Sulphate transport by rat ileum. Effect of molybdate and other anions. Kinetic constants for SO4-2- transport by upper and lower rat ileum in vitro have been determined by computer fitting of rate vs concentration data obtained using the everted sac technique. MoO-4-2- inhibition of this transport is competitive, and kinetic constants for the inhibition were similarly determined. Transport is also inhibited by the anions WO4-2-, S2O3-2- and SeO4-2-, in the order S2O3-2- greater than SeO4-2- greater than or equal to MoO4-2- greater than WO4-2-. These anions have no effect on the transport of L-valine. Low SO4-2- transport rates were observed in sacs from animals fed a high-molybdenum diet. The significance of the results with respect to the problem of molybdate toxicity in animals is discussed, and related to the known protective effect of SO4-2-.


Investigations of thermotropic phase changes in peripheral nerve of frog and rat. A spin label study. The temperature-dependent fluidity of myelin of frog and rat peripheral nerve (Nervus ischiadicus) was studied using the spin label technique. In frog nerve a phase change was detected at 38 degree C. In rat nerve no sharp phase change could be established, and the lipid-depleted frog and rat nerve also showed no transition. From the spectral data, it was concluded that in frog and rat nerve the lipid-protein interactions are different, i.e. species dependent. Ca-2+-depletion of frog nerve caused a loss of transition, while rat nerve remained unaffected. Thus it was indicated that, in frog nerve, Ca-2+ is involved in the phase change. In the total lipid extract of frog nerve a phase change centered at 32 degree C occurred, while the total lipid extract of rat nerve again showed no transition. It is suggested that a connexion exists between our results and investigations on the temperature dependence of an axonal conduction block of nerve.


The optical activity of D-erythro-sphingomyelin and its contribution to the circular dichroism of sphingomyelin-containing systems. Circular dichroism studies on bovine brain sphingomyelin show the presence of a strong negative cotton effect below 200 nm, the position and magnitude of which depend on the physical state of the lipid. This cotton effect is thought to arise from the pi-pi transition of the amide group in the sphingomyelin backbone. The sphingomyelin contribution to the observed ellipticity of membranes and lipoprotein complexes depends on the mol fraction of amide groups present as sphingomyelin: this contribution is calculated to be less than 2% in the case of serum high density lipoprotein and the order of 20% below 200 nm in the case of the erythrocyte ghost membrane. Due to the similarity of the CD spectrum of sphingomyelin to that of a random coil polypeptide, use of uncorrected ellipticity data is expected to lead to an overestimate of the random coil content of proteins in systems containing a high sphingomyelin content.


Spin label evidence for the role of lysoglycerophosphatides in cellular membranes of hibernating mammals. The phospholipid composition of ground squirrel heart muscle changes during hibernation: more lysoglycerophosphatides are found in the hibernating state than in the active state. Phase transitions inferred from spin label motion occur in the usual manner typical of mammalian mitochondria for the mitochondria and mitochondrial lipids from active squirrels. However, a conspicuous absence of a spin label-detectable phase transition is observed in equivalent preparations from hibernating animals. The addition of lysolecithin to preparations from active squirrels removes the break and induces a straight line in the Arrhenius plot. The lack of a spin label-detectable phase transition in hibernating animals, therefore, is attributed to an increased content of lysoglycerophosphatides present in the phospholipids during hibernation.


Studies on the regulation of the concentration of androgens and androgen receptors in nuclei of prostatic cells. Experiments were performed to assess the effect of intracellular androgen metabolism and the availability of cytoplasmic receptors on the concentration of androgens and androgen receptors in nuclei of prostatic cells. It was found that androgens are incorporated into the nucleus by a regulated, selective process which appears to limit the type and amount of androgen transported across the nuclear membrane. The metabolic conversion of testosterone to dihydrotestosterone which takes place in cytoplasm does not reduce transport and, very likely, affects only the ratio of testosterone and dihydrotestosterone transferred into the nucleus. In vivo, when the intranuclear concentration of androgens approaches 250 nM (8 pmol per mg DNA), an apparent concentration ceiling is reached even in the presence of a downward concentration gradient that would be expected to promote further transport across the nuclear membrane. This finding strongly suggests that in vivo the nuclear membrane acts as a barrier to the passage of androgens and, therefore, mitigates against the possibility that passive diffusion is an important mechanism of afferent transport of androgens into the nucleus. The ability of the nucleus to concentrate testosterone and dihydrotestosterone was clearly demonstrated in vivo when cytoplasmic concentrations of androgens of approximately 20 nM were accompanied by intranuclear concentrations in the vicinity of 250 nM. Since the measured concentration of testosterone and dihydrotestosterone in prostate of several species fall within the 5-20 nM range, it is evident that androgen concentrations in the nucleus as high as 250 nM may be typical of the physiological steady state. At the latter concentration the nucleus contains 60 000 androgen molecules: in approximate terms one third of this total is bound to a large molecular weight component of the nucleus, one third is bound to a 3.3 S receptor and one third is free or loosely bound. Since 60 000 androgen molecules and 20 000 receptor molecules appear in the nucleus before transport stops, it seems that the quantity of 4.4 S cytoplasmic receptor estimated at 174 plus or minus 24 pmol per mg protein (equivalent to about 8000 molecules per cell) is insufficient to account for the total influx of androgens and androgen receptors into the nucleus. Thus, although these results support the view that cytoplasmic receptors and the capacity to transport androgens are closely linked phenotypic markers of intracellular steroid hormone action, they suggest that the control of androgen concentration in the nucleus is achieved in a more intricate fashion than simply through a dependence on the presumed translocation of 4.4 S androgen-receptor complex into the nucleus.


The control of anaerobic glycolysis by glucose transport and ouabain in slices of hepatoma 3924A. 1. The activities of glycolysis and K-+ transport have been studied in slices of Morris hepatoma 3924A incubated under anaerobic conditions in the presence of different concentrations of glucose (1-50 mM). 2. Ouabain-sensitive net transport of K-+ was observed at all glucose concentrations greater than 1 mM; ouabain reduced the rate of glycolysis by about 25% at all glucose concentrations able to support ion transport. 3. The net entry of glucose into the intracellular phase was studied at varying glucose concentrations. The rate of glucose entry was similar to the rate of glucose utilisation by anaerobic glycolysis at medium concentrations of 10 mM and less, but exceeded the rate of glycolysis at 20 mM and above. 4. The glucose entry was not Na-+-dependent and was not inhibited by ouabain. 5. The results suggest (a) that the reduction in glycolytic activity caused by ouabain is not due to an inhibition of glucose transport and (b) that the glucose transport system of this poorly differentiated hepatoma has properties similar to that of normal liver.


Chaotropic anions and the surface potential of bilayer membranes. The chaotropic anions perchlorate and thiocyanate adsorb to artificial phospholipid membranes. The negative electrostatic potential they produce at the surface of the membranes was measured by two independent techniques. The conductance produced by neutral carriers of cations and anions was measured to estimate changes in the surface potentials of planar black lipid films and the electrophoretic mobility of phospholipid vesicles was used to monitor changes in the zeta potentials of spherical bilayer membranes. Qualitatively similar results were obtained with the two techniques. The results, moreover, agreed with the change in surface potential produced by these anions at an air water interface, as measured directly with an ionizing electrode (Randles, J. E. B. (1957) Discuss. Faraday Soc. 24, 194-199). The results obtained with artificial bilayers may explain the observation (Wieth, J. O. (1970) J. Physiol. 207, 581-609) that thiocyanate increases the sodium or potassium and decreases the sulfate permeability of erythrocyte membranes.


Selective excision of gamma ray damaged thymine from the DNA of cultured mammalian cells. Three mammalian cell lines (WI-38, SV40-transformed WI-38 and Chinese hamster ovary) were exposed to high doses of 137-Cs gamma rays and their DNA analysed, following various periods of postirradiation incubation, for products of the 5,6-dihydroxy-dihydrothymine type. Within fifteen minutes of incubation at 37 degrees C 70 to 90 percent of these radiation products were removed from acid-precipitable material in all three cell lines. The amount of DNA degradation induced by radiation varied from approximately one percent in WI-38 cells to 15 percent in SV40-transformed WI-38 cells. Comparison of DNA degradation with the amount of thymine radiation product removed indicates that a selective gamma ray-induced excision repair capability exists in mammalian cells. Because of its more rapid kinetics, gamma ray excision repair is probably a distinct process as compared with ultraviolet-induced pyrimidine dimer excision.


Polynucleotide duplexes based on poly(7-deazaadenylic acid). In order to find a poly(A)-poly(U) analog which could not form a triple-stranded complex and which would have a sufficiently high thermal stability to survive under physiological conditions, the interaction of poly(7-deazaadenylic acid) (poly(c-7A)) with modified polyuridylic acids was examined. Mixing curves constructed by the method of continuous variation, isosbestic points and thermal melting profiles proved that poly(c-7A) formed only 1:1 complex with polyribothymidylic acid and poly(5-bromouridylic acid) (Tm values of 50 and 72 degrees C respectively, in 0.15 M NaCl, 0.01 M KH2 PO4, 0.001 M MgCl2, pH7). In addition poly(c-7A) formed a 1:1 complex with poly(I) (Tm equals 22 degrees C in 0.46 M salt, pH 7), and presumed duplexes were observed in the interaction of poly(c-7A) with poly(dT), poly(2'-azido-2'-deoxyuridylic acid) and poly(2'-O-methyluridylic acid) (Tm values of 35, 32 and 41 degrees C respectively, in 0.10 M NaCl, pH7).


Stimulation of nuclear poly (adenosine diphosphate-ribose) polymerase activity from HeLa cells by endonucleases. Isolated nuclei from HeLa cells can incorporate labeled ADP-ribose from NAD into an acid-precipitable product, poly(ADP-ribose). This reaction is stimulated by 4-6-fold by the addition of deoxyribonuclease I to the complete reaction mixture. If the nuclei are treated first with deoxyribonuclease I, no effect is seen; the stimulation is only apparent when the two enzymes deoxyribonuclease I and poly(ADP-ribose) polymerase, are operating at the same time. After making several minor modifications in the assay mixture, it was found that another endonuclease, micrococcal nuclease, can also stimulate the poly(ADP-ribose) polymerase activity of HeLa nuclei. A comparison of the two stimulatory effects indicated that the two endonucleases activated to the poly(ADP-ribose) polymerase activity of HeLa nuclei in the same way. Overall this evidence suggests that poly(ADP-ribose) polymerase may have a functional role in the process of DNA repair.


[Interaction of rhodopsin with quinone]. The reaction of rodopsin extracted with cetyltrimethylammonium bromide with p-benzoquinone was studied with spectrophotometric and ESR methods. In the dark quinone reacted with three SH-groups of rodopsin, in the light with three more SH- and two unknown groups of decolourized rodopsin. The interaction of quinone with SH-groups liberated under light effect on rodopsin proceeds with the formation of intermediate free-radical products.


[EPR study of the mechanism of photooxidation of chlorophyll a by nitro compounds]. An ESR signal with g-value of 2,0026 and peak-to-peak with 8,5 Oe was observed in ethanole solutions of chlorophill a and nitroderivatives of naphtalene under red light illumination. The ESR signal consisted of chlorophyll cation radical resonance, superimposed on an nitronaphtalene anion radical. Less than the ratio greater than between the quantities of these radicals and the character of decay kinetics are influenced by the medium acidity. An analysis of kinetic curves is performed, and the effect of the medium acidity on the mechanism of one-electron transfer reaction between chlorophyll and nitronaphtalene molecules is explained.


[Pharamagnetic centers developing during irradiation of gamma-phos]. The ESR spectra's of X-irradiated radioprotector gamma-phos have been studied. Using the analysis of relaxation effects in this sample irradiated at room temperature three paramagnetic centers have been discovered. One of these centers is classed with the free radical approximately CH2--CH--CH2 approximately (R1)-G(R1) value decreases with a decrease of irradiation temperature. Free radicals R1 are not oxidized in the air. Two other of paramagnetic centers have not been identified.


Tissue fractionation in rat brain, kidney and liver. I. Intracellular localization of a 5-methyltetrahydrofolic acid requiring enzyme. The intracellular distribution of N-methyl-transferase requiring 5-methyl-tetrahydrofolic acid (5 MT-NMT) was studied in brain, kidney and liver of rats. Among these different tissues, the kidney displayed the highest enzyme activity, more than 20 times the activity detected in the brain. As the striatum and, to a lesser extent the hypothalamus, were found to contain slightly higher 5 MT-NMT than other cerebral regions, they were also selected for the study of the subcellular localization. Tissue fractionation was performed by differential centrifugation yielding five different fractions which were analyzed for their enzymatic content not only of 5 MT-NMT but also of marker enzymes, such as cytochrome oxidase, acid phosphatase and inosine diphosphatase. In all the tissues studied, 5 MT-NMT was recovered in the supernatant fraction. Therefore one may consider this enzyme to belong to the cytosol. Although a neuronal localization cannot be excluded, it is beyond doubt that the enzyme is contained in other cellular types. In the brain fractionation, the five fraction procedure seems to be very useful especially when the subcellular distribution of a given enzyme is compared to that obtained in other tissues like liver or kidney. Finally 5 MT-NMT may be considered a good marker enzyme for the supernatant fraction.


[Metabolic synapse. I. Effect of intracellular microinjection of 3'-5'-AMP]. Cyclic 3'-5'-AMP was injected into neurons of Helix pomatia (theta greater than 50 mu) by the current of 10(8) minus 10(9) A. This current was passed from one intracellular electrode to another by means of a photogenerator very well isolated from the ground. The injection of 3'-5'-AMP results in a depolarization and increase of the frequency of spontaneous activity. These changes continue for several seconds after the current is switched off. The intracellular injection of 5'-AMP or 5'-ATP and also extracellular application of 3'-5'-AMP by either the same or tenfold current does not change the spontaneous activity of these neurons.


[Wave regime of neuronal net activity].

[Wave regime of neuronal net activity]. The problem of activity wave propagation in homogeneous isotropic neural net is analyzed. All the neurons of the net are considered in the subthreshold region as summators of synaptic currents with finite memory. The expression for synaptic current on a single neuron in the steady-state regime of the wave activity propagation is found. The equation for the velocity of the wave propagation is derived and analyzed; possible limiting correlations are investigated. The results are compared with the analysis of Beurle.


Effect of chronic marijuana administration of stages of primate sleep-wakefulness. The effects of the repeated administration of delta-9-tetrahydrocannabinol on sleep-wakefulness patterns were studied in adult male squirrel monkeys. The percent time spent in slow wave sleep was reduced with chronic treatment and failed to return to base line levels after 30 days of recovery. The amount of time spent in Stage 1 or drowsy state increased with repeated treatment and remained elevated through recovery. Changes observed in other stages of sleep-wakefulness were sensitive to repeated treatment with marijuana but were found to return to base line levels during recovery.


[Hydrolysis of the methyl esters of the N-arylsulfonyl derivatives of L-arginine by thrombin and trypsin]. Trypsin- and thrombin-catalysed hydrolysis of the Nalpha-aryl-sulfonyl-L-arginine methyl esters was studied. Michaelis constants and kcat were determined. The hydrolysis of the substrates by trypsin appears to depend on the polar nature of substitutes. The thrombin- catalysed reaction was believed to be sensitive to steric effects of the introducted groups.


[Conformational transitions in Na, K-dependent ATPase]. Conformational transitions in membrane preparations of Na, K-dependent ATPase were studied by means of mono-radical and bi-radical hydrophobic spine probes. A decrease in micro-viscosity of non-polar membrane regions in Na, K-ATPase preparation was observed under the formation of phosphorylated N, K-ATPase intermediate in the presence of ATP and Na+ ions. K+-catalysed dephosphorylation of the intermediate resulted in opposite changes. Bi-radical probes of a certain chemical structure turned to be the most sensitive indicators of conformational changes in membranes of Na, K-ATPase preparation. Na+-, K+- and ATP-induced conformational transitions are blocked with suabaine, a specific inhibitor of Na, K-ATPase, with Ca2+ oligomycin and p-chloromercuric benzoate (PCMB). K+-dependent transitions are more sensitive to oligomycin and PCMB as compared with those induced by ATP and Na+. Possible mechanisms of conformational transitions in Na, K-dependent ATPase are discussed.


Energy metabolism in human erythrocytes: the role of phosphoglycerate kinase in cation transport. Three models of disturbed erythrocyte metabolism, triose-depleted normal, phosphoglycerate kinase (PGK)-deficient, and pyruvate kinase (PK)-deficient cells, have been studied to examine further the role of PGK in erythrocyte cation transport. Sodium (Na-+) and potassium (K-+) transport were reduced only in cells fully depleted of triose. In such cells the PGK step presumably was inoperative due to total lack of substrate; 2,3-diphosphoglycerate (2,3-DPG) then became the sole substrate source for remaining steps in glycolysis. At increased intracellular Na-+ concentrations which normally stimulate transport and glycolysis, triose-depleted cells had marked impairment of cation transport and ouabain-inhibitable lactate and pyruvate production from 2,3-DPG. PGK-deficient cells and normal cells with high intracellular Na-+ concentrations had similar increases in transport and ouabain-inhibitable lactate production. PK-deficient cells with high intracellular Na-+ concentrations showed an appropriate increase in transport but less stimulation of lactate production. Transport was not related to total cellular adenosine triphosphate (ATP) concentration. These data suggested that normal coupled cation transport occurred despite diminished metabolite flow through PGK, as in PGK- or PK-deficient cells. Transport was diminished only in triose-depleted cells where metabolite flow through PGK was presumably absent. These data, therefore, support the concept that transport and glycolysis interact at the PGK step, although impairment of PGK must be profound before its effect on transport is evident.


The actions of dihydroxyphenylalanine and dihydroxyphenylserine on the sleep-wakefulness cycle of the rat after peripheral decarboxylase inhibition. 1. The actions of dihydroxyphenylalanine (DOPA) and dihydroxyphenylserine (DOPS) were assessed on the sleep-wakefulness cycle of male Wistar rats. 2. In comparative studies the extracerebral decarboxylase was inhibited with serinetrihydroxybenzylhydrazide (RO 4-4602) before injection of DOPA or DOPS. 3. DOPA (80-160 mg/kg, i.p.) with or without previous inhibition of the peripheral decarboxylase gave rise to an initial significant increase of slow wave activity, which may be related to a release of 5-hydroxytryptamine. 4. During the subsequent 8 h sessions, DOPA significantly decreased slow wave sleep and rapid eye movement sleep (REM) and increased wakefulness. 5. DOPS (80-160 mg/kg, i.p.) did not significantly modify the sleep-wakefulness cycle apart from a decrease of the latency for the first REM episode after 160 mg/kg in the RO 4-4602 pretreated animals.


Acetylcholine-sensitive cells in the caudal medulla of the rat: distribution, pharmacology and effects of pentobarbitone. 1. The distribution of cholinoceptive and non-cholinoceptive cells in various nuclei of the caudal medulla of the rat is described. 2. The nature of the responses of cells of the paramedian reticular nucleus and of the perihypoglossal nuclei to electrophoretically applied acetylcholine (ACh) was investigated. 3. In unanaesthetized decerebrate preparations ACh responses were usually of a "fast onset-fast offset" nature. Dihydro-beta-erythroidine was a more effective antagonist than atropine. 4. In rats anaesthetized with barbiturate nearly all the tach responses showed a slower onset and prolonged action. Atropine was the more effective antagonist. 5. The synaptic responses of cells of the paramedian reticular and perihypoglossal nuclei to stimulation of glossopharyngeal, superior laryngeal, lingual and hypoglossal nerves were investigated. It is concluded that ACh does not mediate the responses at the level of these nuclei.


Modulation by acetylcholine of adrenergic transmission in the rabbit ear artery. 1. Low concentrations of acetylcholine (4 times 10(-11) and 1 times 10(-10) M) increase the vasoconstrictor response of the isolated ear artery of the rabbit to stimulation of the periarterial sympathetic nerves. Higher concentrations (4 times 10(-8) M and greater) decrease the response. 2. Low concentrations of acetylcholine (1 times 10(-11) and 1 times 10(-10) M) increase the stimulation-induced efflux of radioactivity from artery segments previously incubated with [3H]-noradrenaline. Higher ocncentrations (3 times 10(-8) M and greater) decrease the efflux. 3. Neither atropine nor hexamethonium affects the facilitatory action of low concentrations of acetylcholine on adrenergic transmission in the rabbit ear artery. 4. Atropine antagonizes the inhibitory effect of higher concentrations of acetylcholine on adrenergic transmission.


The effects of cyclic adenosine 3',5'-monophosphate and guanosine 3',5'-monophosphate and theophylline on renin secretion in the isolated perfused kidney of the rat. 1. The influence of cyclic adenosine 3',5'-monophosphate (cyclic AMP), cylic guanosine 3',5'-monophosphate (cyclic GMP) and theophylline on renin secretion was examined in the isolated kidney of the rat perfused with Krebs dextran solution. 2. Neither cyclic AMP (10(-6) to 10(-4) M) nor dibytyryl cyclic AMP (10(-5) M) produced an increase in renin secretion. 3. Cyclic GMP and 8 Br-cyclic GMP caused a small rise in renin secretion in some experiments but this effect was independent of the dose and its physiological significance is uncertain. 4. Theophylline (10(-6) to 10(14) M) caused a significant elevation in renin secretion which was not blocked by (+)-propranolol. Theophylline with cyclic AMP or cyclic GMP did not produce an amplified effect. 5. Despite previous suggestions that cyclic AMP stimulated renin secretion, this could not be confirmed in the present preparation. Since there is no evidence that cyclic AMP or cyclic GMP (or their derivatives, dibutyryl cyclic AMP and 8 Br-cyclic GMP) enter the cells, it will be necessary to study their activity in isolated juxtaglomerular cells to define a possible role.


Interaction of competitive antagonists: the anti-curare action of hexamethonium and other antagonists at the skeletal neuromuscular junction. 1. In the rat isolated diaphragm preparation hexamethonium and other low potency competitive antagonists of acetylcholine (ACh), including gallamine and hyoscine butylbromide, reverse block by the potent antagonists tubocurarine, pancuronium and alcuronium. 2. In the presence of tubocurarine, hexamethonium increases the amplitude of the end-plate potential without increasing the quantal content. It enhances the response to ACh applied iontophoretically to the end-plate but does not enhance the response to ACh applied in the bath. 3. The anti-curare effect of hexamethonium is abolished in the diaphragm of the rat, guinea-pig and mouse by inhibitors of acetylcholinesterase. The effect is not observed in the indirectly stimulated toad sartorius muscle. 4. The effect is explained if tubocurarine does not dissociate appreciably in the time taken for ACh to achieve high occupancy of receptors, so that a fraction of receptors is completely excluded from occupation by ACh. Equilibration with hexamethonium reduces the fraction excluded by tubocurarine and the transmitter now competes with hexamethonium for more receptors and produces a larger response. 5. On the basis of this explanation the half-time for dissociation of tubocurarine must be about 1 millisecond. It follows that tubocurarine does not act competitively with ACh at synapses when transmitter action is sufficiently brief, and that its binding to the receptor is probably diffusion-limited.


Effect of infection with gonococci on myeloperoxidase activity of leucocytes. Lyosomal myeloperoxidase activity in human phagocytic leucocytes was stimulated by incubation with virulent (T1) and avirulent (T4) forms of Neisseria gonorrhoeae. The amount of activity, assayed by bacterial iodination (125-iodine) after 30 min. exposure to the pagocytes in the absence of serum, was about fifty times greater in cells infected with T4 strains. In the presence of heated human serum, or its IgG component, myeloperoxidase activity increased, but T1-Stimulated activity was significantly less than that of T4 and was not proportional to multiplicity of infection. From these results and from those of a previous study we conclude that T1 can stimulate leucocyte myeloperoxidase activity from an extracellular location, that for this activity a serum fraction is required, and that this may be a mechanism responsible for some of the killing of the membrane associated T1.


[Hypersomnia by isthmic lesion in cat. II. Neurophysiological and pharmacological study]. (1) The bilateral destruction of the region of the dorsal norepinephrine bundle in the isthmus or in the mesencephalon induces a significant increase of both paradoxical sleep (PS) (up to 400%) and of slow-wave sleep (SWS). The increase of PS lasts for 4-5 days while the increase of SWS lasts at least 8-10 days. The same phenomenon is observed after total hypophysectomy. (2) control lesions effectuated in the isthmus outside the area of the dorsal norepinephrine bundle do not affect significantly the slee-walking cycle. (3) pretreatment with p-chlorophenylalanine suppresses the increase of sleep if the lesion is effectuated at the time of maximum insomnia, while a significant increase of PS is still observed if the lesion is made at the time of the recovery of sleep. (4) the mechanisms of this hypersomnia are discussed in light of the biochemical results showing an increase of the biosynthesis of serotonin in the rostral raphe system. It is postulated that some neurons of the dorsal norepinephrine bundle might tonically control the activity of the raphe system.


Cholinergic agents affect two receptors that modulate transmitter release at a central synapse in Aplsia californica. A unitary, monosynaptic and presumably cholinergic EPSP recorded in cell R15 of the abdominal ganglion of Aplysia californica undergoes depression followed by facilitation when the presynaptic axon is repetitively stimulated at a rate of 1-3 pulses/sec.  During trains of stimulation which produced this sequence of phenomena, the effects of a large number of agents known to affect cholinergic transmission in other systems were studied.  The agents could be divided into 4 classes: (1) agents having no effect upon transmission at this cholinergic junction; (2) agents of a class typified by curare, which depressed all EPSPs of a train to the same extent, and which are believed to be acting in this system solely as competitive postsynaptic blockers; (3) agents typified by acetylcholine and carbachol (ACh class), which selectively depressed earlier EPSPs of a train more than later EPSPs and which appear to act by reducing the fractional release of transmitter; (4) agents typified by trimethidinium (trimethidinium class), which selectively depress later EPSPs of a train more than earlier EPSPs and which appear to act by reducing the rate of transmitter supply into the readily releasable pool.  Neither the ACh class nor the trimethidinium class produced these selective effects on different pulses in the train by changes in the postsynaptic membrane potential or membrane resistance.  Nor did they act by stimulating or inhibiting other recorded inputs onto R15.  Iontophoretic application of acetylcholine onto R15 indicated that the effect of trimethidinium could not be explained by an alteration in desensitization of a postsynaptic acetylcholine receptor.  The structural specificity of the presynaptic receptors mediating the action of the ACh and trimethidinium classes was demonstrated by the use of a larger number of structurally related compounds.


The influence of nerve growth factor, potassium depolarization and dibutyryl (cyclic) adenosine 3',5'-monophosphate on explant cultures of chick embryo sympathetic ganglia. Optimal nerve growth factor (NGF) and potassium concentrations for the culture of explants of single chick embryo sympathetic ganglia in Leighton tubes are described. NGF increases ganglionic monoamine oxidase (MAO; E.C.1.4.3.4) activity in a dose dependent fashion with maximum effects at a concentration of about 8 U/ml NGF. Peak activity compares closely with values seen in chick ganglia in vivo. Potassium-induced depolarization (45 mM K-+) also increases MAO activity; but the extent of the increase depends upon (NGF concentration, for there is little or no increase at high NGF concentrations. Dibutyryl adenosine 3',5'-cyclic monophosphoric acid (db cyclic AMP) can mimic the potassium-induced increase in MAO.


A study of inhibitory antagonism in cat visual cortex. Since there seems to be good evidence that GABA may act as an inhibitory neurotransmitter in the mammalian cortex, we tested the effects of an antagonist of GABA, namely the alkaloid bicuculine, on the response properties of visual cortex neurons, using a computer-controlled stimulus presentation system to assess quantitatively the changes in receptive field organization after the drug. Complex cells were most affected, increasing both evoked and spontaneous activity and losing some of their specificities for stimulus parameters such as orientation and direction. Hyper-complex cells lost their inhibitory flanks, responding equally well to long and short bars after the drug. Simple cells were the least affected, usually becoming somewhat depressed after the drug. Preliminary tests with another inhibitory amino acid antagonist, strychnine, showed that it excited simple cells, indicating that possibly more than one inhibitory transmitter is at work in the cortex. The results are discussed with relation to the synaptic anatomy of the cortex, and it is concluded that a class of stellate cells, using GABA, is a likely candidate for the transmitter of some intracortical inhibition.


The response of the immature fowl to multiple injections of adrenocorticotrophic hormone. 1. Treating chicks, from 1 d or 5 weeks of age with ACTH three times weekly for 3 weeks depresses growth and causes adrenal hypertrophy at dose rates of 30 IU/kg or more, and depletion of adrenal cholesterol (greater than 10 IU/kg). 2. Treating chicks five times weekly at a dose rate of 30 IU/kg was as effective as a dose of 120 IU/kg three times weekly. 3. Plasma glucose and FFA concentrations of chicks treated thrice weekly with 120 IU ACTH/kg for 3 weeks were within the normal range. 4. Rhode Island Reds were more sensitive to ACTH than Light Sussex. 5. Adrenal cholesterol stores in normal chicks show significant variations with season.


The effects of various treatments on the microbial flora of whole poultry carcasses with particular reference to Staphylococcus aureus contamination. Experiments are described in which the effects of dipping defrosted eviscerated poultry carcasses, for various periods, in acetic, lactic, succinic and citric acids, hypochlorite solution containing 200 ppm free Cl2 and in water at temperatures from 70 degrees C to 100 degrees C on the microbial flora were determined. 2. Hot (70 degrees C) water dipping for 2 min appeared to be the treatment most likely to succeed in reducing Staph. aureus, giving reductions of staphylococci of from 63 to greater than 99%, without completely eliminating other organisms. 3. A storage experiment showed this method to be relatively safe, provided the carcasses were refrigerated to less than 3 degrees C as soon as possible after treatment and held at this temperature until adequately cooked.


The effects of ascorbic acid deficiency on calcium and collagen metabolism in cultured fetal rat bones. The effects of ascorbic acid deficiency on growth and calcification of bone were studied in whole 18-day fetal rat radii and ulnae cultured in a chemically defined medium. Ascorbic acid deficiency decreased the formation of labeled hydroxyporline from labeled proline in both bone shafts and cartilage ends while incorporation of tryptophan was maintained. Dry weights and collagen content of bone and cartilage were decreased, but calcification was not affected. The optimun initial concentration of ascorbic acid for collagen synthesis was 200 mug/ml. The effect of ascorbic acid was not antagonized by glucoascorbic acid or replaced by dithiothreitol. Decreased collagen synthesis in ascorbic acid deficiency could not be ascribed to loss of available peptidyl proline hydorxylase. Formation of underhydroxylated collagen and its release into the medium accounted for much of the decrease in hydroxylated collagen in ascorbic acid deficient bones. Nevertheless, the total newly synthesized collagen, as measured by collagenase digestion, was still decreased. Similar effects were exerted by alpha, alpha'-dipyridyl which also inhibited general protein synthesis. Ascorbic acid did not stimulate proline incorporation into collagen in the presence of alpha, alpha'-dipyridyl.


Mitochondrial granules in human osteoblasts with a reference to one case of osteogenesis imperfecta. Electron-dense granules in mitochondria from prenatal human osteoblasts, postnatal human osteoblasts, and from osteoblasts derived from a child with osteogenesis imperfecta congenita are described. The mitochondrial granules were of about 600 A in diameter and were attached to the mitochondrial cristae. Sections of mitochondria from prenatal osteoblasts showed an average number of 10 granuales per mitochondrial section, whereas sections of mitochondria of postnatal osteoblasts showed only occasionally 1-2 granules per mitochondrial section. Mitochondria from osteoblasts derived from the child with an untreated osteogenesis imperfecta congenita showed an average number of 10 granules per mitochondrial section.


Relationship between RNA polymerase and protein kinase activities in rat mammary gland nuclei. 1. Extracts from rat mammary gland nuclei contain cyclic AMP -independent protein kinases which phosphorylate casein rather than histone. 2. A major increase in nuclear protein kinase activity occurred during late pregnancy and was maintained with the onset of lactation. 3. Two major peaks of activity were resolved by chomatography of nuclear extracts on DEAE-Sephadex; the first (NI) appeared in the void volume and the second (NII) was eluted by 0.05-0.12 M ammonium sulfate. Several other regions of lesser activity were also present. 4. Protein kinases in the cytosol 105,000 times g supernatant, precipitated by 70 percent ammonium sulfate, dialyzed against buffer, and chromatographed on DEAE-Sephadex, yielded a major components phosphorylated histone in preference to casein, and this was stimulated by cyclic AMP if histone was the substrate, but only the first (void volume) fraction was cyclic AMP-dependent when casein was used. 5. Most of RNA polymerases Ib and II, derived from the nucleolus and nucleoplasm, respectively, appeared in column fractions distinct from those containing the major NI and NII protein kinases. 6. Cyclic AMP altered the amount of RNA product synthesized by polymerases Ib and II, but the explanation for this is unknown. Due to their elution profiles and cyclic AMP-independence, protein kinases NI and NII are excluded from playing a catalytic role in these effects; participation of quantitatively minor protein kinases which co-elute with polymerase Ib and II is not yet excluded.


Effects of intraventricular injections of N-6,O-2-Dibutyryl adenosine 3,5-monophosphate on the rat brain cholinergic system. Intraventricular injections of dibutyryl cyclic AMP effected increases of acetylcholine in the cerebral cortex-striatal and subcortical regions. No increase in choline acetyltransferase activity was observed; however, there was a significant decrease in acetylcholinesterase activity in the subcortical region.


Observations on abortions in cattle: a comparison of pathological, microbiological and immunological findings in aborted foetuses and foetuses collected at abattoirs. Fifty nonaborted and 50 aborted bovine foetuses were examined utilizing histology, immunoelectrophoresis, bacteriology and the fluorescent antibody technique. Lesions were observed in 12 of the nonaborted foesuses and in four of these immunoglobulins were demonstrated. In addition, two of the nonaborted foetuses had immunoglobulins in the absence of observed lesions. Lesions were observed in 48 of the aborted foetuses and immunoglobulins were detected in 22 of these. An etiological diagnosis was arrived at in 24 of the 50 aborted foetuses. The tissues most frequently observed to have lesions of diagnostic significance were eyelid, intestine, liver, lung and placenta. Intestinal lesions were observed in several foetuses in association with a variety of agents including infectious bovine rhinotracheitis. Foetuses diagnosed as aborting because of mycotic infection consistently displayed lesions in their eyelids. The value of taking eyelid sections in cases of suspected mycotic abortions, the significance of foetal intestinal lesions, the evaluation of abomasal aspirates and the diagnostic importance of immunoglobulin determinations in aborted foetuses are discussed.


Electron microscopic findings of cells with inclusion bodies in experimental hemorrhagic enteritis of turkeys. The spleen, liver, bone marrow and intestines of two turkeys in which hemorrhagic enteritis of turkeys was experimentally reproduced were examined electron-microscopically. Intranuclear inclusion bodies as described in a previous report were found in all the tissues examined. These occupied most of the area in affected nuclei and were composed of viral particles with morphological characteristics of an adenovirus. The cells with the inclusions were divided into two types of cells, immature and reticular cells. There was some variety in the stage of differentiation of the former cells. As the viral particles developed the cells degenerated and disintegrated. A few particles had been released into the cytoplasm of the degenerated cells but no particles were present in intercellular spaces.


Effect of pasteurization and evaporation on foot-and-mouth disease virus in whole milk from infected cows. The effects of pasteurization and evaporation on foot-and-mouth disease virus in whole milk from infected cows obtained one day postinoculation were studied. Virus survived the heating of milk at high temperature-short time pasteurization at 75 degrees C for 15-17 seconds. In addition, virus from infected milk survived heating at 80 degrees C for the same time. Infective virus also survived in the pasteurized milk after evaporation at 65 degrees C to 50% of the original volume. The bovine udder was found to be highly susceptible to foot-and-mouth disease virus replication. Seven log10 plaque-forming units/ml of virus were recovered in whole milk 24 hours postinoculation, and decreasing titers were recovered for as long as seven days postinoculation.


Isolation of equine herpesvirus type 1 from a horse with an acute paralytic disease. A Standardbred mare became paralyzed shortly after showing signs of an upper respiratory infection. The mare was euthanized and equine herpesvirus type 1 was isolated from the brain and spinal cord.


Unique immunobiological aspects of head and neck squamous carcinoma. The immune reactivity of patients with squamous carcinoma of the head and neck region was compared with that of patients with squamous carcinoma of the female pelvic organs and patients with adenocarcinomas, melanomas, and sarcomas. The reactivity of patients with clinically localized tumors was compared with that of cured patients and a large normal population. Patients with squamous carcinomas of the head and neck region and female pelvic organs displayed higher incidences of impaired cellular immune competence than patients with malignancies of other histologic types. Among cured patients, those previously treated for squamous carcinoma were unique in that they displayed cellular immune defects and serum suppressants of in vitro immune reactivity similar to tumor-bearing patients. Antibodies to herpes simplex virus nonvirion antigen was found in high incidence only among patients with squamous carcinomas, and the incidences in tumor-bearing and cured patients were similar. The persisting immune defects in cured squamous carcinoma patients give importance to the determination of the role of genetic and environmental factors in the induction of these tumors. The associations made between herpes virus and squamous carcinoma offer an explanation for the defects and also an approach for the definition of the factors involved in squamous carcinogenesis. The findings are clinically relevant to the isolation of population groups at high risk for the development of squamous carcinoma, as a rational basis for the development of prophylactic measures, and as a basis for more effective therapeutic regimens.


Adenocarcinoma of the larynx.

Adenocarcinoma of the larynx. Adenocarcinomas make up less than one per cent of laryngeal malignancies. Nonspecific adenocarcinomas and adenoid cystic carcinomas comprise most of the glandular neoplasms. Adenoid cystic carcinoma has a slight preponderance in females, and often follows a long course similar to this tumor when it occurs in salivary glands. The nonspecific adenocarcinomas occur predominately in males and have a low survival with most patients dying within two years. Less common adenocarcinomas include oncocytoid carcinoma, mucoepidermoid carcinoma, and metastatic adenocarcinoma.


Adenocarcinoma of the larynx.

Adenocarcinoma of the larynx. Nine patients with primary adenocarcinoma of the larynx representing 0.1 per cent of 888 patients with primary laryngeal malignancy treated between 1955 and 1971 are reported. Following surgical therapy, four patients are alive without disease and five patients are dead due to their disease. Survival time was not related to histopathologic type, stage of disease, or mode of therapy. We cautiously advocate the use of the most conservative surgery feasible as the primary treatment of this poorly understood and highly fatal disease.


Comparative pharmacological studies on butriptyline and some related standard tricyclic antidepressants. Butriptyline was compared with imipramine and other tricyclic antidepressants for its ability to modify: (a) contractions of the cat nictitating membrane induced by noradrenaline (NA) and 5-hydroxytryptamine (5-HT), (b) the adrenergic neuron blocking action of guanethidine in the guinea pig vas deferns, (c) the rabbit's electroencephalogram (EEG) and physostigmine arousal, and (d) the sleep pattern of the rat. Imipramine and amitriptyline potentiated the NA and 5-HT effects on the nictitating membrane and antagonized the inhibitory actions of guanethidine in the guinea pig vas deferens, whereas iprindole and butriptyline were ineffective. These results are consistent with the ability of these drugs to block the neuronal uptake of catecholamines. Butriptyline was a potent blocker of the arousal reaction induced by physostigmine. Butriptyline (20--30 mg/kg) and amitriptyline (10--20 mg/kg) reduced rapid eye movement sleep with a conmitant increase in non-rapid eye movement sleep. This may be a reflection of the dual activity observed in the clinic with these compounds, namely, antidepressant and antianxiety effects.


Response of hepatic carbohydrate and cyclic AMP metabolism to cadmium treatment in rats. Daily intraperitoneal injection of cadmium chloride (0.25 or 1 mg/kg) for 21 or 45 days into rats significantly stimulated the activities of hepatic pyruvate carboxylase, phosphoenolpyruvate carboxykinase, fructose-1, 6-diphosphatase, and glucose-6-phosphatase, increased the concentrations of glucose and urea in the blood, and decreased the levels of glycogen in the liver. Whereas chronic cadmium treatment failed to alter adenosine-3',5'-monophosphate phosphodiesterase (phosphodiesterase) activity, the endogenous levels of cyclic AMP (cAMP) and the activity of basal- and fluoride-stimulated forms of hepatic adenylate cyclase (AC) were markedly increased in cadmium-injected animals. Treatment with the higher dose (1.0 mg/kg) of cadmium chloride for 45 days produced greater metabolic alterations in hepatic tissue than those seen with the lower dose (0.25 mg/kg) given for a shorter period of time (21 days). Discontinuation of cadmium administration for 14 days in rats previously injected with cadmium chloride (1 mg/kg per day) for 21 days, failed to reverse the observed changes in hepatic cAMP or carbohydrate metabolism. A similar persistence of metabolic alterations was noted in rats treated with cadmium (1 mg/kg per day) for 45 days and subsequently maintained without additional treatment for 28 days. Administration of an acute dose of cadmium chloride (60 mg/kg) decreased hepatic phosphodiesterase activity and glycogen content 1 h after the injection. In addition, acute cadmium exposure increased blood glucose, serum urea, and hepatic cAMP levels, and produced an augmentation of basal- and fluoride-activated AC. However, the activities of various hepatic gluconeogenic enzymes remained unaffected in animals given an acute dose of cadmium chloride (60 mg/kg). Data provide evidence that suggests that the gluconeogenic potential of liver is markedly enhanced following chronic exposure to cadmium and that the cadmium-induced changes in carbohydrate metabolism may be associated with an enhanced synthesis of cAMP. In addition, the present study shows that the cadmium-induced metabolic alterations persist even after the cessation of cadmium treatment for a period of 28 days.


Further observations on the response of the glomerular filtration rate to glucagon: comparison with secretin. Glucagon causes marked elevations of glomerular filtration rate (GFR) in dogs when administered intravenously (i.v.) in small doses. The associated natriuresis is thought to be entirely due to increments in the filtered sodium load. In this study, renal denervation, thyroparathyroidectomy, and blockade of cholinergic, alpha- and beta-adrenergic, dopaminergic and histaminergic receptors did not prevent the usual glucagon-induced elevations of GFR or rate of sodium excretion (UNaV). This effect of glucagon was not mediated through the release of cyclic AMP, or by plasma compositional changes of Ca-2+, K+, or amino acids. Pure porcine secretin, in doses of 5--10 mug/min delivered either i.v. or into the left renal artery did not alter GFR; clearance of the p-aminohippurate (CPAH) or UNaV in either hydropenic or saline-loaded dogs. Nor did this polypeptide, structurally very similar to glucagon, abolish the effect of glucagon on GFR. It did, however, partially inhibit the glucagon-induced natriuresis, presumably by preventing a previously undetected glucagon action on tubular reabsorption of sodium.


Conjunctival papillomas in norther Canadian natives. Cases of conjunctival papilloma evaluated at a hospital that acts as a referral centre for native people (Indian and Inuit) from northern Alberta and the Northwest Territories were compared with those from other hospitals in the same region whose referral base is primarily non-native. When the data were standardized for population unitsof 100,000 the tumours appeared to be occurring with a greater frequency among lnuit of the Western Arctic. These findings are supported statistically, although it is possible that bias may have been introduced by the small number of cases and other related factors.


Structural studies of the capsular polysaccharide of Klebsiella type 81. The structure of the capsular polysaccharide from Klebsiella Type 81 has beeninvestigated. Methlylation analysis, uronic acid degradation, Smith degradation, andgraded acid hydrolysis were the principle methods used. Theanomeric nature of theglycosidic linkages was determined by characterization of fragments obtained from thevarious methods of degradation used. One of the L-rhamnosidic linkages was notpresent in any of these fragments, but is assumed to be an alpha-linkage from consideerationsof optical-rotation data. These studies show that they polysaccharide consits  of thefollowing hexasaccharide repeating-unit: yield2)-alpha-L-Rhap-(1yields3)-alpha-L-Rhap-(1-yields4)-beta-D-GlcAP-(1-yields2)-alpha-L-Rhap-(1-yields3)-alpha-L-Rhap-(1-yields3)-beta-D-Galp-(1-yields.


Structural studies of the capsular polysaccharide of Klebsiella type 59. The structure of the capsular polysaccharide from Klebsiella type 59 has been investigated by methylation analysis, a modified Smith-degradation procedure, and uronic acid degradation followed by oxidation and elimination of the substituents of the oxidized residue. The oligomeric fragments produced by these degradative procedures were isolated and characterized. O-Acetyl groups were identified and their position determined. The polysaccharide consists of the following pentasaccharide repeating-unit (dotted lines indicate that only some of the residues carry the O-acetyl substituent). See article.


Structural studies of the capsular polysaccharide of Klebsiella type 28. The capsular polysaccharide from Klebsiella type 28 has been studied by methylation analysis, a modified Smith-degradation procedure, and uronic acid degradation with subsequent oxidation and elimination of the substituents of the oxidized residue. The polysaccharide contained the hexasaccharide repeating-unit shown below. The terminal D-glucopyranose residue was hydrolysed by emulsin, indicating a beta linkage. The anomeric natures of other glycosidic linkages were determined by characterization of fragments obtained during the degradative studies. The D-galactopyranose residue was not present in any fragment, but is assumed to be alpha-linked from optical-rotation considerations. (see article)


Acyloxy neighboring-group participation in the acid-catalyzed cleavage of methyl 2,3-anhydro-beta-D-ribofuranoside. The reaction of methyl 2,3-anhydro-beta-D-ribofuranoside with hydrogen bromide in an acetic acid-acetic anhydride solution leads to the formation of methyl 2,3-di-O-acetyl-5-bromo-5-deoxy-alpha,beta-D-xylofuranoside. Similar treatment of methyl 2,3-anhydro-5-O-benzoyl-beta-D-robofuranoside provided methyl 2-O-acetyl-3-O-benzoyl-5-bromo-5-deoxy-alpha,beta-D-xylofuranosides. The position of halogen substitution was ascertained by hydrogenolysis to the resultant 5-deoxy sugars, which were characterized by their n.m.r. spectra. Confirmation of the structural assignment for methyl 2-O-acetyl-3-O-benzoyl-5-deoxy-alpha,beta-D-xylofuranoside was obtained by synthesis from 1,2-O-isopropylidene-alpha-D-xylofuranose. The formation of the 5-bromo derivatives under the reported conditions probably occurred through the intermediacy of the 3,5-acyloxonium ions. Similar conversions were observed when the starting compound was treated with hydrogen chloride, acetyl bromide, or acetyl chloride in acetic acid-acetic anhydride solutions.


In vitro polyoma DNA synthesis: self-annealing properties of short DNA chains. Short DNA chains, isolated from in vitro pulse-labeled replicating polyoma DNA, exhibit some degree of self-complementarity (28% resistance to S1 nuclease after self-annealing to plateau levels). This level of self-annealing is not increased if short DNA chains present as free single-stranded DNA after extraction are included in the hybridization, excluding a selective loss of chains from one side of the growing fork and supporting a semi-discontinuous mode of chain growth. This mode also applies to restricted synthesis conditions under which a relative excess of short chains is made, since no increase in the self-annealing of such short chains is observed. The self-annealing that can be measured is higher for the faster sedimenting portion (46%) of the short DNA chains than for the slower sedimenting portion (18.5%), indicating that it is most likely due to contaminating continuously growing strands from the other side of the fork. High self-annealing values (up to 60%) are obtained if virus stocks generating defective DNA are used for infection. Restriction endonuclease (Hpall) characterization of such DNA shows evidence for the presence of multiple origins of replication. One of several possible mechanisms is discussed by which replicating defective DNA can generate self-complementary short chains despite a semi-discontinuous mode of replication.


Effect of a single treatment with the alkylating carcinogens dimethynitrosamine, diethylnitrosamine and methyl methanesulphonate, on liver regenerating after partial hepatectomy. I. Test for induction of liver carcinomas. A  single injection of dimethylnitrosamine (DMN), 12.0-15.6 mg-kg, given to 100 g female rats 24 h after partial hepatectomy, induced hepatocellular carcinoma. No animals receiving DMN without partial hepatectomy developed liver carcinomas. Previous evidence had suggested that the incidence of tumours was highest when DMN was administered during the wave of DNA replication which follows partial hepatectomy. The present experiments made this suggestive evidence statistically significant. A single treatment with diethylnitrosamine (DEN) induced liver cell cancer when given to intact or to partially hepatectomised rats. No tumors developed when another alkylating carcinogen, methyl methanesulphonate (MMS), was administered after partial hepatectomy. The significance of these results in relation to the mechanism of initiation of carcinogenesis is discussed.


Effect of a single treatment with the alkylating carcinogens dimethylnitrosamine, diethylnitrosamine and methyl methanesulphonate, on liver regenerating after partial hepatectomy. II. Alkylation of DNA and inhibition of DNA replication. Experiments were carried out to determine whether replication of alkylated DNA could be involved in the initiation of hepatocellular carcinoma which results from a single administration of dimethylnitrosamine (DMN) given after partial hepatectomy. The incidence of tumours is higher when DMN is given during the wave of DNA synthesis induced by the operation than when given in the early prereplicative stage. Therefore the alkylation of DNA in the regenerating liver by DMN given at these times and the effect of DMN on DNA synthesis were investigated. The extent, duration and pattern of alkylation of DNA, including the formation of 0-6-methylguanine, were similar whether DMN was given in the early pre-replicative stage (6 h after the operation) or during the period of DNA synthesis (at 24 h). DMN given a 6 h very greatly reduced the wave of DNA replication which would otherwise have ensued. When given at 24 h, by which time DNA synthesis was already taking place, DMN reduced the rate of incorporation of (-3H)thymidine after 1-2 h delay. However, in neither case was DNA synthesis reduced to the level occurring in normal intact liver.  Treatment with diethylnitrosamine (DEN) at 6 h or at 24 h had a similar effect to DMN on the wave of DNA replication induced by partial hepatectomy. Methyl methanesulphonate (MMS given in the early pre-replicative stage delayed the wave of DNA synthesis by about 8 h, but when it did take place the extent of synthesis was as great as in untreated animals. When given during the period of DNA replication, MMS rapidly reduced the rate of synthesis. As in the case of the nitrosamines, synthesis was not reduced to the level occuring in normal intact animals. The difference from the nitrosamines lies in the nature of the alkylated bases formed in DNA. The fact that a single treatment with DMN induces cancer in partially hepatectomised animals but not in intact adult animals is not considered to be due to a gross difference in the nature of the alkylation of DNA. The experiments described support the concept that replication of DNA containing bases which are likely to mispair during replication may be necessary to 'fix' the lesion and thus cause a permanent inheritable change in the genetic material.


Effect of adenosine 3',5'-cyclic monophosphate on the RNA and DNA synthesis and cell proliferation of rat hepatocytes in primary culture: a radioautographic study. The effect of a 24-h treatment with various doses (from 1.5-10-minus 8 to 3.0-10-minus 3 M) of adenosine 3',5'-cyclic monophospahte (cAMP) on morphometric parameters, [5--3H]uridine radioactivity concentration (URC), [methyl--3H]thymidine [Me--3H]-Tr) labelling index per hour (L.I./h) and per cent mitotic index (M.I.%) of young rat differentiated hepatocytes in primary tissue culture were investigated by morphometric and radioautographic methods. In such cells cAMP was found to induce: (1) a reduction of the apparent surface area (ASA) of total nucleoli, karyoplasm and cytoplasm; (2) significant increases in URC of all the subcellular compartments at all the dosages employed (only cAMP at 1.5-10-minus 8 M did not change karyoplasmic and cytoplasmic URC values); (3) marked increments in [Me--3H]Tdr L.I./h and M.I.% from the lowest dose up to 1.5-10-minus 4 M; at higher doses the L.I./h and M.I.% were less stimulated or approached control values. In cultured rat hepatocytes, adenosine-5'-phosphate (5'-AMP) (1.5-10-minus 4 M per 24 h) increased the karyoplasmic and total cell ASA, the lone total nucleolar URC and both the L.I./h and M.I.%. However, these metabolic effects were significantly less intense than those elicited by isomolar cAMP. Theophylline (Theo) (5.5-10-minus 5 M per 24 h) reduced the in vitro rat hepatocyte total nucleolar ASA but affected neither other morphometric nor any of the URC values. The same dose of Theo plus cAMP (1.5-10-minus M) had no morphometric effect but significantly increased the URC values of all primary rat hepatocyte compartments. Actinomycin D (DAct) (0.1 mug/ml per 24 h) plus cAMP (1.5-10-minus 4 M) decreased the cultured rat hepatocyte total nucleolar ASA but enlarged that of karyoplasm and cytoplasm and, further, markedly curtailed all the compartmental URC values. These data support the hypothesis that cAMP amplified the template activity of the liver chromatin and accelerates the flow of differentiated primary young rat hepatocytes into the various stages of the mitotic cell cycle.


[Different action of alpha-methyl-DOPA on sleep and labyrinth learning in 2 inbred strains of mice]. In two inbred mice strains C 57 BR and C 57 BL/6 presenting the same type of sleep, but a different capacity of learning, Alpha-Methyl-Dopa (100 mg/kg) injected after every session, suppresses paradoxical sleep completely for 9 to 11 h. Maze-learning performance is retarded in C 57 BR mice, but facilitated in C 57 BL/6.


Acute subendocardial myocardial infarction in patients. Its detection by Technetium 99-m stannous pyrophosphate myocardial scintigrams. Eighty-eight patients admitted to a coronary care unit with chest pain of varying etiology but without ECG evidence of an acute transmural myocardial infarction had myocardial scintigrams using technetium-99m stannour pyrophosphate (99m-Tc-PYP). Seventeen of these patients had ECG and enzymatic evidence suggestive of acute subendocardial myocardial infarction. In each of these the scintigrams were postivie demonstrating increased 99m-Tc-PYP uptake either in a faintly but diffusely positive pattern or in a well-localized strongly positive one. The remaining 71 patients did not evolve ECG or enzymatic evidence of acute myocardial infarction. In each of these patients the myocardial scintigram was negative. Thus 99m-Tc-PYP myocardial scintigrams are capable of identifying the presence of acute subendocardial myocardial infarction in patients. The absolute frequency with shich subendocardial myocardial infarction can be recognized utilizing this technique will have to be established in a larger number of patients in the future.


Possible identity between the hepatoma alkaline phosphatase and an isozyme of human amniotic membrane (FL cells). The electrophoretically faster migrating alkaline phosphatase isozyme in FL cells derived from the human amniotic membrane was similar in some of the enzymatic and immunochemical properties to the variant alkaline phosphatase which occurs in serum and cancerous tissue of some patients with hepatocellular carcinoma. These facts suggest that the hepatoma-associated alkaline phosphatase may be the resurgence of the isozyme originally present in the amniotic membrane.


Serum lipid and lipoprotein profile in school children from a rural community. In 447 school children 5--16 years old, in one community, serum lipid and lipoprotein levels were estimated, including total cholesterol, triglycerides, beta- and pre-beta-lipoprotein cholesterol, alpha-lipoprotein cholesterol, and the serum concentrations of beta-, pre-beta-, and alpha-lipoproteins. Mean serum cholesterol of the entire group was 188 mg/100 ml, and alpha-lipoprotein cholesterol constituted 68 mg/100 ml of this total. The levels of serum triglycerides (48.9 mg/100 ml) and pre-beta-lipoprotein (39.8 mg/100 ml) are low in children in comparison to adults. Except for pre-beta-lipoprotein levels, there was very little correlation between age and all lipids and lipoproteins in this age span. Slight differences in values were noted at certain age intervals, but there were no significant differences in values between sexes. These observations emphasize the need for evaluating lipid abnormalities in individual children by quantitating specific serum lipoproteins.


Delayed adolescence.

Delayed adolescence. Delayed adolescence has several causes. Most frequent is the physiological or constitutional (hereditary) delay of growth and adolescence. This is a normal variation of growth and development, with growth, bone age and puberty retarded in a harmonious way. It carries a good prognosis with late but normal puberty and late catch-up growth leading to normal adult height. It manifests itself long before puberty by short stature and retarded bone age. True endocrine defects with permanent hypogonadism (hypothalamic-pituitary deficiency of the gonadotropinds and primary gonadal failure) are rare. Differential diagnosis before puberty is not always possible on clinical grounds alone. The most useful laboratory test consists of the LH-RH test. The i.v. injection of the recently introduced hypothalamic LH-releasing hormone, LH-RH, is followed by an age-dependent increase of the plasma gonadotropinds LH and FSH. This test allows differentiation, before puberty, between constitutional delay of growth and adolescence with a normal response for bone age, true hypothalamic-pituitary insufficiency with no response, and primary gonadal failure with an increased response. True hypogonadism requires permanent sex hormone replacement therapy. Constitutional delay of growth and adolescence in boys may present a psychosocial indication for temporary hormone therapy with testosterone.


Metastatic malignancy of the hand.

Metastatic malignancy of the hand. Metastatic malignancies of the hand are infrequent and seldom initially diagnosed and often taken for infections. They usually occur at the late stage of metastatic generalization and indicate an ominous prognosis and justify surgical amputation. A case of metastasis to the distal phalanx of the left ring finger of a woman who had undergone radical mastectomy for breast cystosarcoma phylloides is reported. The evolution of this lesion, initially taken for a whitlow, lasted 45 days and eventually was treated by amputation. A review of literature confirms the rarity of the lesion, and reveals the pathological, clinical and radiographic characteristics.


Pyrophosphate and diphosphonates in skeletal metabolism. Physiological, clinical and therapeutic aspects. Pyrophosphate and diphosphonates produce striking results on calcium metabolism in experimental animals and man. Compounds containing P-O-P- bonds (e.g. inorganic pyrophosphate [PP-ii1 or P-C-P bonds (diphosponates) inhibit both the formation and dissolution of calcium phosphate crystals in vitro. PP-i may have a physiological function in regulating calcification and bone turnover, and obnormalities in its metabolism may occur in some human diseases notably hypophosphatasia and pseudogout. Diphosphonates inhibit ectopic calcification, and slow down resorption and bone turnover in several experimental systems in vivo. They have helped in studies of various aspects of the regulation of calcium metabolism. The diphosphonate, disodium ethane-1-hydroxy-1,1-diphosphonate (EHDP) has been shown in clinical studies to be effective against ectopic calcification particularly in myositis ossificans progressiva and in disorders of increased bone resorption such as Paget's diseases and some types of osteoporosis. -99mTechnetium complexes of EHDP, PP-i and other polyphosphates have also recently been used successfully as bone scanning agents.


Leoprosy- clinical aspectos of nerve involvement. Leprosy is the cause of the commonest peripheral neuropathy. The predilection of Mycobacterium leprae for nerve tissue accounts for the clinical features that are most dreaded and most characteristic of the disease. Were it not for the progressive destruction of peripheral nerve trunks and the consequences of this, leprosy would largely remain a cutaneous condition of cosmetically unsightly hypopigmented or erythematous areas and aggregations of nodular thickenings. The neurologic damage in leprosy is confined to postganglionic changes. The central nervous system is protected, although rarely in experimental situations organisms have been reported in cerebral tissues.


Leprosy--histopathologic aspects of nerve involvement. The most striking single feature of the clinical manifestations of leprosy is the very wide range of appearances shown by the skin lesions. These include the vague, hypopigmented macules of indeterminate leprosy; the large, sharply defined hypopigmented anaesthetic lesions of tuberculoid leprosy; the nodules and diffuse infiltration of lepromatous leprosy; and a wide range of plaques and annular lesions of the intermediate (borderline or dimorphous) types of disease. From superficial appearances it would be impossible to say that these were manifestations of the same infection. Moreover, histologically there is a similar wide range of appearances, including minimal lymphocyte infiltration around the neurovascular bundles in indeterminate leprosy; epithelioid granulomata with Langhans' giant cells and virtually no bacilli to be seen in tuberculoid cases; and in lepromatous leprosy, histiocytic infiltration occupying the whole thickness of the dermis, with massive parasitisation by Mycobacterium leprae, of which there may be up to 10-9 per gram of tissue. Strain differences have been recognised, but appear not to be responsible for this remarkable range of manifestations; they are caused by variations in the soil, not in the seed. Leprosy is almost unique among infectious diseases in the importance that host factors play in its development. The different clinical appearances shown by different types of leprosy can be graded into a continuous series, forming a spectrum from the single or scanty lesions of tuberculoid leprosy to the total body involvement o lepromatous; and the patient with leprosy can indeed truly be said to wear his skin lesions like medals displaying his capacity to resist Myco. leprate. The histopathologic features of the developed disease have a similar spectrum, from an epithelioid granuloma to infiltration with histiocytes that are full of bacilli and are ultimately converted to foam cells (Virchow cells)...


Upper respiratory infection of lactating sows with transmissible gastroenteritis virus following contact exposure to infected piglets. Ten breeding sows were left in direct contact with their newborn piglets that had been experimentally infected with transmissible gastroenteritis (TGE) virus. All sows became infected with the virus. The sows developed fever and showed mild clinical signs of the disease for a few days. The sows excreted virus in the nasal secretion, feces, and milk during the acute febrile phase of illness. Virus was isolated from the nasal secretion of one sow as early as 20 hours after contact exposure to the infected piglets. At necropsy, the virus was more frequently isolated from the tissues of the upper respiratory tract than from small intestines; this finding indicated that the TGE coronavirus replicated in the upper respiratory tract and induced an acute respiratory infection in susceptible adult swine. Neutralizing antibody was present in the sera 8 sows after 12 to 36 days during the convalescent period. From these results, we conclude that susceptible sows in direct contact with ill piglets can become infected and by excreting virus can serve as a source of TGE virus for other susceptible pigs on the premises.


Adenoviral pneumonia in a foal.

Adenoviral pneumonia in a foal. A three-week-old Arabian filly was admitted to the Large Animal Hospital with a respiratory disorder and died despite symptomatic treatment. The necropsy lesions were suggestive of viral pneumonia. An equine adenovirus were isolated from nasal and pharyngeal swabs and from several tissues after death. Typical adenovirus virions were demonstrated by electron microscopy.


Red cell enzymes.

Red cell enzymes. As compared to other cells of the body, the mammalian red cell has one of the simplest structural organizations. As a result, this cell has been extensively used in studies involving the structure, function, and integrity of cell membranes as well as cytoplasmic events. Additionally, the metabolic activities of the red blood cell are also relatively simple. During the past quarter century or so, an ocean of knowledge has been gathered on various aspects of red cell metabolism and function. The fields of enzymes, hemoglobin, membrane, and metabolic products comprise the major portion of this knowledge. These advances have made valuable contributions to biochemistry and medicine. Despite these favorable aspects of this simple, anucleated cell, it must be conceded that our knowledge about the red cell is far from complete. We are still in the dark concerning the mechanism involved in several aspects of its membrane, hemoglobin, enzymes, and a large number of other constituents. For example, a large number of enzymes with known catalytic activity but with unknown function have eluded investigators despite active pursuit. This review will be a consolidation of our present knowledge of human red cell enzymes, with particular reference to their usefulness in the diagnosis and therapy of disease. Owing to the multitude of publications by prominent investigators on each of the approximately 50 enzymes discussed in this review, it was impossible to cite a majority of them.


Contraceptive steroid binding to the human uterine progesterone-receptor. Using an equilibrium dialysis method, the competition was investigated between tritated-progesterone and a range of synthetic  steroids for binding to 20,000 g. supernatants of human uterine endometrium and myometrium. Both types of uterine tissue showed similar patterns of progestogen binding and probably contain similar, or even identical, progesterone receptors. The gonane progestogen norgestrel binds strongly to the uterine receptors, but only the d-isomer is active. All three pregnane progestogens used in this study (chlormadinone acetate, medroxprogesterone acetate, megestrol acetate) also showed significant binding to receptors, but for the estrane progestogens, norethisterone was the only compound to show a high binding capacity. Other estranes (ethynodiol diacetate, lynestrenol, norethisterone acetate, norethynodrel) showed insignificant binding to the receptors and probably require prior metabolic activation to norethisterone before they can induce progestogenic effects.


Examples of adolescent group homes in alliance with larger institutions. Use of the group home in combination with the services of larger institutions for disturbed adolescents has many treatment advantages if the home program is tailored to the specific needs of the youngsters served.


Perianal Paget's disease: report of two cases. A patient with extensive perianal extra-mammary Paget's disease was treated by wide local excision of the perianal skin. No underlying associated adenocarcinoma was identified, and the patient is clinically free of disease five years after treatment. A second patient had a small focus of Paget's disease confined to a hemorrhoid. The necessity for histologic evaluation of all anal lesions is emphasized.


A pharmacological model of the pathophysioloyg of schizophrenia. A pharmacological model of the pathophysiology of schizophrenia based upon receptor site theory has been presented. This model proposes that, like any neuronal function, schizophrenia is triggered by the action of a small molecule at a specific receptor site. This interaction elicits the abnormal behavior. The model permits analysis of all existing theories of the pathogenesis of schizophrenia and furthermore makes it possible to view the pathophysiologic implications of these theories. It also mkaes it possible to understand the mechanism of action of neuroleptic agents used in the treatment of schizophrenia, regardless of which theories of pathogenesis are valid.


[Nerve lesions following total hip replacement and other hip-joint operations]. Nerve lesions occurred in 17 patients who had undergone hip-joint operations (especially total replacement). The femoral nerve was most commonly affected, followed by sciatic and gluteal nerves, with combined pareses in six. As differential diagnosis from pseudoparesis is not always possible, electromyography should be performed in all cases. Stretching and/or squeezing are the most important causes. The operation results are significantly impaired by such  nerve lesions. However, long-term results are usually satisfactory in the majority of cases. Early diagnosis and treatment are important in ensuring good therapeutic results.


A method for determination of the specific activity of receptor-bound human chorionic gonadotropin (hCG). Precise knowledge of the specific activity (S.A., muc/mug) of the receptor bound radiolabelled hormone is required for study of the stoichiometry of peptide hormone-receptor interactios. A radioligand receptor assay using 131-I-hCG as tracer and transplantable mouse luteoma homogenates (Biol. Reprod. 8:550, 1973) as a source of receptor was used as a model to determine the specific activity of receptor bound 125-I-hCG. Progressive saturation of the gonadotropin receptor by 125-I-hCG suggests the presence of a high affinity-low capacity binding event (saturating between 14 and 37 ng/100 mg homogenate) that does not distinguish between non-radioactive hCG and 125-I-hCG, and a low affinity-high capacity binding event (saturating between 240 and 270 ng/100 mg homogenate) that shows a preference for non-radioactive hCG over 125-I-hCG. Parallelism between bound 125-I-hCG and non-radioactive hCG in terms of competition with tracer 131-I-hCG could only be demonstrated for the high affinity event.


Assay of lactogenic hormones using receptors isolated from rabbit liver. Using 125-I-labelled ovine prolactin and receptors isolated from the livers of rabbits, a sensitive method has been developed suitable for the assay of ovine, bovine, porcine, human and rat prolactins. These hormones showed competitive displacement of 125-I-ovine prolactin which was in general agreement with their respective activities in the pigeon crop sac bioassay. Human and monkey growth hormones and human placental lactogen, which have marked prolactin-like actions on mammary tissue were also effective competitors. Non-primate growth hormones (ovine, bovine, equine and canine) which do not have prolactin-like activity gave little if any displacement as did human FSH, LH, TSH, ACTH and bovine insulin. Preparations of equine and canine prolactin of varying purity gave dose-response curves although their activity as competitors relative to ovine prolactin was poor and not related to their pigeon crop stimulating activity. This indicates species differences between prolactins in hormone-receptor interaction. Experiments with antiserum to human growth hormone have suggested an effective method of making the assay specific in species such as man in which prolactin is not the sole hormone with lactogenic activity.


The effect of LH on cyclic AMP and progesterone in rat ovaries in vivo. LH injected into the abdominal vein of immature female rats increased ovarian cyclec AMP levels within 1 min, and the effect lasted at least 2 hr.  In PMS stimulated rats, a maximal effect on cyclic AMP levels was obtained with 5 mug of LH 10 min after injection.  Significant effects were obtained with as little as 0.1 mug.  The minimum effective dose required to increase ovary cyclic AMP levels in proestrus rats was 0.05 mug of LH.  While 0.01 mug did not affect cyclic AMP levels, an increase in ovary progesterone was found.  In other experiments, the response of cyclis AMP to LH in vivo was greater in immature ovaries than in those from animals pretreated with PMS and hCG.  Small increases in cyclic AMP levels were also seen with FSH in PMS and PMS + hCG treated animals and with prostaglandin E2 and prolactin in PMS treated rats.  These results parallel, to some extent, previous results found in vitro, and show that very small doses of LH can stimulate ovary cyclic AMP increases in vivo.


Antiestrogen action in the uterus: biological ineffectiveness of nuclear bound estradiol after antiestrogen. These studies attempt to analyze the basis of the estrogenic and antiestrogenic action of three nonsteroidal clomophene-type compounds as monitored by their ability to bind to immature rat uterine cytoplasmic estrogen receptor, transfer receptor sites to the nucleus, and elicit estrogenic responses (increased uterine weight and induction of the synthesis of a specific uterine protein, called induced protein, or "IP"), and by their ability to antagonize the effects of estradiol on these receptor interactions and uterine responses. Both CI-628 (CI) and U-11, 100A (UA) [50 mug] elicit slight IP induction at 1-2 hand give pronounced uterine weight increases at 24 h but feeble increases at 72 h (3 single daily injections). Both bind to cytosol, and effect the transfer of receptor sites to the nucleus, which may account for the estrogenicity of these compounds. Both CI and UA give rapid (by 2-4 h), prolonged (for over 24 h), and complete blockage of estradiol-stimulated treatment abolishes short-term estradiol-stimulated uterine weight increase and antagonizes the 72 h estradiol-stimulated uterine weight response to the level attributable to the antiestrogen alone. MER-25, at the same dose (50 mug), had no estrogenic or antiestrogenic activity. Both CI and UA rapidly deplete the cytoplasmic estrogen binding capacity, reducing it to barely detectable levels for 24-42 h. Although during this period, no IP or uterine wet weight response can be elicited by estradiol, administration of saturating levels of [3H]estradiol in vivo or in vitro results in the appearance of considerable [3H]estradiol in the nucleus, bound to a macromolecule sedimenting identically with that of the nuclear receptor-estradiol complex (5.5S) formed in the absence of prior antiestrogen exposure. Hence, the estradiol which becomes bound in the nucleus after antiestrogen is biologically ineffective. The return of IP responsiveness after antiestrogen correlates well with the level of cytoplasmic receptor capable of translocation to the nucleus, and not with the nuclear estradiol uptake capacity, Presumably, then, the antiestrogenic action of CI and UA results from their depletion of cytoplasmic receptor sites and not from their ability to block specific estradiol-nuclear receptor binding per se. These studies indicate that one should be cautious in assuming that the magnitude of an estrogen response is necessarily related to the level of estrogen receptor complex in the nucleus.


Stimulation of division of Y1 adrenal cells by a growth factor isolated from bovine pituitary glands. The effect of Fibroblast Growth Factor (FGF) on the initiation of DNA synthesis and on the rate of division of the Y1 adrenal cell line has been investigated. In sparse populations of Y1 cells (4 times 10- minus 3 cells/cm2) maintained in 0.2 percent calf serum, FGF was able to initiate DNA synthesis to the same extent that an optimal concentration of serum could. Cells maintained in 0.2 percent calf serum sustained continuous growth when given 5 ng/ml of FGF daily. Cultures fixed and stained with crystal violet showed FGF colonies to be of equivalent size and quantity as those with serum alone. Insulin had no mitogenic activity of its own at concentrations as high as 500 ng/ml nor did it have any potentiating effect on the mitogenic activity of FGF. Glucocorticoids (0.1 mug/ml to 1 mug/ml) inhibited (25 percent) the initiation of DNA synthesis as well as the rate of division induced by FGF. ACTH (0.75 IU/ml) was clearly inhibitory. Not only did it reduce the rate of division of cells in serum but it also reduced the rate of DNA synthesis and inhibited division in the presence of FGF.


The effect of LH/FSH, dibutyryl cyclic AMP, and prostaglandins on the production of estrogens by rabbit granulosa cells in vitro. The purpose of this study was to establish the effects of trophic hormones on the production of estrogens by rabbit granulosa cells. A pure population of these cells was isolated from preovulatory follicles (1-15 mm in diameter) of estrous rabbits, and cultured for 6 days with either one or a combination of the following hormones: androstenedione, Pergonal (LH/FSH), dibutyryl cyclic AMP (Bu2cAMP), prostaglandin F2alpha (PGF2alpha), or prostaglandin E2 (PGE2). The medium was collected every 2 days and progesterone (P), estrone (E1) and estradiol-17 beta (E2beta) were measured by radioimmunoassay. Granulosa cells cultured as controls (i.e., without exogenous trophic hormones) secreted P spontaneously and its secretion was stimulated 100 to 1,000 fold with LH/FSH and Bu2cAMP, but not with PGF2alpha or PGE2. Androstenedione, either alone or with trophic hormones had no apparent effect on the cytology of the granulosa cells or their ability to secrete P. In the absence of exogenous androstenedione, the cultures produced very small amounts of E1 or E2beta (smaller than 100 pg/ml), either spontaneously or in response to LH/FSH, B12cAMP, PGF2alpha, or PGE2. Incubating granulosa cells with exogenous androstenedione (1 mug/ml) resulted in a 30- to 150-fold increase in E2beta production, which was stimulated an additional 3- to 5-fold with LH/FSH and Bu2cAMP, but not with PGF2alpha or PGE2. In most cultures, E2beta production was restricted to the first 2 days in vitro. Bu2cAMP, however, maintained E2beta production at relatively high levels throughout the duration of the experiment, but there was a progressive decrease in its production. The production of E1 was only 5 percent of E2beta, but the pattern of secretion was similar for both estrogens. These results suggest that cyclic AMP could have a role in regulating the synthesis of estrogens by rabbit granulosa cells.


Comparison of the binding of human chorionic gonadotropin to isolated bovine luteal cells and bovine luteal plasma membranes. The specific binding of [125-I]iodohCG to intact luteal cells obtained from bovine corpus luteum by enzymatic treatment, or to purified plasma membranes obtained from bovine corpus luteum has been compared. It was a saturable process with respect to the [125-]iodohCG concentration. Specific binding could be detected at concentrations as low as 1.8 ng of HCG per ml and saturation achieved at 92 ng/ml. The [125-I]iodohCG specifically bound to the luteal cells or to the plasma membranes was displaced by increasing concentrations of native hCG. Subunits hCGalpha and beta had respectively 200- and 800-fold less activity than hCG. Four to 10 times more ovine LH than hCG was required to displace an identical amount of bound [125-I]iodohCG. The binding of hCG to its receptor site was a function of time and temperature. The affinity of hCG for its receptor sites in luteal cells or plasma membranes of luteal cells was similar (dissociation constants of 5.3 and 3.8 times 10- minus 10 M, respectively). The number of sites per luteal cell was 5 times 10-4 and the capacity of plasma membranes to bind hCG was 140 fmol per mg of protein at saturation. The data does not, however, allow a comparison between the number of binding sites in the two preparations. It is concluded that the enzymatic treatment necessary to obtain a suspension of viable luteal cells does not affect the kinetic characteristics of the binding of hCG to receptor sites since they are similar to those of plasma membranes not treated with proteolytic enzymes.


The mechanism of ACTH stimulation of adrenal ornithine decarboxylase activity. The mechanism of action of adrenocorticotrophin (ACTH) stimulation of rat adrenal orticotrophin (ACTH) stimulation of rat adrenal ornithine decarboxylase activity was investigated. ACTH induction or ornithine decarboxylase activity was not prevented by administration of drugs that inhibit adrenal steroid biosynthesis. A dose of ACTH that produced maximal levels of adrenal cyclic AMP did not induce ornthine decarboxylase activity. Ovine growth hormone, which caused no increase in adrenal cyclic AMP, stimulated adrenal ornithine decarboxyase activity. These observations suggest that the increase in adrenal ornithine decarboxylase activity stimulated by ACTH is not dependent upon steroidogenesis, nor is it dependent on the early peak of cyclic AMP, although it  may be influenced by the sustained levels of tissue cyclic AMP that follow the administration of large doses of ACTH. Furthermore, it appears there may be a pathway of ornithine decarboxylase activation in the adrenal which is entirely independent of cyclic AMP mediation. The effects of hypophysectomy on adrenal ornithine decarboxylase response to ACTH were examined. In rats given ACTH 16 h after hypophysectomy, the increase in ornithine decarboxylase activity was delayed when compared with the response in animals given ACTH 1 h after hypophysectomy. Actinomycin D given during the first 3 h after ACTH in the 16 h hypophysectomized rat abolished the expected increase in ornithine decarboxylase activity. Thereafter, a progressive increase in ornithine decarboxylase activity was observed as the interval between ACTH and Actinomycin D administration was further increased. In contrast, Actinomycin D administered 15 min before ACTH in the 1 h hypophysectomized  rat had no effect on the subsequent increase in ornithine decarboxylase activity, and actually progressively enhanced the response the longer its administration after ACTH was delayed. Cycloheximide abolished the response to ACTH in both the 1 h and the 16 h hypophysectomized rat. Thus, it appears that ACTH stimulates a post-transcriptional mechanism regulating ornithine decarboxylase activity in the acutely hypophysectomized animal, whereas, in the chronically hypophysectomized rat, ACTH must first stimulate transcription of new messenger RNA which is involved in regulation of adrenal ornithine decarboxylase synthesis.


Relation between thyroid iodine content and the accumulation and oxidation of [35-S] Methimazole in the rat. The thyroid accumulation and oxidation of a single intraperitoneal dose of [35-S] methimazole has been studied in iodine-deficient, normal and iodine-treated rats. A highly significant positive linear correlation was found between the thyroid oxidation of methimazole to sulfate and intrathyroidal iodine content. A single dose of potassium iodide given intraperitoneally (ip) to rats 1 h before administration of [35-S] methimazole (1 mg/kg ip) increased the thyroid accumulation and oxidation of methimazole. Conversely, the thyroids of rats maintained on a low iodine diet for 21 days showed a markedly reduced capacity to accumulate and oxidize methimazole. The level of oxidation found in the iodine-deficient, normal and iodide-treated groups was 0.21, 4.15 and 12.6 nmol sulfate/g thyroid respectively. The animals maintained on the low iodine diet for 21 days showed significant increases in thyroid weight and thus the decrease in methimazole oxidation occurred in spite of increased stimulation by endogenous TSH. These results show that the intrathyroidal iodine content is a critical factor in the metabolism of methimazole in the thyroid.


Effects of TSH on iodide transport by mouse thyroid lobes in vitro. Thyroidal iodide concentration by in vitro mouse thyroid lobes was studied by determining the equilibrium tissue/medium iodide, concentration ratio (T/M[I-]), THYROIDAL I- influx, and thyroidal I-efflux in the presence and absence of ClO4-. Chronic thyroid stimulation by low dietary iodine increased the T/M[I-], and the increase was linearly related to I- influx. There was no difference in efflux rate when animals fed low and high iodine diets were compared, although the efflux with C104- added to block influx was increased by low iodine diet. Addition of TSH in vitro caused a delayed fall in T/M[I-] with a similar TSH concentration-dependence as colloid droplet formation. The TSH effect was mimicked by exogenous cyclic AMP but could be dissociated from stimulation of hormone release by colchicine. Thyroidal I- efflux was increased up to 20-fold by C104-. In the presence of C104- short-term and equilibrium-labeled I- exited at the same rate, but in the absence of C104- short-term-labeled I- efflux was consistently higher. The TSH-induced fall in T/M I- could be accounted for by increased iodide efflux. As TSH also increased efflux when influx was blocked by C104-, the TSH effect would seem due to an increased intrinsic follicular leakiness.


Effects of lithium on vasopressin-dependent cyclic AMP in rat renal medulla. A vasopressin resistant urinary concentrating defect has been described in patients receiving lithium salt for affective disorders. For the pathogenic mechanism of the concentrating defect it has been postulated that lithium inhibits the vasopressin-dependent cyclic AMP system. However, the results of indirect studies on the lithium effect are equivocal. Therefore, the effect of lithium specifically on the vasopressin-dependent cyclic AMP system was investigated in rat renal medulla. The increase of cyclic AMP concentration by vasopressin was inhibited by lithium. But lithium had no effect on the PTH-dependent cyclic AMP concentration in renal cortical slices. Regardless of magnesium concentrations from 0-10 mM in the incubation media, 10 mM lithium had no moeasurable effect on the vasopressin-dependent adenylate cyclase of rat renal medulla. However, 10 mM lithium augmented the cyclic AMP-phosphodiesterase activity in renal medulla in the high Km system. These results suggest that lithium inhibits the vasopressin-dependent cyclic AMP concentration in renal medulla via the augmentation of its catabolism, rather than via the inhibition of cyclic AMP generation.


New evidence for an acute role of protein kinase in hCG action. The present experiments were designed to study whether exogenous hCG could elicit acute changes in the ovarian concentration of soluble cAMP-dependent protein kinases temporally related to binding of hCG and intracellular accumulation of cAMP. Cyclic AMP dependent protein kinase activity decreased five-fold within 5 to 30 min after intravenous administration of highly purified hCG to pseudopregnant rats. Moreover cAMP dependent protein kinase activity was totally suppressed with 0.5 IU hCG, whereas tissue concentration of cAMP continued to increase throughout the dose range (0.05-5.0 IU) of hCG used in the present studies. A marked fall in cAMP-dependent protein kinase activity had occurred before there was a significant change in intracellular accumulation of cAMP, possibly reflecting intracellular compartmentalization of cAMP. Inhibitors of protein did not affect the hCG-induced changes in tissue concentrations of cAMP and soluble cAMP dependent protein kinase activity but did suppress the recovery of cAMP dependent protein kinase activity to pretreatment levels. Cyclic AMP dependent protein kinases appear to play a significant role in mediating hormonal action in vivo. In addition the present studies suggest that, protein kinases may protect the cell from excessive hormonal stimulation.


Relationship between ACTH release and corticosterone binding by the receptor sites of the adenohypophysis and dorsal hippocampus following infusion of corticosterone at a constant rate in the adrenalectomized rat. The amount of corticosterone bound to proteins in the adenohypophysis and dorsal hippocampus was studied concurrently with the plasma ACTH concentration in 4-week adrenalectomized male rats under steady-state conditions achieved by infusing the steroid at a constant rate for 45 min. Corticosterone binding was measured by gel chromatography on Sephadex LH-20, using a TE buffer containing 0.4 m NaCl. No saturation was observed in whole homogenates and supernatants with increasing plasma corticosterone concentrations. However, corticosterone binding by the pituitary and by one of two types of hippocampal receptors evidenced saturation within our range of corticosterone infusion rates, indeed. Scatchard  plots allowed us to distinguish two types of binding sites in the hippocampus. The first, saturated at a low corticosterone concentration has an association constant of 3.0 times 10-8 M-1 with a number of binding sites estimated at 150 times 10- minus 15 mol/mg protein, whereas the second is associated with non-specific binding. The adenohypophysis shows only one kind of binding site with an association constant of 3.6 times 10-8 m- minus 1 and a number of sites estimated at 989 times 10- minus 15 mol/mg protein. A suggestive relationship was observed between ACTH inhibition by corticosterone and saturation of the pituitary binding sites. Our data are consistent with the possible involvement of corticosterone binding by specific sites in the negative feedback regulation of ACTH release.


Inhibition of anterior pituitary estrogen-receptor complex formation by low-affinity interaction with 5 alpha-dihydrotestosterone. A possible role for 5alpha-dihydrotestosterone in the anterior pituitary of the rat has been investigated by determining the  influence of this androgen upon the binding 17beta-estrasiol to its cytoplasmic receptor. Under equilibrium binding conditions, both testosterone and 5 alpha-dihydrotestosterone are very poor competitors for the estrogen binding sites, as compared to estrogenic compounds. 5alpha-Dihydrotestosterone, however, inhibits the initial rate of formation of the estradiol-receptor complex in a concentration-dependent manner. Steroidal specificity of this incubation is indicated by the failure of progesterone to elicit the same effect. The androgen inhibition appears to involve low-affinity interaction with the estrogen receptor, since no high-affinity binding of 5alpha-dihydrotestosterone could be detected. Kinetic analysis indicated that the inhibition is competitive, involving interaction at the estrogen-binding site of the receptor.