 s, and a disubstituted benzoquinone with a half-life of approximately 5 min. The concentrations of each intermediate were determined quantitatively as a function of time, and a complete oxidation mechanism is proposed. At pH 7, the yield of 7,8-dioxochlorpromazine is less than at pH 2, and an additional reaction pathway involving direct hydroxylation of the quinone imine is observed. The relationship of these reactions to the pharmacology of the hydroxylated chlorpromazine metabolites is discussed.


The incidence of mosquitoes feeding on mothers and babies at Kisumu, Kenya. Blood fed mosquitoes were collected inside four bed nets in which mother/child pairs, having different haptoglobin types, slept. The bloodmeals were analysed by gradient gel electrophoresis to determine on which person the mosquitoes had fed. The results suggest that the mothers are fed on much more than babies by both Anopheles gambiae s.l. and Culex fatigans. In one hut at least 15.2 per cent of the mosquitoes had taken all or part of their bloodmeal elsewhere. A greater number of double feeds were detected from engorged Cx. fatigans (12.6%) than An. gambiae s.l. (2.7%).


Experience with 425 subfertile male patients. Subfertility was evaluated in 425 men. Varicocele were diagnosed in 37.4 per cent of these patients and surgical correction of the varicocele in 68 men resulted in a 65 per cent improvement in semen quality. Idiopathic abnormalities were found in 25.4 per cent of the patients. Since the study group was heterogenous setereotyped or empirical treatment cannot hope to be successful if uniformly applied. Other specific causes of male subfertility are identified and therapeutic options are discussed.


Effects of Kö 1366 on the hemodynamics of the normotensive and hypertensive elderly subjects. The hemodynamic effects of a new adrenergic beta-receptor blocking agent, Kö 1366 were investigated in normotensive and hypertensive elderly subjects. This study, in its important part, was carried out to clarify whether the intrinsic sympathomimetic activity of Kö 1366 modified the hemodynamic changes due to beta-adrenergic blocking action or not. Hemodynamic changes elicited by intravenously administered Kö 1366 (0.05 mg/Kg) were correlated with age, cardiac index, and electrocardiographic findings, respectively. With reference to age in normotensive subjects, Kö 1366 caused no significant differences in rate of changes of various hemodynamic items between 3 age groups. Normotensive elderly subjects were divided into 3 groups according to cardic index. Cardiac index decreased greatly in high cardiac index group, slightly in normal cardiac index group, whereas it increased greatly in low cardiac index group. Stroke volume index showed the same tendency as cardiac index. Hypertensive elderly subjects were, also divided into 2 groups according to cardiac index. Again, cardiac index decreased in higher cardiac index group, while it increased in lower cardiac index group, though a statistical evaluation showed only a tendency of significant difference between 2 groups. In the next, hypertensive elderly subjects were divided into 2 groups according to electrocardiographic findings of left ventricular hypertrophy. Cardiac index decreased in the subjects without electrocardiographic changes, while it increased in the subjects with electrocardiographic changes. Stroke volume index of both groups increased: a greater increase in the subjects with electrocardiographic changes and a tendency of significant difference between 2 groups was recognized. These results suggested that Kö 1366 exhibited the intrinsic symathomimetic activity on the inotropism in the hearts with depressed contractile function of both normotensive and hypertensive subjects, thereby canceling the negative chronotropic action of this agent. This could be a more advantageous point of Kö 1366 in the treatment of elderly subjects with cardiac diseases than other agents without the intrinsic sympathomimetic activity.


[Metabolic changes in the retina after experimental microembolism in the miniature pig (author's transl)]. Experimental microembolisation provokes hemodynamics and metabolic changes in the inner retina. The metabolic modifications are the consequence of focal tissue ischaemia and have as a main feature, the activation of anaerobic glycolysis. This activation produces an increase in the production of lactic acid which, in turn, causes a drop of tissular pH. Choroidal circulation maintains photoreceptors in a physiological state.


[Apparatus for the multichannel pH measurement of the upper section of the gastrointestinal tract]. To study simultaneously the acid-base processes in the esophagus, stomach and duodenum a procedure involving the use of multichannel pH measurements is proposed and a setup designed whose basic elements are a 6-channel pH-probe and a registering device. A visual and graphic registration of the investigation results is envisaged.


Tardive dyskinesia and antihistamines.

Tardive dyskinesia and antihistamines. A case of tardive dyskinesia due to the prolonged administration of antihistamines is reported. Clinical features are discussed, as well as the alleviation of these by haloperidol.


Labetalol in the treatment of hypertensive renal patients. The efficacy of labetalol in lowering blood pressure was assessed in 18 patients with chronic renal failure and hypertension. Before the start of labetalol therapy, all patients were receiving combined antihypertensive therapy, the most common being a beta-blocker and hydrallazine. Over the period of about four weeks labetalol was substituted for the prior therapy. 51Cr edetic acid (EDTA) estimations of glomerular filtration rate were performed before labetalol therapy, and then again after one and six months. Before the therapy with labetalol, 12 of the 18 patients had supine diastolic blood pressures of 100 mm Hg or more. At six months, 14 patients remained in the trial and, of these, only four had a supine diastolic blood pressure of 100 mm Hg or more. In the supine position there was a significant reduction of systolic, but not of diastolic, blood pressure. However, in the erect position there was a significant reduction both in systolic and in diastolic blood presure. Pulse rate did not vary significantly. Few side effects were encountered, transient postural dizziness being the most common side effect. Labetalol seems to be an effective substitute for the beta-blocker plus hydrallazine therapy. However, it is not as potent as minoxidil.


[Acidophilic thermotolerant Candida utilis yeast strain obtained by continuous cultivation]. An acidophilic thermotolerant strain of Candida utilis 1668-3-37 was produced under conditions of continuous cultivation during 128 days. The optimum pH is 3.0-4.5 at 32 degrees C. The strain grows with mumax=0.30 hr(-1) at 37-38 degrees C and the same pH values if dry yeast autolysate (0.05%) is added to the medium.


[Metal content in Bacillus megaterium cells under different cultivation conditions]. The content of K, Na, Mg, Fe, Al, Ba, Sr and Cu was studied in the cells of a chemostal culture of Bacillus megaterium whose growth was limited with citrate, the growth rates being D=mu=0.2, 0.4 and 0.7 hr(-1). The growth was affected also by acid and alkaline values of pH. The content of metals in the cells could change tenfold, depending on their physiological state. The content of Ca, Mg and Fe increased with the growth rate while that of K remained constant. The content of metals changed even more sharply, depending on pH. In the alkaline medium, the content of K, Na, Mg, Ca and Fe decreased, in contrast to that of Al, Ba, Sr and Cu, probably as a result of damaged transport.


Specific inactivation of heterospecific transforming DNA by a factor derived from Streptococcus sanguis lysates. A heat-sensitive factor obtained from lysates of competent Streptococcus sanguis cells reacts specifically with native DNA of heterospecific (S. pneumoniae or calf thymus) origin. In vitro it does not alter the double or single strand length of the DNA, nor does it affect uptake of the DNA by compentent S. pneumoniae cells in DNase I-resistant form. Following uptake, however, DNA previously exposed to the factor loses over 90% of its biological activity. Reaction of heterospecific DNA with the factor is competitive, suggesting a competition for binding to the factor. Heating treated DNA prior to its reaction with recipient cells, apparently by irreversibly dissociating the factor, restores to the DNA its original potential transforming activity. Specific activity of the factor can be increased in cells grown under certain conditions; this increase is blocked by erythromycin.


The properties of mnemiopsin, a bioluminescent and light sensitive protein purified by hollow fiber techniques. A calcium activated photoprotein, termed mnemiopsin, which emits bioluminescence upon the addition of calcium ion, has been isolated from the Ctenophore, Memiopsis leidyi, and purified by hollow fiber techniques. The system is similar to aequorin, from the jellyfish Aequorea, except that mnemiopsin can be light-inactivated. Separation of mnemiopsin from the dilute and large volume animal homogenate proved difficult with conventional biochemical techniques. A continuous flow process utilizing large surface area hollow fibers for filtration, concentration, and dialysis was developed which may also be applicable to the purification of other proteins. The resulting mnemiopsin concentrate, after further purification, was judged to be about 90% pure by its gel electrophoretic profile. Estimates by molecular sieve chromatography and SDS gel electrophoresis gave a molecular weight of about 23,000 daltons. A calcium specificity for triggering light emission was studied by comparison of triggering with a variety of cations and anions and by investigating the effects of calcium ionophores and antagonists. The activity of mnemiospin was characterized with respect to pH, temperature and ionic strength. The stability of mnemiopsin activity after exposure to proteases, denaturants, protein group specific reagents, detergents, elevated temperatures and light was determined. Some years ago our laboratory reported that the bioluminescence reaction in the ctenophores which had long eluded definition involved a calcium activated photoprotein similar in many respects to that found in other coelenterates, notably Aequorea. We found, moreover, that the systems differed in that the bioluminescent activity of the isolated protein was lost following exposure to light. The purification and characterization of this biochemical system was undertaken both in our laboratory and by Ward and Seliger. These latter reports provide a detailed and firm foundation for the understanding of the components and mechanisms involved. While many of our results are in agreement with theirs, our approaches, inquiries, and results differed in several significant ways, the description of which forms the basis for this report. In particular, we took a different approach in the purification of the Mnemiopsis photoprotein which in itself is rather a formidable task. The technique was successful and may point the way to other applications where large volume dilute solutions prove cumbersome. Secondly, our study of the effects of salts, proteases, detergents, and other agents indicate that the protein, though sensitive to calcium and visible light inactivation, is relatively resistant to some agents which commonly inactivate proteins.


Extraction of tyrosine aminotransferase mRNA by polyribosome immunosorption on sepharose 4B. Sepharose 4B column with antibody to tyrosine aminotransferase (E.C. 2.6.1.5) (TAT) covalently bound can selectively remove a specific fraction of TAT polysomes from rat liver homogenates. From the polysomes which was adsorbed by immunosorbent one may obtain mRNA containing a segment of poly-adenilate-rich residues, having sedimentation constant of 11--16S. This mRNA may program synthesis of the specific protein in a cell free system. Near 70% of the protein was synthesized in such a system, may react with the antibody to TAT.


The binding of intravenous and oral biliary contrast agents to human and bovine serum albumin. The binding of two homologous series of oral and intravenous biliary contrast agents to human and bovine serum albumin was investigated using the gel filtration technique. All intravenous compounds are bound to human serum albumin via one high affinity and several low affinity binding sites. Within the concentration range investigated, about 3--5 high affinity binding sites for the oral compounds were found on human serum albumin. In general, the intravenous compounds have a greater affinity for human serum albumin than the oral compounds. No significant differences were found for the binding of the oral compounds to human or bovine serum albumin, while the intravenous compounds have a higher affinity for bovine than for human serum albumin. The significance of the plasma protein binding of the biliary contrast agents for the hepatic uptake is discussed.


[Hemorrhagic rectocolitis. Attempt at programmed therapy]. Planned treatment of 30 cases of haemorrhagic colitis is reported. Its protocol included salazopyrine and benzodiazepine per os, salazopyrine and prednisone per rectum, and parenteral ACTH depot. Administration for 60 days brought good results in cases of slight or average severity. Recurrences, albeit less severe, were not prevented, however. Attention is drawn to the need to follow up this  group of patients for a longer period. Even so, it is felt that the effect of this form of treatment is primarily symptomatic.


Expression of hormonally induced tyrosine aminotransferase in host liver and Morris hepatoma No. 7777 during cofactor depletion. Cytoplasmic tyrosine aminotransferase (L-tyrosine: 2-oxoglutarate aminotransferase, EC2.6.1.5) was partially purified from host liver and Morris hepatoma No. 7777 grown in pyridoxine depleted rats. The animals were sacrificed six hours following the intraperitoneal administration of hydrocortisone hemisuccinate. Enzyme preparations were subsequently resolved by electrophoresis on polyacrylamide gels. Enzyme activity was detected histochemically in situ on the gels. Six and at least three enzymatically active protein peaks were detected in host liver and the hepatoma, respectively, by this method.


[The detection of bacterial antigens by counter-immunoelectrophoresis in N. meningitidis, H. influenzae serotype b, S. pneumoniae infections. Diagnostic value and evolutive aspect (in 216 cases) (author's transl)]. Using counterimmunoelectrophoresis (CIE) the authors have assayed for soluble bacterial S. pneumoniae, N meningitidis group A, B, C. H. influenzae type b antigens, biological fluids in 216 patients (meningitis: 136; pneumonia: 76; miscellaneous: 4) during 16 months. Because of heterogeneous recruiting (the bacteriology was carried out by different laboratories) the increase in aetiological diagnosis given by CIE is only statistically valid for the bacteriologic negative group when blind antibiotic therapy had already been given. In this group, CIE makes a notable increase in diagnosis of 22,1 % +/- 10,1 in meningitis and 25,5% +/- 12,7 in pneumonia. Various physiopathological aspects are considered concerning soluble bacterial antigens detection during the course of the disease. This method seems very useful and accurate; and therefore should be used in every microbiologic laboratory.


A new method for the purification and identification of covalently closed circular DNA molcules. A new technique has been developed for the rapid isolation of covalently closed circular DNA molecules. The procedure is a selective extraction based on differences in the partitioning of covalently closed circular DNA molecules and noncovalently closed species between phenol and water at acid pH and low ionic strength. Under the conditions described, linear as well as nicked circular DNA is extracted into phenol, while covalently closed circular DNA molecules remain in the water phase. The method permits the quantitative isolation of covalently closed circular DNA from either total cellular DNA or partially purified preparations, to a degree of purity comparable with buoyant density procedures.


Synthesis and properties of 8-azido-1, N6-etheno adenosine triphosphate--a fluorescent and photosensitive ATP analog. 8-Azido-1,N6-etheno-ATP--a fluorescent and photoreactive ATP analog has been synthesized and characterized by elementary analysis, thin layer chromatography, infrared spectroscopy, proton resonance spectroscopy, UV absorption spectroscopy, and fluorescence spectroscopy. The photolytical decomposition upon irradiation at different pH values is tested.


Short RNA chains synthesized at low pH are initiated at promoter sites. Under non optimal conditions- either with limiting substrate concentrations (1) or at low pH (2)- RNA polymerase of Escherichia coli synthesizes very short RNA chains. By sequencing one RNA species synthesized at pH 5.8 upon T7 DNA we were able to demonstrate that under these conditions transcription is initiated at a normal promoter site (here A1) but however is terminated soon afterwards at specific artificial sites not used in vivo.


[Polarographic method of determining phenol in the air]. A specific, highly sensitive and exact polarographic method of phenol determination in the air is developed. The conditions of sample taking are outlined, and the effect of polarographic solution pH on the height of step and semi-wave potential is verified. The suggested method is readily applicable simple for execution, and provides for a good reproducibility and promptness of determination during productional tests.


Minichromosome from BK virus as a template for transcription in vitro. BK virus DNA can be extracted from virions as a nucleoprotein complex containing about 20 nucleosomes. Transcription of this "minichromosome" with Escherichia coli RNA polymerase indicates that both initiation and elongation of RNA chains are reduced by the presence of nucleosomes. Hybridization analysis of RNA made on the complex shows preferential transcription of one region of BK virus genome. No increase in strand selection is observed with respect to transcription of purified superhelical BK virus DNA.


Transglutaminase activity in human lymphocytes: early activation by phytomitogens. Transglutaminase activity is present in human peripheral lymphocytes and is enhanced up to 15-fold within 10-30 min after treatment of the cells with concanavalin A. Phytohemagglutinin has a similar effect. The enzyme is not detected when intact cells are assayed; it is detected only in cell lysates. Concanavalin A enhances transglutaminase activity only when it is incubated with intact cells; concanavalin A treatment of cell lysates has no effect, alpha-Methyl-D-mannoside specifically inhibits the enhancement of transglutaminase activity in cells treated with concanavalin A results from the specific interaction of the lectin with its saccharide binding site on the cell surface, rather than by direct interaction with the enzyme itself. The increased activity of transglutaminase in cells treated with concanavalin A, as compared to unstimulated cells, is maintained under assay conditions in which saturating levels of Ca2+ are present. Transglutaminase may be involved in early cellular events leading to lymphocyte blastogenesis.


Topographical analysis of regulatory and metal ion binding sites on glutamine synthetase from Escherichia coli: 13C and 31P nuclear magnetic resonance and fluorescence energy transfer study. The paramagnetic effect of Mn(II) on (13)C and (31)P nuclear magnetic resonance signals from the [2-(13)C]ATP adenylylated glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming); EC 6.3.1.2] from Escherichia coli was measured. This effect permitted the determination of distances from the 2-C position and the phosphorus of covalently bound AMP to the two Mn(II) binding sites, n(1) and n(2). Binding of Mn(II) to the n(1) site converts an inactive apo-enzyme to its active form, while the metal ion bound at n(2) occupies the metal-nucleotide substrate site. The distances from Mn(II) at the n(1) and n(2) sites to phosphorus are approximately 10 and approximately 7 A and to the 2-C position of the adenine ring are approximately 12 and approximately 11 A, respectively. The fluorescence energy transfer method was used to determine distances between Co(II) at n(1) and n(2) and the adenylyl site. For this experiment the enzyme was adenylylated with epsilon-ATP. The distances between epsilon-adenine and Co(II) at n(1) and n(2) are approximately 13 and approximately 11 A, respectively. Quantitation of the paramagnetic effect due to Co(II) on the (31)P nuclear magnetic resonance signal yielded values of 8 and 6 A for the distances between the phosphorus of the covalently bound AMP and the n(1) and n(2) sites, respectively. The results reveal that the covalent modification site is very close to the catalytic center of the enzyme. In this study both nuclear magnetic resonance and fluorescence energy transfer techniques have been used to determine distances between the same set of sites on an enzyme surface.


Composition of vacuoles and sarcoplasmic reticulum in fatigued muscle: electron probe analysis. Electron probe analysis, cryo-ultramicrotomy, and freeze-substitution were used to determine the nature of vacuolation and the subcellular composition in fatigued frog skeletal muscle fibers. The vacuoles caused by fatigue were part of the T-tubule system and contained high concentrations of NaCl. The calcium concentration in the terminal cisternae was higher than previously measured normal resting values. Mitochondrial calcium content was relatively low (mean +/- SEM, 2 +/- 2 mmol/kg dry weight). Fiber NaCl was increased. It is concluded that fatigue is not due to the depletion of calcium stores from the terminal cisternae or to uncoupling of mitochondria due to calcium loading but may be caused by multiple mechanisms including failure of the T-tubule action potential.


Regulation of glutamine synthetase in cultured 3T3-L1 cells by insulin, hydrocortisone, and dibutyryl cyclic AMP. The 3T3-L1 mouse fibroblast cell line develops morphological and biochemical characteristics of adipocytes when maintained at confluence. This conversion to adipocytes is accelerated by addition of insulin to the culture medium [Green, H. & Kehinde, O. (1975) Cell 5, 19-27]. During the course of the insulin-mediated adipocyte conversion, the specific activity (units/mg of protein) of glutamine synthetase [L-glutamate:ammonia ligase (ADP-forming), EC 6.3.1.2] increases more than 100-fold. The specific activities of hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) and glucose-6-P dehydrogenase (D-glucose-6-phosphate:NADP(+) 1-oxidoreductase, EC 1.1.1.49) also increase but less dramatically (1.5- to 3-fold). In contrast, confluent cells maintained in the absence of insulin for the same time (12-20 days after confluence) display only minimal increases in the activity of these enzymes. Maintenance of confluent cells in culture medium lacking added L-glutamine has little, if any, effect on glutamine synthetase activity in either control or insulin-treated cultures. Treatment of confluent 3T3-L1 cultures with hydrocortisone (1 mug/ml) for 3 days prior to harvesting results in an increase in glutamine synthetase specific activity of 12-fold for control cultures maintained for 13 days in the absence of insulin and 1.4-fold for adipocyte cultures maintained for 13 days in the presence of insulin (10 mug/ml). Treatment of 3T3-L1 control cells and adipocytes with dibutyryl cyclic AMP (1 mM) plus theophylline (1 mM) decreases the glutamine synthetase specific activity and almost completely reverses the insulin- and hydrocortisone-mediated increases in enzyme activity. In contrast, treatment with dibutyryl cyclic AMP plus theophylline has relatively little effect on the specific activities of hexokinase or glucose-6-P dehydrogenase or on the protein content of the cultures. These data indicate that glutamine synthetase activity is hormonally regulated in 3T3-L1 cells.


Crosslinking of the nearest membrane protein neighbors in ATP depleted, calcium enriched and irreversibly sickled red cells. ATP depleted or Ca2+ (0.1 mM) enriched normal red cells (greater than 80% echinocytes III) subjected to crosslinking by catalytic oxidation contain a greater than 1,000,000 daltons spectrin rich polymer cleavable by dithiothreitol (DTT) reduction. Similar complex is seen after glutaraldehyde crosslinking suggesting spectrin rearrangement into closer contacts or aggregation. In addition, a non-reducible greater than 1,000,000 daltons polymer is produced in fresh rbc or ghosts by high (greater than 0.5 mM) Ca2+ conc. and ionophore A23187. This complex is attributed to endogenous membrane protein crosslinking, catalyzed by a Ca2+ stimulated rbc transglutamidase. ISCs exhibiting a 4 fold increase in Ca2+ and a decrease in ATP do not exhibit these polymers. However, ISCs have an increased propensity to form the spectrin rich polymer during a subsequent ATP depletion and this is associated with a transformation of greater than 60% ISCs into spheroechinocytes. Similar cells are occasionally noted (greater than 4%) in the densest ISC rich fractions separated from fresh blood. We conclude that neither Ca2+, ATP dependent spectrin aggregation nor a spontaneous irreversible crosslinking underlie the membrane lesion of ISCs. Accelerated calcium gain and ATP depletion in ISCs leads to spectrin rearrangement and transformation of ISCs into spheroechinocytes which may represent an end stage ISC lesion resulting in an ISC removal from circulation.


Pipradrol enhances reinforcing properties of stimuli paired with brain stimulation. The hypothesis that a psychomotor stimulant drug (pipradrol) enhances the reinforcing effects of stimuli paired with reinforcing brain stimulation was tested using a conditioned reinforcement paradigm. Rats were trained to discriminate between two stimuli (S+ and S-) to obtain ICS in the lateral hypothalamus by pushing a panel in the presence of S+. In a subsequent preference test, ICS was no longer available, but responding on one of two novel levers now produced S+, whereas responding on the other lever produced S-. Four groups of four rats received 0, 5, 10 or 15 mg/kg pipradrol. Doses of 5 and 10 mg/kg significantly enhanced the preference for S+ over S-. These doses increased responding for S+, but had no effect on responding for S-. These results support the hypothesis tested, and suggest that pipradrol potentiates the effects of conditioned reinforcement.


Nucleosome structure.

Nucleosome structure. Electron microscopic and biochemical results are presented supporting the following conclusions: (1) Two molecules of each histone H2A, H2B, H3 and H4 are necessary and sufficient to form a nucleosome with a diameter of 12.5 +/- 1 nm and containing about 200 base pairs of DNA. (2) H3 plus H4 alone can compact 129 +/- 8 DNA base pairs into a sub-nucleosomal particle with a diameter of 8 +/- 1 nm. In such a particle the DNA duplex is under a constraint equivalent to negative superhelicity. (3) Chromatin should be viewed as a dynamic structure, oscillating between a compact structure (the nucleosome) and more open structures, depending on the environmental conditions.


Relation of nucleosomes to nucleotide sequences in the rat. The relation of nucleosomes to nucleotide sequences is random for most single copy sequences in rat liver. This could be due to variation in the DNA content of nucleosomes, and a procedure for detecting such variation is described.


Conformational flexibility in DNA structure and its implications in understanding the organization of DNA in chromatin. X-ray crystallographic studies of drug-nucleic acid crystalline complexes have suggested that DNA first bends or 'kinks' before accepting an intercalative drug or dye. This flexibility in DNA structure is made possible by altering the normal C2' endo deoxyribose sugar puckering in B DNA to a mixed sugar puckering pattern of the type C3' endo (3'-5') C2' endo and partially unstacking base pairs. A kinking scheme such as this would require minimal sterochemical rearrangement and would also involve small energies. This has prompted us to ask more generally if a conformational change such as this could be used by proteins in their interactions with DNA. Here we describe an interesting superhelical DNA structure formed by kinking DNA every ten base pairs. This structure may be used in the organization of DNA within the nucleosome structure in chromatin.


The structure of SV 40 chromatin.

The structure of SV 40 chromatin. Simian virus 40 (SV40) nucleoprotein complexes were studied with the electron microscope. Depending on the isolation procedure, SV40 chromatin has two different conformations: complexes isolated in the presence of 0.15 M NaCl appeared as very compact globular structures, while those isolated in the presence of 0.6 M NaCl had the typical 'beads-on-a-string' appearance of the primary nucleofilament. Concomitant with this structural change was a variation in the histone pattern and sedimentation behaviour of the complexes: with NaCl at 0.15 mol 1(-1) the isolated complexes contained both the nucleosomal histones and histone H1, and sedimented in sucrose gradients at 70S. Increasing the ionic strength to 0.6 M NaCl resulted in the removal of histone H1 from the complexes and in a decrease of the sedimentation coefficient to 40S. DNA relaxing enzyme is associated with the SV40 nucleoprotein complexes. The numbers of superhelical turns in DNA from compact and open types of complexes were found to be the same. Therefore the transition from the condensed to the open structure of viral chromatin does not require a change in the topological winding number of its DNA.


Aspects of the regulation of histone genes. Sequencing of cloned histone DNA of the sea urchin Psammechinus miliaris has confirmed the map of the histone genes obtained earlier by rather less refined techniques. Sequencing of spacer has revealed that it is unlikely to code for protein. Some interesting sequences in the prelude regions to the structural genes have been found. The technique of injecting DNA into the germinal vesicle of the Xenopus oocyte has been greatly simplified, so that now many of the parameters governing the transcription of the injected genes can be investigated. Some mRNA-like molecules appear when circular histone DNA is inserted into the oocyte nucleus. We are cautiously optimistic that the technique can be further developed and will provide a useful tool for the study of the molecular mechanisms governing the expression of structural genes coding for proteins.


Organization of transcribed regions of chromatin. The endonuclease DNase II preferentially attacks a limited and tissue-specific portion of chromosomal DNA. This material may be separated from the bulk of chromatin DNA by virtue of its solubility in 2 mM MgCl2. The Mg2+ soluble fraction forms a specific subset of DNA sequences and is enriched four to sevenfold in sequences coding for cytoplasmic poly(A)-containing RNA and globin messenger RNA (in globin-producing cells). The bulk (70--90%) of rapidly labelled RNA is found associated with the Mg2+-soluble fraction. Transcriptionally active, Mc2+-soluble chromatin is organized into repeating subunits of DNA (200 +/- 5 base pairs) and histone. Mc2+-soluble active subunits differ from the subunits or nucleosomes of non-transcribed regions in many respects: namely, chemical composition (non-histone protein and RNA), sedimentation properties, differential sensitivity to DNase I and the single-strand-specific nuclease S1, and optical melting behaviour. These results suggest that chromatin subunits adopt a new configuration during the process of transcription.


The organization of transcription on lampbrush chromosomes. The meiotic lampbrush chromosomes of amphibian oocytes display readily distinguishable regions of transcription (lateral loops) which extend from axial condensates of chromatin (chromomeres). The chromomeres contain most of the chromosomal ENA which, along with histone, is tightly compacted as regular arrays of DNP. Many RNA transcripts are generated on the lateral loops, and heterogeneous non-histone proteins associate with these transcripts, forming periodic condensates of 20--30 nm ribonucleoprotein (RNP) particles. These unit particles aggregate in various ways and to varying degrees and thereby confer distinctive gross morphologies to particular loops. There are about 10(4) lateral loops per haploid complement of newt chromosomes and this figure is similar to the experimentally derived number of different messenger RNA sequences found in oocytes. From cytological and biochemical studies it is now possible to consider individual lateral loops from various aspects: as morphologically distinct units; as units of inheritance; as units of functional activity; as units of transcription; as units of transcribed repetitive sequences; and as units containing one coding sequence. The difficulties in arriving at a simple explanation of the organization of transcription in lampbrush chromosomes are discussed.


The 75 S RNA transcription unit in Balbiani ring 2 and its relation to chromosome structure. A defined transcription unit in the Balbiani ring 2 (BR 2) region of chromosome IV in the salivary glands of Chironomus tentans has been characterized on the basis of analysis of the corresponding primary transcript, 75S RNA, and its functional significance. The available information on the transcription unit and its relations to chromosome structure can be summarized in the following way: 1. The size of the 75S RNA transcription unit in BR 2 is on the order of 30 000 base pairs. 2. The unit is likely to contain a long coding segment (at least 6000 base pairs), probably corresponding to information for salivary polypeptides. 3. The sequences are distributed in more than one chromomere (probably in 3--5 chromomeres). Further studies are needed before it can be stated whether or not there is a simple one-to-one relation between chromomeres and transcription units in the BR 2 region.


Heat shock of Drosophila melanogaster induces the synthesis of new messenger RNAs and proteins. The heat shock proteins, labelled in vivo with [35S]methionine, were separated by sodium dodecylsulphate-polyacrylamide gel electrophoresis and fingerprinted after tryptic digestion. Eight distinct heat shock polypeptides are characterized in this way. Heat shock messenger RNAs were isolated and partially purified. Assayed in vitro for protein synthesis, they were found to code for heat shock polypeptides. Some parameters of the kinetics of in vivo synthesis of the heat shock proteins are presented.


Transcription in the interbands of Drosophila. Many interbands of Drosophila contain morphological evidence for transcription. Decondensed band material makes no detectable contribution to certain transcriptionally active interbands.


Antihypertensive drugs and depression: a reappraisal. Eighty-nine new referral hypertensive out-patients and 46 new referral non-hypertensive chronically physically ill out-patients completed a mood rating scale at regular intervals for one year. The results showed a high prevalence of depression in both groups of patients, with no preponderance in the hypertensive group. Hypertensive patients with psychiatric histories had a higher prevalence of depression than the comparison patients. This was accounted for by a significant number of depressions occurring in methyl dopa treated patients with psychiatric histories.


The behavioural actions of the hypothalamic peptides: a review. Recent work has shown that the hypothalamic peptides commonly associated with the control of pituitary function have important behavioural actions of possible psychiatric significance. Thus, vasopressin, ACTH and like peptides may influence memory processes, and ACTH and MSH given intracranially induce a peculiar stretching and yawning syndrome accompanied by penile erection and ejaculation. Thyrotrophic hormone-releasing factor potentiates behavioural excitation, somatostatin is depressive, while luteinizing hormone-releasing hormone facilitates sexual behaviour and the newly identified endorphins are markedly opioid in character. These and other activities of the hypothalamic peptides are reviewed and assessed alongside the clinical information available.


Factor analysis of a sleep evaluation questionnaire. A self-completion sleep evaluation questionnaire (SEQ), consisting of 10 cm line analogue rating scale questions, was constructed to investigate subjects' responses to aspects of sleep and early morning behaviour. The questions were grouped into 4 chronological areas: the ease of getting to sleep (GTS), the perceived quality of sleep (QOS), the ease of awakening from sleep (AFS), and the integrity of early morning behaviour following wakefulness (BFW). Five hundred and one SEQs were completed during several investigations into the comparative effectiveness of hypnotic drugs. The classical factor analysis produced 4 factors which corresponded to the 4 aspects of sleep and early morning behaviour listed above. The GTS and QOS factors were positively correlated (+0.57), as were the AFS and BFW factors (+0.48). The 2 sleeping state factors (GTS and QOS) were orthogonal to the 2 waking state factors (AFS and BFW).


Some altered responses in rats formerly dependent on morphine. The investigation examined whether or not physical dependence or other abnormalities were detectable 1--3 months after withdrawal in dependent rats that had been treated with the morphine (amintenance dose of 100 X 2 mg/kg/day, s.c.) for 7 weeks. When narcotic antagonists were administered on the 32nd day after withdrawal, nalorphine caused a dose-dependent increase in spontaneous locomotor activity and a complete inhibition of wet-dog shakes and the writhing syndrome. Naloxone was ineffective. A remarkable increase in spontaneous locomotor activity on the 67th day and a significant increase in body weight on the 69th and 92nd day after withdrawal occurred after an acute injection of morphine (10 mg/kg, s.c.). When morphine (10 mg/kg) was administered for 3 days from the 92nd day after withdrawal, withdrawal from morphine produced a significant decrease in body weight. When morphine (10 mg/kg) was administered for 3 days from the 102nd day after withdrawal, a levallorphan injection caused a significant decrease in spontaneous locomotor activity and an increase in the frequency of the diarrheal syndrome. These abnormal responses, not observed in the naive rats, suggest the remains of some behavioral and biochemical abnormalities 3 months after morphine withdrawal.


Comparison studies of chlorazepate administered as a divided daily dose and as a single dose at night. The residual effects of dipotassium chlorazepate administered as either a single daily dose of 20 mg at bedtime or a divided daily dose (5 + 5 + 10 mg) were studied in a placebo-controlled, double-blind trial comprising 12 out-patients. The following tests were used to determine changes in perceptual wakefulness, performance ability, fine motor skills, and coordination: critical flicker fusion test, car driving in a simulator, and the "bead and needle tests." In addition, the patients underwent a clinical assessment and also filled out a self-rating scale for judging factors related to the tests. No significant differences were found between the dosage schedules or between the active medication and the placebo. The clinical results were not dependent on the dosage schedule.


A survey study of the use of electropupillogram in predicting response to psychostimulants. To confirm the conclusions from a previous study supporting the usefullness of electropupillogram (E.P.G.) in predicting clinical response, data from three separate studies with hyperkinetic and learning disabled (L.D.) children treated with stimulants were surveyed. Change in extent of pupillary contraction (E.C.) after a test dose of stimulant as measured by E.P.G. did not correlate significantly with actual clinical rating change (with one exception out of 14 correlations calculated). These negative results are reported with a reservation regarding their validity because of technical difficulties in data collection.


Neuroleptic drugs block both the hyperactivity and the increase in caudate nucleus cyclic AMP concentration produced by the administration of tranylcypromine and L-dopa to rats. Injection of rats with tranylcypromine and L-dopa increased brain dopamine concentrations and produced a behavioural syndrome that includes hyperactivity. It also elevated caudate nucleus cyclic AMP concentrations by approximately 50% in vivo, probably by stimulating dopamine receptors. Pretreatment with chlorpromazine inhibited both the tranylcypromine/L-dopa-induced behaviour and elevated cyclic AMP concentrations in a dose-dependent manner. Haloperidol and alpha-flupenthixol also inhibited both effects, while beta-flupenthixol and pimozide were without effect. Since none of these drugs altered the tranylcypromine/L-dopa-induced rise of brain dopamine, it is likely that they produced their effect by inhibiting dopamine-sensitive adenylate cyclase. A good correlation was found to exist between the neuroleptic inhibition of both the increased behavioural activity and the increased caudate nucleus cyclic AMP concentrations produced by tranylcypromine and L-dopa.


Labilization of brain lysosomes by trichloroethylene metabolites and peroxide anion in vitro. Lysosomes were isolated from adult male Wistar rat brain and incubated in vitro with 10 micron trichloroethylene and microsomes or postmicrosomal supernatant with added NADPH. Freezing and incubation with the solvent alone served as controls. The results were compared with the effect of peroxide anion from the xanthine-xanthine oxidase reaction. Mere solvent did not cause an appearance of acid proteinase activity in the incubation medium whereas other treatments induced lysosomal labilization.


Changes in plasma high density lipoproteins in chronic male alcoholics during and after abuse. Alterations in plasma high density lipoproteins (HDL) were studied in thirty-eight male chronic alcoholics. Twenty-four (63%) of the patients had increased HDL protein (measured immunochemically) and twenty-five (66%) had increased HDL cholesterol (determined after polyanion precipitation of very low density lipoproteins (VLDL) and low density lipoproteins (LDL)). A statistically significant correlation was found between HDL protein and HDL cholesterol (r = 0.39). gamma-Glutamyltransferase (GT) was elevated in eighteen (47%) of the alcoholics. No significant correlations were found between GT and HDL protein or HDL cholesterol. The increase in HDL, as studied by rate zonal ultracentrifugation, was heterogeneous with changes in the HDL2 as well as HDL3 subfractions. It is suggested that determination of HDL total cholesterol, in combination with GT, may represent a valuable and sensitive test for detection of alcoholism.


Evidence for two types of non-specific suppressor cells activated by the graft-versus-host reaction in mice. In vitro experiments were carried out to investigate the immunosuppressive efect of different populations of spleen cells obtained from animal experiencing a graft-versus-host reaction (GVHR). To induce the GVHR, parental lymphoid cells were injected into adult F1 hybrid mice. GVHR-activated spleen cells (GVH-sc) taken at different times post-GVHR induction were separated into adherent and non-adherent fractions and treated with anti-theta serum plus complement. The different types of GVH-SC were added to either parental (donor-type) of F1 (host-type) normal spleen cells and cultured with sheep erythrocytes in a modified Marbrook chamber. It was found that both adherent and non-adherent 10-day GVH-SC significantly inhibited the plaque-forming cell response of F1 normal spleen cells but not that of parental normal spleen cells. Significant suppression of the parental response was observed following the addition of 15-day GVH-SC or anti-theta treated adherent and non-adherent 7-day GVH-SC. The results suggest that the non-specific immunosuppressive effect of GVH-SC is mediated by GVHR-activated macrophages and B lymphocytes found in the anti-theta treated adherent and non-adherent fractions of the GVH-SC suspensions respectively.


The rapidity of bronchodilatation. A comparison of isoprenaline, terbutaline and rimiterol. The onset of action of three sympathomimetic drugs (rimiterol 0.2 mg, terbutaline 0.25 mg and isoprenaline 0.08 mg) administered by metered dose inhaler was compared in 12 asthmatics. In order to evaluate side effects of the drugs, a second dose of the aerosols, 8-10 times higher than the first one, was given after 30 min. The effects were measured in terms of VC, FEV1.0, PEF and heart rate. All the substances caused a significant increase in spirometric values within 1.5 min, the results with isoprenaline being slightly better than those with terbutaline. An increased heart rate was observed after the first dose of isoprenaline whereas no further increase was found after the higher dose. Two patients complained of palpitations, one after isoprenaline and one after rimiterol. No other side effects were recorded. It is concluded that there is a small, clinically insignificant, difference in the onset of action of isoprenaline compared with that of the selective beta2-stimulating aerosol, terbutaline.


[Role of metabolites in the relationship between pharmacokinetics and the effect of beta blockers. Studies on tolamolol and bufuralol]. Plasma concentrations of tolamolol and bufuralol (beta-blocking agents) were measured after oral and intravenous administration to healthy volunteers. The plasma levels of their main metabolite was also determined. Simultaneously, the effect of the drugs on the heart rate and blood pressure was monitored under various stimuli (isoproterenol, exercise or orthostatism) and Valsalva maneuver. When given orally, the two drugs are extensively metabolized by a hepatic first-pass effect. After reaching the systemic circulation, they are metabolized in the liver to hydroxylated derivatives with similar pharmacologic activity as the parent molecule. For tolamolol it is possible to demonstrate a good correlation between parent drug blood levels and the pharmacodynamic effect; this relation is less evident for bufuralol. The pharmacokinetic analysis of the behaviour of the two beta-blocking agents and their main metabolite makes it possible to explain this difference in part. The results of the present study emphasize the importance of measuring metabolites when dose-action relationships are investigated.


Graft-versus-host reactions within the thyroid gland may evoke pari passu autoimmune thyroiditis. Unilateral, intraorganal injection of parental lymph node cells into thyroid lobes of F1 hybrid rats elicited an intense, local graft-versus-host reaction within the injected lobes containing massive numbers of lymphocytes. Surprisingly, significant mononuclear cell infiltration was also seen within the contralateral (noninjected) thyroid lobes, even when the isthmus had been divided. The local graft-versus-host reaction spread from the injected gland to involve the regional lymph nodes and spleen. Migration of 51-cr-labeled donor lymph node cells was documented in these same tissue sites; lesser numbers of labeled cells were found in distant lymph nodes, including the contralateral cervical nodes. However, no significant label was found in the contralateral thyroid lobes. Microscopic evaluation of noninjected glands revealed intense infiltration of mononuclear cells around and within follicles, reminiscent of lesions induced by immunization with thyroid extract in Freund's complete adjuvant. The absence of donor cells in the inflammatory reactions within noninjected thyroid lobes raises the possibility of an autoimmune rather than graft-versus-host pathogenesis.


[Risk of surgery at the height of a gastroduodenal hemorrhage]. The authors believe that the decrease of mortality in operations for gastroduodenal bleeding does not depend on the time interval between the start of bleeding and surgical interventions, but it depends upon the conditions under which these interventions are carried out. The conditions should approximate the conditions existing in planned gastric surgery as much as possible. In this regard the results of the treatment of 188 cases with massive gastroduodenal bleeding (69 cases aged over 60) are analyzed. The applied method permitted to ensure a sufficient volume of the examination of a case and of his preparation to the operation to avoid "desperate operations" and to decrease considerably the mortality in gastroduodenal bleeding. Among patients under 65 there were no fatal cases.


[Haemodynamic characterization of a new beta-receptor blocker celiprolol (st1396) at rest and during ergometer exercise compared with propranolol (inderal) (author's transl)]. A new beta-receptor blocker (Celiprolol) was characterized in man by haemodynamic studies carried out on a random cross-over basis in 6 volunteers before and after intravenous administration of the drug or propranolol. The studies were performed at rest and in response to ergometer exercise. The studies showed that: 1. Celiprolol is a cardio-selective beta-receptor blocker with pronounced intrinsic sympathomimetic activity. Hence, Celiprolol increases the heart rate and cardiac output at rest. 2. The heart rate was reduced by Celiprolol during pronounced physical exercise. 3. Celiprolol probably has only a very slight blocking effect on those, beta1-receptors which mediate a positive inotropic effect. 4. The increase in blood pressure during exercise was less pronounced during Celiprolol medication than with propranolol.


[Experimental study concerning the psychological preparation to parturition (author's transl)]. The following study reports an application to primae gravidae of two different psychological treatments aiming at reducing the fear of childbearing and facilitating the delivery. The treatments under comparison were systematic desensitization and modification of self instruction, a more cognitive approach. The effects of the two treatments proved to be identical; nevertheless there were significant differences between the results of the women who underwent treatment and those of the control group. Moreover, in our study a certain  number of socioeconomic factors turned out to be correlated with the treatment effects.


[The influence on efficacy of formaldehyd and phenol against bacterial cells I. Effect of drying, cations and pH-value (author's transl)]. 1. When cells of E. coli and Staph. aureus are dried, the efficacy of phenol is reduced but the efficacy of formaldehyde is increased. 2. A change of the aw-value by addition of salts changes the efficacy of formaldehyde as well as that of phenol. For these tests the chlorides of various alkali and alkaline earth metals were used in different concentrations. 3. The optimal efficacy of phenol and formaldehyde within the tested concentration gradientis caused by different concentrations of the ions: In low concentrations the efficacy of formaldehyde is optimally increased, the efficacy of phenol, however, attains a minimum at this concentration range. By higher concentrations of ions the efficacy of phenol is increased, that of formaldehyde is reduced. 4. The position of the cations in the periodic system also plays a role in the influence on the efficacy of the phenol and formaldehyde respectively. The cations of the lower periods from the main groups I and II favour the damage to the cells by phenol, the cations of the higher periods favour the damage by formaldehyde.


Changes in gastric secretion following administration of preparations affecting the parasympathetic and sympathetic nervous system. The changes in gastric secretion following administration of different preparations have been studied in  a group of 180 patients with duodenal ulcer and chronic gastritis. In patients with hypersecretion the medication with bishpan and inderal as compared with atropine caused considerable shifts in the indices of gastric secretion. Isoproterenol exerted a stimulating influence both on initial hyper- and hyposecretion, while the other preparations studied were shown not to cause any essential rise in gastric secretion in patients with secretory deficiency. Reliable changes of the stimulated gastric secretion were observed only after inderal. The data obtained can be useful when choosing a rational method for the therapy of gastric scretory disorders.


Cardiovascular effects of local adrenaline infiltration during halothane anaesthesia and adrenergic beta-receptor blockade in man. Adrenergic beta-receptor blocking agents, alprenolol, propranolol and practolol were given as a prophylactic measure to patients undergoing middle-ear microsurgery where adrenaline was deliberately infiltrated during halothane-N2O/O2 anaesthesia. These three beta blockers did not differ in their action on heart rate, arterial blood pressure, right ventricular pressure, CVP or peripheral pulse wave in equipotent doses, which were 0.04 mg/kg for alprenolol and propranolol and 0.4 mg/kg for practolol in this study. Atropine caused a highly significant increase in heart rate in the spontaneously ventilating group and maintained blood pressure at higher levels, although the latter difference was not significant. Alprenolol and propranolol offered partial protection for over 40 min against adrenaline-induced arrhythmias. Occasionally occuring tachyarrhythmias were easily terminated with a further dose of a beta blocker. The effective half-life of practolol was less than 15 min and doses up to 0.4 mg/kg were unable to prevent arrhythmias during adrenaline challenge.


Cardiovascular effects of local adrenaline infiltration during neurolept analgesia and adrenergic beta-receptor blockade in man. The adrenergic beta-receptor blocking drug, alprenolol, was given together with atropine to 15 patients during neurolept analgesia (NLA) in order to prevent adrenaline-induced cardiac arrhythmias. Four patients operated on without adrenergic beta-receptor blockade formed the control group. Three of them developed tachyarrhythmias, ventricular tachycardia, multifocal ventricular extrasystoles and supraventricular tachycardia. All these arrhythmias were terminated by the adrenergic beta-receptor blocking drug, propranolol. The fourth patient developed a temporary sinus tachycardia which did not require any treatment. The adrenergic beta-receptor blockade induced a slight increase in pulse rate with almost no change in arterial blood pressure. The incidence of arrhythmias due to adrenaline infiltration was much less in this group. There was one supraventricular tachycardia and scattered ventricular extrasystoles in two other patients. NLA alone does not protect the heart against adrenaline-induced arrhythmias. It does, however, seem to be compatible with an adrenergic beta-receptor blockade, which in turn provides partial protection against adrenaline-induced arrhythmias. The occasionally developing arrhythmias can then be terminated with specific adrenergic beta blockers.


Qualitative differences between beta-adrenergic and alpha-adrenergic inotropic effects in rat heart muscle. If beta- and alpha-adrenergic inotropic effects are cyclic AMP dependent and cyclic AMP independent, respectively, they may be qualitatively different. The inotropic effects of beta-receptor stimulation (isoprenaline) and alpha-receptor stimulation (phenylephrine combined with propranolol) were characterized in isolated perfused rat hearts, rat atria and rat papillary muscles. The beta-effect reached its maximum before the alpha-effect. The alpha-effect followed a three-phasic time-course indicating both stimulatory and inhibitory components. The aortic pressure wave (perfused heart) indicated a shorter contraction phase after beta-stimulation than after alpha-stimulation. The time to peak tension (atrium, papillary muscle) was relatively shorter after isoprenaline than after alpha-stimulation, which tended to prolong it. The contraction-relaxation cycles (atrium, papillary muscle) were examined by recording the isometric tension (T), its first (T') and second (T'') deri derivatives. alpha and beta-stimulation both increased Tmax, T'max (maximal rate of tension rise), T'min (maximal rate of tension decline) and T''min (maximal rate of transition from rise to decline of tension). Isoprenaline increased T'min (papillary muscle) and T''min (atrium, papillary muscle) relatively more than did alpha-stimulation, i.e. the relaxing processes were activated relatively more by beta-stimulation. The results indicate different mechanisms for the two adrenergic inotropic effects. The relatively larger activation of relaxation by beta-stimulation is assumed to be caused by clic AMP.


The effect of nifedipine on the sinus and atrioventricular node of the dog heart after beta-adrenergic receptor blockade. The effect of nifedipine (BAY 1040), a calcium-antagonistic inhibitor of the electromechanical coupling process was tested on atrioventricular conduction and refractoriness of the dog heart in situ by means of His-bundle electrography and programmed electrical stimulation. The animals were anaesthetized with sodium pentobarbital. As the basic effects of the compound can be altered by release of catecholamines from sympathetic nerves of heart and vessels, the dogs were pretreated with acebutolol, a beta-adrenergic receptor blocking agent, which decreased heart rate and prolonged atrioventricular conduction and refractoriness. Nifedipine 1,6 and particularly 30 microgram/kg body weight increased the heart rate and decreased atrioventricular conduction time during atrial pacing, whereas atrioventricular conduction time during sinus rhythm and atrioventricular refractoriness were only affected by nifedipine 30 microgram/kg. In this respect, nifedipine differs distinctly from another calcium antagonistic compound, verapamil.


Myocardial effects of lithium in vitro.

Myocardial effects of lithium in vitro. The myocardial effects of increasing concentrations of ionic lithium were investigated in isolated, spontaneously beating rabbit hearts, which were retrogradely perfused with a modified Krebs-Henseleit solution containing lithium in concentration of 1, 5 or 10 mM. The following functional parameters were continuously recorded: heart rate, amplitude and rate of contraction, coronary flow, myocardial oxygen consumption and ECG. The lowest concentration of 1 mM lithium did not cause any measurable changes in these parameters. A concentration of 5 mM lithium produced a rather slowly developed increase in amplitude and rate of contraction of about 22% without any concomitant increase of oxygen consumption. A delayed increase in the oxygen consumption of about 21%, but accompanied by a significant decrease in contractility however, occurred much later in the experiments, probably as an expression of a decrease in myocardial efficiency. The highest lithium concentration produced similar effects. None of the other parameters measured were markedly influenced by lithium. The positive inotropic effect of lithium on myocardial contractility was very convincingly verified in isolated electrically stimulated rabbit papillary muscles by isometric measurements of the maximum tension produced and the rate of its development. An inhibitory influence of lithium on the positive inotropic effect of increasing frequency of stimulation was observed. Adrenergic beta-receptor blockade did not abolish the positive inotropic effect of lithium. A possible causative influence of lithium on the cellular calcium metabolism is discussed.