Lack of additive effect between mechanisms of resistance to carbapenems and other beta-lactam agents inPseudomonas aeruginosa

Eighty-nine clinical isolates resistant (n=61) or susceptible (n=28) to imipenem and exhibiting the main patterns of susceptibility to other -lactam agents (wild type pattern, penicillinase pattern, constitutive cephalosporinase pattern) were studied in order to investigate (i) the mechanism of resistance involved and (ii) whether resistance to carbapenems affects the level of resistance to other -lactam agents and, conversely, if resistance to other -lactam agents affects the level of resistance to carbapenems. For this purpose, the presence of OprD protein in the cell wall was detected by Western blot and -lactamase activity by spectrophotometric assay and isoelectric focusing. OprD expression was not detectable in the imipenem-resistant (MIC16 g/ml) strains. It was decreased in half the strains for which MICs of imipenem were 2 to 8 g/ml and was close to a normal level in the most susceptible strains (MIC 1 g/ml), thus demonstrating a direct correlation between the level of susceptibility to imipenem and the level of OprD expression. No imipenemase activity was detected in imipenem-resistant strains. Synergy between imipenem or meropenem and BRL42715 was observed for all of the strains, demonstrating the role of cephalosporinase in carbapenem resistance. Within each pattern of susceptibility, the mean MICs of -lactam agents other than carbapenems were similar, whether the strains were susceptible or resistant to imipenem. Conversely, the mean MICs of imipenem or meropenem for either the imipenem-resistant or the imipenem-susceptible strains were similar, regardless of the susceptibility of these strains to the other -lactam agents. Thus, when several mechanisms of resistance to -lactam agents are present in the same strain ofPseudomonas aeruginosa, there is no additive effect between these mechanisms.     

Distal dendrites of frog motor neurons: a computer-aided electron microscopic study of cobalt-filled cells

Summary With the aid of the cobalt labelling technique, frog spinal cord motor neuron dendrites of the subpial dendritic plexus have been identified in serial electron micrographs. Computer reconstructions of various lengths (2.5–9.8 m) of dendritic segments showed the contours of these dendrites to be highly irregular, and to present many thorn-like projections 0.4–1.8 m long. Number, size and distribution of synaptic contacts were also determined. Almost half of the synapses occurred at the origins of the thorns and these synapses had the largest contact areas. Only 8 out of 54 synapses analysed were found on thorns and these were the smallest. For the total length of reconstructed dendrites there was, on average, one synapse per 1.2 m, while 4.4% of the total dendritic surface was covered with synaptic contacts. The functional significance of these distal dendrites and their capacity to influence the soma membrane potential is discussed.     

Effect of lysolecithin on blood vessel reactivity and of some ethers and esters of glycerophosphocholine and phosphocholine,respectively, on aggregation of rat platelets

Infusion of lysolecithin (LPC; e.g. 88 g/ml for 0.5–1.0 min) did not significantly impair the vasopressor action of norepinephrine (NE), prostaglandin F₂ (PGF₂) and extract of posterior pituitary (EPP) in the isolated perfused hind legs of rats. In other words, vascular smooth muscle behaves differently from the smooth muscle of the guinea-pig small intestine, since, in the latter, contractions evoked by acetylcholine, prostaglandins etc., are inhibited by LPC. Triton X 100 which, by comparison, was used as a detergent effective on the guinea-pig small intestine, depressed the vasopressor effect of NE, PGF₂ and EPP.LPC, at low concentrations (40 mol/l), potentiated (15% max.) ADP-induced platelet aggregation (PA) in rat PRP but, at high concentrations, inhibited PA (IC₅₀=390 mol/l). 2-Hexadecylglycerophosphocholine and its short-chain 1-alkyl ethers, which are structurally related to platelet-activating factor, as well as some long-chain alkanol phosphocholine esters, were somewhat more active than LPC. Dipalmitoyllecithin (4–700 mol/l) was without any effect.     

Fibre types inLimulus telson muscles: morphology and histochemistry

Summary Using a variety of techniques, we have demonstrated the presence of at least two fibre types inLimulus median telson levator muscle. By light and electron microscopy, large (21 56 m² mean cross-sectional area) fibres have A-bands of 4.1 m, one-half I bands of 2.15 m and Z lines 0.5 m in width. Few mitochondria are found in these fibres, which comprise 54% of those present in a given microscope field and which occupy 82% of the total cross-sectional area. Small fibres (484 m² mean cross-sectional area) have A bands of 6.3 m, one-half I bands of 3.1 m and Z lines between 0.5 and 1.0 m in width and are rich in mitochondria. Although small fibres comprise nearly one-half (46%) of the fibres in a field, they occupy only 18% of the total cross-sectional area.Histochemical staining for alkaline-stable myofibrillar ATPase activity and mitochondrial reduced -nicotinamide adenine nucleotide (-NADH) tetrazolium reductase activity confirms the presence of two fibre types. The large fibres react positively for the myofibrillar ATPase activity and negatively for the mitochondrial enzyme activity. The reverse is seen with the small fibres. Some fibres of intermediate size, having intermediate staining characteristics, were also observed. Native gel electrophoresis of both myofibrillar and purified myosin preparations supports the observed differences in myofibrillar ATPase activity in that two myosin isozymes are resolved on pyrophosphate gels. Although the thick filaments isolated from unstimulated small fibres are longer (>6.0 m) than those isolated from unstimulated large fibres (4.26 m), all have a similar appearance with respect to the arrangement of myosin heads on their surfaces, and similar diameters. The implications of the observed heterogeneity of fibre types is discussed with reference to previously reported phenomena inLimulus telson muscle, including changes in length of thick filaments on fibre stimulation and the shape of the length-tension curve obtained from fibre bundles.     

On the regulation of the expressed L-type calcium channel by cAMP-dependent phosphorylation

The Ca²⁺ channel subunits _1C-a and _1C-b were stably expressed in Chinese hamster ovary (CHO) and human embryonic kidney (HEK) 293 cells. The peak Ba²⁺ current (I _Ba) of these cells was not affected significantly by internal dialysis with 0.1 mM cAMP-dependent protein kinase inhibitor peptide (mPKI), 25 M cAMP-dependent protein kinase catalytic subunit (PKA), or a combination of 25 M PKA and 1 M okadaic acid. The activity of the _1C-b channel subunit expressed stably in HEK 293 cells was depressed by 1 M H 89 and was not increased by superfusion with 5 M forskolin plus 20 M isobutylmethylxanthine (IBMX). The _1C-a·₂·₂/ complex was transiently expressed in HEK 293 cells; it was inhibited by internal dialysis of the cells with 1 M H 89, but was not affected by internal dialysis with mPKI, PKA or microcystin. Internal dialysis of cells expressing the _1C-a·₂·₂/ channel with 10 M PKA did not induce facilitation after a 150-ms prepulse to +50 mV. The Ca²⁺ current (I _Ca) of cardiac myocytes increased threefold during internal dialysis with 5 M PKA or 25 M microcystin and during external superfusion with 0.1 M isoproterenol or 5 M forskolin plus 50 M IBMX. These results indicate that the L-type Ca²⁺ channel expressed is not modulated by cAMP-dependent phosphorylation to the same extent as in native cardiac myocytes.     

Präzisierung der Reizwirkung mittelfrequenter Wechselströme

Zusammenfassung Bei der Mittelfrequenz-Impuls-Reizung ist streng darauf zu achten, daß keine polaritären Reizkomponenten auftreten. Die diesbezügliche Kontrolle wird am besten mit Hilfe des Konvertibilitätstestes vorgenommen, d. h., es darf beim Vertauschen der Zuführungen zu den Reizelektroden weder die Reizschwelle bzw. die Größe des kollektiven Reizerfolges noch dessen Latenzzeit eine signifikante Änderung erfahren. Auf diese Weise wird die Phasenunabhängigkeit des echten Mittelfrequenz-Reizeffektes nachgewiesen.Diesen Anforderungen entsprechen Mittelfrequenz-Impulse, deren Trägerfrequenz über einige wenige Perioden sich aufschaukelt und ebenso wieder abklingt. Demgegenüber sind Mittelfrequenz-Stromstöße mit phasenstarrem Einsatz und Ende nicht unbedingt frei von polaritären Ein- bzw. Ausschalt-effekten, indem sowohl die erste als auch die letzte Trägerperiode einen polaritären Wechselimpuls-Reizeffekt ergeben kann, je nach Phasenlage bezogen auf die wirksame Reizelektrode und Art der Ansprechbarkeit des Reizobjektes (Nerv) auf entsprechend kurze gleitspiegelsymmetrische Wechselimpulse. Für eine echte Mittelfrequenz-Stromstoß-Reizung ist demnach ebenfalls ein Aufschaukeln und Abklingen der Trägerfrequenz über einige wenige Perioden erforderlich.Es besteht ein prinzipieller Unterschied zwischen der echten Mittelfrequenz-Reizung, die phasen -bzw. periodenunabhängig ist und schon früher als apolaritär bezeichnet wurde, und der konventionellen polaritären Reizung, die als polaritäre Komplikation der Mittelfrequenz-Reizung auftreten kann.Diese Präzisierung der Reizwirkung mittelfrequenter Wechselströme wurde angeregt durch zwei im Text erwähnte Publikationen, in denen in keineswegs überzeugender Weise versucht wird, die Mittelfrequenz-Reizung letzten Endes auf das polare Gesetz der Erregung zurückzuführen.

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Summary The particular excitatory action exerted by middle-frequency alternating current can only be revealed if care is taken to eliminate the occurrence of so-called polarity effects. Such effects are produced by the short alternating impulses represented by the first and the last period of a middle-frequency current pulse and are based on the polar law of excitation.In order to prevent such polarity intrusions, it is necessary to increase and decrease the amplitude of the middle-frequency current pulses over a few carrierperiods, or, to use amplitude-modulated middle-frequency impulses of variable shape and duration of envelope.A true middle-frequency excitatory effect is easily demonstrated by resorting to the convertibility test. It will then become evident that stimulation threshold, magnitude as well as latency of response do not change during reversal of the stimulating poles. This means, that no significant phase change of the response with regard to the carrier-frequency occurs when the leads to the stimulating electrodes are commuted, and that, as a result, true middle-frequency effects do not depend upon one particular catelectrotonic variation among the carrier-periods of a middle-frequency current pulse.It can thus be concluded that a fundamental difference exists between true middle-frequency stimulation, which is based on a non-polarity or apolarity principle, and the conventional stimulation of the polar or polarity type.This paper has been written in the hope of dispelling some errors of interpretation (discussed in the text) tending to ascribe the excitatory effects of middle-frequency impulse stimulation to the classical polar law of excitation.

     

Intrinsic cholinergic excitation in the rat neostriatum: Nicotinic and muscarinic receptors

Summary An in vitro slice technique was employed to study the receptors involved in intrinsic cholinergic excitation in the rat neostriatum. The locally evoked synaptic potentials were suppressed by antinicotinic agents, mecamylamine (10 M), d-tubocurarine (3 M) or hexamethonium (100 M), but not by the antimuscarinic agent atropine (100 M). If the slices were exposed to an acetylcholinesterase (AChE)-inhibitor (paraoxon 1–20 M, physostigmine 0.1–0.5 M), the synaptic potentials were potentiated. The amplitude of the orthodromic population spike increased, and it was further facilitated when the stimulus frequencies were raised from 1–3 Hz to 10–30 Hz. The frequency facilitation following exposure to an AChE-inhibitor was blocked by atropine (1–100 M). Intracellular recording indicated that a slow depolarizing potential caused the frequency potentiation of the orthodromic discharges. Apparently rat neostriatum is similar to cholinergic systems in sympathetic ganglia and spinal Renshaw cells, in that nicotinic receptors mediate fast excitation and muscarinic receptors mediate slow excitation.     

Sodium salicylate: alternate mechanism of central antipyretic action in the rat

Infusion of sodium salicylate (50.0 or 100.0 g/l) into the ventral septal area (VSA) of the rat brain suppressed Prostaglandin-E₁-induced hyperthermia. Infusion of artificial cerebrospinal fluid (aCSF) or 10.0 g doses of salicylate did not. The suppression of intracerebroventricularly-induced (icv) Prostaglandin E₁ (PGE₁) hyperthermia was not due to a hypothermic action of salicylate since salicylate infusions given during cold exposure (10.0°C) did not lower core body temperatures. A possible interaction between salicylate and endogenous arginine vasopressin (AVP) was investigated. Infusion of both salicylate (50.0 g/l) and either AVP antiserum or AVP antagonist into the VSA resulted in PGE hyperthermias occurring at levels which were not different from control levels as opposed to enhanced hyperthermia (antiserum or antagonist alone) or suppressed hyperthermia (salicylate alone). These results are consistent with the notion that sodium salicylate infusions within the VSA enhance AVP action and thus bring about the attenuation of PGE-induced hyperthermia.     

Effects of ryanodine on skinned skeletal muscle fibers of the rabbit

The mechanism(s) of ryanodine-induced contracture of skeletal muscle were studied in skinned fibers from soleus (SL) and adductor magnus (AM) (slow- and fast-twitch skeletal muscles) of rabbits. Pieces of SL or AM were homogenized (sarcolemma disrupted). Single fibers were dissected from the homogenate and mounted on photodiode force transducers. At concentrations 1–50 M, ryanodine slightly but significantly increased the submaximal Ca²⁺-activated tension development of the contractile proteins in skinned fibers of AM but not of SL. Ryanodine in uptake phase or release phase increased caffeine-induced tension transients in the SR of both muscle types; however, no dose-response relation was found. Ryanodine 1 M decreased, however, the second control tension transients in a dose-dependent manner. The depression was nearly irreversible and activity-dependent. The concentrations of ryanodine that inhibited the second control tension transients by 50% were 10 M and 5 M for SL and AM, respectively, following ryanodine administration in the release phase, and 100 M and 30 M, respectively, for these preparations after the drug was present in the uptake phase. The quantity of calcium released from the SR by Triton X-100 and caffeine in the second control tension transient was unchanged by ryanodine at all concentrations tested when compared with that of the absence of ryanodine. The present findings suggest that the ability of ryanodine to increase immediate calcium release from the SR, and in AM but not SL, to increase the sensitivity of the contractile proteins to Ca²⁺ underlies the contracture caused by this agent in intact skeletal muscles. The delayed decreased Ca²⁺ efflux by caffeine, as evidenced by depression of tension transient with no change in the calcium content may be responsible for the decreased twitch tension caused by this agent.     

Heterogeneity of circular mitochondrial DNA molecules from sugar beet with normal and male sterile cytoplasms

Summary Mitochondrial (mt) DNAs from normal (N) and male sterile (S) cytoplasms of sugar been have been isolated and investigated by electron microscopy. The results showed that mtDNA was composed of a heterogeneous population of circular molecules. Their contour lengths varied from 0.28 to 51 m, but unlike in the case of maize, a large difference was not observed in the distribution of molecular classes greater than 1.0 m between N and S cytoplasms of sugar beet. On the other hand, N and S cytoplasms were shown to contain their own characteristic combinations of small circular mtDNA species with lengths between 0.28 m and 0.6 m. Mitochondrial DNAs from various sources of male-sterile cytoplasms were analyzed by agarose gel electrophoresis to determine the extent of cytoplasmic variation. Additional low molecular weight DNA bands appeared in all male-sterile lines examined, and as a result, three distinctive banding patterns were recognized. These data are in general agreement with those based upon restriction endonuclease digestion of mt and chloroplast DNAs and the genetic analysis of fertility restoration in test crosses.     

Effect of noradrenalin on the collateral coronary circulation

Summary A study was made of the action of noradrenalin (in doses of 0.5 g/kg and 4 g/kg) on collateral coronary circulation of dogs. Administration of g/kg caused a brief increase in the retrograde coronary flow in direct proportion to its pressor effect. Low doses of noradrenalin induced stable increase in the retrograde blood flow in the descending branch of the left coronary artery against the background of the normalized general blood pressure.(Presented by Active Member AMN SSSR V. A. Sanotskii) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 58, No. 8, pp. 72–74, August, 1964     

Effect of thyrotropin releasing hormone and thyroxine on cytochrome oxidase activity in the rat adenohypophysis

Thyrotropin releasing hormone (TRH), in a dose of 0.01 and 1.0 g/ml, sharply increased cytochrome oxidase activity in the adenohypophysis of rats fed for 6 weeks with methylthiouracil. This effect of TRH on enzyme activity was blocked by thyroxine (T₄), if added to the incubation medium in a concentration of 20 g/ml. Actinomycin D (20 g/ml) prevented the blocking of cytochrome oxidase by T₄. TRH in a concentration of 0.01 g/ml and T₄, in a dose of 2.0 g/ml, caused no change in cytochrome oxidase activity in the adenohypophysis of intact and partially thyroidectomized rats.Laboratory of Biological Standardization of Hormones, Institute of Experimental Endocrinology and Hormone Chemistry, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR N. A. Yudaev.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 11, pp. 559–562, November, 1977.     

In vitro efficacy and fungicidal activity of voriconazole againstAspergillus andFusarium species

Twenty-nineAspergillus isolates and 25Fusarium isolates underwent in vitro antifungal susceptibility testing by a broth macrodilution procedure adapted from the National Committee for Clinical Laboratory Standards guidelines. The MIC50s of both voriconazole and amphotericin B were 0.5 g/ml and 1 g/ml against species ofAspergillus andFusarium, respectively, while the MIC90s of both agents were 1 and 2 g/ml. Voriconazole was more active in vitro than amphotericin B: the geometric mean MICs of voriconazole and amphotericin B againstAspergillus spp. were 0.36 g/ml and 0.64 g/ml, respectively. Voriconazole also demonstrated fungicidal activity againstAspergillus spp., with 86% (24/29) of isolates exhibiting minimum lethal concentrations of 4 g/ml.     

Histamine H2-receptor blocking activity of ranitidine and lamtidine analogues containing aminomethyl-substituted aliphatic systems

The possibility that the aromatic component in the classical H₂-antagonists might not be essential for histamine H₂-receptor blockade has been investigated. In the ranitidine series the removal of the furan ring is accompanied by a drastic decrease in H₂-blocking activity, but not by its disappearance (compound HB5: K_B on guinea pig isolated atria 31.6 M) whereas in the lamtidine analogues the substitution of the phenyl moiety with the more reduced -bonded CH₃-C=N-area generates a compound whose activity is comparable to that of cimetidine (K_B on atria 1.12M; ID₅₀ in the lumen-perfused stomach of the anaesthetized rat 3.61 mol/kg i.v.). The results also indicate that the diaminofurazan group confers high affinity at the histamine H₂-receptor.It is concluded that the aromatic portion of H₂-antagonists related to ranitidine and lamtidine is not a minimal requisite for activity when an appropriate polar group is used as an urea equivalent moiety.     

On three new species of the genus Criconemoides Taylor, 1936 (Nematoda: Criconematidae) from North India

Summary The definition of the genus Criconemoides should be extended so as to include those aberrant forms which possess slight cuticular ornamentation. Three new species of the genus Criconemoides Taylor, 1936 are described and illustrated from North India. Criconemoides aberrans n. sp. is distinctive in having 38–43 body annules, rough cuticular outgrowths on the body, 68–78 long spear, bluntly rounded tail, absence of spermatheca and males unknown. Criconemoides neoaxeste n. sp. has 45–49 body annules marked with faint longitudinal lines and rough posterior margins, 65–75 long spear, rounded tail terminus, vulva located at 7th or 8th annule from the posterior end and larva having a cuticular flap with rough margins on each annule. Criconemoides macrolobatus n. sp. is distinguished by 81–86 body annules, very large oval sublateral lobes, 71–75 long spear, tail terminus rounded and the absence of spermatheca and males unknown.

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Zusammenfassung Die Definition der Gattung Criconemoides sollte so erweitert werden, daß auch solche aberranten Formen eingeschlossen werden können, die leichte cuticulare Ornamente besitzen. Drei neue Arten der Gattung Criconemoides Taylor (1936) aus Nordindien werden beschrieben und abgebildet. C. aberrans n. sp. ist deutlich charakterisiert durch 38 bis 43 Körperringe, cuticulare Auswüchse am Körper, 68–78 langen Lanzen, stumpf-abgerundeten Schwanz; Spermatheke fehlt, Männchen nicht bekannt. C. neoaxeste n. sp. hat 45–49 Körperringe mit feiner longitudinaler Linienzeichnung und groben hinteren Rändern, 65–75 lange Lanzen, abgerundetes Schwanzende. Vulva im 7. oder 8. Ring vom hinteren Ende lokalisiert; die Larven haben eine cuticulare Klappe mit groben Rändern an jedem Ring. C. macrolobatus n. sp. ist gekennzeichnet durch 81–86 Körperringe mit sehr breitovalen sublateralen Loben, 71–75 lange Lanzen, Schwanzende abgerundet, Spermatheken fehlen und Männchen unbekannt.

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With 17 Figures in the Text     

Activation of atrial muscarinic K+ channels by low concentrations ofβγ subunits of rat brain G protein

Effects of G protein subunits from rat brain on cardiac K⁺ channel was examined in single atrial cells of guinea-pig, using patch clamp techniques. We found that 10 pM concentration of rat brain subunits preparation could activate the atrial muscarine receptor-gated K⁺ channel (I_K.ACh). Neither the detergent, CHAPS, used to suspend nor the boiled preparation activated I_K.ACh. Furthermore, preincubation of subunits preparation in Mg²⁺-free solution, which easily inactivated -GTP-S, did not affect -activation of I_K.ACh. We concluded, therefore, that subunits themselves can activate I_K.ACh.Supported by the grants from the Ministry of Education, Culture and Science of Japan and from the Calcium Signal Workshop on Cardiovascular Systems     

Naegleria fowleri andN. lovaniensis: Differences in sensitivity to trimethoprim and other antifolates

The growth ofNaegleria fowleri cultures in a BCS medium was not affected either by trimethoprim at 400 g/ml or by aminopterine, 3,5-diaminopterine and methotrexate at 500 g/ml.N. lovaniensis propagation in the same medium was inhibited with 10 g/ml of trimethoprim, 50 g/ml methotrexate and 100 g/ml 3,5-diaminopteridine. Aminopterine was ineffective at a concentration of 500 g/ml. The inhibitory effect of trimethoprim onN. lovaniensis cultures depended on the medium composition and could be neutralized by an addition of folic or tetrahydrofolic acids and a suspension of heat-killedEnterobacter aerogenes. Thymine, thymidine, hypoxantine and 2-amino-4-hydroxy-6-(tatrahydroxybutyl)-pteridine did not have an adverse effect. Trimethoprim activity inN. fowleri cultures could not be enhanced by the addition of Triton X-100 and Polymyxine B. Cryolyzate ofN. fowleri amoebae did not influence the trimethoprim inhibition ofN. lovaniensis cultures. Deviation in dihydrofolatereductase chemical structure or thymine dependency seems to be the probable explanation forN. fowleri antifolate resistance.     

Inhibition of chemiluminescence in granulocytes and alveolar macrophages by azelastine

The effect of azelastine, an orally effective antiasthmatic/antiallergic drug, on the generation of oxygenderived free radicals in phagocytes was investigated using different chemiluminescence-assays. The chemiluminescence (CL) of both human polymorphonuclear granulocytes (PMNL) and guinea-pig alveolar macrophages (AM) was induced either by phorbol myristate acetate (PMA) or zymosan and amplified either by lucigenin or DMNH (7-dimethylamino-naphthalene-1,2-dicarbonic-acidhydrazide). The inhibitory effect of azelastine was dependent on the inducer employed and the condition and type of cells used. Azelastine reduced PMA-induced CL concentration-dependently in both PMNL (IC₃₀=3.9 M) and AM (IC₃₀=9.8 M). In AM zymosan-induced CL was inhibited 21.7% by 10 M azelastine, whereas in PMNL it remained unchanged up to 10 M azelastine. Azelastine has a significantly stronger inhibitory effect (IC₃₀=4.2 M) on oxygen free radical generation in AM primed by fetal calf serum than in unprimed AM. Based on present results it is likely that azelastine inhibitis oxygen-derived free radical generation by interaction with protein kinase C.     

Expression of surface membrane IgG on pokeweed mitogen-reactive anti-tetanus toxoid antibody-producing cells

Anti-tetanus toxoid antibody-producing cells, differentially expressing surface membrane IgM, were analyzed for the additional expression of surface membrane IgG. ⁺ and ^– cells were rosetted with anti--ox red blood cells and separated by density centrifugation into fractions enriched or depleted or ⁺ cells. These B-cell subsets were assayed for the production of IgM and IgG anti-tetanus toxoid antibody and total IgM and IgG. The results indicated that the majority of anti-tetanus toxoid antibody synthesis in the ^– fraction was by ⁺ cells. In the ⁺ fraction, however, both IgM and IgG anti-tetanus toxoid antibody production was detected in the ^+– and ⁺⁺ fraction. The inclusion of isotype-specific antisera during the first 2 days of culture further established that was expressed on the surface of the majority of the precursors for IgG anti-tetanus antibody productionin vitro. Studies performed to determine the culture requirements of ^– and ⁺ cells revealed that production of IgG anti-tetanus toxoid antibody by both cell subsets was dependent on T cells and pokeweed mitogen. However, some ^– cells could produce IgG in the presence of T cells alone.     

The control of cell proliferation by preformed purines: A genetic study II. Pleiotropic manifestations and mechanism of a control exerted by adenylic purines on PRPP synthesis

When plated in medium containing 0.5 g/ml coformycin and adenosine (or adenine) fibroblasts were killed, even if pyrimidines were supplied. Measurements of N-formylglycine amide ribonucleotide synthesis showed that lethality is a manifestation of purine starvation. In the case of adenosine kinase deficient cells, growth was restored by hypoxanthine. The adenylic derivatives block only purine biosynthesis, presumably by inhibition of PRPP-amidotransferase. In this same medium, wildtype cells exhibited symptoms of PRPP deprivation: purine and pyrimidine syntheses were both shut off and HGPRT was simultaneously inactivated. The pleiotropic control by adenosine was abolished in adenosineresistant mutants that behaved as PRPP overproducers. These mutations conferred partial resistance to various toxic purine and pyrimidine analogs and preserved HGPRT activity in adenosinecontaining medium. This permits selection against these mutants. Evidence suggesting that adenosine kinase products may fulfill a specific function in the regulation of PRPP synthesis is discussed.Abbreviations AK adenosine kinase - APRT adenine phosphoribosyltransferase - HGPRT hypoxanthine guanine phosphoribosyltransferase - AD adenosine deaminase - OPRT orotate phosphoribosyltransferase - PRA phosphoribosylpyrophosphate amidotransferase - A adenine - rA adenosine - Hx hypoxanthine - dA 2-deoxyadenosine - rU uridine - dT thymidine - PRPP phosphoribosylpyrophosphate - Pala N(phosphonacetyl)-l-aspartate - FGAR -N-formyl-glycinamide ribonucleotide - Amp aminopterin - Amp + dT medium standard ERH medium supplemented with Amp and dT - HC-medium ERH supplemented with 0.5g coformycin/ml.