Acute renal failure after cis-dichlorodiammineplatinum(II) and gentamicin-cephalothin therapies. Combined gentamicin-cephalothin therapy in four patients after treatment with cis-dichlorodiammineplatinum(II) (CPDD) for advanced solid tumors was complicated by severe acute renal failure. The total dose of gentamicin varied from 240 to 945 mg and that of cephalothin varied from 28 to 48 g. Low-dose (0.5 mg/kg x 8) CPDD was given to one patient, high-dose (3 mg/kg) CPDD was given to two patients, and very high-dose (5 mg/kg) CPDD was given to one patient. The high and very high doses of CPDD were given with concomitant mannitol diuresis. CPDD therapy was complicated by mild transient azotemia in three patients and by severe acute renal failure in one. In the latter, the azotemia began to improve on Day 7 after CPDD treatment. Following gentamicin-cephalothin therapy, all patients developed severe acute renal failure which persisted until death. At autopsy, all patients had extensive renal tubular necrosis at various stages. These findings indicate that gentamicin-cephalothin therapy after treatment with CPDD can be severely nephrotoxic, and that this antibiotic combination should be given with great caution, if at all, to patients receiving CPDD treatment for malignancy.


Intra-arterial radionuclide infusion: a new technique to assess chemotherapy perfusion patterns. A mechanical infusion pump was used to introduce 99mTc-sulfur colloid at a rate of 1 ml/minute into the hepatic arteries of six patients. The radiotracer distribution patterns were compared to those obtained with "bolus" radionuclide angiography and those seen with standard contrast angiography. In all cases, the slow flow rate radionuclide studies evidenced dramatic differences from the patterns obtained with rapid flow rate injections. The results indicate that standard, rapid-injection contrast techniques may not produce the most reliable information regarding potential flow distribution. It is concluded that radiotracers introduced at flow rates approximating those attained with infusion pumps will offer the best estimates of both initial catheter placement and subsequent patterns of hepatic distribution of chemotherapeutic agents.


Antimetastatic action of some triazene derivatives against the Lewis lung carcinoma in mice. Two dimethyltriazenoimidazoles, DTIC and BRL 51308, and a benzenoid dimethyltriazene, CB 10286, have been examined for their effects in mice bearing Lewis lung carcinoma. A slight reduction of primary tumor growth was found after treatment with DTIC and BRL 51308, whereas CB 10286 caused no significant effect. On the contrary, all the tested compounds sharply reduced the number of lung metastases and also resulted in a high proportion of animals free of metastases at death. No significant cytotoxic effect of the triazenes was observed in small established pulmonary tumors, as determined by evaluating the effects of treatment on the fractional incorporation of 3H-TdR into DNA of the lung colonies. These results are in contrast to those obtained with a purely cytotoxic agent, cyclophosphamide, and indicate that all three triazene derivatives tested have selective antimetastatic properties.


Therapeutic implications of cell kinetic changes after cyclophosphamide treatment in "spontaneous" and "transplantable" mammary tumors. The present studies were initiated to investigate the cell kinetics in spontaneous and transplantable mammary tumors after single doses of cyclophosphamide (Cp). The 3H-thymidine (3H-TdR) labeling index (LI), the DNA synthesis times, and the primer-dependent DNA polymerase labeling index (PDPI) were determined by in vitro methods for spontaneous mammary tumors (SMT) in C3H/HeJ retired breeders and 13762 transplantable rat mammary tumors (RMT) in Fischer 344 rats from 6 hours to 14 days after Cp treatment. The perturbations in the cell kinetics, although quantitatively different in the two tumor models, were qualitatively similar in that transient changes in both PDPI and 3H-TdR LI were observed within the first 24 hours after treatment. These changes were followed by a variable period during which cell proliferation was suppressed. Increases in the PDPI and 3H-TdR LI, presumably reflecting proliferative recovery, were observed prior to the initiation of tumor regrowth. Increases in the 3H-TdR LI and PDPI within the first 24 hours after Cp in C3H/HeJ SMT indicated an interval of increased sensitivity to adriamycin. In the 13762 RMT, combination chemotherapy protocols designed to exploit changes in cell kinetics at early times (24 hours) and during the recovery phase of the response resulted in greater long-term tumor-free survival.


Effect of a transplantable leukemia on hematopoietic stem cells in AKR mice. The number of colony-forming units-spleen (CFU-S) and colony-forming units-culture in the femoral marrow of mice bearing an AKR transplantable leukemia remained relatively unchanged up to the death of the host despite the almost complete displacement of the normal differentiated hematopoietic cells by infiltrating leukemia cells. The proliferative state of the CFU-S was not changed significantly by the presence of the leukemia cells as determined by using 5-fluorouracil as a proliferative probe. The implications of these findings are discussed with regard to possible control mechanisms governing the interaction of leukemia cells with normal marrow cells and their potential importance for cancer chemotherapy.


Therapeutic value of mepitiostane in the treatment of advanced breast cancer. A new, orally active antiestrogenic steroid, mepitiostane (20 mg/day), was given to 45 patients with advanced breast cancer. The regression rate was 31.1%, or 14 of 45 patients, and a duration of regression of greater than 6 months was obtained in seven patients. Virilizing effects such as hoarseness, hirsutism, and acne were observed relatively often, but there was no evidence of abnormality in the liver function tests or in the serum calcium level.


Rate of DNA synthesis as an indication of drug toxicity and as a guide for scheduling cancer therapy. The rates of incorporation of [C3H3]thymidine into the DNA of tissues of normal mice at various times after single doses of several drugs (adriamycin, vincristine, methotrexate, 5-fluorouracil [5-FU], melphalan, CCNU, and chlorozotocin) that have some clinical activity against cancer have been determined and are considered to be indicative of the "degree of normalcy" of the respective tissues at those times. These experiments show that there are some differences and some similarities in the effects of the agents upon the various tissues of the mouse. Each of the agents caused early inhibition of incorporation of [C3H3]thymidine followed by stimulation of such incorporation by intestinal mucosa and marrow, but the timing and the extents of inhibition and stimulation differed for the various agents. Except for CCNU, the preceding description would also apply to the effects upon liver and lungs, but the stimulation was delayed in comparison to that of intestinal mucosa and marrow. Early inhibition with little subsequent stimulation occurred in the spleen, kidneys, and heart. For all tissues, recovery was slower and less extensive after CCNU than after the other agents. The data obtained with the single agents were used for setting up experiments in which a single dose of one agent was followed after selected intervals of time with a single dose of a second agent. The combination of adriamycin plus 5-FU was less toxic when the two agents were given simultaneously than when the 5-FU was given 24-96 hours after the adriamycin, while for the combination of adriamycin plus CCNU there was less toxicity if the CCNU was administered 24 hours after the adriamycin rather than simultaneously with it. When a combination of a single dose of methotrexate and a single dose of 5-FU was administered to mice, the lethality was greater when the two agents were administered 120 hours apart than when the two agents were given simultaneously or were separated by 24 or 48 hours; this was true regardless of which agent was administered first. These results are consistent with what might be predicted upon the basis of the data obtained for the single agents in this study and of what is known about the mechanisms of action of these agents and of the effects of the agents in relation to cell and tissue kinetics. These results indicate that patterns of thymidine fixation by tissues following single doses of agents might be useful in scheduling multiple-drug therapy to minimize host toxicity.


Comparison of adriamycin-induced nucleolar segregation in skeletal muscle, cardiac muscle, and liver cells. Male Sprague-Dawley rats were treated with either 2.5, 3.5, or 5.0 mg/kg of adriamycin by iv injection. After 1 or 3 hours of treatment, samples of liver, cardiac muscle, and skeletal muscle cells were examined by electron microscopy. The changes in ultrastructure observed in these tissues after the first hour included nucleolar segregation and altered distribution of the perinucleolar chromatin. However, no alterations in the ultrastructure of either the nucleus or cytoplasm were observed in tissues examined 3 hours after a 5-mg/kg dose of adriamycin. The doses at which nucleolar alterations occurred varied between tissues. In skeletal and cardiac muscle cells, marked alterations in nucleolar ultrastructure were observed at doses of 3.5 and 5.0 mg/kg of adriamycin. Liver cell nucleoli, however, exhibited few structural aberrations at these doses. The similarities in response of skeletal and cardiac muscle suggest that ultrastructural analysis of skeletal muscle biopsies may be useful in evaluating adriamycin cardiotoxicity.


Distribution and antitumor activity of adriamycin given in a high-dose and a repeated low-dose schedule to mice. Experimental studies on the distribution of adriamycin (AM) under different treatment conditions and possible correlations between tissue and plasma levels and chemotherapeutic activity are discussed. C57BL/6J mice bearing im Lewis lung carcinoma and (C3H x O2O)F1 mice bearing mammary carcinoma were injected iv with AM at a single dose of 15 mg/kg or with the same total amount of drug administered in spaced doses of 3.75 mg/kg for 4 consecutive days. In the two experimental systems studied, the drug reached approximately the same value in the tumor and spleen with both types of treatment, but with the 3.75-mg/kg x 4 schedule much lower AM concentrations were observed in the heart than with the single high-dose treatment. The therapeutic activity of the two treatments also differed: the antitumor and antimetastatic effect was the same in the two tumor systems, but with the 3.75-mg/kg x 4 schedule, increased survival and somewhat lower toxicity were observed. Daunorubicin, tested in the mammary carcinoma system with the two schedules of treatment, behaves very similarly to AM in terms of both distribution and chemotherapeutic effect.


Evaluation of VP-16-213 in malignant lymphoma and melanoma. Phase II studies utilizing VP-16-213 in the treatment of 56 patients with malignant lymphoma and 29 patients with malignant melanoma were carried out by the Southwest Oncology Group. All patients had received extensive prior therapy. The initial dose of VP-16-213 administered was 45 mg/m2 by iv infusion over 30-60 minutes on Days 1-5 every 3 weeks but, because, of severe myelosuppression in the lymphoma group, the dose was subsequently reduced to 35 mg/m2. Only three partial regressions lasting 6, 2, and 1 months were noted in 17 patients with Hodgkin's disease. No favorable responses were noted in 35 patients with non-Hodgkin's lymphoma including 16 with the diffuse histiocytic type. No responses were noted in patients with melanoma. The major toxic effect was myelosuppression. VP-16-213 appears to lack significant effectiveness in these previously treated disease.


Peptichemio in advanced neuroblastoma.

Peptichemio in advanced neuroblastoma. Peptichemio (PTC) is a mixture of six synthetic peptides of m-L-phenylalanine mustard. It acts with both alkylating and antimetabolic effects, interfering with the synthesis of DNA, RNA, and proteins. PTC was administered iv to 18 previously untreated children with advanced neuroblastoma at a dose of 1-1.5 mg/kg/day for one to three cycles of 5-6 consecutive days each. Eleven of 12 patients (92%) experienced both objective and subjective improvement; complete remission was achieved in two of them. In spite of the high remission rate, the median duration of remission has been short (4 months) and the overall survival (median, 6 months) did not seem to be influenced by the use of PTC. The primary toxic effects were, in order of importance, bone marrow depression, phlebosclerosis, nausea and vomiting, and alopecia. Chronic use of PTC seems limited by two major factors: profound long-lasting thrombocytopenia and severe phlebosclerosis.


Phase II study of triazinate and pyrazofurin in patients with advanced breast cancer previously exposed to cytotoxic chemotherapy. The purpose of this phase II clinical trial was to evaluate the effectiveness of triazinate and pyrazofurin in the treatment of advanced breast cancer. Although an occasional patient experienced stability of disease, neither agent, as employed in this study, was able to produce an objective antitumor effect in this group of patients with previous chemotherapeutic exposure. At the dosages used in this study, neither agent can be recommended for further treatment of advanced breast cancer.


Junctional escape rhythm in the sick sinus syndrome. In 21 patients with and 31 without junctional escape beats, a comparison of the symptoms, revealed that there were no significant differences (p greater than 0.05) in the symptoms between these two groups manifesting a sick sinus syndrome. Thus, the occurrence of junctional escape beats did not provide the anticipated shortening of the asystolic pauses to prevent the occurrence of symptoms related to the period of electrical silence. In 7 patients with an escape junctional rhythm, the long pause was interrupted by the occurrence of a regular junctional rhythm. The pause was not preceded by a supraventricular tachycardia, ruling out physiologic overdrive suppression of the junctional pacemaker. The period of asystole preceding the junctional rhythm was not multiple of the R-R interval of the junctional rhythm. 4 additional patients demonstrated long periods of asystole, uninterrupted by junctional escape beats and at other times exhibited shorter pauses which were terminated by junctional escape beats. These findings can be explained by the presence of a 'junctional arrest' which is analogous to sinoatrial arrest. The phenomenon of 'junctional arrest' may be one tenable explanation to account for the lack of protection by junctional escape activity against the symptoms associated with the sick sinus syndrome.


Normalization of the QRS complex by concealed conduction in a patient with Wolff-Parkinson-White syndrome. In a patient with Wolff-Parkinson-White syndrome, transient normalization of the QRS complex occurred as a result of retrograde concealed conduction into the accessory pathway from an A-V junctional escape beat. This patient also had a bradycardia-tachycardia syndrome.


Sick sinus syndrome due to cardiac amyloidosis. In a patient suffering from cardiac amyloidosis a case of sick sinus syndrome, manifested by markedly prolonged recovery time of the sinus node, was documented by an atrial pacing study. The first A-V junctional escape interval was markedly prolonged following the termination of the atrial pacing, pointing to a coexisting A-V nodal dysfunction. The patient required a permanent artificial pacemaker implantation.


A comparison of hemodynamic and angiographic indices of left ventricular performance in patients with coronary artery disease. We compared ejection fraction, left ventricular end-diastolic pressure, cardiac index and the relation of left ventricular stroke work index to left ventricular end-diastolic pressure during rest and exercise in 60 patients with coronary artery disease. Left ventricular end-diastolic pressure was usually normal at rest (48/60) and abnormal during exercise (46/60) and did not correlate with ejection fraction. Cardiac index was insensitive, usually remaining normal until ejection fraction was less than 0.40. Patients with a normal left ventricular stroke work index response to exercise had higher ejection fractions than those with an abnormal response (p is less than 0.05). However, 9 patients with normal ejection fractions had an abnormal exercise response. This may reflect loss of left ventricular reserve, abnormal compliance or clinically silent ischemia during exercise. Different indices of left ventricular performance may be widely disparate in coronary artery disease, and abnormalities are frequently apparent only during exercise.


Transtelephone adjustment of antiarrhythmic therapy in ambulatory patients. The electrocardiograms of 80 ambulatory patients receiving antiarrhythmic therapy were supervised with the help of a transtelephone monitoring system. The patients used a pocket-size modulator and reported several times daily to the receiving center integrated into the intensive cardiac care unit. The surveillance lasted for 5-28 days during which various drugs in varying dosages were administered to suppress or prevent dysrhythmias. In 94% of the patients, a satisfactory therapeutic achievement was obtained. The transtelephone system provides easy diagnosis, immediate pattern recognition, direct and frequent contact with the ambulatory patient and long periods of follow-up. During this time, the proper antiarrhythmic agent can successfully be defined, its effective dose can safely be determined and unnecessary hospitalization can thus be prevented.


Scanning electron microscopic studies on cultured heart cells after the induction of arrhythmias and fibrillation. Scanning electron microscopic observations were made of mouse embryonic heart cells in culture to clarify the interrelationship between morphological damage and cardiac arrhythmias and fibrillation induced by changes of ion concentrations. They were studied with particular reference to the abnormalities in the heart cell and the intercellular connections. The cells exhibited arrhythmias with any alteration in ion concentration (MgSO4, CaCl2, KCl, NaCl) in the culture medium, and they developed fibrillation-like beating under certain conditions of lowered potassium, or sodium, and increased sodium with a reduction in calcium or potassium. The heart cells showing arrhythmias were revealed by scanning electron microscopy to have more or less fine morphological changes in their slender processes, and detached or raised cell edges. Fold-like undulations in the cell body and cell ledge were also numerous in cells exhibiting fibrillation-like beating. These cell body changes may, however, be characteristic of cells maintained in cultures with lowered potassium content, since only these cultures showed such changes. Changes of cell ledge were also demonstrable in cultures with altered MgSO4, CaCl2 content or increased KCl or NaCl content although the cells in these cultures did not develop fibrillation. It was noted that there was a tendency for the changes of cell body and cell edge to be conspicuous in cells exhibiting fibrillation. The results obtained indicated that fibrillation developed only in the presence of altered concentrations of certain ions, in association with subtle morphological changes in the cells and intercellular connections.


Ventricular arrhythmias associated with coronary artery occlusion and reperfusion in the isolated perfused rat heart: a model for assessment of antifibrillatory action of antiarrhythmic agents. The isolated Langendorff-perfused rat heart, subjected to coronary artery occlusion and reperfusion consistently developed ventricular arrhythmias. The incidence of hearts with ventricular premature extrasystoles, ventricular tachycardia, and fibrillatioon ventricular fibrillation could be abolished in the presence of an extracellular potassium concentration of 12 mmol-litre--1, whereas with 4.5 mmol-litre--1 all hearts developed all the ventricular arrhythmias including fibrillation. This predictable occurrence of ventricular arrhythmias was used to assess the antiarrhythmic effect of lignocaine with particular reference to its effect on the spontaneously occurring reperfusion ventricular fibrillation. Lignocaine decreased the incidence of all ventricular arrhythmias both during ischaemia and on reperfusion in a concentration-related fashion.


Ventricular refractoriness during acute myocardial ischaemia and its relationship to ventricular fibrillation. Studies were made in anaesthetised dog of the effects of repeated acute occlusions of a branch of the anterior descending coronary artery on ventricular refractory periods in adjacent ischaemic and non-ischaemic myocardium. Differences occurred in refractoriness between normal and ischaemic areas in the ventricle. This was greatest 2.5 min after occlusion, and on release of occlusion, ventricular refractory periods reverted to normal within 5 min. Spontaneous ventricular fibrillation was directly and significantly related to the degree of dispersion of refractoriness in a given dog immediately preceding release and following release of occlusion. Infusion of isoprenaline caused significant shortening of refactory period and increased dispersion of refractoriness during ischaemia. Studies of dispersion of refractoriness should prove valuable in assessing the efficiency of metabolic or antiarrhythmic protection against ventricular fibrillation.


Maturational changes in left and right ventricular electromechanical intervals in the newborn lamb. Electromechanical intervals of right and left ventricles were examined in newborn lambs (9 hours to 42 days). The electromechanical interval of the left ventricle did not change with maturation, while that of the right ventricle became progressively longer, resulting in an increasing ventricular mechanical asynchrony. We postulate that the pattern of developing mechanical asynchrony described may occur secondary to postnatal changes in ventricular morphology.


The magnitude of the bradycardia induced by face immersion in patients convalescing from myocardial infarction. Twenty patients who had suffered myocardial infarction within the previous 3 to 13 days performed face immersion with breath-holding. The maximum reduction in heart rate was 17 +/- 9 beats-min--1 (mean +/- SD). When compared with 24 patients without evidence ischaemic heart disease, they behaved more like younger patients (22 +/- 6) than patients in the same age range (7 +/- 6). A greater vagal response for age after infarction may reflect persistent hypersensitivity of a reflex designed to protect the myocardium during cardiogenic shock.


Relation of platelets to catecholamine induced myocardial injury. To test the thesis that myocardial injury, induced by catecholamines, is ischaemic in origin due to platelet accumulation in the coronary microvasculature, sustained left intracoronary and systemic infusion of catecholamines in toxic dosage was given to dogs previously infused with autologous 51Cr-labelled platelets. Subsequent determination of tissue radioactivity and electrolytes, as well as electronmicrography, indicated that the induced myocardial injury was not related to microvascular occlusion by platelets.


Transmural distribution of intrinsic and transmitted left ventricular diastolic intramyocardial pressure in dogs. The left ventricular transmural distribution of steady diastolic blood flow was measured with microspheres at three constant coronary perfusion pressures and two constant diastolic transmural pressures. The pressure-flow ratio in each layer was near linear allowing calculation of an apparent downstream pressure at zero diastolic transmural pressure (or intrinsic myocardial pressure) of 2.0 to 2.4 kPa (15 to 18 mmHg). When the transmural pressure was 2.67 kPa (20 mmHg), the increment of pressure in the outer 4/5 of the ventricular wall was 0.80 kPa (6 mmHg) (interpreted as transmitted intramyocardial pressure). Thus, intrinsic intramyocardial pressure is large and is a major determinant of diastolic coronary blood flow, and transmitted intramyocardial pressure has its major effect in the subendocardium.


Failure to find a humoral factor which influences the compensatory response after resection of the rat small bowel. Small bowel resection was performed in one parabiont of a parabiotic rat pair to determine whether there was a "humoral" or other circulating factor present which could influence the compensatory response seen in the remainder of the small bowel. Following intestinal resection in one parabiont, there was an increase in DNA synthesis in the intestine of that member provided at least 40% of the small intestine was removed. The compensatory response seen in a resected animal was not observed in its parabiotic partner, even though an attempt was made to make the potentially responding animal more sensitive to the influence of any circulating "factors" by resecting its intestine as well. The results obtained are in disagreement with the one closely related report in the literature; it is suggested that there is no good evidence for the existence of a resection-induced circulating factor with the capability of crossing a vascular anastomosis and controlling proliferation of small intestinal epithelium.


G2 sub-population in mouse liver induced into mitosis by lead acetate. A single intracardiac dose of lead acetate (40 microgram lead/g body weight) induced a 25-fold increase in mitosis of mouse hepatocytes 5 hr after injection, as determined by autoradiography. The prompt appearance of a mitotic wave and the relatively large number of mitoses suggest that the mitotic cells were derived from a hepatocyte sub-population arrested in the G2 phase. The injection of lead also stimulated a small increase in labeled hepatocytes within 6 hr. Analysis of grain counts gave no evidence for unscheduled DNA synthesis. The incremental labeled cells may have originated from a small fraction of the G1 population that was ready to enter the S phase without the usual pre-synthetic delay.


Growth fraction and cycle duration of hepatocytes in the three-week-old rat. The proliferation of hepatocytes in the liver of 3-week-old rats has been investigated by autoradiographic methods. This investigation is a continuation of earlier work on the same topic (Schultze & Maurer, 1972; 1973). 21 days after birth, 102 rats received a single injection of 3H-TdR. The percentage of labelled mitoses was then determined 1 hr later and at various times throughout the interval up to 12 days after application of 3H-TdR. In agreement with earlier work, a first peak of labelled mitoses was found 7 hr after 3H-TdR injection. The area under the peak indicates an S phase duration of 8 hr. In addition a second very broad peak of labelled mitoses was found between 2 and 12 days after pulse labelling. The analysis of the results leads to the conclusion that the hepatocytes of the 3-week-old rat have a growth fraction close to 1 and a doubling time of 6-7 days. This is at variance with earlier results of Post, Huang & Hoffman (1963) and Grisham (1969) who had derived a value of 21.5 hr for the duration of the cell cycle and a value of only 0.1-0.2 for the growth fraction of the hepatocytes.


Defect of erythropoiesis in non-leukaemic AKR mice. The kinetics of the CFU population and of erythropoiesis were investigated in the AKR strain mouse prior to the onset of thymic leukaemias: haemopoiesis was compared in syngeneic AKR, semi-allogenic C3H and (C3H X AKR) F1 mice injected with AKR stem cells. These experiments demonstrate that the reduction in The number of spleen colonies previously described by Perkins et al. (1971) in syngeneic hosts, as compared to semi-allogenic C3H hosts, is actually related to defective erythropoiesis resulting from a dysfunction of the AKR haemopoietic inductive microenvironment (HIM). Erythropoietin secretion is normal in AKR mice. The early haemopoietic events related to the stem cell: lodgement of the CFU ('f' factor) and doubling time, are not disturbed, but the onset of CFU proliferation is markedly delayed in the AKR strain. The main expression of the AKR HIM dysfunction is a significant reduction in the number of erythroid (E) colonies and an impaired output of red blood cells per E-colony in the syngeneic host as compared to the allogenic one. In addition, data indicate that a weakly histo-incompatible system, such as that in C3H and hybrid hosts, does not interfere with the stages of haemopoiesis except by lengthening the doubling time of the CFU. The results, on the whole, emphasize the prevalent influence of HIM.


Cell proliferation in EMT6 tumors treated with single doses of x-rays or hydroxyurea. I. Experimental results. EMT6 mouse mammary tumors were treated in vivo with 5 mg/mouse of hydroxyurea (HU) or 300 rads of X-rays. The proliferation of the tumor cells was followed for 28 hr after treatment. Changes in the 3H-TdR labeling index, the mitotic index, the specific activity of the 3H-TdR-labeled DNA, and the proportion of suspended, clonogenic cells in the S phase of the cell cycle were examined and compared. Evidence was found for reassortment of the surviving cells in treated tumors into partially synchronous cohorts. The partial synchrony in the proliferation of the surviving cells was not accurately predicted by the changes in the labeling index and the mitotic index. The changes in DNA specific activity proved unacceptable as an indicator of cell proliferation in solid EMT6 tumors treated with low doses of radiation or HU.


On the existence of 'arrested G2 cells' in mouse epidermis. It has been postulated that mouse epidermis contains two populations of resting cells, one of which is blocked at the G1-S boundary and the other between G2 and mitosis. The 'arrested G2 cells' were estimated, by the labelled mitosis method, to comprise 5-10% of the epidermal population and presumed to function as a 'reserve pool' which could be activated by wounding. A comprehensive search has now been carried out for arrested G2 cells in mouse epidermis using the direct methods of single cell and flow through cytophotometry. No evidence was obtained which supports the existence of such a cell compartment. Suitable control experiments were carried out to ensure that G2 cells were not lost during the isolation of epidermal nuclei.


Cell kinetics in the erythroid compartment of guinea pig bone marrow: a model based on 3H-TdR studies. A model of steady-state erythropoiesis in the guinea pig is described. The model incorporates an unidentified progenitor compartment, as well as compartments representing proerythroblasts, basophilic, polychromatic and orthochromatic cells. A computer representation of the model permits a simulation of the labeling curves obtained in pulse and intermittent labeling regimes. It was found that a reasonable fit to the data can be achieved when the parameters for the various compartments are essentially identical. The results of a preliminary sensitivity analysis, carried out by perturbing the duration of S phase from the best fit value, are reported. The fit achieved to the data supports the hypothesis underlying the model that each compartment corresponds to one generation and that the flux within and between compartments is sequential.


The development of a zonula occludens in peripheral myelin of the chick embryo. A freeze-fracture study. Sciatic nerves of chick embryos, 12 to 18 days incubation, were examined in freeze-fracture replicas with special emphasis placed on the development of tight junctional contacts in the myelin sheaths. In stages of beginning myelination short isolated particulate chains (focal tight junctions) appear in fracture faces of the adjacent membranes in the outer myelin lamellae, i.e., the outer mesaxon. In stages of progressing myelination these tight junctional elements elongate and become more numerous. They can also be found in the membranes of the inner mesaxons, the paranodal loops and the intramyelinic cytoplasmic inclusions. In fibers of advanced myelinogenesis a fusion of these isolated tight junctions--either end-to-end or at an angle--gives rise to continuous zonulae occludentes. This contact zone extends in the mesaxonal membranes along the direction of the fiber, whereas in paranodal myelin it acquires a helical course joining the membranes of the paranodal loops. It is proposed that this zonula occludens, which seals the cytoplasmic border of the Schwann cell, separates an intramyelinic from an extramyelinic, extracellular space already during the developmental stages of myelinogenesis.


Origin of the glial processes responsible for the spontaneous postnatal phagocytosis of boutons on cat spinal motoneurons. Previous studies have demonstrated that astrocyte processes are responsible for a spontaneously occurring phagocytosis of boutons on cat spinal motoneurons during the second postnatal week. In the present investigation, the astrocytes and the astrocyte processes in contact with the motoneurons were studied qualitatively and quantitatively during the early postnatal period. It could be concluded that the cells responsible for the phagocytosis of boutons are immature astrocytes. These cells were present not only during the period of phagocytosis but also prior to this period. The type of process responsible for the phagocytosis was present not only during the period of phagocytosis but also prior to and after that period although the relative contribution of such processes to the glia-covered membrane area of the motoneurons was reduced in the older animals. On the basis of these results, the possible specificity of the immature astrocyte as the element responsible for the phagocytosis of boutons during normal development is discussed.


Effect of pimozide on the cytology of the eel pituitary. I. Prolactin-secreting cells. Pimozide, a specific blocker of dopaminergic receptors, was injected for 4 to 9 days in freshwater (FW) eels or eels acclimated to sea water (SW), for 10 to 30 days. The daily dose was 100 or 200 microgram/100 g. In FW, pimozide induces a nuclear hypertrophy in the prolactin (PRL) cells of eels; these elongated cells increase in height. The amount of erythrosinophilic granules in the cytoplasm, initially reduced, increases. Plasma electrolyte values are not modified: only the plasma sodium level slightly rises with the higher dose. In SW, PRL cells appear less active. After 10 days, this hypoactivity is not yet fully evident; pimozide stimulates PRL cells without affecting electrolyte values. After 1 month in SW, PRL cells are stimulated with pimozide and a slight regranulation may occasionally occur. The response in SW is never as marked as it is in FW; a high dose is not more effective than a low one. The higher dose significantly raises Na+, Ca2+ and Cl- plasma levels. These data suggest that prolactin synthesis and release increase with pimozide. They corroborate the hypothesis of a hypothalamic inhibitory control on PRL secretion mediated through dopaminergic fibers in the eel, but other factors may also be involved in this regulation in addition to the effect of salinity.


Effect of pimozide on the cytology of the eel pituitary. II. MSH-secreting cells. Pimozide, a specific blocker of dopaminergic receptors, was injected for 4 or 9 days in freshwater (FW) eels or eels acclimated to sea water (SW) for 10 or 30 days. The daily dose was 100 or 200 microgram/100 g. Melanophore index values increase in FW and in 1 month-SW injected eels. All the treated fish react by a total or subtotal degranulation of the lead-hematoxylin positive cells in the pars intermedia. These cells were previously identified as alpha-MSH-secreting cells. The MSH cell nuclear area is significantly increased, nucleoli are larger and the endoplasmic reticulum more developed. The intensity of the response is similar in FW and SW eels, but it does not increase with the higher dose. The rapid release of pituitary alpha-MSH is also visualized by immunofluorescence and immunoenzymologic techniques. No effect on the second cell type of the pars intermedia (PAS-positive cell) is detected. The amount of neurosecretory material is often reduced in the neurohypophysis. These results suggest that the hypothalamic inhibitory control of MSH release and synthesis is mediated through dopaminergic fibers in the eel, but other factors cannot be ignored in this regulation.


Distribution of fibres reacting with an alpha-endorphin antiserum in the neurohypophysis of Carassius auratus and Cyprinus carpio. A strong positive immunoreaction with an alpha-endorphin antiserum occurs in two distinct sites of the goldfish and carp neurohypophysis. Fluorescent nerve terminals are found in the laminar nerve processes located in the rostral pars distalis, but the immunocytological reaction is mainly localised on the nerve processes of the posterior neurohypophysis lying between the intermediate lobe cells. Almost all the digitations of the neurohypophysis are strongly fluorescent. The immunoreactive fibres probably originate from the hypothalamus, where perikarya displaying the same immunoreaction have been found in the pars lateralis of the nucleus lateralis tuberis and in some minor centres. The possibility that the immunoreactive substances revealed on the neurohypophyseal processes may originate in the intermediate lobe cells is also discussed. It has now to be established if this hypothalamo-hypophyseal system contains a substance with endorphic properties or only some immunologically related substance devoid of the corresponding physiological activities.


Catecholaminergic innervation of muscles in the hindgut of crustaceans. An ultrastructural study. The crustacean species Pacifastacus leniusculus and Gammarus pulex were investigated by electron microscopy in a search for possible neuromuscular junctions in the hindgut, which has a rich supply of catecholaminergic fibres. True neuromuscular synapses were found in both species between nerve terminals containing dense-core vesicles (80-110 nm in diam.) and muscle fibres. We suggest that the dense-core vesicle terminals contain a catecholamine, and this is supported by ultrahistochemical tests for monoamines. Two types of junctions are found: one in which the nerve terminal is embedded in the muscle cell (both species) and one in which protrusions from the muscle cell meet nerve terminals (Pacifastacus). Gammarus pulex, which has only circular muscles in the hindgut, has only catecholaminergic innervation, whereas Pacifastacus leniusculus has circular and longitudinal muscles both with at least two types of innervation.


Influence of maternal adrenalectomy on the ultrastructure of the adrenal gland in neonatal rats. Maternal adrenalectomy at 7 or 14 days of gestation produced increased cell necrosis within zona reticularis cells on the day of birth and at 24 or 48 h after birth. Small remnants or large portions of adrenocortical cells were present within macrophages. In otherwise normal adrenocortical cells, lipid droplets were incorporated within some mitochondria. Autophagocytosis of single mitochondria was observed within adrenocortical cells. Undoubtedly ultrastructural changes represent stimulation of adrenocortical cells in neonatal rats in response to maternal adrenalectomy.


Quantitative ultrastructural studies on rat gastric zymogen cells under different physiological and experimental conditions. Quantitative electron microscopical measurements performed on gastric zymogen cells were aimed at determinations of size and volume density of the zymogen granules, and size of cell and nuclear profiles. 17 groups of rats each comprising 6-8 animals were investigated: five of these groups were used to study the influence of fasting and feeding, three groups were killed at different intervals after a pilocarpine injection, and four other groups were investigated after an atropine injection. The remaining five groups of rats were operated on: vagotomy was performed on one group, vagotomy + pyloroplasty on another, pyloroplasty on the third group, and antrectomy on the last two groups. The operated rats were sacrificed 4 or 10 weeks after the operations. Pilocarpine was more effective than feeding in reducing the size and the amount of zymogen granules. After atropine the size and amount of zymogen granules tended to increase. Ten weeks after pyloroplasty, vagotomy + pyloroplasty, or antrectomy the mean size of the zymogen cells was reduced. Loss of trophic vagal impulses, duodenal regurgitation, and abnormal serum gastrin levels are factors which might be responsible for the zymogen cell hypotrophy in operated rats.


The proliferation of adrenal medullary cells in newborn and adult mice. A light and electron microscopic autoradiographic study. The proliferative activity of newborn and adult mouse adrenal medullary cells was determined with light and electron microscopic autoradiography. The H3 thymidine labelling index of 2 weeks old mice adrenal medullary cells was about 9.4% and declined to less than 1% in adult mice. In electron microscopic autoradiography labelled norepinephrine as well as epinephrine cells could be seen. Only in 1 and 2 weeks old mice some morphologically undifferentiated cells were visible. In formaldehyde induced fluorescence combined with light microscopic autoradiography the fluorescence intensities of labelled and unlabelled medullary cells were measured. On average the fluorescence intensity of labelled cells was lower than that of unlabelled cells. The differences could be explained by a higher number of autoradiographic silver grains laying on the cytoplasm of labelled cells. These results give evidence that fully differentiated adrenal medullary cells are capable of division.


The reticular formation of lampreys (Petromyzonidae)--a target area for exohypothalamic vasotocinergic fibres. The octapeptide vasotocin, which is formed in the classical neurosecretory nuclei of lampreys (Petromyzonidae), is transported, bound to the carrier protein neurophysin, not only to the neurohypophysis but also to various other regions of the brain via exohypothalamic fibres. A target area of this exohypothalamic vasotocinergic system is, in the brook lamprey (Lampetra planeri Bloch), a relatively well circumscribed area in the isthmus region of the rhombencephalic tegmentum motoricum, which is called area lateralis tegmenti. In this area, which belongs to the reticular formation, the vasotocinergic fibres form synaptic contacts with nerve cell perikarya and processes. The vesicles contained in the fibres were identified, ultrahistochemically, as neurophysin vesicles. They correspond to the neurophysin vesicles observed in the neurohypophysis of the same species. The functional significance of the vasotocinergic supply to portions of the reticular formation in lampreys is open to discussion.


The ultrastructure and function of follicle cells in Foucartia squamulata (Herbst) (Curculionidae). The follicle cells of Foucartia squamulata are involved in the formation of both vitelline membrane and chorion. Precursors for these egg coverings are synthesized by the rough endoplasmic reticulum and condensed within dictyosomes. The vitelline membrane and the chorion appear on the oocyte surface simultaneously, which is an unusual phenomenon for insects. The follicular epithelium has not been found to contribute to vitellogenesis in the species under study.


Histogenesis of the pituitary in rainbow trout (Salmo gairdneri) in different ambient salinities with particular reference to the rostral pars distalis. The pituitary gland is first evident in rainbow trout two weeks before hatching. Differentiation of prolactin and ACTH cells is not marked until 3-4 days after hatching when the follicular arrangement of the prolactin cells become apparent. There is no difference in the time of development of either prolactin or ACTH cells in larvae reared in different ambient salinities despite marked changes in tissue ion and water content. This suggests that prolactin and ACTH do not play the osmo- and iono-regulatory roles in larval rainbow trout that they are considered to play in adult salmonids.


A study of the fine structure of the pit organ of the common Japanese sea eel Conger myriaster. The fine structure of the pit organ of the sea eel has been studied by means of electron microscopy. The sensory epithelium of the pit organ consists of sensory cells and supporting cells. The apical surface of the sensory cell is studded with sensory hairs consisting of a kinocilium and stereocilia. The sensory cells are divided into two groups. In one, the kinocilium points dorsally and in the other the kinocilium points ventrally. The total number of sensory cells in one pit organ is about 100, and the ratio of cells with opposite polarity is about 1:1. On the basis of these structural features, the pit organ is considered to be a mechano-receptor sensitive to the movement of liquid in a dorso-ventral direction. It may also serve as an ion receptor, sensitive to environmental ion concentration. Efferent nerve terminals make rare synaptic contacts on the afferent nerve fiber.


Cell to cell relationships in the seminiferous epithelium in the mouse embryo. Germ cells and Sertoli cells in embryonic mouse testes (day 14 to 20 of gestation) were examined by sectioning and freeze-fracture. Intercellular cytoplasmic bridges between the germ cells are observed in day 14 and older embryos. Membrane specializations with dense fuzzy material similar to the so-called "desmosome-like structures" are found between Sertoli cells and germ cells. A cell contact area with dense opposed membranes is also found between adjacent germ cells. Asymmetrical dense fuzzy lining of both Sertoli and germ cell membranes is noted. Pinocytotic pits or caveolae are frequently found in the Sertoli cell membrane. Between adjacent Sertoli cells, gap junctions of various sizes and focal meshworks of the occluding junctions are found. Most of the occluding junctional particles are located in the center of the grooves in the E face, and are similar to those in postnatal and adult Sertoli cell junctions. In addition, on both fractured faces there are ridges and grooves devoid of particles which are continuous with occluding junctions with particles, suggesting an initial stage in the formation of occluding junctions of the Sertoli cells. Particles gathered at the site of desmosome-like structures are present on the P face of the Sertoli cell.


Synaptic junctions in the sinus gland of the freshwater prawn Palaemon paucidens. Two types of neurosecretory fibers, designated as Type 5 and Type 6 axons, in the sinus gland of the freshwater prawn, Palaemon, establish contact with other neurosecretory axons by means of synaptic junctions. This finding strongly supports the view that release of some neurohormones from the eyestalk may be regulated by neurosecretory neurons through synaptic transmission.


The sites of encephalic photoreception in phosoperiodic induction of the growth of the testes in the white-crowned sparrow, Zonotrichia leucophrys gambelii. In bilaterally enucleated White-crowned Sparrows, Zonotrichia leucophrys gambelii, testicular growth and development at rates similar to those of intact birds subjected to long days, can be induced by illumination of sites within and near the ventromedial hypothalamus from the tips of single, chronically implanted, light-conducting fibers. Essentially identical results are obtained with the same mode of encephalic illumination of birds with intact eyes held on short days that are otherwise nonstimulatory. Examination of the distribution of the sites of the fibers from which weak illumination induced testicular growth, increase in plasma level of luteinizing hormone or increase in plasma level of testosterone suggests that most of the photoreceptors must lie either within the ventromedial hypothalamus or in sites ventral thereto, such as the tuberal complex. For the time being, at least, we find it necessary to assume that some photosensitive elements of the system occur outside of the ventral hypothalamus.


The rhombencephalic recess in the rat. A light and electron microscopic study. The rhombencephalic recess, an ependymal organ, has been studied for the first time by light- and electron microscopy. It is situated mediosagitally on the floor of the rhomboid fossa at the level of the colliculus facialis. The recess and the superimposed tissue are built up by tanycytes, their apices being connected by tight junctions. HRP, injected into the c.s.f., does not penetrate into the intercellular clefts of the recess area. The recess area reveals a certain autonomy regarding its supply with arteries and capillaries. A blood-brain barrier exists, but shows slight leakage in circumscribed areas as a result of intense transendothelial vesicular transport. The organization of the recess area is compared with that of other ependymal organs, especially circumventricular organs.


Anastomosing tubular arrangement of dog exocrine pancreas. Wax reconstructions of zymogen granule-containing cells in dog show the arrangement of the exocrine pancreas to be a branching, anastomosing system of tubules which vary in diameter and end blindly. This arrangement is not that of a true acinar gland.


Avian enteric nerve plexuses. A histochemical study. The enteric nerve plexuses of the domestic fowl (Gallus domesticus) were investigated in sections and stretch preparations by means of the cholinesterase and glyoxylic acid fluorescence histochemical techniques. Cholinesterase-positive and varicose and non-varicose fluorescent nerve fibers were distributed at all levels of the gut in myenteric, submucosal, muscle and mucosal plexuses, and in a perivascular plexus. The density of the innervation and the detailed distribution of the nerves varied in different parts of the intestinal tract. All nerve plexuses appeared to be best developed in the rectum. Whereas the circular muscle coat contained a substantial number of nerves at all levels of the gut, the longitudinal coat was well innervated only in the rectum. The major portion of the mucosal plexus appeared to be associated with the intestinal glands. The nerve cell bodies were restricted to the myenteric and submucosal plexuses and were mainly cholinesterase-positive. Fluorescent ganglion cells were not observed. Pretreatment of stretch preparations with NADH: Nitro BT to stain ganglion cells showed that the majority of the cells were surrounded by a meshwork of fluorescent varicose fibres, although none of the fibres appeared to be associated with individual cells. The perivascular plexus was mainly associated with the arteries. The functional significance of the innervation is discussed.


Evolution of a human Y chromosome-specific repeated sequence. The structure and evolution of a repetitive sequence on the human Y chromosome has been studied by restriction enzyme analysis of both total DNA and the isolated sequence. The sequence is shown to cross-hybridize to sequences in female DNA forming unstable duplexes. Mouse/human cell hybrids have been used to investigate the pattern of sequence homology on the X chromosome and some autosomes. We conclude that this sequence is related to human satellite III, but shows considerable differences in structure.


Role of carbohydrates in protein secretion and turnover: effects of tunicamycin on the major cell surface glycoprotein of chick embryo fibroblasts. Using tunicamycin, we have investigated the role of glycosylation in the biosynthesis, processing and turnover of CSP, the major cell surface glycoprotein of chick embryo fibroblasts (CEF). This antibiotic specifically inhibits glycosylation mediated by dolichol pyrophosphate and consequently inhibits the glycosylation of asparaginyl residues of glycoproteins. Tunicamycin inhibited the incorporation of 3H-mannose into CSP by 92--98% and 14C-glucosamine by 84--96%, whereas total protein synthesis was decreased by only 15--45%. Tunicamycin treatment decreased total amounts of CSP by approximately 50--65%, with equal decreases in CSP occurring on the cell surface and in culture medium, whereas intracellular pools of CSP were not substantially affected. In contrast to CSP, three other membrane-associated proteins of apparent molecular weights 75,000, 95,000 and 150,000 daltons were found in increased amounts. Procollagen secretion was not inhibited by tunicamycin. Both procollagen and CSP secretion into culture medium were also not increased in AD6, a glycosylation-deficient, mutant mouse 3T3 cell line compared to wild-type cells. We examined the mechanism of the decrease in CSP after tunicamycin treatment. The rate of CSP biosynthesis as measured by pulse-labeling with 14C-leucine was not altered. Tunicamycin had only a slight effect on the initial times and rates of CSP appearance on the cell surface; some apparent intracellular redistribution of CSP was detected by immunofluorescence. The major effect of tunicamycin treatment was to accelerate the rate of degradation of CSP 2--3 fold. This increase is sufficient to account for the observed decreases after tunicamycin treatment. Our results suggest that carbohydrates may not be essential for CSP or procollagen synthesis, intracellular processing and secretion, but that carbohydrates may help stabilize CSP against proteolytic degradation.


Coordinate regulation of contractile protein synthesis during myoblast differentiation. The synthesis of contractile proteins has been studied during the differentiation of quail skeletal muscle myoblasts in culture. Myoblast differentiation was synchronized by transferring secondary cultures of rapidly dividing myoblasts into medium lacking cell division-promoting factors. Cultures at various stages of differentiation were then pulse-labeled with 35S-methionine, and cell extracts were resolved by electrophoresis on two-dimensional gels. Incorporation into specific proteins was quantitated by autoradiography and fluorography using a scanning densitometer. Contractile proteins synthesized by muscle cultures were identified by their co-electrophoresis on two-dimensional gels with contracile proteins purified from quail breast muscle. Our results show that the synthesis of myosin heavy chain, two myosin light chains, two subunits of troponin and two subunits of tropomyosin is first detected at the time of myoblast fusion and then rapidly increase at least 500 fold to maximum rates which remain constant in muscle fibers. Both the kinetics of activation and the molar rates of synthesis of these contractile proteins are virtually identical. Muscle-specific actin (alpha) synthesis also increases at the time of myoblast fusion, but this actin (alpha) is synthesized at 3 times the rate of other contractile proteins. The synthesis of 30 other muscle cell proteins was quantitated, and most of these are shown to follow different patterns of regulation. From these results, we conclude that the contractile proteins are regulated coordinately during myoblast differentiation.


The relationship between transformation and somatic mutation in human and Chinese hamster cells. The frequencies of transformations of primary human and Chinese hamster fibroblasts have been compared with the spontaneous and induced frequencies of mutation for resistance to thioguanine and ouabain, and for ability to use fructose, using the carcinogens benzo (alpha) pyrene and urethane. Whereas the rates and frequencies of mutation were similar in the two cell systems, transformations to morphologically altered cells was observed only in hamster cells. The frequency of this latter transformation event in hamster cells was abour 10(3) greater than the frequencies of mutation in these cells. The morphologically altered cells formed in the above transformation process cannot grow in agar (aga-) and do not produce tumors when injected into animals. The frequency of transition of these latter cells to aga+ cells which produce tumors in animals is similar to the mutation-like events.


Dexamethasone-resistant cystic fibrosis fibroblasts show cross-resistance to sex steroids. Diploid skin fibroblasts derived from individuals with the autosomal recessive disease, cystic fibrosis (CF), were shown previously to be significantly  more resistant to the cytotoxicity of dexamethasone, a glucocorticoid hormone, than were normal human fibroblasts. Here cystic fibrosis fibroblasts are also shown to be more resistant than normal human fibroblasts to the cytotoxic effects of the sex hormones, 17 beta-estradiol, dihydrotestosterone and progesterone. Since cells are believed to contain different receptors for each of the steroid hormones, it is not probable than the resistance of CF cells to these hormones results from a receptor deficiency. This was shown by the fact that CF cells were found to exhibit the same receptor activity as normal cells for 3-H-dexamethasone. Furthermore, neither normal human nor CF fibroblasts could be demonstrated to contain detectable receptor activity for 3H-17 beta-estradiol. In addition, the studies of fibroblast killing by hormones led to the further interesting observation that normal human diploid fibroblasts, regardless of the sex of the tissue donor, are sensitive to killing by each of the sex hormones. These findings suggest that the cytotoxic effects of the steroid hormones may be observed independently of the specific hormone receptors. The studies reported here thus suggest that the resistance of CF cells to the different steroid hormones is probably the result of a defect in a pathway in cellular steroid hormone metabolism other than that involving receptors.


Structure of chromatin containing extensively acetylated H3 and H4. I have grown HeLa cells in 5 mM sodium n-butyrate leading to extensive in vivo histone acetylation, and have characterized the structure of chromatin containing the modified histones. Nuclear DNA in butyrate-treated cells is digested 5-10 fold more rapidly by DNAase I than the DNA of control cells. Staphylococcal nuclease degrades the two nuclear samples to acid-soluble material with identical rates; this nuclease, however, does excise nucleosomes with extensively acetylated histones from the nucleoprotein chain preferentially. The physical properties of unsheared chromatin and isolated core particles from control and butyrate-treated cells are closely similar, as are the rates of digestion of core particles from the two cell preparations by DNAase I. Determination of the relative susceptibilities of cleavage sites for DNAase I demonstrates that the site 60 bases from the ends of the DNA resistant in control cells, becomes susceptible to the nuclease in core particles containing acetylated histones. Similarly, the 5' terminal phosphate at the end of DNA in core prticles is removed by staphylococcal nuclease 2-3 fold faster in particles containing acetylated histones than in particles from control cells.


Nucleotide sequence heterogeneity of an RNA phage population. The nucleotide sequence of 32P-RNA from Q beta phage clones was sampled by two-dimensional polyacrylamide gel electrophoresis of the RNAase T1-resistant oligonucleotides (T1 fingerprinting). About 15% of the clones derived from a multiply passaged Q beta population showed fingerprint patterns which deviated from that of the RNA from the total population. All deviations examined could be attributed to one and, less frequently, to two or more nucleotide transitions. Since the fingerprinting technique allows the analysis of only about 10% of the RNA sequence, we estimate that each viable phage genome in a multiply passaged population differs in one to two positions from the "average" sequence of the parental population. Several deviant clones were tested by growth competition against a "wildtype" population, after 10-20 generations, the resulting phage showed the "wild-type" T1 fingerprint pattern. We propose that a Q beta phage population is in a dynamic equilibrium, with viable mutants arising at a high rate (Batschelet, Domingo and Weissmann, 1976; Domingo, Flavell and Weissmann, 1976) on the one hand, and being strongly selected against on the other. The genome of Q beta phage cannot be described as a defined unique structure, but rather as a weighted average of a large number of different individual sequences.


Structure of the genome of Moloney murine leukemia virus: a terminally redundant sequence. The genome of the Moloney strain of murine leukemia virus (Mo-MuLV) has been analyzed by digestion with ribonuclease T1 and separation of the digestion products by two-dimensional gel electrophoresis. Thirty large oligonucleotides isolated from such a fingerprint have been characterized. One of these oligonucleotides (number 21) was found to be present in twice the molar yield of the rest. The 30 oligonucleotides were mapped on the genome by determining their yields in various size classes of 3' terminal fragments of Mo-MuLV RNA. The physical map obtained in this way suggested that oligonucletoide 21 was present very near the 3' end of the geome as well as in another location near or at the 5' end. The genome structure suggested by these results was confirmed by analyzing oligonucleotides in Mo-Mulv RNA complementary to strong stop DNA, which is shown to be a copy of the 5' terminal 134 nucleotides of the MoMuLV genome. Some of the oligonucleotides in the RNA protected from RNAase digestion by hybridization to this DNA, including oligonucleotide 21, were present near both the 3' and 5' ends. Comparison of these with the nucleotide sequence of strong stop DNA shows that there is a terminal redundancy of 49-60 nucleotides in the Mo-MuLV genome RNA.


The mobilization of maternal histone messenger RNA after fertilization of the sea urchin egg. The extent of protein, RNA and DNA synthesis in early cleavage stages of the sea urchin embryo (Parechinus angulosus) was determined. A histone mRNA specific cDNA was used in hybridization experiments to investigate the cytoplasmic localization of maternal histone mRNA in the unfertilized sea urchin egg and first cleavage stage embryo. In the unfertilized egg histone mRNA was localized exclusively in ribonucleoprotein particles with none in ribosomes or polyribosomes. This distribution changed after fertilization, in particular, coupled with the first cleavage telophase there was a significant transfer of histone mRNA from the ribonucleoprotein fraction to the polyribosomes. The results indicate mRNA specific translational control mechanisms.


Developmental studies on an apparent cell-lethal mutant gene-ut-in the Mexican axolotl, Ambystoma mexicanum. The discovery of a new mutant gene in stocks of the Mexican axolotl derived from breeding stock of the Hubrecht Laboratory, the Netherlands, is described. The gene appears to be a simple recessive and displays complete penetrance. ut/ut larvae develop normally through hatching, but begin to lag in growth and display characteristics defects in gill and limb formation shortly thereafter. The results of parabiosis of normal and mutant embryos, as well as embryological transplants of mutant limb and branchial rudiments, support the conclusion that the gene ut is expressed as an 'autonomous-cell lethal'. Despite gross morphological defects in ut/ut larvae, comparisons between normal and mutant animals of the protein spectra of various tissues and organs revealed no substantial differences. A subtle change in the metabolism of ut/ut larvae apparently, therefore, leads to developmental arrest.


Metabolically stable non-histone chromosomal proteins in growing maize roots. DNA and non-histone chromosomal proteins (NHCP) of meristematic cells of maize primary roots were double labelled in vivo with [3H]- or [14C] thymidine and [14C]- or [3H]-tryptophan respectively. The ratio of labelled tryptophan to labelled DNA was followed during the transition of the meristematic cells of the root tip into the distal zones of finally differentiated cells. It was found that only 20% of the newly synthesized NHCP in the proliferating cells were turned over, while the rest were preserved and found as metabolically stable proteins in the zone of final differentiation. This result is consistent with the hypothesis that some NHCP remain permanently associated with chromatin of non-dividing differentiated cells in order to maintain the genomic characteristics of a given cellular type.


Repair of DNA damaged by mutagenic metabolites of benzo(a)pyrene in human cells. The repair of human DNA after damage by known and potential metabolites of benzo(a)pyrene has been examined utilizing the bromodeoxyuridine photolysis assay. Repair was characterized as either ultraviolet ("long") or ionizing radiation type ("short") repair utilizing normal cells and cells deficient in ultraviolet-type repair endonuclease from a patient with xeroderma pigmentosum (XP). We have found that only (+/-)-7beta,8alpha-dihydroxy-9beta,-10beta-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (BP diol epoxide 1) and its disastereomer, (+/-)-7beta,8alpha,-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (BP diol epoxide 2) elicit damage to DNA which is recognizable by the ultraviolet excision repair system in normal human cells. Benzo(a)pyrene 4,5-, 9,10-, 11,12-oxides do not elicit damage which is repairable by this repair system. The 1,2-diol-3,4-epoxides from naphthalene have no measurable activity in our assay. These results indicate that both the benzo(a)pyrene ring structure and the diol epoxide groups are important in causing the damage to DNA which is repairable by the ultraviolet excision repair system. These results parallel the reported high mutagenic activity of these compounds and support the concept that benzo(a)pyrene 7,8-diol-9,10-epoxides may be the ultimate, metabolically activated forms of benzo(a)pyrene.


Postreplication repair of DNA in human fibroblasts after UV irradiation or treatment with metabolites of benzo(a)pyrene. We have examined the ability of normal fibroblasts and of excision-deficient xeroderma pigmentosum (XP) and XP variant fibroblasts to perform postreplication DNA repair after increasing doses of either ultraviolet (UV) irradiation or mutagenic benzo(a)pyrene derivatives. XP cells defective in the excision of both UV-induced pyrimidine dimers and guanine adducts induced by treatment with the 7,8-diol-9,10-epoxides of benzo(a)pyrene were partially defective in their ability to synthesize high molecular weight DNA after the induction of both classes of DNA lesions. This defect was more marked in XP variant cells, despite their ability to remove by excision repair both pyrimidine dimers and the diol epoxide-induced lesions to the same degree as observed in normal cells. The benzo(a)pyrene 9,10-oxide had no effect in any of the 3 cell lines. The response of the excision and postreplication DNA repair mechanisms operating in human fibroblasts treated with benzo(a)pyrene 7,8-diol-9,10-epoxides, therefore, appears to resemble closely that seen after the induction of pyrimidine dimers by UV irradiation.


Formation of DNA-binding products from isolated benzo[a]pyrene metabolites in rat liver nuclei. Liver nuclei from 3-methylcholanthrene-treated rats in the presence of NADPH metabolized 3- and 9-hydroxybenzo[a]pyrene and 7,8-dihydro-7,8-dihydroxybenzo[a]pyrene to products that bound to DNA. Maximal binding was obtained with the dihydrodiol which was approximately 3-fold that with 9-hydroxybenzo[a]pyrene, and 60-fold that with 3-hydroxybenzo[a]pyrene, as substrates. Both 4,5-dihydro-4,5-dihydroxybenzo[a]pyrene and 9,10-dihydro-9,10-dihydroxybenzo[a]pyrene were also extensively metabolized by the nuclear fraction but did not give rise to DNA-binding products. The available evidence suggests that the DNA binding species derived from 9-hydroxy-benzo[a]pyrene is 9-hydroxy-benzo[a]pyrene-4,5-oxide and from 7,8-dihydro-7,8-dihydroxybenzo[a]pyrene, as previously observed in different systems, 7,8-dihydro-7,8-dihydroxy-benzo[a]pyrene-9,10-oxide.


Nitrofurazone-induced DNA damage to tissues of mice. Cytotoxicity and DNA damage by nitrofurans has previously been correlated with metabolic reduction of these drugs in vitro. In the present study, nitrofurazone increased the rate of disappearance of stable [3H]thymidine labelled DNA from tissues of mice fed 0.1% nitrofurazone in the diet. Significant loss of labelled DNA occurred within 25 days after the start of the diet in all tissue observed, and loss was in relation to the rate of metabolic reduction of nitrofurazone. A similar correlation was found when another endpoint for DNA damage was used; nitrofurazone reduced by mouse tissue slices caused DNA single-strand breaks in cultured mouse L cells incubated in vitro with the tissues. Again, the ability of each tissue to produce toxic nitrofurazone metabolites determined the amount of DNA damage to the L cells.


Metabolism of N-nitrosomorpholine by the rat in vivo and by rat liver microsomes and its oxidation by the Fenton system. N-Nitrosomorpholine is converted into N-nitroso-2-hydroxymorpholine by rat liver microsomes and by the Fenton oxidation system. The hydroxy derivative was also synthesised by the oxidation of N-nitrosodiethanolamine with permanganate and characterized as the methoxime and the 2,4-dinitrophenylhydrazone. The Fenton system also afforded products believed to be N-nitroso-2-morpholone, and the 2-hydroperoxy- and 2-peroxy-derivatives of N-nitrosomorpholine. The only urinary metabolite definitely identified was N-nitrosodiethanolamine. The significance of metabolic 2-hydroxylation in relation to the carcinogenic action of N-nitrosomorpholine is discussed.


The identification of 3-methylcholanthrene-9,10-dihydrodiol as an intermediate in the binding of 3-methylcholanthrene to DNA in cells in culture. Two metabolites of 3-methylcholanthrene (3MC), previously shown to be precursors of the DNA-bound form of 3MC observed in embryo cells in culture, were prepared from 3MC by microsomal metabolism and isolated by high pressure liquid chromatography (HPLC). From HPLC analysis of the metabolites of 3MC, from mass spectrometric analysis and from comparison with the fluorescence spectra of all 5 possible dihydrodiols of the alkylated benzanthracenes, it was deduced that one of the precursor metabolites was a 9,10-dihydrodiol of 3MC while the other was a 1 or 2-hydroxy derivative thereof.


Modification by metals, sulphydryl reagents and cyanide of the particle stimulated enhancement of oxygen consumption in bovine granulocytes. A simple, rapid method for the isolation of neutrophil-enriched leucocyte suspensions from bovine blood is described. The capacity of these cells to produce a particle stimulated increase in oxygen consumption deteriorated during a period of storage of the cells whilst the viability of the cells remained unchanged. Potassium cyanide inhibited the basal oxygen consumption but enhanced the stimulated respiratory burst. Zinc ions also enhanced this respiratory burst but ferric and manganous ions did not. The lipid-soluble non-haem iron chelator, 4,4,4-trifluoro-1-(2 thienyl)-1,3-butanedione, preferentially inhibited the particle stimulated type of oxygen consumption, as did the sulphydryl reagents, N-ethyl maleimide and diazine dicarboxylic acid bis-dimethyl amide. These data allow us to consider that zinc ions may play a role in the respiratory burst associated with phagocytosis and that iron-sulphur interactions may be important in oxygen consuming reactions.


[Castleman's benign lymphoma].

[Castleman's benign lymphoma]. The Authors, when presenting a personal case of CBL, make a critical revision of the literature, summing up the evolution of knowledge of this morbid picture. They analyse and discuss the aetiological theories suggested by other Authors on the basis of clinical and anatomopathological criteria. Lastly they evidence the difficulty of diagnosis both for the clinical physician and for the anatomopathologist, since there are numerous pathological pictures which may simulate CBL in the symptoms when present, the x-ray aspects and even the anatomohistological examination.


[The amylase-creatinine clearance ratio in the differential diagnosis of pancreatitis and gastroduodenal ulcer with hyperamylasemia]. The Authors followed 29 patients, hospitaled with a diagnosis of pancreatitis. They all presented the same sympotomatology and a considerable increase of the serum amylase ad urinary amylase. The examination of the ratio between the clearance of amylasis and creatinine permitted to make a differential diagnosis for 8 cases (4rd group) that were nothing but peptic ulcera. Such a diagnosis was confirmed by the radiological contrastographic examination or by the intraoperative report. So if the ratio between the clearance of amylase and creatinine is normal we must think about a pathological situation were the iperamylasemia has a pathogenetic cause different from pancreatitis.


[Our experience in the surgical treatment of perforated duodenal ulcer]. The Authors present their surgical experience of more than a decade with regard to emergency therapy of perforated duodenal ulcers. The paper covers the case of 124 patients hospitalized urgently and variously subjected to the aspirative method according to Taylor, raphe, vagotomy and pyloroplasty according to Judd, or immediate gastric resection. With regard to the short, medium and long term results, they conclude in favour of immediate vagotomy, free from dangers and mortality, the fairly positive nature of immediate gastric resection, and the failure or at any rate low reliability of the other two methods used.


Follow-up to 16 years of school children who had marked speech defects at 7 years. In this paper we present the results of a follow-up study to age 16 years of 215 children in the National Child Development Study who were reported to show marked speech defects with normal hearing at 7 years. At 16 years information was available for 180 (85%) of the surviving 212 children. 30% of these children were in special schools and of the 70% in ordinary schools, 51% were reported still to have a residual speech problem. Additional health and social problems were frequent among the study group children and with very few exceptions they did badly at school. The findings at 16 years strongly reinforce our earlier conclusions that markedly defective speech at 7 years indicates the likelihood of continued backwardness in the development of acceptable verbal communication, social maturity and scholastic attainments.


The magnitude of the electromotive force of canine ventricular myocardium. The isolated and perfused dog heart was placed in a cubic container filled with Tyrode's solution. Ventricular ectopic beats were produced by electrical stimulation of the left ventricular wall, and initial QRS vectors of these beats were determined with orthogonal leads from the surface of the container. At the same instants, the activated area on the epicardial surface was measured by means of a large number of contiguous bipolar leads from the epicardial surface. The QRS vector and the activated epicardial area were found to be nearly porportional. By use of these results and a calibration system with artificial dipoles, the double layer moment of the ventricular activation wave was calculated as 0.13 mA.cm per unit area. This value corresponds to 60% of the maximal possible strength of the tissue electromotive force. Lowering the conductivity of the surrounding solution increased the QRS voltage but not as much as the potential caused by a constant-current dipole within the solution. The relationship between the QRS voltage and the conductivity of the medium was analyzed by a simplified model of the system and was found to correspond approximately to that of a constant-current source within a spherical heart with a resistivity 2 to 3 times that of Tyrode's solution.


Action research in day nurseries: evaluating programmes through staff perceptions and attitudes. Day nurseries in deprived inner-city areas have to cope with a wide range of problems which are presented by the young children and their families. Many of the children can be delayed in some area of their development or have emotional problems which require sensitive handling. These challenges are in addition to the expected difficulties of helping young children to adjust to a new situation and the presence of unfamiliar adults and children. The staff of day nurseries tend to be young and the 2-year training has a great deal to cover on normal development. The consequence is that staff in their late teens and early twenties have to face the demands of children whose development is often far from 'normal' in terms of the average range. It was a concern about this situation in a very disadvantaged area of London and the fact that nurseries have traditionally been fairly isolated institutions that led to the planning of a 2-year exploratory project with the objective of introducing new ideas and approaches to nursery staff which would help them provide a better quality of care to socially disadvantaged under-5s.


Whole body vascular reactivity during the development of deoxycorticosterone acetate hypertension in the pig. A change in vascular reactivity was characterized in the pig during the development of deoxycorticosterone acetate (DOCA) hypertension. Pigs, 8-12 weeks of age, were subjected to unilateral nephrectomy and instrumented with an electromagnetic flowprobe on the ascending aorta and with Tygon catheters in the aorta and thoracic vena cava. Approximately 2 weeks after surgery, Silastic strips impregnated with DOCA (100 mg/kg) were implanted subcutaneously. Within the first 3 days after implantation, mean arterial pressure (MAP) began to rise and reached levels approximately 40% greater than control during the 4th week. Graded intravenous infusions of norepinephrine or angiotensin II were given to unanesthetized pigs before, and at intervals during, the development of hypertension. Changes in total peripheral resistance were calculated from recordings of MAP and cardiac output. Comparison of pre-DOCA response curves with those obtained at intervals following DOCA implantation demonstrated a significant increase in vascular smooth muscle sensitivity (decrease in threshold infusion rates) to both drugs post-DOCA. There also was a shift to the left in the dose-response curves. These changes in systemic vascular reactivity occurred at the time the arterial pressure began to rise. The temporal relationship suggests that the increase in vascular reactivity may initiate the increase in mean arterial pressure.


Services for the mentally handicapped in Sweden. This paper, based on a visit by the authors to Sweden, describes Swedish services for mentally handicapped people and discusses the viability of the Swedish model in contrast with services in Britain.


Inhibition of protein degradation in mouse hearts by agents that cause lysosomal dysfunction. Although the heart contains lysosomes, it has been uncertain whether these organelles and their proteolytic enzymes can play an important role in cardiac protein degradation. Recent studies have demonstrated that fetal mouse hearts in organ culture sustain selective derangements in lysosomal structure and function during exposure to chloroquine or nonmetabolizable sugars. Accordingly, we tested the effects of these agents on cardiac proteolysis under controlled conditions in vitro using two techniques (measurement of loss of radioactivity from trichoroacetic acid-precipitable protein after prelabeling with tritiated phenylalanine and measurement of loss of cold phenylalanine after blockade of protein synthesis with cycloheximide). Chloroquine (0.1 mM) reduced the average rate of protein breakdown in hearts of matched littermates from 45%/24 hours to 32%/24 hours (P less than 0.01) and decreased the release of cold phenylalanine by 31 +/- 5% (0.108 vs 0.075 nmol/mg per hour, P less than 0.01). Exposure to 100 mM sucrose for 24-48 hours reduced the rate of breakdown from 44%/24 hours to 33%/24 hours (P less than 0.01) and decreased the release of cold phenylalanine by 35 +/- 9% (0.092 vs. 0.060 nmol/mg per hour, P less than 0.01). The results suggest that interference with lysosomal function in cultured fetal mouse hearts causes a significant reduction in the cardiac capacity to degrade proteins.


Hydrolase activities in the rat aorta. II. Effects of hypertension alone and in combination with diabetes mellitus. Hypertension is an important risk factor for atherosclerosis and often occurs in association with diabetes mellitus. Specific activities of hydrolases in homogenates of aortas from rats with renal-clip hypertension, normotension following a period of hypertension, and hypertension combined with streptozotocin-induced diabetes mellitus were measured. Enzymes included: neutral alpha-glucosidase, and lysosomal N-acetyl-beta-glucosaminidase, beta-galactosidase, cathepsin C, acid alpha-glucosidase, and acid cholesteryl esterase. After 6 or 12 weeks of hypertension, specific activities of all enzymes measured were significantly increased, levels ranging from 24% above normal for cathepsin C to 351% above normal for N-acetyl-beta-glucosaminidase. Six weeks of normotension following 6 weeks of hypertension resulted in restoration to normal of four of the six enzyme activities; the remaining two enzymes were significantly below normal levels. Combined hypertension and diabetes mellitus showed smooth muscle cell levels of four of the five hydrolases measured to be significantly lower than those present with hypertension alone. In every instance, histochemical studies of aortas showed acid phosphatase and N-acetyl-beta-glucosaminidase activities which corresponded to the biochemical findings. These findings indicate profound and discrete effects of two clinical risk factors on vascular smooth muscle cell lysosomes.


The pressor response to angiotensin II in patients with low renin essential hypertension. The etiology of low renin essential hypertension (LREH) has not been established with certainty, but mineralocorticoid excess has been implicated frequently in its pathogenesis. The finding of several investigators of a normal exchangeable sodium space and extracellular fluid volume, however, does not support this hypothesis. To evaluate the possible role of sodium and water retention in LREH, the pressor response to infused angiotensin II (A II) was determined and compared to that of normal subjects and that of subjects with normal renin essential hypertension (NREH). This approach was based on the known suprasensitivity of vascular receptors to A II in situations in which sodium and water compartments are expanded as they are, for example, in proven hypermineralocorticoid states such as primary aldosteronism. In this study, we found that subjects with LREH demonstrated no increased pressor response to graded doses of A II; this suggests that LREH is not primarily mediated by sodium and water retention.


Clonal markers in the study of the origin and growth of human atherosclerotic lesions. The X-linked enzyme, glucose-6-phosphate dehydrogenase (G-6-PD) was used as a cellular marker to study the clonal characteristics of human atherosclerotic lesions from females heterozygous for G-6-PD isoenzymes. Portions of uninvolved aortic wall contained both isoenzyme types (A and B), and their isoenzyme patterns were used to establish criteria for polyclonal lesions. Portions of uterine leiomyomas contained predominantly one isoenzyme type (either all A or all B) and their isoenzyme patterns were used to establish criteria for monoclonal lesions. These techniques were used to address three questions concerning atherogenesis. First, evidence for the monoclonal origin of fibrous-capped plaques was provided by the findings that small plaques had G-6-PD isoenzyme distributions similar to those of leimyomas; that in large plaques with multiple portions assayed for G-6-PD, a large proportion (25 of 26, 96%) of plaques had monoclonal characteristics; and that multiple monoclonal portions were present in the same plaque. Second, the role of the fatty streak  as a precursor of fibrous plaques was supported by the demonstration that a proportion (11 of 66, 16.7%) of fatty streaks contained isoenzyme patterns intermediate between those of polyclonal uninvolved aortic wall and monoclonal leiomyomas. Increased cellularity of fatty streaks correlated with increased deviation of isoenzyme pattern toward monoclonality. Third, the assay of portions of both small and large plaques provided no evidence for clonal selection as plaques increase in size.


Suppression of calcific fibrous-fatty plaque formation in rabbits by agents not affecting elevated serum cholesterol levels. The effect of thiophene compounds. We tested the suppressive effect of antihypercalcemic-hyperphosphatemic agents on atherogenesis. We studied five groups of rabbits for 8 weeks, one control group and four groups on a fibrogenic atherogenic diet. One group received the atherogenic diet alone, and the remaining three atherogenic groups were treated simultaneously with 2-thiophenecarboxylic acid (ThCA), 5-methyl-2-thiophenecarboxylic acid (5-CH3-ThCA), and 5-bromo-2-thiophenecarboxaldehyde (5-Br-ThCA). Rabbits receiving the atherogenic diet alone developed: (1) elevations of serum cholesterol, calcium, and phosphorus; (2) massive fibrous-fatty aortic plaques with excessive accumulation of aortic collagen, elastin, and lipids; (3) marked deposition of calcium and phosphorus in both aortic tissue and elastin; and (4) severe lipid infiltration of the liver. Treatment with all three drugs normalized the elevated serum calcium but not the cholesterol levels, and effectively inhibited all aspects of the atherosclerotic process as determined morphologically and biochemically. The order of effectiveness was: 5-CH3-ThCa greater than 5-Br-ThCA greater than ThCA. No bone resorption occurred in the treated groups. The normalizing effects of the thiophene compounds on serum phosphorus levels were not significant at the dosages used.


The relationship of excitability to conduction velocity in canine Purkinje tissue. The relationship between interelectrode conduction time and "take-off" potential (TOP) was studied with microelectrode techniques in isolated canine false tendons. Conduction of regular or test beats initiated during phase 4 dopolarization or late phase 3 repolarization speeded as TOP decreased. Similarly, beats initiated during digitalis-induced oscillatory after potentials demonstrated more rapid conduction at lower TOP. Because of the frequency-coupled nature of the oscillations, conduction times became rate dependent. Phenytoin antagonized digitalis oscillations and reversed speeding of conduction attributable to the oscillations. No uniform relationship between speed of conduction and maximum upstroke velocity of the action potential could be demonstrated in the above experiments or when speed of conduction was varied by changes in concentration of K+ or Ca2+. However speed of conduction could be demonstrated to vary directly with changes in excitability as measured by intracellular current injection.


Evidence for a splanchnic sodium input monitor regulating renal sodium excretion in man. Lack of dependence upon aldosterone. Eight normal male subjects were placed on a constant 10 mEq sodium, 60 mEq potassium diet for 5 days. At 8:00 a.m. on the 5th day, the subjects were given a standard dose of 100 mEq of sodium orally or intravenously. Subjects receiving oral sodium also received 200ml of 5% dextrose in water intravenously, and those receiving intravenous sodium also received placebo capsules orally. Water intake and posture were controlled. The subjects then returned to a free diet for 1 month and subsequently were restudied by using the opposite route of sodium administration. The subjects given the oral sodium load excreted greater quantities of sodium in their urine than those repleted intravenously. The differential natriuresis was significant as early as 2 hours after sodium loading. Plasma aldosterone concentration was similar irrespective of the route of sodium administration. Six patients with primary adrenocortical insufficiency  and documented hypoaldosteronism were studied with the same protocol after 5 days of 50 mEq sodium, 60 mEq potassium intake. They also had significantly greater natriuresis after oral than intravenous sodium administration. The data suggest the presence of a splanchnic input monitor for sodium which partially regulates renal sodium excretion and is not dependent upon a turn-off mechanism for aldosterone secretion.


Effect of l-dopa on sympathetic nerve activity and blood pressure in the spontaneously hypertensive rat. We examined the effect of L-Dopa, after peripheral L-amino acid decarboxylase inhibition, on sympathetic nerve activity (SNA) and blood pressure in spontaneously hypertensive rats (SHR) and in normotensive control rats. L-Dopa reduced SNA in both groups of animals. The SHRs were significantly more sensitive to the depressor effect of L-dopa than were the control animals, the threshold dose for reduction of SNA being 3 mg/kg in the SHR and 15 mg/kg in control rats. Similarly, the magnitude of inhibition of SNA was substantially greater in the SHR than in normotensive rats. The reduction in SNA in the SHR accompanied by a parallel fall in blood pressure. In contrast, blood pressure in control rats did not change significantly, even though SNA was diminished. Studies of the penetration of L-dopa into the cerebral parenchyma revealed that equivalent amounts of the amino acid entered the brains of the two groups of rats. These results suggest that the SHRs are more sensitive to the SNA-inhibiting effects of L-dopa than are normotensive rats. In addition, they confirm our previous suggestion that excessive SNA plays a causative role in the hypertension of the SHR.


The effects of acetylcholine on the electrical activity of canine cardiac Purkinje fibers. We studied the effects of acetylcholine (ACh) on small bundles of canine cardiac Purkinje fibers exposed to normal, or low-chloride (isethionate) Tyrode's solution in a rapid superfusion system. In superfusate containing 4 mM K+, the resting potential of Purkinje fibers may be either "low," near -40 mV, or "high," near -90 mV. ACh, at 10(-6) to 10 (-5) M, increased the membrane potential from both the low and high resting levels and, in low-Cl solution, often induced a maintained shift in potential from the low to the high level. The increase in membrane potential caused by ACh was greater at the low than at the high level. ACh, at 10(-6) to 10(-5) M, reduced action potential duration in both normal and low-Cl Tyrode's solution, the effect being more marked in the latter. These effects of ACh were reversibly abolished by atropine (5 X 10(-5) M), indicating that they were mediated via muscarinic ACh receptors, and they probably result from an increase in membrane K+ conductance since 10(-5) M ACh reversibly reduced, by 13% on the average, the amplitudes of the steady changes in membrane potential evoked by applying small current pulses (-5 to -25 nA, 200 msec). ACh (10(-5) M) also diminished the rate of, or stopped, spontaneous activity arising from either level of membrane potential. The cessation of spontaneous slow response activity, arising from the low level, sometimes was accompanied by a maintained shift of the membrane potential to the high resting level. It is concluded that the action of ACh on Purkinje fibers is qualitatively similar to its action on sinoatrial nodal and atrial cells.


Renal function and renin secretion after administration of ouabain and ouabain plus furosemide in conscious sheep. The effects of ouabain or ouabain and furosemide on renal function and renin secretion were studied in conscious isovolemic sheep. The sheep received a continuous renal arterial infusion of papaverine, 7 mg/min, throughout the experiment. Ouabain alone (7 X 10(-7) M in the renal plasma) produced significant decreases in glomerular filtration rate (GFR) and renal plasma flow (RPF) but not in renal perfusion pressure. Plasma [K+] rose after ouabain administration. Fractional (FENa) and absolute (UNaV) Na+ excretion were 2.9 +/- 1.0% (mean +/- SE) and 78 +/- 54 muEq/min, respectively, during the papaverine infusion and rose to 19 +/- 5.1% (P less than 0.05) and 528 +/- 116 muEq/min (P less than 0.01) after ouabain administration. Despite the large changes in Na+ reabsorption, renin secretion was not stimulated. During the control period, renin secretion was 281 +/- 131 ng/min and the average renin secretion after ouabain administration was 310 +/- 78 ng/min (not significant). A smaller dose of ouabain (2 X 10(-7) M) infused into the renal artery with 40 mg of furosemide, iv, did not decrease GFR but RPF was suppressed. FENa and UNaV averaged 4.4 +/- 1.6% and 121 +/- 44 muEq/min, respectively, while papaverine was infused into the renal artery and increased to 18 +/- 4.8% (P less than 0.05) and 636 +/- 209 muEq/min (P less than 0.05) after ouabain and furosemide were infused. Renin secretion was 118 +/- 62 ng/min during the control period and averaged 240 +/- 67 ng/min after ouabain plus furosemide. The difference was not statistically significant. Thus ouabain alone does not stimulate renin secretion in the conscious, isovolemic sheep despite a presumed increase in [NaCl] at the macula densa and inhibition of NaCl transport by the loop of Henle. Ouabain also blocks the normal stimulatory effects of furosemide on renin secretion.


Sequential unipolar strength-interval curves and conduction times during myocardial ischemia and reperfusion in the dog. Computerized techniques were employed to generate alternating anodal and cathodal or sequential anodal strength-interval curves during and following 15-minute coronary artery ligations in 14 anesthetized dogs. The right atrium was paced at 2.5 Hz, and unipolar ventricular strength-interval curves with simultaneous conduction times were recorded every 45-120 seconds during ischemia and reperfusion. Within 1--2 minutes of ligation, anodal midcurve and late diastolic thresholds fell sharply, and cathodal thresholds fell slightly or changed little. After 5 minutes of ischemia, anodal thresholds remained low, cathodal thresholds rose, and conduction times increased. At 10--15 minutes of ligation, if the ischemic zone was small, anodal thresholds were low, often approaching cathodal values, and conduction returned toward control values. When the ischemic zone was large, unipolar thresholds and conduction times increased late during the ligation period. Throughout the course of ischemia, the falling limb of the strength-interval curve shifted progressively to the left indicating shorter refractory periods. Following abrupt reperfusion, anodal phase 3 dips promptly reappeared; refractory periods returned toward control, and supernormal conduction was noted. By 3--5 minutes of reperfusion, the falling limb of the strength-interval curve had shifted to the right of control and conduction times increased. Thus, vulnerability to arrhythmias during early ischemia (i.e., 5 minutes) is characterized by low anodal midcurves and late diastolic thresholds, short refractory periods, and slow conduction. During the first minute of reperfusion, anodal excitability is increased during the early dip and conduction times are supernormal. Increases in anodal excitability correlate better with the peak incidence of early ligation and reperfusion arrhythmias than do changes in cathodal excitability.


Morphological changes in isolated perfused dog lungs after acute hydrostatic edema. Morphometric data from stable (nonedematous) isolated dog lungs, perfused with nearly cell-free perfusates, were compared to similar stereological evaluations of isolated dog lungs after induction of severe acute hydrostatic edema. In the edematous lungs, capillary surface and volume densities were substantially increased. Alveolar surface density was also increased. Thicknesses of the endothelial and type I epithelial cellular compartments of the air-blood barrier were unchanged. Thickness of the interstitial compartmemt of the air-blood barrier was substantially increased and this, in turn, caused an overall increase in mean thickness of the barrier. Volume densities of the nonparenchymal connective tissue spaces surrounding the extra-alveolar vessels and airways were also increased. In both the endothelial and type I epithelial cells cytoplasmic volume densities of pinocytotic vesicles were increased. In addition, the number of vesicles opening onto the luminal and albuminal cellular surfaces increased signficantly. Transendothelial vesicular passage may contribute to interstitial edema formation, and transepithelial vesicular transport may contribute to alveolar flooding in isolated perfused dog lungs.


Initial myocardial adjustments to brief periods of ischemia and reperfusion in the conscious dog. The changes in left ventricular (LV) dynamics induced by brief periods of ischemia (100 seconds) and subsequent reperfusion were analyzed in conscious dogs. Global LV ischemia, induced by partially occluding the left main coronary artery, reduced LV flow homogeneously and impaired LV function as reflected by decreases in LV stroke "work" (89 +/- 4% M +/- SE), systolic shortening (72 +/- 4%), velocity of shortening (56 +/- 6%), LV systolic pressure (34 +/- 5%), and dP/dt (59 +/- 6%). Regional LV ischemia, induced by occluding either the left circumflex or anterior descending coronary artery completely, reduced flow to the ischemic segment (82 +/- 3%) while decreasing segment work (96 +/- 5%), shortening (82 +/- 3%), and velocity of shortening (70 +/- 5%), with minimal depression of overall LV function. In both groups the extent of shortening was reduced more rapidly and greater (P less than 0.01) than shortening velocity. Moreover, with localized ischemia, segment work was reduced more (P less than 0.01) than shortening. With reperfusion, a transient overshoot in function above preischemic control levels was observed in both groups (global work increased by 60 +/- 12% and regional work by 28 +/- 4% above control). This overshoot was not dependent on adrenergic mechanisms, but was prevented by inhibiting reactive hyperemia. Thus myocardial ischemia induces a dissociation between extent and rate of myocardial shortening. A further dissociation between shortening and work is apparent with regional ischemia. After reperfusion there is a transient overshoot in function which appears to be dependent upon the associated reactive hyperemia.