Emergency management of radiation accidents. The Joint Committee on Accreditation of Hospitals requires that hospitals have a written procedure to cope with radiation accidents. Emergency personnel, specifically, should be prepared to manage radiation accident victims initially. Guidelines currently in use in over 15 hospitals in the United States include 1) advance notification to the hospital of arrival of such patients; 2) designated treatment area; 3) operating room attire; 4) perform life-saving functions first; 5) cover open wounds and remove clothing; 6) search for contamination with radiation detector, or 7) flush all wounds if no radiation detector is available; 8) wash patient; 9) evaluate internal and external radiation exposure, and 10) obtain medical consultation. For community hospitals, a less elaborate procedure will suffice for adequate handling of radiation accident victims and accreditation.


Nonresuscitative aspects of the management of severe injuries. Although resuscitation of seriously injured patients in the emergency department is the physician's primary consideration, there are other important aspects of overall care. These include attention to the patient's family and friends and their reaction to a loved one's serious injury or death, as well as the effect of the injuries and initial treatment upon the patient, particularly if he regains consciousness in an apparently unreal world, connected to machines by various tubes and unable to speak.


Torsion of the fallopian tube.

Torsion of the fallopian tube. Although an infrequent cause of acute lower abdominal pain, fallopian tube torsion should be considered whenever a woman presents with this complaint. Early diagnosis and surgical intervention may save a tube that has not yet infarcted, yet initial presentation may give little indication of the seriousness of the condition until peritonitis develops. Definitive treatment is removal of the affected adnexa. Diminished fertility and an increased incidence of ectopic pregnancy in the remaining fallopian tube follows unilateral salpingectomy. A case history of fallopian tube torsion is presented.


Shock and resuscitation. II: Volume repletion with minimal edema using the "HALFD"(Hypertonic Albuminated Fluid Demand) regimen. A hypertonic albuminated fluid demand regimen (HALFD) for resuscitation has been used in burn patients since January 1, 1976. The effects of the HALFD method were compared with hypertonic fluid and Ringer's lactate resuscitation. Specific attention was directed to fluid, colloid, and volume changes. Resuscitation was guided by maintaining the mean arterial pressure between 60 to 110 torr, and urine volume at 30 to 50 ml/hr. Patients treated with the HALFD method fared significantly better clinically, needed less fluid, had less weight gain and plasma leak, and experienced slower plasma volume repletion than those treated more traditionally. We conclude that the HALFD method is a physically and physiologically appropriate paradigm for resuscitating the volume-depleted patient.


The microcomputer in the emergency department: a clinical information center. A microcomputer system was developed for the Tri-City Hospital Emergency Department to serve as a clinical information center. The system is being used to better allocate resources, both personnel and materials, to accurately assess the nature of the problems presenting to the emergency department, to aid in the diagnosis and laboratory test interpretation, to improve the quality of care, to serve as an epidemiological tool and to help in the education of emergency care personnel.


Scientific communications.

Scientific communications. Scientific communications have two main forms, oral presentation at scientific meetings, accompanied by audio-visual aids, and manuscripts prepared for publication in scientific journals. In an oral presentation, the author should be prepared to speak for ten minutes, without depending on notes, and have prepared answers to anticipated questions. Slides are standard aids to summarize and reinforce the material. Preparation of manuscripts follows well-established guidelines available from scientific journals. Manuscripts should proceed from introduction to methods, results, discussion, and conclusion. Prior to writing, the author should answer the following: 1) What is the purpose of this communication? 2) How is this communication different from other data already in the literature? What is its unique contribution? 3) Who is the audience? Manuscripts will go through as many as seven drafts.


[Anatomic and histologic studies of a Triatominae subfamily (Heteroptera, Reduviidae). Triatoma infestans' brain and its nerves]. To give a concrete aspect of structure, topography and function of the different nervous systems of the Triatominae, we begin this study with a description of the senso-motor system. As the first part we choose the study of the brain and its nerves. The external form of the brain is conditioned by the anatomy and morphology of the head. The brain is situated in the posterior part of the head beneath and behind the ocelli. It is characterized by the reduction of its longitudinal extension and, by this, the connectives between Syncerebrum and Gnathocerebrum are extremely short, but large, and the third commissure is included in the mass of the mandibular segment of the Gnatocerebrum. The optical masses are of the typical form, also the optical commissure. The Central Corps is volumous and is directly linked with the majority of the centres of the Protocerebrum and Deutocerebrum. The Pedunculated Corps has only one glomerulus that looks like a pedunculated mushroom without any signal of the formation of a calyx, found in ants and other more evoluted insects. Beneath the Central Corps, the extremities of the pedunculus disintegrate and form a number of claviform annexes like in the primitive Machilidae. The Deutocerebrum is characterized by volumous antennal centre, composed by numerous little glomeruli in peripheral situation. The Tritocerebrum is very reduced and has in its anterior part two very short frontal connectives and, by this, the frontal ganglion lies near the Tritocerebrum. The compact mass of the Gnathocerebrum shows its composition by three pairs of ganglions only by observation of series of preparations. With exception of the Lobi optici, 8 pair nerves and 3 impair nerves come from the brain: From the Protocerebrum: the nerves of the ocelli and the nerve of the Corpus allatum. From the Deutocerebrum: the nerves of the antennae and those of the pharynx. From the Tritocerebrum, the nerves of the labrum and, by the frontal ganglion, the Nervus recurrens and the nerve of pharyngeal muscles. From the Gnathocerebrum: the nerves of the amndibulae, maxillae, labium and of the salivary glands.


[Complement fixation reaction in the typing of influenza viruses isolated in Rio de Janeiro]. The author studied by the complement fixation test the influenza virus strains isolated in Rio de Janeiro during the 1973 epidemic. He prepared immunesera in hamsters by the inoculation of the allantoic fluid from infected chick embryos with each of the 7 isolated strains and the standard strains. The soluble antigens were prepared with the allantoic fluid of infected chick embryos. The tests were identically positive with the A2/Hong Kong/68 and A2/England/72 antigens and negative with the B/Mass/66. The tests were type specific and the behaviour of the A2/Hong Kong/68 and the A2/England/72 and the 7 strains of the isolated viruses was almost the same. They fixed 3 or 4 units of complement. The variants PR8, FM1 and Asia fixed only 2 units of complement.


[Immunoglobulins in the chronic phase of Chagas disease]. Since divergent results were obtained by different authors about immunoglobulins concentration in sera from acute and chronic cases of Chagas disease, the problem was submitted to statistical approach in field conditions in the endemic zone of Bambui, state of Minas Gerais, Brazil. Positives cases were considered in all individuals presenting evidences in parasitological, immunological or electrocardiographical examination living exactly in the same conditions, but without any evidence of infection under the same criteria. The sample consisted of 32 chronic cases and no significant difference in the immunoglobulins levels were detected in chronic cases of Chagas disease as compared with the control group. Heterophili antibodies are being determined in chronic cases and apparently there is no relationship between its concentration and pathogenic effects.


Fibrinolysis in the rat lung after intravenous tumour cell injection. Intravenous injection of a tumour cell suspension in rats induced an early tendency to increased fibrinolysis followed by a temporarily, significantly decreased fibrinolytic activity in lung vessels. These reactions were similar to those previously found after a large trauma and might reflect the vascular reaction to thrombotic material trapped in the lung microcirculation.


Karyology of a human colonic carcinoma (GW-39) serially xenografted in hamsters and in nude mice. A karyological analysis of the G-banded chromosomes of the serially transplantable GW-39 human colonic carcinoma was undertaken. GW-39 cells harvested from hamsters, nude (athymic) mice and cell cultures from 1972 to 1976 revealed a hypodiploid stemline of 45 human chromosomes, with a consistent loss of a 21-chromosome. No apparent alterations of the near-diploid stemline resulted during long-term xenogeneic propagation of this human cancer line.


Carcinogenic effect of di-isopropanolnitrosamine on the rat thyroid gland. Up to a 50% rate of thyroid gland tumours was induced in Sprague-Dawley rats after s.c. administration of di-isopropanolnitrosamine (DIPN). The neoplasms occurred both unilaterally and bilaterally, and were identified histologically as adenomas and adenocarcinomas of follicular epithelium origin. In a few instances, adenomas were observed as early as 18 weeks after commencement of treatment. Most adenocarcinomas could be observed macroscopically, and frequently demonstrated infiltration into the surrounding muscles.


Partial inhibition of primary lung tumourigenesis by tetramisole. Continuous treatment of mice with tetramisole (TMS, approximately 8 mg/kg/day) resulted in a highly significant (p < 0.002) reduction in numbers of primary lung tumours forming in response to neonatal urethane injection. Kinetic study indicated the slopes of the oncogenesis response curves differed, but not significantly. Therefore inhibition of primary lung tumourigenesis was only partial, even though TMS was given during the early stages of tumour development, continuously, and at near maximum tolerable dosage.


Blastogenic response and EBNA induction in human lymphocytes by Epstein-Barr virus only requires B cells but not macrophages. The postulated role of macrophages in the primary infection of human lymphocytes by Epstein-Barr virus (EBV) was examined. Macrophage removal had no effect on the blastogenic response but slightly reduced the percentage of EBNA-positive cells induced by the B 95-8 virus strain. Both the rate of DNA synthesis and the appearance of EBNA-positive cells could rather be related to the number of B cells in the infected populations. Macrophages are thus not required for the initiation of EBV-induced lymphocyte transformation.


Mutagenicities of smoke condensates and the charred surface of fish and meat. Smoke condensates obtained from broiling fish showed mutagenic activity for Salmonella typhimurium TA100 and TA98. Metabolic activation was required to induce mutagenic activity of smoke condensates of some species of fish. The smoke condensate obtained during charcoal broiling of beefsteak was far less mutagenic than that of fish, with or without metabolic activation. Extracts of the charred surface of broiled fish and meat also contained mutagenic substances. These extracts needed metabolic activation to exhibit mutagenicities on TA98. The mutagenic activity of the smoke condensate obtained from one sardine weighing 100 g was equivalent to that of 132 micrograms benzo(a)pyrene and that of the charred surface of the sardine was equivalent to 358 micrograms benzo(a)pyrene. One piece of beefsteak weighing 190 g, contained mutagenic activity equivalent to that of 855 micrograns benzo(a)pyrene.


The comparative toxicity and therapeutic efficacy of adriamycin and the adriamycin-DNA complex in the chemotherapy of C3H mice with transplanted mammary adenocarcinoma. Tissue analyses and tumour regression studies using the oncolytic antibiotic, adriamycin (14-hydroxydaunomycin), and its DNA complex at adriamycin dosages of 5 mg/kg and 10 mg/kg were made on C3H mice with transplanted mammary adenocarcinoma. Chemical analysis indicated a significantly lower (P < 0.05) uptake of adriamycin into cardiac tissue for the adriamycin-DNA complex. Tumour regression was comparable for both the complex and free adriamycin. Results suggest an advantageous role for the adriamycin DNA complex in the chemotherapy of metastatic breast carcinoma.


Improvement of pancreatic cancer model by modified treatment with N-nitroso-bis (2-oxopropyl) amine. Comparative studies were conducted in 2 groups of Syrian golden hamsters treated with N-nitroso-bis (2-oxopropyl)amine (BOP) weekly for life (group A) or weekly for 6 weeks (group B), and sacrificed at 2-week intervals. Pancreatic neoplasms developed as early as 8 weeks (group B) and 10 weeks (group A); however, in group B there were fewer, smaller lesions, well-differentiated morphologically. Liver neoplasms occurred only in group A, while gallbladder and kidney tumors were seen in both groups. A lower incidence of pulmonary adenomas was found in group B than in group A, which also had pulmonary carcinomas. The results indicate a further advance in the development of a pancreatic cancer model.


The deficiency of cyclic AMP responsive G1 controls in cultured malignant liver epithelial cells. Normal rat liver epithelial cells in culture were blocked outside the S phase by dibutyryl cyclic AMP while malignant rat liver epithelial cells were slowed in S phase. It is suggested that this lack of normal response to a growth inhibitory factor by malignant cells may underly their relatively autonomous growth behavior.


Cytoplasmic receptors for 17 beta-estradiol, 5 alpha-dihydrotestosterone and progesterone in normal and abnormal human uterine tissues. Determinations of specific cytoplasmic receptors for 17 beta-estradiol (E), 5 alpha-dihydrotestosterone (DHT) and progesterone (P) in normal and abnormal endometrium are reported. The standardization of methodology with particular emphasis on specificity trials is outlined. Receptors were present in all but one case, a moderately differentiated endometrial adenocarcinoma. Generally speaking, steroid and peptide hormone plasma content in patients with malignant conditions were at the lower limit values of normal, except for follicle-stimulating hormone which had values significantly higher than normal. The question of E competition with DHT in binding DHT-receptor and the therapeutic implications of P-receptor estimation are discussed.


Binding of N-hydroxy acetylaminofluorene to eu- and heterochromatin fractions of rat liver in vivo. The in vivo binding of N-hydroxy acetylaminofluorene (N-OH AAF) to rat liver DNA was studied in hetero- and euchromatin fractions prepared by sedimentation through sucrose gradients. The greater transcriptional capacity of the slowly sedimenting (euchromatin) fractions was confirmed by their enhanced incorporation of radioactive precursors into RNA in vivo and their increased template activity for in vitro RNA synthesis by purified RNA polymerase. N-OH AFF was bound in 4- to 5-fold greater amounts to euchromatin DNA than to heterochromatin 2 h after a single injection of the compound. However, the bound carcinogen appeared to be eliminated more rapidly from euchromatin than from heterochromatin by DNA repair processes.


A novel class of antitumour agents. I. Chemistry and animal experiments. Newly synthesized tertiary amino steroids, among them 2 beta,16 beta-dipiperidino-5 alpha-androstane-3 alpha,17 beta-diol dipivalate (DAP), were tested in three animal species as to their antitumour activity against transplantable tumours. The acute toxicity of DAP was similar to that of cyclophosphamide and considerably lower than vinblastine. Rats with Walker ascites, treated intraperitoneally with DAP, survived up to one year without ascites; untreated animals died within 10 days of transplantation. Intragastric and intraperitoneal application of DAP caused side effects suggesting a local toxicity.


A novel class of antitumour agents. II. In vitro testing. Suspensions of Walker and HeLa cells, rat fibroblasts and human stimulated lymphocytes were incubated over various periods with the synthetic amino steroid, 2 beta,16 beta-dipiperidino-5 alpha-androstane-3 alpha,17 beta-diol dipivalate (DAP). The acute cell destroying effect was found to be exponentially time and dose dependent within a certain range. With lower nontoxic doses, decreases in the mitotic and labelling indices were observed, and clotted mitotic figures occurred. The effects were essentially the same for all experimental cell types; chromosome alterations were not seen.


Protein bound carboxyl-methyl ester as a precursor of methanol formation during oxidation of dimethylnitrosamine in vitro. Protein modification with dimethylnitrosamine was studied in vitro in the presence of hamster liver microsomal fraction. Incorporation of radioactive methyl groups from dimethylnitrosamine into the exogenously added protein was dependent on the microsomal mixed function oxidase system. The methylation yielded chemically labile and stable products. The former was completely hydrolyzed by the mild alkaline treatment, pH 7.4, 100 degrees C, for 5 min and the hydrolytic product was identified as methanol indicating that the activated methyl groups from dimethylnitrosamine were incorporated into a protein as a carboxyl-methyl ester. Thus, it is suggested that methanol, recovered as one of the products during the biodegradation of dimethylnitrosamine [8], is derived, at least in part, from protein carboxyl-methyl ester which is unstable under physiological conditions.


Comparison of the biological activity of the tumor promotor phorbol myristate acetate and a metabolite, phorbolol myristate acetate, in the cell culture. Phorbolol myristate acetate, a metabolite of the tumor promotor phorbol muristate acetate in mouse skin, has one-fiftieth the potency of the parent molecule for the induction of cell division in stationary cultures of BALB/c-3T3 mouse embryo cells. Similarly, in a mixed cell culture assay devised for detection of tumor-promoting agents, phorbolol myristate acetate exhibited only a small fraction of the activity of unmetabolized phorbol ester. The results indicate that the biological activity of phorbol esters either does not require metabolic conversion or is converted by the cells used in this system and that phorbolol myristate acetate would be a tumor promotor of low potency for mouse skin compared to phorbol myristate acetate.


Metabolism of oestradiol by human mammary tumour 800 x g supernatants pretreated with dihydrolipoic acid. Incubation of human mammary tumour 800 x g supernatants with [3H]oestradiol after preincubation for 10 min at 0 degrees C with 10 mM dihydrolipoic acid resulted in metabolism of oestradiol. This was noted in 12 malignant tumours, 8 of which contained measurable oestrogen receptor levels. In 2 benign tumours lacking measurable levels of receptor, dihydrolipoic acid pretreatment had no effect. This metabolism was further stimulated by pretreatment with NAD, NADP and the anti-oestrogen Tamoxifen (ICI 46,474). When incubations were carried out in an O2 atmosphere using oxygenated buffers, the effect was suppressed.


General practitioners and postgraduate education in the Northern Region. We describe a survey of general practitioners in the Northern Region which was carried out during the summer of 1977 as the result of a commission from the Education Committee of the North of England Faculty of the Royal College of General Practitioners. Seventy-five per cent of a one in two random sample of general practitioner principals returned a postal questionnaire in which their perceptions of postgraduate education were sought and their behaviour measured by the number of sessions they attended during the previous year at their 'usual' and other postgraduate centres. Almost half the respondents had more than a basic qualification and all but three per cent had held full-time hospital appointments, two thirds of them at the level of senior house officer or above. Their experience as general practitioner principals averaged 15 years and 57 per cent held part-time appointments outside their practices. Only four per cent had not attended any postgraduate events during the previous year but the remaining respondents had attended eight sessions on average, six of which were at their usual centres. Those attending more than the average number of sessions tended to have registered between 1950 and 1969, to work in larger practices, to hold additional appointments, or to be trainers or College tutors. Most of the respondents were conservative in their perceptions of teaching methods, the topics discussed at meetings, and the contributors to postgraduate education but the younger general practitioners and a group of established general practitioners affiliated to the Royal College of General Practitioners held more radical views. They agreed about the primacy of traditional clinical topics but were sceptical of the value of ward rounds and formal lectures and favoured the seminar and clinical attachments. They saw a need for more material about practice management and wanted experienced general practitioners and community paramedical staff as teachers in addition to hospital consultants. Most of the respondents believed that their usual centres were well organized and managed but failed to cater for the special requirements of general practitioners in non-clinical aspects of practice. Lunchtime and evenings were seen as the most convenient for weekday meetings and Sunday as the most convenient day of the week. A majority of respondents believed that post-graduate education had altered their practice of clinical medicine but only 15 per cent believed it had caused changes in their practice organization.


Proposed knobs-into-holes packing for several membrane proteins. We investigated the possible side chain/side chain interactions of four potential transmembrane proteins. The basic assumptions are that the proteins are alpha-helical, and that the proteins aggregate with knobs-into-holes packing. It was found that these four proteins can be assembled into stereochemically feasible bundles of alpha-helices with hydrophobic exteriors and with hydrogen bonds between the side chains of one alpha-helix and the side chains of its knobs-into-holes packed neighbors.


Binding of mercurials to membrane suspensions and undenatured proteins. Binding capacities of membrane suspensions and dissolved compounds for mercurials were titrated by a new potentiometric method. Critical steps included a silver electrode of new design, the use of L-cysteine as a thiol buffer, a nitrogen atmosphere, and pretreatment of samples with equimolar mercurial and cysteine. Titrations had a sharp endpoint, accurate +/- 26 nmole methylmercury or +/- 8 nmole mercuric salt. Measurements of binding capacity of bovine serum albumin averaged 93% of the titer predicted for one SH group per molecule; those of human hemoglobin yielded 86-91% of the titer predicted for two SH groups per molecule. Yields dropped with exposure of protein solutions or membrane suspensions to atmospheric oxygen. Brain microsomes had significantly higher binding capacities (per milligram of protein) than red blood cell ghosts. The ratio of endpoint titers of CH3HgCl to HgCl2 averaged 2:1 in assays of cysteine, proteins, and membranes, showing that the assay was free of denaturation artifacts and protein-protein interference. Solutions of EDTA showed measurable binding of Hg2+ but not of CH3Hg+. Satisfactory titrations were also obtained with N-ethylmaleimide.


Small intestinal sugar and amino acid transport in semistarvation. The effect of semistarvation on small intestinal transport of D-glucose, L-valine, and NaCl was studied in an in vitro system of isolated rat brush border membrane vesicles. Whereas semistarvation enhanced the transport rate for L-valine by 19-29%, there was no change in D-glucose transport. When energy in the form of a NaSCN gradient was supplied to the membrane vesicles prepared from semistarved animals, L-valine was concentrated to a greater extent than those from well-fed animals. Strain differences were observed in the manner semistarvation affected NaCl transport across the brush border membrane. Semistarvation increased the NaCl transport rate by a factor of 3.5 in one rat strain and not at all in another. These results provide a partial explanation for the cellular basis of elevated neutral amino acid absorption by the small intestine in semistarvation.


Reconstitution of neutral amino acid transport system from Ehrlich ascites tumor cells. Amino acid transport systems for alanine and leucine have been reconstituted into artificial lipid vesicles. Purified plasma membrane vesicles from Ehrlich ascites cells were dissolved in 2% sodium cholate, 1 mM dithiothreitol, 0.5 mM EDTA, a mixture which solubilized approximately 50% of the membrane protein. This solubilized protein fraction was further purified by a combination of ammonium sulfate precipitations, gel filtration, and DEAE-cellulose chromatography. A fraction containing approximately 15 Coomassie blue staining bands on sodium dodecyl sulfate gels was obtained. This material was reconstituted into liposomes, and preliminary results demonstrated transport of alanine and leucine dependent on a sodium gradient. In addition, an electrogenic gradient mediated by valinomycin-induced potassium diffusion seemed to stimulate alanine uptake further.


Characterization of HeLa 5'-nucleotidase: a stable plasma membrane marker. 5'-Nucleotidase, assayed as 5'-AMPase, has been extensively characterized and established as a stable, quantitative plasma membrane marker in HeLa S3 cells. The membrane 5'-AMPase has a Km of 7.0 microM. Relative affinities of the other 5'-mononucleotides for the enzyme are 5'-GMP > 5'-TMP > 5'-UMP > 5'-CMP. There are activity optima at pH7 and 10; the latter is Mg(2+)-dependent. The membrane preparations have a small amount of acid phosphatase activity that is distinct from 5'-AMPase activity but no alkaline phosphatase. AOPCP, ADP, and ATP are strongly inhibitory. Mg2+, Ca2+, or Co2+ additions do not affect the pH 7.0 activity; Mn2+ activates slightly, whereas Zn2+, Cu2+, and Ni2+ are inhibitory. EDTA slowly inactivates, but removal of the EDTA without the addition of divalent cations restores activity. The inactivation is also substantially reversed by Co2+ or Mn2+, but reactivability by divalent cations decreases with time in EDTA. ConA strongly inhibits, and alpha-methyl-D-mannoside or glucose (the latter much less efficiently) relieves the inhibition, indicating that the 5'-AMPase is a glycoprotein. Histidine is also inhibitory. Ouabain, phloretin, cytochalasin B, cysteine, phenyl-alanine, MalNEt, and IAA are without effect. 5'-AMPase activity codistributes with pulse-bound [3H]ouabain when either of two cell fractionation procedures are used. The 5'-AMPase activity per cell is constant at different cell densities in exponentially growing cells, and activity per unit cell volume remains constant throughout the cell cycle. These properties, together with its absence in other organelles, its stability to storage, its insensitivity to certain experimental manipulations, and its general insensitivity to inhibitors of specific transport systems, make 5'-AMPase a useful quantitative marker in studies on the regulation of HeLa membrane transport systems. Key Words: HeLa, 5'-nucleotidase, plasma membrane marker, non-specific phosphatases, divalent ions, ConA, AOPCP, cell cycle, mitochondria, transport inhibitors.


Asynchronous termination of plasma membrane protein synthesis in erythroid cells. In this study we focused our attention on the terminal stages of cellular differentiation and asked the question whether the turning off of gene activity is via a general mechanism whereby all proteins are turned off synchronously, or if it is regulated specifically. We examined this problem by measuring the relative rates of synthesis of the plasma membrane proteins in cells that are near the final stages of erythroid differentiation. Our results show that although the rates of synthesis of all proteins decline during maturation the relative rates of decline are different among the various membrane proteins, indicating that the termination of plasma membrane protein synthesis during terminal differentiation is asynchronous.


Cation-sugar cotransport in the melibiose transport system of Escherichia coli. The entry of Na+ or H+ into cells of Escherichia coli via the melibiose transport system was stimulated by the addition of certain galactosides. The principal cell used in these studies (W3133) was a lactose transport negative strain of E. coli possessing an inducible melibiose transport system. Such cells were grown in the presence of melibiose, washed, and incubated in the presence of 25 microM Na+. The addition of thiomethylgalactoside (TMG) resulted in a fall in Na+ concentration in the incubation medium. No TMG-stimulated Na+ movement was observed in uninduced cells. In an alpha-galactosidase negative derivative of W3133 (RA11) a sugar-stimulated Na+ uptake was observed in melibiose-induced cells on the addition of melibiose, thiodigalactoside, methyl-alpha-galactoside, methyl-beta-galactoside, and galactose, but not lactose. It was inferred from these studies that the substrates of the melibiose system enter the cell on the melibiose carrier associated with the simultaneous entry of Na+ when this cation is present in the incubation medium. Extracellular pH was measured in unbuffered suspensions of induced cells in order to study proton movement across the membrane of cells exposed to different galactosides. In the absence of external Na+ or Li+ the addition of melibiose or methyl-alpha-galactoside resulted in marked alkalinization of the external medium (consistent with H+-sugar cotransport). On the other hand TMG, thiodigalactoside, and methyl-beta-galactoside gave no proton movement under these conditions. When Na+ was present, the addition of TMG or melibiose resulted in acidification of the medium. This observation is consistent with the view that the entry of Na+ with TMG or melibiose carries into the cell a positive charge (Na+) which provides the driving force for the diffusion of protons out of the cell. It is concluded that the melibiose carrier recognition of cations differs with different substrates.


The role of creatine phosphokinase in supplying energy for the calcium pump system of heart sarcoplasmic reticulum. An investigation of isolated and purified heart sarcoplasmic reticulum performed in the current study indicates the presence of significant creatine phosphokinase (CPK) activity in this preparation. The localization of CPK on the membrane of sarcoplasmic reticulum has been revealed also by an electron microscopic histochemical method. Under the conditions of the Ca(2+)-ATPase reaction in the presence of creatine phosphate, the release of creatine into the reaction medium is observed, the rate of the latter process being dependent on the MgATP concentration in accordance with the kinetic parameters of the Ca2+-ATPase reaction. CPK localized on the reticular membrane is able to maintain the high rate of calcium consumption by the sarcoplasmic reticulum vesicles. The results obtained demonstrate the close functional coupling between CPK and Ca2+-ATPase in the membrane of sarcoplasmic reticulum and indicate the important functional role of CPK in supplying energy for the Ca(2+)-ATPase reaction and ion transport across the membrane of heart sarcoplasmic reticulum.


Thiol-dependent subunit interactions of a major fat cell plasma membrane glycoprotein. A major glycoprotein fraction of the isolated adipocyte plasma membrane migrated in the 200,000-dalton region of dodecyl sulfate polyacrylamide gels following solubilization in 2% dodecyl sulfate/4M urea at room temperature in the absence of reductant. Limited heat treatment allowed resolution of the glycoprotein into two distinct bands in the same high-molecular-weight region plus a new 94,000-dalton glycoprotein band. Prolonged incubation of solubilized plasma membranes at 100 degrees C for 15-30 min or incubation with reductant resulted in complete conversion of the high-molecular-weight bands to the 94,000-dalton region, indicating dissociation of dimers to the monomeric form. When the dimer bands on column gels were electrophoresed in the second dimension on slab gels in the presence of reductant, no low-molecular-weight bands were observed other than that in the 94,000-dalton region. The effects of limited heat treatment to permit resolution of the two dimers and the extended treatment to convert the dimers to the monomeric form were markedly inhibited by alkylation of the solubilized membrane protein with N-ethylmaleimide or oxidation with H2O2 or diamide. However, these latter treatments did not prevent complete dissociation of dimers due to addition of reductants. These results suggest that two glycoprotein fractions may exist as dimers in the native fat cell plasma membrane. The data are consistent with a model in which the glycoprotein subunits are linked by hydrophobic bonds that are sensitive to reduction of intramolecular disulfides but are stabilized by alkylation or oxidation of the glycoprotein sulfhydryls.


A micromethod for preparation of human macrophage cultures for the study of lymphocyte-macrophage interaction in immune interferon production and blastogenesis. A new economical and reproducible micromethod for the preparation of human macrophage cultures in wells of a microtiter plate is described. The technique has been employed for the study of events which occur in the interaction of lymphocytes with macrophages in PHA-stimulated immune interferon production and blastogenesis. By comparison with the current Leighton tube macroculture systems, the microculture technique yielded a 7-fold increase in the number of macrophage cultures and a 5-fold increase in the number of T lymphocyte macrophage cultures from a given volume of blood. The replicability from sample to sample with regard to 3H-thymidine incorporation and amount of interferon produced is better in the microculture system than in the Leighton tube system. Such a microculture technique will thus provide a system whereby ready analysis of monocyte-macrophage function and interaction with lymphocytes in numerous disease states can now be realized.


Modulation of the acquisition and expression of immunity by Tilorone: I. Delayed-type hypersensitivity responses. The chronic administration of Tilorone [(2,7-bis(diethylamino)ethoxy]fluoren-9-one) with antigen seems to prevent the generation of DTH effector cells. The administration of Tilorone proximal to DTH challenge blocks the expression of DTH. By local transfer of DTH effector cells, this latter effect can be shown to be due to the lack of nonspecific inflammatory cells. By the use of both systemic and local transfer systems, it is also shown that specific DTH effector cells from animals treated with Tilorone are unable to circulate in the normal manner. Overall, these experiments suggest that Tilorone does not block the generation of DTH effector cells, but it does greatly alter the expression of DTH. The relation between these effects and the known interferon induction capacity on Tilorone remains unclear.


Modulation of the acquisition and expression of immunity by Tilorone: II. Humoral responsiveness. Tilorone (2,7-bis[(diethylamino)ethoxy]fluoren-9-one) can enhance the humoral immune response of mice to SRBC although Tilorone has no effect on functional activity of splenic B cells in the absence of immunogen. Tilorone does not alter the effective level of T cell help but does seem to alter the level of effective suppression. Tilorone seems to be an adjuvant only in the sense of removing a normal homeostatic control mechanism, perhaps suppressor T cells.


Quantitative comparisons of various biological responses of neutrophils to different active and inactive chemotactic factors. The effects of chemotactic factors on rabbit neutrophils were evaluated measuring cell migration in modified Boyden chambers and under agarose, in lysosomal enzyme release, leukocyte aggregation, and in vivo neutropenia. Chemotactins employed included the complement-derived C3 and C5 fragments, the bacterial chemotactic factor from culture supernatant fluids of Escherichia coli, and the synthetic chemotactic factors Met-Leu-Phe and formyl-Met-Leu-Phe. A consistent parallelism was found in all the leukocyte responses to a given chemotactic factor. In no instance, with any of the five chemotactic factor preparations, did cells responding in one assay system fail to respond in the four other assay systems, suggesting a common event in all of the cell responses. Boyden chamber chemotaxis was consistently the most sensitive assay; the agarose assay was, in general, less sensitive by a factor of 100 fold. Enzyme release approached, in cell sensitivity to chemotactic factors, that of the Boyden chamber assay. In general, in vitro leukocyte aggregation and in vivo neutropenia were considerably less sensitive assays. Chemotactic factor inactivator (CFI) purified from human serum destroyed in parallel all biological activities of C3 and C5 chemotactic factors but had no effect on the bacterial chemotactic factor and the activities of synthetic chemotactic peptides.


Cellular aspects of conjunctival inflammation induced by the synergistic action of histamine and prostaglandins. Histamine or prostaglandin (PG) E1 or E2 administered to rabbits topically alone in high doses produced conjunctival vasodilation associated with little or no edema while their mixture at lower concentrations produced conjunctival vasodilation associated with profound edema. Sections of tissues treated with the mixture of histamine and PGE1 or PGE2 showed widespread epithelial and subepithelial inflammatory cellular infiltration. Conjunctival smears from eyes treated with the histamine/PG mixture contained small lymphocytes and polymorphonuclear leukocytes, including eosinophilic and occasionally basophilic cells. Differential staining of the polymorphs demonstrated both eosinophils and pseudoeosinophils. Histological examination of the conjunctival smears and sections of the lids obtained from eyes treated with either histamine or PGE1 or PGE2 alone did not show any detectable increase of inflammatory cells when compared to normal controls. The clinical and histological results indicate that the synergistic effect of histamine with PGs of the E-type in the conjunctiva produces an inflammatory response similar to that seen in various clinical forms of human allergic conjunctivitis. Such a response could not be produced by histamine or PGE1 or PGE2 alone even at much higher doses than in the mixture. The data indicate that an interplay of several different mediators may be crucial in the conjunctival response in allergy.


Conditions for generating and assaying human leukocyte inhibitory factor (LIF) induced by purified protein derivative (PPD). This report examines a variety of experimental conditions for the production of human leukocyte inhibitory factor (LIF). The data indicate that the production of LIF as measured in the indirect capillary-tube migration inhibition assay using human polymorphonuclear indicator cells accurately reflects delayed hypersensitivity to purified protein derivative (PPD) as detected by skin-test reactivity. Mononuclear cells and semipurified lymphocytes separated from whole blood by sedimentation in Ficoll-Hypaque were able to generate LIF in response to PPD. The quantity of cells responsible for LIF production were standardized and as little as 1 x 10(6) mononuclear cells were required for detectable LIF production. LIF produced by mononuclear cells required only temporary exposure to PPD, thus eliminating the necessity for a control to which antigen was added at the end of culture to antigen-free supernatants. In addition, the removal of antigen after a brief exposure helps to avoid the possible toxic effect of some antigens on the migrating polymorphonuclear leukocytes (PMNL) population. LIF production did not require extraneous serum protein (i.e., fetal bovine serum). Further, kinetic studies indicated that LIF was detected as early as 8 hr following a 2 hr-exposure to PPD and that even a 1 hr-exposure was sufficient to generate measurable detectable quantities of LIF. After 48 hr of culture, supernatants were found to contain considerable amounts of LIF, with reactivity at dilutions as high as 1:40.


Enhancement of macrophage adenylate cyclase by microtubule disrupting drugs. The accumulation of cyclic adenosine 3',5'-monophosphate (cAMP) in guinea-pig macrophages exposed to the adenylate cyclase (AC) stimulators prostaglandin E1 (PGE1) and isoproterenol (IP), was markedly enhanced by pretreatment of the cells with colchicine, vinblastine, and podophyllotoxin--agents which prevent microtubule assembly. The same agents did not augment basal cAMP levels. The facilitating effect of the drugs on the response to PGE1 and IP developed both in the absence and presence of a phosphodiesterase (PDE) inhibitor. The same drugs also enhanced the accumulation of cAMP induced by cholera toxin (CT) but the presence of a PDE inhibitor was required for such enhancement to become evident. Pretreatment of macrophages with cytochalasin B, an agent interfering with microfilament function, had no effect on the responsiveness of the cells to AC stimulators. The microtubule stabilizer, deuterium oxide (D2O) partially reversed the colchicine effect. Microtubule disrupting drugs did not block the release of cAMP from the cells into the surrounding medium. Macrophages incubated as monolayers or in suspension showed the same degree of increased responsiveness to stimulators after preexposure to colchicine. Preincubation with the ionophore A23187, which elevates the intracellular concentration of Ca2+, also enhanced the stimulation of AC by PGE1 and IP. Microtubule disrupting agents did not potentiate AC activity in broken cell preparations, whether added to the intact cells before disruption or directly to the enzyme assay mixture, nor did they affect PDE activity of macrophage sonicates. Moderate enhancement of PGE1-induced cAMP formation was also seen in colchicine- and vinblastine-treated lymphocytes. It was concluded that microtubules control the activity of AC by restricting the mobility of membrane receptors. Disruption of microtubules by drugs results in the removal of such restraints and an augmented chance of productive interactions between receptors and catalytic units of AC.


Physical mapping of the restriction fragments obtained from bacteriophage T4 dC-DNA with the restriction endonucleases SmaI, KpnI and BglII. The cytosine-containing DNA of a mutant of bacteriophage T4 was digested with restriction endonucleases SmaI, KpnI and BglII producing 5, 7 and 13 fragments respectively. Complete physical maps of the T4 genome were constructed with the enzymes SmaI and KpnI and an almost complete map with the enzyme BglII.


Enzyme immunoassay of a beta-adrenergic agent using beta-galactosidase as label. Antisera against tetrahydronaphthalenols, which are conformationally rigid derivatives of adrenergic catecholamine, were produced in rabbits immunized with trans-5-amino-6-hydroxy-2-isopropylamino-1,2,3,4-tetrahydronaphthalene-1 -ol (I) conjugated to succinylated bovine serum albumin at the C5 position on the tetralin ring. Antisera were screened by immunodiffusion and further characterized by passive hemagglutination assay using erythrocytes sensitized with trans-I-ovalbumin conjugate and by enzyme immunoassay using trans-I-beta-galactosidase conjugate. Cross-reactivity studies indicated that the antiserum was highly specific for the tetralin structure and for substitution at the C2 position. The antiserum also selectively discriminated the stereoisomers about the C1-C2 bond. The anti-trans-I serum was used to develop EIA for trans-5-hydroxymethyl-6-hydroxy-2-isopropylamino-1,2,3,4-tetrahydronapht halene- 1-ol (IIb), which exhibited strong beta-stimulating activity fairly selective to tracheal muscle, since it recognized trans-IIb to the same degree as trans-I. The assay could detect as little as 100 pg of this compound. The mean recovery of trans-IIb added to plasma was 105%, and values for plasma trans-IIb determined by this immunoassay correlated well with those determined by gas chromatography-mass spectrometry.


Effect of preconditioning unconditioned stimulus experience on learned taste aversions. One taste-aversion study using male Long-Evans rats in which ethanol was the unconditioned stimulus (UCS) and six studies in which lithium chloride (LiCl) was the UCS demonstrate that (a) exposure to the UCS prior to conditioning retards subsequent acquisition of learned taste aversions; (b) a single preconditioning UCS exposure is sufficient to attenuate conditioning; (c) the preconditioning UCS exposure must occur within a limited period prior to conditioning to attenuate learning; (d) repeated conditioning trials will override the effect of prior exposure to the UCS; (e) tolerance to the UCS is not a necessary condition for the attenuation effect to occur; (f) pairing the preconditioning UCS with a novel flavor other than the CS does not remove the preexposure effect, although it may reduce its magnitude; and (g) the degree of disruption is a positive function of preconditioning UCS dosage and an inverse function of conditioning UCS dosage.


Factors affecting the measurement of classically conditioned fear in rats following exposure to escapable versus inescapable signaled shock. Three experiments are reported in which rats first received 50 escapable or inescapable signaled-shock trials. Experiment 1 (n = 22) employed an acquired-drive paradigm and found inescapable shock subjects learned a hurdle-jump response to escape the signal less rapidly than did escapable-shock subjects. Experiment 2 (n = 24) employed a conditioned emotional response paradigm and found inescapable-shock subjects suppressed more when the signal was introduced in the appetitive bar-pressing task. Both experiments measured spontaneous activity immediately following conditioning and found no group differences. Experiment 3 (n = 39) employed the same activity task and found no difference between escapable- and inescapable-shock groups when the signal was introduced into the activity task. Both groups displayed less activity than a nonshock control group during the signal. The results suggest that lack of control over the shock in the conditioning phase did not result in an increase of conditioned fear. The results are discussed in terms of a learned active-inactive predisposition to respond.


A possible case of subcorneal pustular dermatosis in an Egyptian mummy. Examination of the mummy of an Egyptian who died in 892 BC +/- 53 years showed the skin to be well preserved, with inguinal subcorneal vesicles of the type seen in subcorneal pustular dermatosis. This condition apparently antedates by almost three millenia its description as a clinicopathological entity by Sneddon and Wilkinson in 1956.


The platelet defect in leukemia.  Platelet ultrastructure, adenine nucleotide metabolism, and the release reaction. The ultrastructure and adenine nucleotide metabolism of platelets from patients with acute leukemia were studied to elucidate possible mechanisms for the platelet dysfunction observed in this clinical setting. Nonstimulated (resting) platelets from leukemic patients varied greatly in size; exhibited marked variation in the number of alpha granules present per cell; had poorly delineated circumferential bands of microtubules; and often grossly dilated open channel systems or cytoplasmic vacuolization. The intracellular concentrations of ATP and ADP were significantly below normal, and the specific radioactivity of ATP and ADP of nonstimulated platelets in leukemia was equivalent to or exceeded that seen in stimulated normal platelets. Addition of ADP or collagen to platelets from leukemic patients was followed by retarded and incomplete shape change, delayed and incomplete centripetal migration of subcellular organelles, impaired degranulation, and the formation of loose aggregates composed of relatively few platelets. Stimulation of "leukemic" platelets with collagen led to the release of significantly subnormal amounts of ATP and ADP and no significant change in the specific radioactivity of the intracellular nucleotides. In contrast to the results in normal platelets, the conversion of ATP to inosine monophosphate and hypoxanthine in platelets in leukemia failed to increase significantly with collagen stimulation. The results indicate that abnormalities exist in the storage pool of adenine nucleotides and the release mechanism of platelets in acute leukemia. These defects appear to contribute to an impairment in the release reaction in these platelets. Many of the ultrastructural and metabolic defects seen in acute leukemia occur in platelets in preleukemia.


Nonmechanical factors affecting aortocoronary vein graft flow. To study aortocoronary vein graft flow, hemodynamic measurements were made in 17 patients with 22 grafts while on cardiopulmonary bypass during ventricular fibrillation and sinus rhythm and 15 minutes after termination of cardiopulmonary bypass. When compared with graft flow to the stable working heart, flow to the nonworking heart in sinus rhythm was reduced 50%. When fibrillating, the nonworking heart received 33% more flow than when in sinus rhythm but still significantly less flow than the working heart. All hemodynamic parameters measured, including flow in 12 grafts, remained stable between 15 minutes and 60 minutes following cardiopulmonary bypass. In contrast, heart rate and flow in 10 grafts fell within five minutes after 1 mg of propranolol was given intravenously. Although it was still reduced, heart rate had begun to rise 30 minutes after propranolol, but graft flow did not begin to return toward predrug control levels until over 60 minutes after drug administration. Further reduction in aortocoronary vein graft flow after heart rate had started to increase supports the possibility of a direct propranolol action reducing myocardial oxygen consumption in addition to its bradycardiac effect. This study shows the sensitivity of intraoperative vein graft flow to physiological and pharmacological factors and the need to standardize the technique and timing of flow measurements if their value for predicting long-term graft patency is to be determined.


Results of reoperation for unsuccessful coronary artery bypass surgery. Twenty-one reoperations for relief of angina have been performed at Stanford Medical Center in patients who had had previously unsuccessful direct coronary artery surgery. Complete releif of angina was obtained in seven (33%), improvement of angina in six (29%), and no change in angina in eight (38%) after these operations. Reoperation was least successful in relieving angina when performed in patients with graft occlusion and most successful when performed in patients with new critical lesions or with previously unbypassed critical lesions (P less than 0.02). Reoperation was of greater benefit in patients with unstable angina than in those with stable angina (P less than 0.05). Four of the seven patients having no angina after reoperation sustained perioperative infarctions. Reoperation does not provide the same rate of angina relief as first operations; however, certain subgroups of patients may exhibit more favorable results.


Effect of cardiotonic and vasoactive drugs on transmural flow distribution and ventricular volume in the fibrillating heart on cardiopulmonary bypass. Subendocardial hemorrhagic necrosis in an important cause of death following cardiopulmonary bypass. The transmural distribution of flow across the left ventricle (LV), septum (SP), and right ventricle (RV) is a complex interaction of vascular resistance and myocardial compressive resistance. We studied the change in transmural blood flow in LV, SP, and RV, and left ventricular volume, following administration of cardiotonic and vasoactive drugs in the fibrillating heart. The drugs studied included calcium with and without ATP-induced vasodilation, isoproterenol, epinephrine, angiotensin, and ouabain. Calcium produced underperfusion of LV subendocardium with or without previous ATP vasodilation. Isoproterenol also caused underperfusion of LV subendocardium. Both calcium and isoproterenol decreased ventricular volume. Angiotensin increased resistance in the subepicardium and increased flow in the subendocardium, with no change in ventricular volume. Epinephrine and ouabain caused no consistent changes in transmural flow. The decreased ventricular volume produced by calcium and isoproterenol restricts flow in the subendocardium because of increased compressive resistance. Increased subendocardial flow with angiotensin indicates that subepicardial vasodilation in the fibrillating heart causes epicardial "steal," which contributes to subendocardial ischemia.


Does cardiac transplantation prolong life and improve its quality? An updated report. The current status of the human cardiac transplant experience at Stanford University Medical Center is presented in order to reassess its role in the treatment of end-stage cardiac disease. Of 109 patients undergoing transplantation at Stanford between January 1968 and August 1976, 44 were still alive as of August 1, 1976. The overall 1- and 2-year survival rates for the series are 52% and 43%, respectively. Sixty-nine patients have survived more than 3 months, and their overall 1- and 2-year survival rates are 80% and 66%, respectively. Of the 3-month survivors, 62 (90%) returned to functional Class I New York Heart Association cardiac status and most of these returned to their pre-illness activities. Of 40 patients selected for transplantation for whom a donor did not become available, 38 were dead in less than 6 months. Complications related to immunosuppression with steroids are currently the major barrier to longer survival and improved rehabilitation postransplantation. On the basis of these data we conclude that cardiac transplantation not only prolongs survival, but can return carefully selected recipients to an active life.


The effect of mammalian gonadotropins and homologous gonadotropins on the activity of delta 5-3 beta-hydroxy-steroid-dehydrogenase (HSD) in hypophysectomized Rana esculenta. Mammalian gonadotropins and partially purified gonadotropins (FSH and LH) from an amphibian (Rana esculenta) were used to investigate their action on enzymatic activity of delta 5-3 beta-HSD in male and female hypophysectomized Rana esculenta in stage of genital activity (October) and in stasis or hibernation stage (January). Amphibian and equine LH stimulate delta 5-3 beta-HSD activity especially in females hypophysectomized in the stage of genital activity, while ovine FSH action is very low and amphybian FSH practically inactive. The present data suggested that control of gonadotropic activity in the reproductive cycle in Rana esculenta is LH-dependent.


Histochemical and biophysical study of cuticle sclerotization in Pycnoscelus surinamensis L. (Blattaria). The problem of melaninogenesis and quinone tanning of the cuticle was examined by histochemical and biophysical methods (electroparamagnetic resonance: EPR) on normal subjects of Pycnoscelus surinamensis and on subjects with abnormal cuticular colour. The cuticle of abnormal subjects showed a lower content of polyphenolic substances and a greater positivity for the indole group. This suggests that in these insects tanning products can be synthetized differently and not derived from tyrosine but from tryptophan as postulated by Pryor (1955). Furthermore, a higher number of unsaturated aminic groups is found in abnormal subjects. Granules present only in the cytoplasm of the epidermal cells of the abnormal newly moulted subjects may indicate that the polyphenolic compound of tanning, secreted in an inactive form as 4-0, beta-glucoside, is not freed from the beta-glucosidase and remains as such in the cytoplasm.


Evaluation of cardiovascular and pulmonary changes during meperidine-diazepam anesthesia. Studies were conducted in 189 oral surgery out-patients to evaluate the combination of meperidine hydrochloride and diazepam for anesthesia and to determine its effect on the cardiovascular and pulmonary systems. Results showed that neither ventricular cardiac arrhythmias nor clinically significant respiratory depression occurred in the sedated patients; meperidine-diazepam proved to be satisfactory for oral surgical procedures.


In vitro polymerization of microtubules into asters and spindles in homogenates of surf clam eggs. The eggs of the surf clam Spisula solidissima were artificially activated, homogenized at various times in cold 0.5 M MES buffer, 1mM EGTA at pH 6.5, and microtubule polymerization was induced by raising the temperature to 28 degrees C. In homogenates of unactivated eggs few microtubules form and no asters are observed. By 2.5 min after activation microtubules polymerize in association with a dense central cylinder, resulting in the formation of small asterlike structures. By 4.5 min after activation the asters formed in vitro contain a distinct centriole, and microtubules now radiate from a larger volume of granular material which surrounds the centriole. By 15 min (metaphase I) the granular material is more disperse and only loosely associated with the centriole. Microtubules are occasionally observed which appear to radiate directly from one end of the centriole. The organizing center can be partially isolated by centrifugation of homogenates of metaphase eggs and will induce aster formation if mixed with tubulin from either activated or unactivated eggs. Pretreatment of the eggs with colchicine does not prevent the formation of a functional organizing center. Complete spindles can also be obtained under polymerizing conditions by either homogenizing the eggs directly into warm buffer or by adding a warm high-speed supernate to spindles which have been isolated in a microtubule stabilizing medium. Extensive addition of new tubulin occurs onto the isolated spindles, resulting primarily in growth of astral fibers, although there occasionally appears to be growth of chromosomal fibers and of pole-to-pole fibers. Negatively stained aster microtubules have a strong tendency to associate side by side, and under some conditions distinct cross bridges can be observed. However, under other conditions large numbers of 300-400-A particles surround the microtubules; the presence of stain between particles can give the appearance of cross bridges.


An ultrastructural examination of the role of cell membrane surface coat material during neurulation. Data from neural crest cultures indicate that cell surface coat material (CSM) is directly involved in cellular migration and events surrounding differentiation. To investigate whether the CSM also has a morphogenetic role, embryos of the amphibian Ambystoma maculatum were examined ultrastructurally throughout the stages of neurulation. Segments of the neural axis were fixed in glutaraldehyde-containing Alcian blue 8GX, which reportedly enhances preservation of CSM, and were postfixed in OsO4 containing 1 percent lanthanum nitrate, which stains the CSM. The medial groove formed by the appearance of the neural ridges contains a large amount of CSM and numerous vesicles coated with lanthanum-positive material. In contrast, the lateral ridge surfaces are covered by a small amount of uniformly distributed CSM and a paucity of vesicles. As the ridges begin to fold there is a progressive increase in the amount of CSM within the presumptive neural tube region. Further convergence of the neural folds is accompanied by an increase of CSM at their leading edges. As the folds approximate each other, lanthanum-positive material physically bridges the gap. However, as the apposing tissue actually abuts to form the neural tube, no CSM is observed in the remaining interspace. The specific distribution and sequential accumulation of cell CSM during the events of neurulation strongly suggest its direct participation in the morphogenetic process.


Molecular organization of rat prolactin granules. I. In vitro stability of intact and "membraneless" granules. Studies carried out on a number of secretory cell systems suggest that the specific cytoplasmic granules in which the secretion products are stored before their release are complex organelles which can possess a distinct molecular organization. For instance, it has been reported that in some granules the segregated secretion products are organized into crystalline structures (1-3) or large intermolecular aggregates (4-8). It is likely that all phenomena of this type are favorable to the economy of the cell, in the sense that they reduce the energy required for storage of the secretion products. The prolactin (LTH) granules of the rat pituitary possess a number of morphological features which strongly suggest that the molecules(s) of their content might be arranged in a relatively stable structure. Thus, these granules are remarkably polymorphic in shape, and their membrane is usually separated from their content by a clear space. Furthermore, identifiable LTH granules devoid of their membrane are often seen in the pericapillary space, suggesting that upon discharge by exocytosis they are dissolved only slowly (9). However, no studies specifically concerned with the mechanisms of LTH storage have been reported so far. In order to obtain some information on this question, we have studied the behavior of isolated granule fractions incubated in vitro under a variety of carefully controlled experimental conditions.


The role of cortical orientation in the control of the direction of ciliary beat in Paramecium. The swimming behavior of many ciliate protozoans depends on graded changes in the direction of the ciliary effective stroke in response to depolarizing stimuli (i.e., the avoiding reaction of Paramecium). We investigated the problem of whether the directional response of cilia with a variable plane of beat is related to the polarity of the cell as a whole or to the orientation of the cortical structures themselves. To do this, we used a stock of Paramecium aurelia with part of the cortex reversed 180 degrees. We determined the relation of the orientation of the kineties (ciliary rows) to the direction of beat in these mosaic paramecia by cinemicrography of particle movements near living cells and by scanning electron microscopy of instantaneously fixed material. We found that the cilia of the inverted rows always beat in the direction opposite to that of normally oriented cilia during both forward and backward swimming. In addition, metachronal waves of ciliary coordination were present on the inverted patch, travelling in the direction opposite to those on the normal cortex. The reference point for the directional response of Paramecium cilia to stimuli thus resides within the cilia or their immediate cortical surroundings.


Sequential expression of germ line genes in development of immunoglobulin class diversity. Differentiation of B cells occurs in two discontinuous stages. Primary differentiation of stem cells to B lymphocytes in birds occurs exclusively in the lymphoepithelial bursa of Fabricius; the fetal liver may serve this function in mammals. In chickens both the size of the B-lymphocyte pool and the generation of precursors for cells secreting different immunoglobulin classes is controlled by the bursa. The latter process involves the sequential expression of genes coding for heavy chain constant regions in the order mu, gamma, alpha. The second stage of B-cell differentiation is antigen-driven, and involves proliferation and maturation of B lymphocytes to plasma cells. Ontogenetic development of different classes of B lymphocytes in mammals is orderly, independent of exogenous antigens, and occurs in the sequence mu, gamma, alpha. A developmental switch in expression of Ch genes, beginning with mu, has been experimentally verified. We favor the hypothesis that generation of class diversity of B lymphocytes occurs during the antigen-independent first stage of differentiation, and that the genetic switch in Ch gene expression follows the sequence mu leads to gamma leads to alpha, but evidence of these points remains inconclusive.


Normal and altered phenotypic expression of immunoglobulin genes. Genetically controlled intraspecific differences between immunoglobulins (allotypes) provide valuable markers for the study of the quantitative expression of allelic and nonallelic alternative forms of immunoglobulins (Igs) during the normal development of rabbits. Heterozygous rabbits are mosaics of cells expressing different Ig-genes since fully differentiated productive cells generally secrete only one of alternative forms of Ig. The proportions of cells that differentiate to produce allelic forms of immunoglobulins during normal development depend on the particular heterozygous genotype. The normal proportions of some markers can be drastically altered if the differentiation of lymphoid cells in the young rabbit occurs in the milieu of antibody specific for one form (allotype suppression). An initiating step in the establishment of persistent allotype suppression is probably the interaction of antiallotype antibody with allotype-bearing receptors on lymphoid cell surfaces, but the mechanism for the maintenance of a state of chronic suppression may well be more complex. Allotype suppression can be viewed as one example of numerous immunological phenomena that reflect specific and finely tuned regulatory mechanisms governing the differentiation and clonal expansion of lymphoid cells destined to secrete immunoglobulins.


Expression of specific clones during B cell development. The expression of DNP- and TNP-specific B cells in spleens of neonatal BALB/c mice was analyzed by the in vitro splenic focus technique. B cells of these specificities were found to be present in slightly higher frequency in neonatal than in adult spleens. The parameters of stimulation of neonatal B cells were similar to those of adult B cells but the antibody-forming cell progeny of neonatal B cells produce predominantly gammaM rather than gammaG antibody and produce less antibody than the progeny of adult B cells. Isoelectric focusing analyses of monoclonal antibodies derived from neonatal B cells stimulated in vitro with DNP or TNP revealed that over 90 per cent of the antibodies could be identified as belonging to one of six predominant clonotypes, three specific for DNP and three for TNP. While individual neonates rarely expressed all of the predominant clonotypes, B cells of each of the six clonotypes were found in several donors. When B cells of a given predominant clonotype were present in an individual many such B cells could be found and in many cases the entire DNP- or TNP-specific B cell population of an individual could be accounted for by B cells of a single clonotype. These findings are discussed in terms of the diversity of clonotype specificities available in neonates, the kinetics of development of cells within a clonotype, and factors that may play a role in controlling the expression of B cell clones.


Neurons of the dorsal lateral geniculate nucleus of the albino rat. Light and electron microscopic observations were made on the dorsal lateral geniculate nucleus (DLGN) of 33 young adult male albino rats. Three variants of the Golgi silver impregnation technique were employed in the light microscopic studies. Neurons were classified into three categories based on location, dendritic pattern, and dendritic appendages. Type 1 and type 3 neurons were distributed throughout the DLGN. Type 2 neurons were located in the superficial zone. Dendritic appendages of type 1 and type 2 neurons indicated these cells may function as geniculo-cortical relay neurons. The type 3 neurons had lobulated dendritic appendages and an axon that terminated withinthe nucleus. Type 3 neurons may represent Golgi-type-II interneurons. Camera lucida drawings, photomicrographs, and electronmicrographs illustrate the characteristics ofthe three cell types. The literature on ultrastructural and neurophysiological findings may substantiate the presence of three neuronal types. Initially, the rat DLGN does not appear as elaborately organized as the nucleus observed in cats and primates; however, there are notable similarities in neuronal morphology and synaptology.


Accumulation of lipofuscin (age pigment) in aging skeletal connective tissues as revealed by electron microscopy. An EM survey of aging skeletal connective tissue was made for the presence of lipofuscin accumulation. Twenty-six mice of both sexes, 5 to 130 weeks of age were perfused with glutaraldehyde. Femoral periosteal samples were fixed in cold glutaraldehyde, decalcified in EDTA, and post-fixed in osmium tetroxide. Sections were stained with uranyl acetate and lead citrate. Lipofuscin was absent in all cell types in mice 5 and 26 weeks of age. At 52 weeks, single membrane-bound lipofuscin granules were observed in fibrocytes, precursor osteogenic cells (preosteoblasts), osteoblasts, and osteocytes. With increasing age (78 to 104 weeks) lipofuscin was observed more frequently, as well as the number of granules per cell. At 104 and 130 weeks, many of the remaining cells revealed age pigment. These observations point to a wider distribution of age pigment in various tissues than previously realized. The presence of lipofuscin in connective tissue may facilitate a reevaluation of the role of this pigment in cellular aging.


A new antigen-antibody system. Clinical significance in long-term carriers of hepatitis B surface antigen. A new antigen-antibody system was recently described in hepatitis B surface antigen (HBSAG(-positive sera. Despite indications of heterogeneity in specificity, the designations "e antigen" and "e antibodies" are used for the system as such in this articly. E'IGHT OF 17 long-term carriers of HBSAg with a histological picture of chronic persistent hepatitis or chronic aggressive hepatitis carried the e antigen, while none had demonstrable e antibodies in serum. Ten of 12 healthy carriers with e antibodies were blood donors who had donated 95 units of blood; none of these carriers was associeated with a reported case of posttransfusion hepatitis. In five individuals in the incubation stage of hepatitis B, e antigen appeared simultaneously with HBSAg but before the rise in transaminase levels. This finding further links e antigen to hepatitis B.


Diagnosis of plasma cell myeloma.

Diagnosis of plasma cell myeloma. Diagnosis of myeloma depends on identification of malignant plasma cells and the product of these cells, a monoclonal immunoprotein. Of the clinical manifestations of plasma cell myeloma, skeletal pain and anemia are two of the more common. Unexplained anemia and osteoporosis noted in the elderly should suggest the possibility of myeloma; this combination of symptoms certainly warrants obtaining a protein electrophoresis. Hypercalcemia and renal insufficiency are frequent sequelae of myeloma.


Intracardiac electrography in children and young adults. The interpretation of IE recorded in children has been hampered by a lack of agreement regarding normal values. We recorded IE in 158 children and young adults (ages, three days to 33 years) to define the various conduction intervals in normal and disease states. The HBP was recorded in 156 subjects. In 85 subjects with normal conduction indicated by surface ECG, including 19 subjects with normal hearts, there were no statistically significant age-related differences in internodal, A-V nodal, or His-Purkinje conduction intervals. Therapeutic levels of digitalis did not alter the conduction intervals. In 11 subjects with first degree A-V block and in five subjects with congenital complete A-V block, the site of block as determined by IE could not be predicted from the surface ECG. No abnormalities in conduction intervals were found in 18 subjects with right bundle branch block (surgically induced in 17 cases). Intracardiac electrography with recording of the HBP was found to be a safe, informative technique for electrophysiologic investigations in children and young adults.


An autopsy study of human axillary lymph node histology. This is a histologic study of axillary lymph nodes removed from 487 random autopsies. Histologic features such as germinal centers, deep cortical regions, sinus histiocytosis, hyalin deposition, fibrosis and overall cellularity were evaluated in each case. Results were correlated with the age of the deceased and cause of death. The results a) support the general opinion that germinal centers are more frequently found in children and young adults than in the old, b) showed that hyaline deposits increase with age, c) showed that lymphocyte depletion associated with fibrosis and hyaline deposits was found in patients dying of chronic disease, particularly cancer, d) pointed out a selective depletion of the deep cortical regions in patients dying of hemorrhage, and e) contributed to knowledge of the morphology of axillary lymph nodes in neonates. (Am J Pathol 78:7-22, 1975)


Immunochemical staining of the rat adenohypophysis in organ culture. Rat anterior pituitaries were cytologically studied following cultivation in organ culture, with and without the addition of hypothalamic and cortical extracts. Although five distinct cell types could be identified with classical stains in the uncultivated glands, the peroxidase-labeled antibody technique (using antibodies against STH, LTH, FSH, LH and TSH) showed that not all of the immune-specific cell types were being identified with the classical stains. This discrepancy was magnified following culture as chromophilic cells seen with classic stains decreased in number with an increase in culture time. The peroxidase technique, however, revealed that all cells remained constant in type and number regardless of time in culture. While the addition of either hypothalamic or cortical extract to the culture medium produced cytological alterations demonstrated by the classical dyes, the antibody technique showed no such alterations. Such a comparison of staining techniques emphasizes the hazards of relying solely on histological procedures to reveal the hormonal activity of the pituitary gland.


Junctional complexes in the preimplantation rabbit embryo. The morphology and development of junctional complexes between blastomeres of the preimplantation rabbit embryo were investigated using several approaches. Electron microscopic examination of embryos stained en bloc with uranyl acetate, and the study of junction permeability using horseradish peroxidase and lanthanum nitrate provided information on structure, intermembrane spacing and permeability of the junctional complexes. In addition, the freeze fracture technique was used with day 5 and day 6 blastocysts, since the large size of these embryos facilitated use of this method. These experiments showed that although rudimentary junctions were present between blastomeres of the early cleavage stages, effective tight junctions were not present until the blastocyst stage. Electron microscopic examination of thin sections revealed apical foci of membrane approximation or "fusion" between trophoblast cells by day 4. Freeze fracturing revealed a lattice of interconnecting ridges (on the A face) and grooves (on the B face) in the apical region between trophoblast cells of the day 5 blastocyst. This lattice formed a continuous band along the apical margin of each cell, and therefore constituted a zonula occludens. The zonula occludens of the day 5 blastocyst averages 2-3 ridges per lattice, while day 6 blastocysts had lattices that averaged 5-6 ridges. Also seen in the freeze fracture replicas from the day 5 and day 6 blastocysts were local accumulations of intramembranous particles on the A face. These particles were often observed in aggregates similar to those of previously described gap junctions. It could not be determined whether these small regions of particles were true gap junctions or a possible primitive form of gap junction because the complementary pitted surfaces (B face pits) were not demonstrated.


The variable fine structure of elastin visualized with Verhoeff's iron hematoxylin. Verhoeff's iron hematoxylin (VIH) followed by lead citrate (LC) applied to epoxy thin sections stained the dense component of elastic fibers heavily and the peripheral microfibrillar component lightly in guinea pig trachea and mouse testis fixed with a glutaraldehyde-osmium tetroxide sequence. This method stained large fimbriated fibers beneath tracheal epithelium, small fibers and stacked aggregates thereof in the deep lamina propria, cartilage and adventitia of the trachea and large stacked fibers in the fibroelastic band of the trachea. Fibers of the fetus differed from those of the adult, especially in the subepithelial elastic lamina of the trachea. Elastic fibers were intimately associated with fibroblasts and particularly slender fibroblast processes in tracheal stroma and with chondrocytes in tracheal cartilage. Fibroblasts associated with elastic fibers in the tracheal subepithelial lamina propria were often closely bordered by eosinophils, mast cells, or monocytes. Occasional mast cells extended slender processes around elastic fibers in the subepithelial lamina propria. In mouse testis and in many regions of the trachea, small elastic fibers were identified which were below the limits of resolution for the light microscope and were not apparent at the ultrastructural level in routinely stained thin sections.


The effect of ruthenium red and Concanavalin A on the vitelline surface of fertilized and unfertilized rabbit ova. Fertilized and unfertilized rabbit ova exposed to ruthenium red with the zona intact or after mechanical removal, for the demonstration of mucopolysaccharides on the surface coat. Ova were also exposed to Concanavalin A. Ruthenium red bound strongly to elements of the zona and increased the opacity of the plasmalemma. There was no notable change in staining of the vitellus following fertilization. Although there were ruthenium red stained bodies resembling cortical granules in crypts in the plasmalemma, there was no evidence for cortical granule extrusion of mucopolysaccharides. The Concanavalin reaction was seen over both unfertilized and fertilized ova, demonstrating receptors for terminal alpha-D-mannopyranosyl, alpha-D-glycopyranosyl, beta-D-fructofuranyosyl residues. The deposit on the fertilized egg surface was enormously enhanced indicating a change in the distribution of receptor sites.


Benzopyranopyridine derivatives. 1. Aminoalkyl derivatives of the azaxanthenes as bronchodilating agents. The preparation of the four isomeric azaxanthones 3 and a number of their aromatic ring substituted derivatives is described. These ketones were converted into the title compounds which were examined for their biological properties. The most interesting compound in this series, the 1-methyl-4-piperidylidene derivative of 1-azaxanthene, shows the profile of an orally effective potent bronchodilating agent as well as a moderate antihistamine. Biological properties of this compound were compared to a number of antihistamines as well as known bronchodilating agents. Structure-activity relationships are also discussed.


Quantitative structure-activity relationships for dicoumarol antivitamins K in the uncoupling of mitochondrial oxidative phosphorylation. The dynamic structure of dicoumarols substituted on the methylene bridge has been studied by nuclear magnetic resonance (nmr) spectroscopy. These molecules may be considered as dimers with restricted rotation around the methylene bridge, held by intramolecular hydrogen bonds; the presence of the substituent R modifies this dynamic process, mainly the facility of exchange of the two hydroxyl protons. These compounds have been compared in respect to their potency in uncoupling oxidative phosphorylation in pig heart mitochondria; the data have been correlated with constants used to characterize the substituent R; to make a potent uncoupler, R should be as small and as hydrophilic as possible. These results are discussed in consonance with the postulated mechanisms of action of the uncouplers, but no simple conclusion can be drawn, especially concerning the role of the dissociable protons.


Heterogeneity of morphological, cytochemical, and cytogenetic features in the blastic phase of chronic granulocytic leukemia. Possible predictive criteria of the refractoriness to therapy of the blastic phase of Ph-1-positive chronic granulocytic leukemia (CGL) have been sought. Eight cases in the blastic phase were studied. The blasts were noted to be of two types: some displayed a high nuclear:cytoplasmic ratio with deep blue cytoplasm, while others had a comparatively low nuclear:cytoplasmic ratio and bluish gray cytoplasm containing a few small granules. Electron microscopic studies showed a variety of features, including defective organelles and giant mitochondria. Cytochemical staining revealed the majority of blast cells to be peroxidase- and Sudan black-negative; granular PAS positivity was the rule. Serial cytogenetic studies demonstrated increasing aneuploidy. Bone marrow biopsy showed myelofibrotic changes in two cases. Two patients entered complete remission with prednisone and vincristine and with Ara-C and thioguanine, respectively. It is concluded that the blastic phase of CGL may manifest heterogeneity.


"Pure" monocytic or histiomonocytic leukemia: a revised concept. In a series of 130 cases of acute leukemia studied by cytochemical staining techniques, 10 cases cytochemically diagnosed as "pure" monocytic leukemia were seen. Cytochemical staining of bone marrow aspirates from these patients revealed all leukemic cells to be Sudan black negative. No positive reactions were observed for peroxidase or naphthol AS-D chloroacetate esterase. All cases demonstrated strong alpha-naphthyl acetate esterase positivity; and fluoride-inhibited naphthol AS-D acetate esterase positivity was observed in 8 of 9 cases tested. The P.A.S. reaction showed diffuse fine to coarse granules. Oil red O stain was positive in 8 of 9 cases, and the beta-glucuronidase activity was strong in 5 of 9 cases. Light microscopy revealed cells with monocytic or histiocytic morphology. Electron microscopic studies in 2 cases demonstrated features consistent with leukemic monocytic or histiocytic morphology; none was suggestive of granulocytic or lymphocytic leukemia. Five of 6 patients treated with drug regimens including prednisone and vincristine entered a complete remission; the other obtained a partial remission. Two patients achieved complete remission after treatment with Adriamycin, 1 following a relapse. Three patients who received cytosine arabinoside as their only therapy died soon after treatment was commenced. It is suggested that the cytochemical similarity but morphological differences in those patients may be objectively used to group them as cases of histiomonocytic leukemia.


Assessment of reactivities of natural antibodies to endogenous RNA tumor virus envelope antigens and virus-induced cell surface antigens. The autogenous humoral immune response of mice to their endogenous leukemia virus (MuLV) has been examined with respect to the reactivities of natural antibodies to MuLV envelope antigens and virus-induced cells surface antigens. The natural reactivity of MuLV envelope antigens was evaluated by means of a radioimmune precipitation assay of intact and disrupted virus, as well as by virus neutralization tests. The specificity of natural antibody for MuLV envelope antigens was determined by immunoelectron microscopy and radioimmune precipitation. Antibody reactivity to virus-induced cell-surface antigens was evaluated by immunoelectron microscopy and a complement-dependent cytotoxicity test. The strains of mice seleced for study were C57BL/6, C3H/Anf, and the C57BL/6 X C3H/Anf F1 hybrid. Although there were quantitative differences in the antibody levels among these various strains, the naturally recognized antigenic determinants of the virus were consistent, i.e., gp68, gp-43, and p15. High levels of neutralizing antibody against the xenotropic BALB:virus-2 were detected in these various normal sera with the focus reduction assay; however, only marginal levels of neutralizing activity against Moloney leukemia virus were detected with the XC virus assay. No anticellular antibody could be detected in these normal sera with the complement-dependent cytotoxicity assay.


Different hematological diseases induced by type C viruses chemically activated from embryo cells of different mouse strains. Type C RNA viruses can be induced by certain chemicals from cells of many mouse strains. Both C58 and BALB/c cells have been shown to contain endogenous viruses that are designated N-tropic because they grow preferentially in cells of NIH Swiss mouse origin. While demonstrating many similar biological and immunological properties, the C58-induced virus is around 10-fold more infectious per physical particle than the N-tropic virus of BALB/c cells. In the present studies, inoculation of these viruses into newborn NIH Swiss mice led to the development of diseases associated with splenomegaly and lymphadenopathy at similar frequency in each group. The disease induced by C58-MuLV was histophathologically diagnosed as lymphoblastic leukemia and was highly malignant following transplantation into newborn mice. The histopathological appearance of spleens from BALB/c virus-affected animals was distinguishable, demonstrating instead myeloid metaplasia or myelogenous leukemia. These findings provide evidence that different endogenous mouse type C viruses can induce distinct diseases in the same mouse strain. Furthermore, they implicate the N-tropic virus endogenous to C58 cells as a major factor in the development of lymphoblastic leukemia that occurs at high frequency in that strain.


Biochemical estimation of the basic dye-binding capacity of RNA from rat hepatoma. Areas of hyperplastic livers that acquire hyperbasophilic properties at advanced stages of carcinogenesis apparently represent the sites of neoplastic trasnformation, and hyperstaining of cytoplasmic RNA with basic dyes also characterizes the cancer cells. Estimations of the RNA content of cell fractions from normal rat liver and solid Novikoff hepatoma provided no evidence that the intense staining of cancer cells could be explained on the basis of an increase in cytoplasmic RNA content. The possibility that cytoplasmic fractions of Novikoff hepatoma show greater affinity for basic dyes than corresponding normal fractions has been examined by means of a test-tube toluidine blue-binding assay. The results revealed that the dye-binding capacity of total cytoplasmic fractions from tumors is 75% higher than normal after Carnoy fixation which retains mostly ribosomal RNA. Assays on fresh ribosomes indicated that tumor ribosomes bind 71% more toluidine blue per mg of RNA than the ribosomal preparation from normal liver. This study thus demonstrates a greater affinity of tumor RNA for basic dyes, and a comparison of biochemical and cytophotometric analyses suggests that an increase in basophilia by a factor OF ABOUT 2 WOULD BE DUE TO A qualitative alteration in robosomal RNA molecules and/or ribosome structure in cnacer cells.


Microfluorometric comparisons of heat-induced nuclear acridine orange metachromasia between normal cells and neoplastic cells from primary tumors of diverse origin. Nuclear fluorescence metachromasia of heated fixed cells subsequently stained with acridine orange was compared in smears and isolated nuclei of various types of primary tumors and normal cells from the tissues that gave rise to the tumors. The ratios of fluorescence emission at 590 and 530 nm reflect the thermal stability of chromatin in situ. The results show that the mean thermal stability of the chromatin in neoplastic cells was lower than the stability of their normal counterparts in all cases. This was found in both spontaneous and chemically induced tumors as divergent in type as a dog vaginal tumor and murine lymphocytic leukemia. These data, together with our previous observations in other neoplastic systems, indicate that reduced chromatin thermal stability may be a general characteristic of cells that have undergone neoplastic transformation and is not confined to rapidly growing tumors. The present investigation identifies the sources of variability encountered in measuring fluorescence metachromasia in slide preparations, and methods of minimizing this variability for potential cytodiagnostic application are discussed.


Ultrastructural study of the effect of bleomycin A2 on the nucleolus and its possibly related cytoplasmic constituents in Novikoff hepatoma cells. Treatment of Novikoff hepatoma ascites cells with bleomycin A2, in vivo as well as in vitro, in varying doses produced marked alterations in nucleolar and cytoplasmic ultrastructural organization. A series of changes occur including formation of fibrillar centers, fragmentation of fibrillar nucleolar elements, appearance of microspherules, and the loss of fibrillar elements from these nucleoli. A bleomycin concentration of 10 mug/ml in vitro produced an increased number of fibrillar centers with well-defined nucleolonemas. At a concentration of 50 mug/ml, there was an increase in number and fragmentation of these fibrillar centers and many microspherules were found throughout the nucleolus. Approximately one-fourth of the cells contained cytoplasmic fibrillar bodies and amorphous fibrous tufts around the nuclear envelope. At a bleomycin concentration of 100 mug/ml, the nucleoli contained more granular elements and numerous microspherules. Almost 90% of the cells contained cytoplasmic fibrillar bodies. The effects of bleomycin in vivo (10 mg/kg) closely resemble those found in vitro with concentrations of 50 and 100 mug/ml.


Effect of glucocorticoids on activation of leukemia virus in AKR mouse embryo cells. The effect of glucocorticoids on activation and replication of leukemia virus in AKR mouse embryo cells was analyzed. The number of cells detected as positive by fluorescent antibody techniques as well as the virus production in cells chronically producing virus was doubled at optimal concentrations of glucocorticoids. The effect of the hormones in activated cells was found to be not on the process of activation per se but rather on synthesis of the viral components after activation has occurred. Intracellular reverse transcriptase levels were not changed by hormone treatment. The stimulation of virus synthesis by glucocorticoids requires binding of the steroid to a cytoplasmic receptor protein.


Modification of seizure activity by electrical stimulation: cortical areas. 1. Three different cortical areas were found with respect to the development, by repeated electrical stimulation, of electrographic and motor seizures in rats. Paleocortical areas responded similarly to subcorticallimbic structures. Initial after-discharges (ADs) were accompanied by little or no behavioral response. With spaced reptition ADs became longer, propagated more strongly, and were eventually accompanied by a convulsive response in which the dominant components were forelimb clonus and rearing. Anterior neocortical placements, on the other hand, yielded convulsive responses from the first AD. These convulsive responses became stronger with repeated stimulation until a mild form of extension was triggered. ADs remained relatively short. Posterior neocortical areas showed electrographic developments very similar to those found in anterior neocortical areas but convulsive responses never developed. 2. All areas tested showed similar reductions in AD thresholds when repeatedly stimulated. Initial thresholds were lower in paleocortical than in neocortical areas. 3. Motor seizures developed more rapidly with stimulation of contralateral homologous sites after seizures had been developed in the primary (initially stimulated) focus. AD thresholds, on the other hand, were not significantly affected in contralateral sites after being lowered in primary sites.


Cortical evoked potentials elicited by real speech words and human sounds. Averaged evoked potentials were recorded from PZ and left and right temporo-parietal electodes to real speech words and human sounds in 8 right-handed subjects. Stimuli were presented in a "no task" condition where the subject was instructed to listen attentively, and a vigilance condition where the subject responded to a particular word or sound during a run of such stimuli. The vigilance condition produced two classes of stimuli:signals and non-signals. Evoked potentials to physically identical words or sounds were examined when they were "no task", non-signal and signal stimuli. P300 amplitude increased significantly as a function of increasing task demands going from "no task" to non-signal to signal. When a strict statistical criterion for multiple comparisons (Bonferroni test) was applied in looking for asymmetries between hemispheres, only 2 isolated left greater than right differences turned out to be significant. Review of the literature concerning evoked potential correlates of differential hemispheric processing pointed up flaws in design, statistical technique, and inconsistencies in reported findings which suggested that while evoked potentials may sometimes reflect differences in hemispheric functioning, this effect is marginal at best.


Comatose state maintained during eight years following a vascular ponto-mesencephalic lesion. A 24-year-old man survived for 8 years after a vascular lesion of the pons and midbrain. During these years a subresponsive comatose state with neurological signs extending from the oculomotor nuclei to the trigeminal and facial nuclei levels was present. A decorticated type of rigidity, with bilateral paralysis of the limbs accompanied by pyramidal signs, was also present. The EEG during the first months showed slow activity which afterwards gradually changed towards fast and alpha activities, maintained in the course of the years. The pathological diagnosis was a fibrous  endarteritis, mainly in the territory of the basilar and vertebral arteries. The pons and midbrain, presenting a large cystic infarct, were serially studied to determine the anatomical extent of the lesion. After reviewing other examples from the literature, the clinical features of our case are discussed. The evolution of the EEG is related to different structures of the ponto-mesencephalic region with different functional activities.


Audiometric comparison of the middle and late components of the adult auditory evoked potentials awake and asleep. The middle and the late components of auditory evoked potentials were alternately recorded in sequential sets from 28 adults, both awake and asleep. Sleep was induced by secobarbital and was monitored for depth. 1000 c/sec tone pips in one laboratory or filtered clicks in the other were delivered at 10, 20 or 30 dB sensation level. Control collections without stimulation were included. For the middle responses a single simple scoring template and one set of voltage criteria could be used for all stages of waking and sleeping. For late responses different templates and voltage criteria were needed. Estimates of the threshold of detection of the evoked potentials were based on the percentage of clearly positive responses was often not attained at 30 dB SL, i.e., the threshold was indeterminate. In light sleep and awake the middle responses of most subjects gave thresholds more sensitive than the late by 10-15 dB and also fewer indeterminate trials. The results in our two laboratories agreed closely in spite of differences in equipment and details of procedure. The relatively low thresholds of the middle responses (median 17.5 dB SL awake and 15 dB SL in light sleep) suggest that the middle responses must be considered seriously for use in clinical electric response audiometry, even though one of the 28 subjects failed to yield any identifiable middle responses.


Human sleep and EEG through a cycle of methadone dependence. The effects of oral methadone on EEG and sleep were studied in 6 male postaddicts. Continuous nocturnal measurement of EEG, EMG and EOG was used to define sleep patterns. Period analysis and power spectral analysis were performed on each 8 min sample of daytime (eyes closed) EEG. Both sleep and EEG were studied during a predrug control period, during the methadone induction phase (45-60 mg/day), stabilization phase (100 mg/day), and then 6; 10, 13, 18 and 22 weeks after withdrawal. One subject did not complete the last two withdrawal sessions. While on methadone, subjects reported that they slept more and also showed an increase in slow wave activity and a decrease in fast wave activity of their EEG during this time. Nocturnal sleep was not markedly altered during the chronic administration of methadone. Subjects reported an increase in dreaming soon after withdrawal, and then 3-5 weeks of nocturnal isnomnia. At the 6th week after withdrawal, slow wave activity in the daytime EEG was decreased, fast wave activity was increased, and mean EEG frequency was increased. REM sleep and delta sleep were increased during withdrawal. These data provide further evidence that chronic administration of narcotic analgesics may induce persistent functional changes in the central nervous system.