Rabbit liver pyridoxamine (pyridoxine) 5'-phosphate oxidase. Purification and properties.

Pyridoxamine (pyridoxine) 5'-phosphate oxidase (EC 1.4.3.5) has been purified 2000-fold from rabbit liver. The enzyme preparation migrates as a single protein and activity band on analytical disc gels containing 4,7, or 9 percent acrylamide, and as a single protein band on sodium dodecyl sulfate acrylamide gels. The oxidase is, therefore, homogeneous by these criteria. The pure enzyme catalyzes the following reactions in the presence of FMN: (See journal for formula). These activities copurify in the ratio of 1:1:1. Apparent K-m values are 10 muM for pyridoxamine-P, 30 muM for pyridoxine-P, and 40 nM for FMN. Apparent K-m values for N-(phosphopyridoxyl)amines range from 3.1 times 10-5 M to 1.6 times 10-3 M. The dissociation constant for FMN binding, determined by quenching of protein fluorescence, is 20 nM. The pH optima for all three types of substrates are broad, with maxima near pH 9. The pH dependence of FMN binding, measured by quenching of flavin fluorescence, has the same shape as the substrate activity profile. The holoenzyme has absorption maxima red-shifted from those of FMN to 380 nm and 448 nm, and exhibits spectral changes typical of flavoproteins upon reduction with dithionite. Its oxidation-reduction potential at pH 7 in phosphate buffer is -0.131 volt. The native enzyme has a molecular weight of 54,000 and is made up of two possibly identical polypeptide chains with molecular weights of 27,000. The applicability of proposed mechanisms of flavin catalysis to this flavoprotein is discussed.


A specific enzyme for glucose 1,6-bisphosphate synthesis.

The reaction: glycerate-1,3-P2 PLUS GLUCOSE-1-P YIELDS TO GLUCOSE-1,6-P2 plus glycerate-P is catalyzed by a distinct enzyme of mouse brain. A divalent metal requirement was shown when the enzyme was treated with imidazole and EDTA. Mg2+, Mn2+, Ca2+, Zn2+, Ni2+, Co2+, and Cd2+ were quite effective cofactors. The enzyme, in better than 50 percent yield, has been purified away from 99 percent of the phosphoglucomutase, phosphoglycrate mutase, and phosphofructokinase. Acetyl-P, ATP, enolpyruvate-P, creatine-P, and fructose-1,6-P2 are not phosphoryl donors. Glucose-6-P and mannose-1-P are good alternate acceptors. Mannose-6-P, galactose-Ps, and fructose-Ps have little or no acceptor activity. Strong inhibition was found with fructose-1,6-P2, glycerate-2,3-P2, enolpyruvate-P, and acetyl CoA. From the amount of activity and the kinetic constants of the purified enzyme it seems likely that this enzyme is responsible for the glucose-1,6-P2 synthesis of brain.


Use of the sodium borohydride reduction technique to identify a gamma-glutamyl phosphate intermediary in the Escherichia coli glutamine synthetase reaction.

Incubation of unadenylylated Escherichia coli glutamine synthetase with ATP, L-[14C]glutamate and metal ion results in the formation of gamma-glutamyl-P which can under appropriate conditions be reduced by sodium borohydride. The acyl-P compound is formed catalytically as judged by the quantity of radioactive alpha-amino-delta-hydroxyvalerate produced compared to the concentration of enzyme subunits. Formation of the glutamyl-P compound occurs in the presence of magnesium or manganous ions, and the relation of this apparent lack of metal ion specificity with regard to the highly specific Mg2+-supported biosynthetic activity of the unadenylylated form is discussed.


The esterase activity of bovine carbonic anhydrase B above pH 9. Reversible and cooalent inhibition by acetozolamide.

The reversible complex between the metalloenzyme bovine carbonic anhydrase B and the sulfonamide inhibitor acetazolamide can be "frozen" irreversibly by the addition of a covalent bond between the methyl group of the inhibitor and the tau-nitrogen of histidine-64. In both cases the inhibited enzyme is inactive as an esterase toward p-nitrophenyl propionate at physiological pH but retains activity controlled by an ionization in the protein exhibiting a pK-a greater than 10. Similarly, both the covalently and reversibly inhibited enzymes in which the catalytically essential Zn(II) ion has been replaced with Co(II) display the same visible absorption spectrum which is invariant over the pH range from 5 to 12. The evidence therefore indicates that the position of the acetazolamide moiety in the active site is independent of both pH and the presence of the covalent bond to histidine-64. Moreover, when reversibly bound, this inhibitor has been shown to replace the water molecule (or hydroxide ion) known to occupy the fourth coordination position of the metal ion and frequently implicated in the catalytic mechanism of carbonic anhydrases. Thus, the activity exhibited by the inhibited enzymes and consequently the second rise observed in the pH rate profile of the native enzyme above pH 0 cannot reflect the ionization of such a water molecule in contrast to what has been postulated previously (Pocker, Y., and Storm, D. R. (1968) Biochemistry 7, 1202-1214). Displacement of the zinc-bound solvent molecule rather than the alkylation of histidine-64 is suggested, however, as the cause of the inactivation of the alkylated enzyme round neutrality. Taken together, the biphasic pH rate profile of native bovine carbonic anhydrase B as well as the activity retained by the alkylated enzyme above pH 9 are best described by a model in which two groups in the enzyme ionize independently, thereby raising the possibility that the high pH activity is controlled by an ionization outside the active site region of the enzyme. Above pH 9.5 the pK; for the reversible interaction between native carbonic anhydrase and acetazolamide falls off linearly with increasing pH. The slope of --1.56 suggests that, among other factors, more than one ionization is responsible for the descending limb of the pH-i-pH profile.


Properties of a reconstituted calcium pump.

1. The translocation of 45Ca2+ in vesicles reconstituted with purified Ca2+ ATPase of sarcoplasmic reticulum and phospholipids was dependent on ATP and varied greatly with the composition of the phospholipids. 2. In contrast to sarcoplasmic reticulum fragments, the reconstituted vesicles were impermeable to 14C-labeled oxalate, 3H- or 32P-labeled ATP, or 32P-i. There was no translocation of phosphate from gamma-labeled ATP during Ca2+ uptake. These results are inconsistent with some current formulations of the mechanism of pump action. 3. Reversal of the Ca2+ pump and generation of ATP and ADP and P-i was observed when vesicles loaded with Ca2+ were exposed to ethylene glycol bis(beta-aminoethyl ether)-N,N'-tetraacetic acid. 4. Experiments on the formation of phosphoenzyme with 32P-labeled ATP showed that most if not all functional ATPase molecules in the reconstituted vesicles were oriented in the same direction, as in the case of sarcoplasmic reticulum fragments.


Purification and properties of guanosine triphosphate cyclohydrolase II from Escherichia coli.

An enzyme that uses GTP as substrate for the formation in stoichiometric quantities of formate, inorganic pyrophosphate, and 2,5-diamino-6-hydroxy-4-(ribosylamino)pyrimidine-5'-phosphate has been purified 2200-fold from extracts of Escherichia coli B. This enzyme is named GTP cyclohydrolase II to distinguish it from a previously studied E. coli enzyme, named GTP cyclohydrolase (and called GTP cyclohydrolase I in this paper), that catalyzes the first of a series of enzymatic reactions leading to the biosynthesis of the pteridine portion of folic acid (Burg, A. W., and Brown, G. M. (1968) J. Biol. Chem. 243, 2349-2358). Some of the properties of GTP cyclohydrolase II are: (a) divalent cations are required for activity (Mg2+ is most effective); (b) its molecular weight, estimated by filtration on Sephadex G-200, is 44,000; (c) the K-m for GTP is 41 mum; (d) its pH optimum is 8.5; and (e) its activity is inhibited by inorganic pyrophosphate, one of the products of the reaction. Compounds not used as substrate are: GDP, GMP, guanosine, dGTP, ATP, ITP, and XTP. Properties a, b, c, and e (above), as well as the nature of the products, distinguish this enzyme from GTP cyclohydrolase I. Since GTP cyclohydrolase II apparently is not concerned with the biosynthesis of folic acid, the possible physiological role of this enzyme in the biosynthesis of riboflavin is considered in the light of the present investigations and the previously published work on riboflavin biosynthesis by other investigators.


Carbamyl phosphate: glucose phosphotransferase and glucose-6-phosphate phosphohydrolase of nuclear membrane. Interrelationships between membrane integrity, enzymic latency, and catalytic behavior.

The presence of carbamyl-phosphate:glucose phosphotransferase in liver nuclei of five species of mammals and birds is demonstrated. The activity is confined to nuclear membranes and is due exclusively to multifunctional glucose-6-phosphatase-phosphotransferase (D-glucose-6-phosphate phosphohydrolase; EC 3.1.3.9). The nuclear enzyme constitutes approximately 16 to 19 percent of total hepatic glucose-6-phosphatase-phosphotransferase. Carbamyl-phosphate:glucose phosphotransferase and glucose-6-P phosphohydrolase activities of membrane of chicken liver nuclei are shown to be catalytically identical with the maximally activated microsomal enzyme. A correspondence is seen in two-substrate kinetic double reciprocal plots, K-m or apparent K-m values for the various substrates, K-i values for the competitive inhibitors P-i and ATP, and pH-activity profiles. Comparative studies were carried out with various intact, disrupted, and detergent-dispersed membranous preparations by a combination of enzyme kinetic and electron microscopic techniques. It is concluded that (a) intimate interrelationships exists between catalytic behavior of this enzyme and morphological integrity of membranes of which the enzyme is a part; (b) activities of the enzyme of nuclear membrane appear quite available for physiological phosphorylative functions; and (c) interrelationships between membrane morphology and catalytic behavior of this membrane-bound enzyme may well be involved in the bioregulation of this complex, multifunctional enzyme system.


Genitourinary anomalies associated with Klippel-Feil syndrome.

Of thirty-nine patients with Klippel-Feil syndrome, twenty-five (64 per cent) had significant genitourinary-tract anomalies demonstrated by intravenous urogram and physical examination. The incidence of these anomalies in Feil's three types of the syndrome was essentially the same, unilateral renal agenesis being the most common. A routine intravenous urogram is indicated in patients with this syndrome.


Cell to substrate adhesion and spreading: inhibition by cationic anesthetics.

The plasma membrane is the postulated site of action of anesthetics on nerve or muscle. The drugs may be useful in the analysis of membrane phenomena in other cells. We show here that cationic anesthetics and tranquilizers inhibit cell adhesion and spreading, metabolically dependent processes that involve membrane motility and changes in cell shape. Adhesion was measured by layering 51Cr labeled Sarcoma I (Sa I) cells on glass coverslips for 20 minutes at 34 degrees C, rinsing and estimating the glass-associated radioactivity. Spreading was evaluated microscopically. Both cell adhesion to untreated glass and the Mn2+ dependent adhesion to serum-coated coverslips were inhibited by the drugs, in the following order of increasing activity: tetracaine, promethazine, cyclomethycaine, chlorpromazine and fluphenazine. Similar ranks of drug activity have been reported for nerve blocking, inhibition of cell fusion and inhibition of induced spreading of macrophages. Microscopic observations showed the drugs also inhibited MN2+ INDUCED SPREADING OF Sa I. Drug treated cells were rounded, refractile, devoid of cell processes or ruffles visible by light microscopy. The effects of the drugs on adhesion and spreading were reversible upon washing of the cells. We postulate that the inhibition of adhesion and spreading are a consequence of the inhibition of cell surface motility by the anesthetics.


Reversible alterations in the mitotic cycle of chick embryo cells in various states of growth regulation.

Chick embryo cells which have been kept overnight at pH 6.8 in the absence of serum multiply very slowly. Only a small fraction of cells is in the S period at any given time, and the rate of uptake of 2-deoxy-D-glucose is very low. Upon raising the pH to 7.4 and adding serum ("turn-on") the uptake of 2-deoxy-D-glucose increases immediately; the rate of DNA synthesis increases after a lag of about 4 hours, and represents an increase in the fraction of cells synthesizing DNA. the uptake of 2-deoxy-D-glucose is rapidly returned to its original low rate at any time by again lowering the pH and removing serum ("turn-off"). The synthesis of DNA in the culture remains constant or continues to rise at a markedly reduced rate following the same treatment. Lowering pH or removing serum independently of each other is less efficient at inhibiting the increase in DNA synthesis than the combined treatment but each accomplishes a similar result. Cultures which have been "turned-off" during the early stages of the rapid increase in DNA synthesis, resume their prior rate of increase immediately if "turned-on" again within 2.5 hours. If the cultures have been "turned-off" for 5.5 hours before restoring the "turn-on", there is a 2 hour delay before they resume an increased rate of DNA synthesis. The results until shortly before, or at the time of the onset of the S period. Up to 96% of the cells in post-confluent cultures growing in conventional medium become labeled upon continuous, prolonged exposure to 3H-thymidine. Seventy-eight percent of the cells in serum-deprived cultures growing at a very low rate become labeled. These and other considerations suggest that the inhibition of cell multiplication by high population density or serum deprivation is caused by a lengthening of the time cells remain in the prereplicative G1 period rather than by shifting cells into a qualitatively distinct G0 period. There may, however, be a period common to all cells regardless of growth rate, in which cells are not progressing toward the S period. The length of this variable period would then determine the growth rate of a population of cells.


Analysis of the induction and deinduction of tyrosine aminotransferase in enucleated HTC cells.

Anucleate HTC cells have been used to determine the importance of the nucleus in the regulation of the intracellular levels of tyrosine aminotransferase (TAT) in hepatoma tissue culture (HTC) cells. In the absence of the nucleus, neither the induction of the enzyme by dexamethasone nor its deinduction upon removal of the hormone occurs. Degradation of the enzyme takes place when protein synthesis is inhibited in anucleates by cycloheximide. Therefore, the maintenance of induced levels of enzyme activity after dexamethasone withdrawal from pre-induced anucleates suggest that the nucleus is required for the inactivation of the TAT mRNA.


Response of the kallikrein-kinin and renin-angiotensin systems to saline infusion and upright posture.

The possibility that bradykinin, a potent vasodilator, might be a physiological antagonist of the renin-angiotensin system was investigated. 11 norman subjects, ranging in age from 21 to 33 yr were studied. Seven of the subjects were given a 10 meq sodium, 100 meq potassium, 2500 ml isocaloric diet. After metabolic balance was achieved, they were infused with either 1 liter of 5 per cent glucose over 2 h or 2 liters of 0.9 per cent saline over 4 h. During the infusions, plasma renin activity (PRA), angiotensin II (A II), prekallikrein, bradykinin, and aldosterone levels were frequently determined. Plasma prekallikrein and kallikrein inhibitor did not change during the infusion of either glucose or saline. In subjects receiving saline, plasma bradykinin fell from 3.9 plus or minus 1.5 (SEM) ng/ml at 0 min to 0.93 plus or minus 0.2 at 30 min and 0.95 plus or minus 0.3 at 120 min. These changes paralleled the decrease in PRA over the same period (7.9 plus or minus 1.3 ng/ml/h to 5.6 plus or minus 0.8 at 30 min and 3.5 plus or minus 0.7 at 120 min). Similarly, A II fell from 113 plus or minus 12 pg/ml to 62 plus or minus 10 and 48 plus or minus 5, respectively, at 30 and 120 min. In contrast, the control group infused with glucose showed no change in bradykinin, A II, or PRA. Another four subjects were given a constant 200 meq sodium/100 meq potassium isocaloric diet. After metabolic balance was achieved, they were kept supine and fasting overnight. At 9 a.m. they assumed an upright position and began walking a fixed distance (200 ft) at a normal rate (3-4 ft/s). Plasma prekallikrein and kallikrein inhibitor did not change during the posture study. The plasma bradykinin rose from a base line of 0.54 plus or minus 0.01 (SEM) ng/ml to 0.96 plus or minus 0.13 at 20 min. 0.77 plus or minus 0.18 at 60 min, and 0.96 plus or minus 0.07 at 120 min. These changes parallel the increase in PRA over the same period (1.65 plus or minus 3.3 ng/ml/h to 3.6 plus or minus 0.85 at 20 min, 5.3 plus or minus 0.9 at 60 min, and 5.35 plus or minus 0.55 at 120 min). Likewise, the A II rose from 32.5 plus or minus 1.82 pg/ml to 50.8 plus or minus 3.6 at 20 min, 54.3 plus or minus 3.2 at 60 min, and 61.3 plus or minus 5.9 at 120 min. Thus, in sodium-depleted individuals, saline infusion produces a rapid fall of plasma bradykinin at a rate similar to that observed for a II and PRA. Conversely, in sodium-loaded individuals, assumption of upright posture leads to a parallel rise in A II, TPRA, and bradykinin. These studies indicate that there is a close correlation of bradykinin levels with renin activity and angiotensin II, in both acute sodium loading and assumption of upright posture, suggesting that these two systems may be physiologically interrelated.


NADPH oxidase deficiency in X-linked chronic granulomatous disease.

We measured the cyanide-insensitive pyridine nucleotide oxidase activity of fractionated resting and phagocytic neutrophils from 11 normal donors, 1 patient with hereditary deficiency of myeloperoxidase, and 7 patients with X-linked chronic granulomatous disease (CGD). When measured under optimal conditions (at pH 5.5 and in the presence of 0.5 mM Mn++), NADPH oxidase activity increased fourfold with phagocytosis and was six-fold higher than with NADH. Phagocytic neutrophils from patients with CGD were markedly deficient in NADPH oxidase activity.


An isotopic assay for NADPH oxidase activity and some characteristics of the enzyme from human polymorphonuclear leukocytes.

An isotopic assay for NADPH ixodase that measures the amount of NADP formed by the 6-phosphogluconate dehydrogenase reaction has been developed. Under appropriate conditions, the amount of NADP present is directly proportional to the amount of 14CO2 released from [1-14C]6-phosphogluconic acid. Because this assay employs radioisotopes, it is far more sensitive than conventional assays for the enzyme. The human granule NADPH oxidase, as measured by this assay, is active in the presence of CN minus, is stimulated by Mn-2+, and has a pth optimum of 5.5. Granules isolated from cells that have been allowed to ingest zymosan consistently exhibited more enzyme activity than did granules isolated from either resting cells or cells challenged with zymosan that was not preopsonized. This effect was observed over a wide range of substrate concentrations and could not be explained by differences in protein concentrations between the various samples. If whole homogenates are used in place of isolated granules, the enzyme activity can be observed only with a homogenate of phagocytizing cells and even then only at a high concentration of NADPH. This suggests that an inhibitor of the enzyme might be present within the cell. One patient with chronic granulomatous disease was studied. There was no difference in tnadph oxidase activity of the patients' cells when granules from resting and phagocytizing cells were compared. In contrast, the enzyme activity in granules from two control patients doubled upon phagocytosis. These results are consistent with a role for NADPH oxidase in the initiation of the respiratory burst accompanying phagocytosis by human neutrophils.


Changes in the electroencephalogram in acute uremia. Effects of parathyroid hormone and brain electrolytes.

Studies were carried out in order to evaluate the effects of changes in brain calcium and the influence of parathyroidectomy and administration of parathyroid extract on the electroencephalogram (EEG) of normal and uremic dogs. Manual analysis of frequency and power distribution of the EEG in uremic dogs revealed a significant increase in both the percentage distribution and the area or power occupied by frequencies below 5 Hz. In addition, high amplitude bursts of delta activity were apparent in the uremic dog. These changes were largely prevented by parathyroidectomy before the induction of uremia, but the administration of parathyroid extract to either normal dogs, or to previously parathyroidectomized uremic dogs, induced EEG changes similar to those noted in uremic animals with intact parathyroid glands. In all groups of animals which showed EEG changes, brain content of calcium was significantly higher than in either normal dogs or previously parathyroidectomized uremic dogs. Changes in arterial pH and bicarbonate, or in the concentrations of Na+, K+, urea, or creatinine in plasma or cerebrospinal fluid were similar in uremic animals with intact parthyroid glands and in previously parathyroidectomized uremia dogs. The results indicate that the EEG changes found in dogs with acute renal failure require the presence of excess parathyroid hormone in blood, and they may be related to the observed changes in brain content of calcium.


Ethacrynic acid. Effects on the cochlear potentials in normal and high blood oxygen.

The effect of ethacrynic acid (EA) at different blood O(2) saturations on cochlear potentials of guinea pigs was investigated. All 18 young healthy guinea pigs received 50 mg/kg/h of EA intravenously and were divided into three groups: first group, normal (90.00+/-6.30-86.17+/-4.83 mm Hg); second group, lower Po(2) (78.00+/-4.74-70.00+/-4.42 mm Hg); and third group, high Po(2) (174.40+/-13.41-179.00+/-26.15 mm Hg). The partial pressure of oxygen (Po(2)), the partial pressure of carbon dioxide (Pco(2)), and the pH of the blood were measured before EA administration and at the end of the experiment (3 h later) by drawing blood samples from the contralateral carotid artery. Cochlear potentials-endocochlear potential (EP), cochlear microphonics (CM), and action potentials (AP)-were recorded by standard methods from the first turn of the cochlea. Experimental data seem to indicate that elevation of the Po(2) to 174-179 mm Hg during relatively high doses of EA treatment prevents the declines in cochlear potentials which were observed in the first and second groups (normal and lower Po(2)), and preserves active ion transport which is responsible for the generation of cochlear potentials. These data suggest a means by which to reduce the ototoxic effect of EA and possibly indicates a method of treatment for hearing loss which developed after the administration of EA.


Red cell NADP+ and NADPH in glucose-6-phosphate dehydrogenase deficiency.

Extraction in the presence of sodium hydroxide and cysteine allows estimates of NADPH and total NADP in human red cells without the erroneously high values of NADP+ obtained with earlier methods. An application of this technique to G6PD-deficient cells reveals that most of the nucleotide is in the oxidized form. In contrast, normal red cells have nearly all of the nucleotide in the reudced form. In addition to providing information concerning the intracellular regulation of the hexose monophosphate shunt, these findings support the concept that G6PD deficiency is a product-deficiency disorder.


Energy dependence of urinary bicarbonate secretion in turtle bladder.

Addition of HCO3- to the serosal side (S) of the isolated turtle bladder results in a HCO3- flow from S to the mucosal side (M) which markedly reduces the net rate of acid secretion. To characterize the driving forces for this downhill HCO3- flow, the effects of metabolic inhibitors and substrates were examined. In short-circuited bladders with the M pH lowered  to the point of zero net H+ secretion, the rate of HCO3- entry into M in response to a 20-mM HCO3- gradient was measured by pH stat titration. Deoxygenation reduced the HCO3- flux from 1.24 plus or minus 0.1 mum/h/8 cm2 (SEM) to 0.50 plus or minus 0.1 muM/h with glucose (2 times 10-3 M) AND FROM 1.32 PLUS OR MINUS TO 0.47 PLUS OR MINUS 0.1 MUM/h without glucose. A similar reduction (61 per cent) was observed in the presence of 1 per cent C92. Dinitrophenol (10-4 M), cyanide (10-3 M), and deoxyglucose (10-2 M) inhibited the HCO3- flux by 39 per cent, 37 per cent, and 38 per cent, respectively. The combination of any of these inhibitors with N2 caused the same inhibition as N2 alone. In bladders depleted of substrate, pyruvate (5 times 10-3 M) increased the HCO3- flux from 0.36 plus or minus 0.05 to 0.58 plus or minus 0.01 muM/h (P smaller than 0.005); the increment was abolished by deoxygenation. The results indicate that the bulk of the downhill HCO3- flow in this system is dependent on metabolic energy derived primarily from oxidative sources, and that this energy-dependent flow approximates the electroneutral component of HCO3- secretion that is coupled to Cl- absorption.


Guanosine 3',5'-monophosphate and adenosine 3',5'-monophosphate content of human umbilical artery.

Human umbilical arteries are unique vessels in that they close quickly and completely at birth. It has been suggested that cyclic uanosine 3',5'-monophosphate (cAMP) in relaxation. This hypothesis has been evaluated in term gestational human umbilical artery segments incubated at 37 degrees C and in room air. (a) The basal cGMP content (1 pmol/mg protein) of artery segments incubated in room air was almost twice that of cAMP. (b) Bradykinin, histamine, serotonin, acetylcholine, and K+ ion, which cause umbilical artery constriction, can increase the cGMP content of the artery segments within 30 s of exposure without altering the cAMP content. (c) Prostaglandin E1, but not isoproterenol, caused accumulation of cAMP which is consistent with reports that umbilical arteries lack functional beta-receptors and that only prostaglandin E1 can bring about relaxation of umbilical arteries. (d) 1 muM atropine blocked the effect of 100 muM acetylcholine on cGMP content without altering the responses to histamine, bradykinin, serotonin, or K+ ion. (e) Pyrilamine (an H1 antagonist), but not metiamide (an H2 antagonist), blocked the effect of histamine on cGMP from which it is inferred that histamine causes accumulation of cGMP in umbilical artery via its interaction with H1 receptors. The results are consistent with the view that metabolism of the two cyclic nucleotides is independently controlled in the human umbilical artery and that cGMP is involved in contraction of the artery at birth.


In vivo myocardial cell pH in the dog. Response to ischemia and infusion of alkali.

Myocardial cell pH has been measured with 5,5-dimethyl-2,4-oxazolidinedione (DMO) in intact anesthetized dogs by a transient indicator dilution technique. Bolus injections of labeled DMO, vascular, extracellular, and water indicators were made into the anterior descending coronary artery, and blood samples were collected from the great cardiac vein. The steady-state distribution of DMO between cells and plasma was calculated from the indicator mean transit times, and the plasma pH. Myocardial cell pH was determined from the distribution value and plasma pH. Normal myocardial cell pH averaged 6.94. Changes in myocardial cell pH after infusions of acid or alkali. Myocardial ischemia induced by inflation of a coronary artery balloon resulted in progressive decreases in cellular pH to average values of 6.83 within the initial 15 min and to 6.59 within the interval between 20 and 70 min. Infusions of Na2CO3 tended to diminish intracellular acidosis although these infusions had little effect on the difference in pH between the myocardial cell and extracellular fluid.


Pharyngeal lipase and digestion of dietary triglyceride in man.

Lipolytic activity was studied in esophageal and gastric aspirates obtained with a nasogastric tube from 14 healthy adult subjects. Samples were collected from esophagus, first at 30-35 cm and then at 40-45 cm from the nose, as the subject, after drinking 15-30 ml of a cream-milk mixture, swallowed small amounts of water. The samples from stomach were taken last and usually contained a small amount of cream-milk mixture. Lipolytic activity was assayed using chylomicron, milk, and corn oil triglyceride as substrate. Esophageal and gastric samples both contained lipolytic activity which hydrolyzed long-chain triglyceride to diglyceride, monoglyceride, and FFTA, had a pH optimum of 5.4, and was not affected by either had a pH optimum of 5.4, and was not affected by either 0.5 M NaCl or 4 mM sodium taurodexycholate. The activity, expressed as nanomoles of chylomicron triglyceride hydrolyzed per milliter per minute, ranged from 0 to 145 in upper esophageal, 5 to 303 in lower esophageal, and 50 to 357 in gastric samples. Only a trace of lipolytic activity was found at pH 5.4 in saliva collected from the parotid, submandibular, and sublingual glands, thus excluding those tissues as a source of the activity found in esophageal and gastric aspirates. The findings suggest that in man glands in or near the pharynx secrete a lipase that acts in the stomach to hydrolyze long-chain triglyceride to partial glycerides and FFA. It is proposed this reaction is the first step in the digestion of dietary fat and that the amphiphilic lipids formed by lipolysis facilitate the emulsification of triglyceride in the stomach.


The influence of extraneous factors on Coulter S measurement of the mean corpuscular volume.

Alteration in the temperature of the Isoton diluent in a Coulter model S counter over a range of possible laboratory working temperatures produced a change in the mean corpuscular volume using EDTA and dipotassium acid citrate dextrose blood and a commercial control, 4C. Contamination of the Isoton diluent with hypochlorite solution, used to clean the Coulter model S, may increase the mean corpuscular volume of EDTA blood without altering other measured parameters. Attention to both these points is required in assessing quality and calibrating the machine.


The Compu-pet 100: a versatile dispenser-diluter for the mechanization of microbiological techniques.

The Compu-pet 100 diluter/dispenser is a versatile and accurate instrument which can be employed for a wide variety of microbiological tests and techniques. Time in performing tests is often more than halved when the diluter is used. Large numbers of tests can be performed with minimal fatigue.


Neocortical and hippocampal activation relation to behavior: effects of atropine, eserine, phenothiazines, and amphetamine.

Evidence is presented to suggest that the hippocampus receives 2 nonspecific inputs from the brainstem, each capable of producing rhythmical slow activity. The neocortex appears to receive 2 similar inputs, each capable of producing low voltage fast activity. One input to both hippocampus and neocortex is blocked by atropine and stimulated by eserine, and is essentially unrelated to concurrent motor activity. A second input to both hippocampus and neocortex is resistant to atropine, is depressed by phenothiazines, and is activated by d-amphetamine. Activity in this system is closely related to concurrent "voluntary" movement.


Role of the biogenic amines in the reversal of cycloheximide-induced amnesia.

Amnesia was induced by pretraining injections of cycloheximide (CYC) in a food motivated discrimination reversal task. Magnitude of amnesia varied as a function of the amount of training on both the initial discrimination and the reversal and also as a function of the length of intertrial interval used on both the reversal and the test. Memory spontaneously recovered 48 hr. following reversal training. Recovery from amnesia was induced by pretesting injections of d-amphetamine and 2 monoamine oxidase inhibitors, pargyline and catron. This enhanced performance was a true recovery of the memory and  not a result of enhanced learning or increased arousal. Depletion of catecholamines by alpha-methyl-para-tyrosine, a tyrosine hydroxylase inhibitor, and diethyldithiocarbamate, a dopamine beta hydroxylase inhbitor, resulted in an amnesia quantitatively and qualitatively similar to amnesia induced by CYC. These data support the hypothesis that CYC-induced amnesia is mediated via central catecholamines.


Monoaminergic mediation of female sexual behavior.

Lordotic behavior was facilitated in estrogen-primed female rats by direct application of progesterone or serotonergic or beta-adrenergic receptor blockers to specific telencephalic, anterior hypothalamic-medial preoptic, or posterior hypothalamic sites. Blockade of the alpha-adrenergic system was ineffective in facilitating lordosis, as was the application of the active drugs to control sites in the thalamus or basal ganglia. Female soliciting behavior was not evoked by any of the treatments. It is concluded that the lordotic behavior component of the female rat's estrous behavior pattern is inhibited by a specific central monoaminergic system that also responds to progesterone. Soliciting behavior appears to be mediated by systems that are anatomically, and possibly neurochemically, separable from those regulating lordosis.


Female lordotic behavior mediated by monoamines in male rats.

Direct application of serotonergic of beta-adrenergic receptor blockers to anterior or posterior areas of the hypothalamus induced lordosis in intact estrogen-primed male rats. Such treatment with an alpha-adrenergic blocker or systemic administration of progesterone failed to increase lordosis. Centrally elicited lordosis did not occur without estrogen priming. Anatomical and neurochemical similarity may exist in the brain mechanism mediating lordotic behavior in male and female adult rats.


The nature of the stimulation of the growth of Streptococcus lactis by yeast extract.

Yeast extract was fractionated on Sephadex G-25 into 7 fractions. The fraction most stimulatory to the growth of Streptococcus lactis C10 contained over 70% of the amino N present in yeast extract and consisted of a wide variety of free amino acids and a small amount of peptide material. Examination of possible replacement factors for this fraction revealed that the amino -acid material present was largely responsible for the stimulation of Str. lactis C10. Purine and pyrimidine bases and inorganic constituents also contributed to the stimulation. In addition, yeast extract contained a component which decomposed H2O2, a metabolite which accumulates in the growth medium under aerobic conditions and inhibits growth. The nature of the stimulation was studied by isolating slow and fast acid-producing colonies of Str. lactis C10. It appeared that yeast extract and other amino-acid supplements prevented an observed inhibition of the growth of the slow variants below pH 6.0, apparently by satisfying a nutritional deficiency caused by a drop in pH.


Pptidase activities in group N streptococci.

Several peptidase activites in the 3 species of Group N streptococci were partly separated by gel filtration on Sephadex G-200. The peptidases identified were a general aminopeptidase of wide specificity, a tripeptidase, a proline iminopeptidase (prolyl-beta-napthylamidase), a proline iminodipeptidase and an aminopeptidase-P. The effects of temperature and pH on the stability of the enzyme activities, and the influence of the type of N source used in the growth medium on the elution pattern of the enzymes were examined.


Preparation and properties of beta-casein from buffalo's milk.

Beta-casein from individual buffalo's milk was found to be homogeneous by starch-gel electrophoresis. Beta-Casein was separated from buffalo's milk by the method of Warner (1944) and purified by DEAE-cellulose chromatography. Buffalo beta-casein possesses identical end-groups to those of cow beta-casein; namely N-terminal arginine and assuming a single polypeptide chain a possible C-terminal sequence of Ile-Ile-Val. However, the amino-acid composition and the trypic peptide patterns of the 2 proteins are not the same.


The effect of aging and acute illness on delayed hypersensitivity.

A group of standard antigens (monilia, mosquito, mumps, purified protein derivative [PPD], staphylococcus toxoid, streptokinase-streptodornase [sk-sd] and trichophyton) were applied to 321 normal volunteers, 60 patients admitted to a medical ward, and 41 patients in an intensive care unit (ICU) to evaluate their delayed hypersensitivity skin test response. Eighty-eight per cent of the normal subjects reacted with at least 5 mm. induration to one or more skin tests without any decrease in reactivity occurring with increasing age. In the hospitalized group, only 62 per cent of the ward and 63 per cent of the ICU patients reacted to the same skin tests. All (8/8) anergic ICU patients tested after discharge regained normal reactivity. Acute illness but not increasing age will impair the delayed hypersensitivity skin test response.


Inactivation of slow reacting substance of anaphylaxis by human eosinophil arylsulfatase.

Arylsulfatase preferentially present in the human eosinophil as compared to other leukocytes was isolated by sequential gel filtration and cation exchange chromatography. The apparent molecular weight of 60,000, the preferential cleavage of 4-nitrocatechol sulfate (PNCS) over p-acetyl-benzenesulfonic acid (PABS), inhibition by phosphate ions and pH optimum of 5.7 are characteristics of a type II B arylsulfatase. Eosinophil arylsulfatase inactivated purified human slow reacting substance of anaphylaxis (SRS-A) in a time-dependent reaction with the rate dependent upon the enzyme/substrate ratio. That SRS-A inactivation was the result of intrinsic arylsulfatase activity was indicated by association of PNCS cleavage and SRS-A inactivating activity during chromatography, the similar pH optimum for cleavage of both substrates and the capacity of SRS-A to inhibit PNCS cleavage by arylsulfatase. The finding that eosinophil arylsulfatase inactivates SRS-A suggests that eosinophil ingress into the site of an immediate hypersensitivity reaction in response to ECF-A could represent a regulatory function.


Metabolism of glucose and fatty acid by leukocytes from patients with endogenous hypertriglyceridemia.

The metabolic abnormalities responsible for endogenous hypertriglyceridemia have not been defined. Some in vivo studies have suggested that excessive triglyceride production is the cause of this defect. In an attempt to obtain direct evidence concerning this mechanism, we have compared in vitro the metabolism of radioactive glucose and palmitate by leukocytes from patients with endogenous hypertriglyceridemia and normal subjects. Leukocytes from the patients incorporated 9.82 plus or minus 1.7 (S.E.M.) nanomoles of glucose into cellular lipid per 10-8 cells per hour. When the cell lipid extract was sugjected to mild alkaline hydrolysis, 92 per cent of the glucose radioactivity was recovered in the glycerol backbone of the lipid esters. Comparison of specific yields of CO2 from glucose labeled in the 1- or 6- position revealed that 0.53 plus or minus 0.02 per cent was metabolized via the pentose cycle. The leukocytes from hypertriglyceridemic persons incorproated 140 plus or minus 6.9 nanomoles of [1-14C]-palmitate per 10-8 cells per hour. Eighty-four per cent of the radioactivity was in triglycerides and 14 per cent in phospholipids. The major phospholipid into which palmitate was incorporated was phosphatidyl choline. The leukocytes oxidized palmitate at a rate of 2.88 plus or minus 0.23 nanomoles per 10-8 cells per hour. There were no differences in any of the above values between leukocytes from hypertriglyceridemic patients and normal subjects. Likewise, there was no correlation between the plasma triglyceride concentration and glucose or palmitate incorporation into triglycerides. To the extent that leukocytes reflect systemic metabolic processes, these data provide no support for the interpretation that the mechanism of the plasma triglyceride elevation is excessive biosynthesis.


The sorbitol-oxidizing enzyme of red blood cells.

The sorbitol-oxidizing enzyme of human erythrocytes has been partially purified and characterized. Its kinetic characteristics were found to be similar to the sorbitol dehydrogenase of sheep liver and the DPN-xyluloxe dehydrogenase which have been isolated from sheep liver and guinea pig liver, respectively. Although this enzyme is able to oxidize sorbitol relatively efficiently, it has very little activity toward dulcitol, and therefore cannot account for the high K-m pathway of methemoglobin reduction by galactose.


Erythrocyte delta-aminolevulinic acid dehydratase activity as a measure of lead exposure.

The measurement of erythrocyte delta-aminolevulinic acid dehydratase (ALA-D) activity has been found to be a useful guide to lead exposure for screening purposes. The pH of the test should be well-controlled for adequate clinical interpretation. False-normal tests are rare even with iron-deficiency anemia or macrocytosis. Recent transfusion and recent bleeding or hemolysis with reticulocytosis tend to invalidate the test. ALA-D activity above 600 International Units effectively excludes significant lead exposure and eliminates the need for further testing. ALA-D activity is maximally depressed at a red cell lead concentration of 200 mug per 100 ml. When activity is moderately depressed, interpretation is difficult and the urinary excretion of lead after the administration of a chelating agent is usually required to differentiate the degree and significance of lead exposure.


Characterization of erythrocyte membrane-associated enzymes (glyceraldehyde-3-phosphate dehydrogenase and phosphoglyceric kinase).

The purpose of our study was to determine why several of the glycolytic enzymes of the erythrocyte have the propensity for adhering to erythrocyte membranes while others do not. Two of these membrane-associated enzymes, glyceraldehyde phosphate dehydrogenase (GAPD) and phosphoglyceric kinase (PGK), have been shown to have controlling functions on intra-erythrocytic glycolysis and sodium-potassium transport. In order to test the hypothesis that the membrane-associated fraction of these enzymes consisted of isozymes with increased capacity to become membrane associated, the enzymes from cytosol or membrane sources were partially purified and characterized. Determination of molecular weight by gel filtration chromatography, measurement of certain kinetic parameters, and the curves relating to pH optimal activity indicated that there were no measurable differences between membrane and cytosol PGK and membrane and cytosol GAPD. It was possible to raise inhibitory antibodies to GAPD in certain species of mice, and these antibodies did not distinguish between GAPD isolated from either membrane or cytosol sources. GAPD was firmly bound to membranes and the membrane-associated fraction counted for approximately 60 per cent of total erythrocytic enzyme. Membrane-associated PGK was only loosely adherent, and accounted for only 1 per cent of the total erythrocytic enzyme. The reasons for membrane association have yet to be determined, but these inward facing membrane-associated enzymes appear to function by carrying the organization of the membrane into the cell interior.


Factors affecting the evolution of factor XIa during blood coagulation.

Certain conditions affecting the evolution of factor XI-a activity during blood coagulation have been examined. Earlier data had indicated that calcium ion was not required for the conversion of factor XI to its activated form, but very little XI-a could be isolated from citrated or EDTA plasma, whether or not the plasma had been clotted by recalcification. Conversely, factor XI-a activity was identified in resin-decalcified plasma, again with or without recalcification. This confirmed that calcium is not required for the evolution of factor XI-a. This observation also permitted us to perform experiments which indicate that there is no obligatory participation of cellular elements in the evolution of factor XI-a during blood coagulation.


A comparative chemical and serological study of the full and empty particles of foot-and mouth disease virus.

The chemical and serological properties of the full, naturally occurring empty and artificially produced empty particles of foot-and-mouth disease virus, serotype A(subtype 10, strain 16) have been studies. The full 146S particles comprised the virus RNA, three polypeptides (VP1 to VP3) mol. wt. about 30 X 10-3, one polypeptide (VP4) mol. wt. about 13-5 X 10-3, and a small amount of a polypeptide (VPo) mol. wt. about 43 X 10-3. The naturally occurring 75S empty particles contained no RNA and much less VP1 and VP4 than were found in the fall particles. However they contained a much greater proportion of VPo than the full particles. Dialysis of purified full particles against tris-EDTA, pH 7-6, produced artificial 75S empty particles which contained only a small amount of RNA and no VP4; otherwise the polypeptide composition was similar to that of the full particles. Immunological and serological tests showed that the full particles were antigenically similar to the naturally occurring empty particles but distinct from the artificial empty particles. The latter particles, however, had serological properties similar to those of the 12S protein subunit of the virus. Both the full and naturally occurring empty particles attached efficiently to susceptible cells, whereas the artificial empty particles attached only to a limited extent. The results are related to the function of the individual polypeptides of the virus particle and compared with published work on other picornaviruses.


The phosphorylation of sendai virus proteins by a virus particle-associated protein kinase.

Highly purified Sendai virus contained a protein kinase activity which atatlysed the phosphorylation of endogenous polypeptides or exogenous protamine sulphate. The virus contained very low levels of phosphoprotein phosphatase activity. Polyacrylamide gel analysis of the reaction product indicated that the phosphorylation was specific for certain polypeptides and varied according to whether the virus was grown in eggs or in tissue culture. This variation was partially associated with the difference in the polypeptide pattern that occurred when the virus was grown in eggs or in tissue culture. Characterization of these phosphoproteins demonstrated that the phosphate was incorporated predominantly in a phosphoester linkage with theonine residues. Using a detergent and high salt solubilization procedure, the protein kinase activity was found associated within glycoprotein free virus particles but not with the nucleocapsid-associated polypeptides. In vivo phosphorylation occurred when Sendai virus was grown in eggs or in tissue culture with [32P] and the phosphorylated polypeptides were similar to those of the protein kinase reaction product. Phosphorylation could also be detected in the infected cell and could occur once the virus particle polypeptides were being synthesized. The non-structural polypeptides were not phosphorylated.


Attitudes toward death: a survey of nursing students.

The literature devoted to the topics of death and care of the dying is expanding rapidly. As nurses are inevitably involved in terminal illness, death and grief, their attitudes toward death and factors which affect these attitudes, are worthy of study. This report describes the results of a continuing two-year survey of one class of students in a baccalaureate nursing program. A questionnaire was used at the beginning and end of one academic year. Data were obtained regarding background experiences with death, involvement in the care of dying patients, and common ideas, concerns and feelings about death. Data from the second testing also included perceived changes in "positive" and "negative" attitudes toward death, and the relative effect on attitudes of various factors during the year. Suggestions are offered for curriculum development and research in the challenging area of death education in nursing.


Translocation of the umbilical artery to the lower abdomen: an adjunct to the postoperative monitoring of arterial blood gases in major abdominal wall defects.

A technique is presented for preserving the umbilical arteries for postoperative monitoring of arterial blood gases after repair of giant omphaloceles and gastroschisis. Translocation of the arteries to the lower abdominal wall removes them from the operative field and allows cannulation and long-term blood sampling. Complications have been minimal and the improved application of modern pulmonary support mechanisms has materially contributed to better patient management. The technique is a simple one and may prove applicable to other major newborn medical and surgical diseases involving respiratory insufficiency.


Improvement of solubility and dissolution rate of poorly water-soluble salicylic acid by a spray-drying technique.

Spray drying techniques have been applied to improve the solubility and dissolution rate of poorly water-soluble salicylic acid. Spray drying of the acid dispersed in acacia solutions resulted in as much as a 50% improvement in the solubility of the product. Solubility improvement was closely related not only to the concentration of acacia but also the amount of amorphous material in the spray-dried products. The heat of solution was inversely related to these parameters. The dissolution rate of spray-dried product was almost instantaneous being about 60 times faster than that of the original powder. A great improvement in the wettability of the spray-dried material seemed to be mainly responsible for the increase of dissolution rate.


A crystallographic and theoretical study of the conformation of DOET and its significance for the hallucinogenic amphetamines.

The crystal and molecular structure of 2, 5-dimethoxy-4-ethyl-alpha-methyl-phenylethylamine (DOET) has been determined by X-ray crystallography and the conformation of the side chain has been examined theoretically by the use of a potential energy calculation. There is no indication in the solid state of any intra or inter-molecular hydrogen bonding. The isopropylamine side chain is staggered to the plane of the benzene ring. The alpha-methyl group is fully extended (antiplanar) whereas the amino group is orientated back towards the ring (synclinal). The potential energy calculations show the existence of 6 minima, one of which corresponds to the crystal position. The calculations also show that the energy differences between the various minima of the side chain are very small. The relationship of these data to the conformations of the hallucinogens, mescaline and 2, 4, 5,-trimethoxyamphetamine is discussed.


Some pharmacological actions of 2,5-dimethoxy-4-ethylamphetamine (DOET) in rats and mice.

DOET, like DOM, exhibited pressor action in rats. This increase of blood pressure was blocked by pretreatment with cinanserin. DOET at high doses decreased the spontaneous locomotor activity of mice at the first hour but increased the activity at the second hour; a low dose was less effective. DOET also increased the rectal temperature of rats and this hyperthermic action was suppressed by pretreating the animals with cinanserin or methysergide. These actions of DOET were compared with those of DOM.


Comparison of the beta-adrenoceptors in the myocardium and coronary vasculature of the kitten heart.

The relative potencies of (minus) noradrenaline, (minus)-adrenaline, (plus or minus)-isoprenaline and (plus or minus)-salbutamol have been assessed for their positive inotropic and chronotropic actions on kitten isolated atria. These relative potencies have been compared with those obtained for the relative coronary dilator potencies in two preparations. These were intact hearts perfused by Langendorff's method and isolated perfused coronary arteries from the kitten. The relative molar potencies for inotropic effects were noradrenaline 1: adrenaline1: iso-prenaline 7: salbutamol 0.6. The observed ratios for chronotropic effects were not significantly different from those for inotropic effects. The relative potencies of noradrenaline, adrenaline and isoprenaline as myocardial stimulants were similar to their relative potencies as coronary dilators in the intact heart. Similar relative potencies for noradrenaline and isoprenaline were also found in the isolated coronary artery but adrenaline was only one third as active as noradrenaline on this preparation. In both the intact heart and in the isolated coronary artery salbutamol was about one tenth as active as noradrenaline. It was therefore less active relative to noradrenaline as a coronary dilator than as a myocardial stimulant. In spite of these differences in relative potency ratios for myocardial and coronary dilator effects, similar dissociation constants (Kb values) for practolol against isoprenaline were found in driven atria and in isolated coronary arteries. Myocardial and coronary vascular beta-adrenoceptors thus can both be placed in the general classification of beta1-adrenoceptors.


The effect of 5-hydroxytryptophan on food intake and on the anorexic action of amphetamine and fenfluramine.

5-Hydroxytryptophan (5-HTP), amphetamine and fenfluramine suppressed food intake in normal rats and in aminals with lesions of the lateral hypothalamus. The anorexic effect of amphetamine was reduced in lesioned animals compared with controls while the effect of 5-HTP like that of fenfluramine was increased. When administered in conjunction with anorexic drugs, 5-HTP markedly potentiated the anorexic effect of amphetamine in both control and lesioned animals. However, 5-HTP potentiated fenfluramine anorexia only in lesioned rats. These findings provide further evidence for the role of 5-hydroxytryptamine (5-HT,in the anorexic effect of fenfluramine, and suggest that a 5-HT mechanism, inhibitory for feeding, produces particulary severe suppression of food intake in rats with lateral hypothalamic lesions.


The role of p-hydroxylation of amphetamine in its peripheral mode of action.

Desipramine completely abolished the blood pressure response to (+) amphetamine in the rat, whereas the tricyclic antidepressant iprindole did not reduce this response. Since both tricyclic drugs inhibit the aromatic hydroxylation of amphetamine, these studies do not support the hypothesis that hydroxylated metabolites of amphetamine, p-hydroxyamphetamine and p-hydroxynorephedrine are essential for the peripheral sympathomimetic effects of amphetamine.


Construction and implantation of a cannula system for repeated injections into localized regions of the rat brain.

The construction and method of implantation of a brain cannula for chronic use in rats is described. All the components of the cannula system are easily made from stainless steel tubing and no machined parts are required. The use of a metal key eliminates the need for tapped holes during implantation. Although little time is required to prepare each animal the system remains firmly fixed to the skull and allows injections to be made accurately into selected brain areas.


An automated sedimentation balance.

A method of size grading powders using an automated sedimentation balance is described. The results are recorded as percentages of the total powder present in 1 or 5 mum (intervals) of particle size, after they have been multiplied by a simple correction factor. Analyses of calcium carbonate powder using this balance and the Gallenkamp balance have been compared.


Nerve stimulation-meditated overflow of norepinephrine and dopamine-beta-hydroxylase. III. Effects of norepinephrine depletion on the alpha presynaptic regulation of release.

A frequency-dependent increase in the overflow of norepinephrine (NE), 3-H-NE, total 3-H and dopamine-beta-hydroxylase (DBH) activity per nerve impulse was obtained after electrical stimulation of the isolated, perfused cat spleen. Over the range of frequencies studied, a proportional increase in the overflow of transmitter and DBH activity was observed, suggesting that an increase in the exocytotic release of NE is the primary mechanism by which the overflow of transmitter is enhanced with increasing frequencies of stimulation. The effects of 3 muM phenoxybenzamine (PBA) on the nerve stimulation-mediated overflow of NE, total 3-h, 3-h-ne and DBH activity were studied at two frequencies of stimulation, 1 and 5 Hz, in spleens from control and alpha-methyl-p-tyrosine-treated cats. Similarly, release of DBH activity was studied in spleens of reserpine-treated cats. In spleens from control cats, PBA produced a marked increase in the overflow of transmitter and DBH activity. This enhancement was more pronounced at 5 Hz than at 1 Hz. It was estimated that only 10 and 25% of the overflow of NE obtained in the presence of PBA at 1 and 5 Hz, respectively, could be accounted for by increased exocytosis. Depletion of tissue NE (80%) by pretreatment with alpha-methyl-p-tyrosine considerably reduced the effectiveness of PBA in enhancing the nerve stimulation-mediated overflow of NE, 3-H-NE, total 3-H and, particularly, of DBH activity. After reserpine treatment, the tissue NE content was reduced by more than 99%, yet the nerve stimulation-mediated release of DBH activity was similar to control spleens. PBA failed to enhance the nerve stimulation-mediated overflow of DBH activity after reserpine treatment. Thus it appears that the enhancement in the exocytotic release of transmitter by nerve stimulation observed in the presence of PBA is related to the concentration of NE in the synaptic space. Therefore, the present study is in favor of the notion that presynaptic alphareceptors may form part of a negative feedback control mechanism by which the transmitter may inhibit its own release.


Cardiovascular effects of acute and chronic inhalations of fluorocarbon 12 in rabbits.

The effects of inhaling fluorocarbon 12, a common propellant in household aerosols, were studied in closed-chested rabbits. Inhalation of 10 or 20% fluorocarbon 12 produced a decline in cardiac output and a dose-related depression of peak left ventricular (LV) dP/dt without associated arrthythmias, hypoxemia or significant changes in LV end-diastolicpressue or heart rate. There was a small decline in LV systolic pressure without a significant drop in mean arterial pressure. Breathing fluorocarbon 12 for 30 minutes caused a decline in peak LV dP/dt, cardiac output, LV systolic pressure and mean arterial pressure which was present throughout the exposure period. The effects observed during acute fluorocarbon 12 exposure were not altered by previous chronic, intermittent exposure to 10% fluorocarbon 12.


Effects of exercise metabolites on adrenergic vasoconstriction in the gracilis muscle of the dog.

Comparable and submaximal responses to sympathetic stimulation and to intra-arterial injections and infusions of norepinephrine were elicited in the isolated dog gracilis muscle, perfused with blood at constantflow. The changes in potassium concentration, osmolality, Po2 and inorganic phosphate concentration were determined during exercise. Blood was perfused to the muscle with the concentrations of each of these parameters varied to include that seen in exercise. Hypoxia (average Po2 5.4 mm Hg) produced a marked vasodilatation, and responses to norepinephrine and sympathetic stimulation were both increased to the same extent, due to the lowered vascular tone. Increases in osmolality (up to 50 mOsmol/kg) caused little antagonism of adrenergic responses, but higher increases caused a significant and equal inhibition of responses to norepinephrine and sympathetic stimulation. Increases in phosphate concentration (plus 41.6 mM) had little effect either on muscle resistance or on adrenergic responses. Increases in potassium concentration (plus10 mM) completely abolished responses to sympathetic stimulation, whereas those to norepinephrine were unaffected. This prejunctional effect may result from inhibition of sympathetic neurotransmission due to depolarization of adrenergic nerve terminals.


Effects of prostaglandins B2 and B1 on the pulmonary circulation in the intact dog.

The effects of prostaglandins (PG) B2 and B1 on the pulmonary circulation were studied in the intact spontaneously breathing dog under conditions of controlled blood flow using right heart and transseptal catheterization techniques to isolate and perfuse the left lower lung lobe. Infusion of PGB2 and PGB1 into the left lower lung lobe increased lobar arterial and lobar venous pressure but had no significant effect on left atrial pressure. PGB2 and PGB1 increased pressure gradients between the lobar artery and lobar small vein and between the lobar small vein and the left atrium. PGB2 and PGB1 increased isometric tension in isolated helical segments of lobar vein (3-5 mm diameter) but had little or no effect on lobar arterial segments of the same size. These results indicate that in the intact dog prostaglandins of the B series increase pulmonary vascular resistance by constricting lobar veins and to a lesser extent vessels upstream to lobar small veins, presumably small lobar arteries. PGB2 and PGB1 both produced large increases in pulmonary vascular resistance in the dog with PGB2 being about 10 times as potent as PGB1. It is concluded that PGB2 is one of the most potent pressor substances in the canine pulmonary vascular bed.


Renal mechanisms for the excretion of nicotinic acid.

Nicotinic acid, an essential endogenous organic acid, was studied in free-flow clearance experiments in the dog at plasma concentrations ranging from 1.2 to 600 mug/ml. At all concentrations, net reabsorption was observed. At concentrations of 1.2 mug/ml, the clearance of nicotinic acid as compared to the clearance of insulin was approximately 0.50. As nicotinic acid plasma concentrations were increased to 90 mug/ml, the clearance ratio declined to 0.22. The clearance ratio then steadily increased to 0.75 as plasma concentrations reached 600 mug/ml. Plasma levels of nicotinic acid in excess of 600 mug/ml resulted in renal toxicity as indicated by a marked decrease in the glomerular filtration rate. The decline in the clearance ratio in the presence of 90 mug/ml of nicotinic acid suggested saturation of a secretory system and hence cyanine 863 and probenecid were used to observe their effects on the clearance ratio. The base transport inhibitor was without effect; probenecid decreased the clearance. Alkalinization of the urine had no noticeable effect on nicotinic acid clearance. Protein binding did not occur. The data indicate two important points. First, nicotinic acid is secreted to a limited degree by the organic anion secretory system and is simultaneously reabsorbed. Second, a homeostatic mechanism for conservation of nicotinic acid at low plasma levels does not seem to exist.


Urinary excretion of morphine and its metabolites in morphine-dependent subjects.

Morphine, morphine glucuronide, morphine ethereal sulfate, normorphine and total normorphine in three consecutive 24-hour urines of four morphine-dependent subjects receiving morphine sulfate 60 mg s.c. q.i.d. have been determined with thin-layer chromatography and gas-liquid chromatography. With thin-layer chromatography the mean daily excretion of free morphine was 10% of the administered dose; morphine glucuronide, 65%; total (free and acid hydrolyzable conjugate) morphine 85%; and total normorphine, 3.5%. With gas-liquid chromatography, the percentage excretion for free morphine was 10%; total morphine, 74%; free normorphine, 1%; and total normorphine, 4%. The excretion of total drug was linearly related to the volume of the daily urine output.


Regulation of cyclic adenosine 3',5'-monophosphate levels in guinea-pig cerebral cortex by interaction of alpha adrenergic and adenosine receptor activity.

Direct assay of adenosine 3',5'-monophosphate (cyclic AMP) in guinea-pig cerebral cortex in vitro has shown that an alpha adrenergic receptor that was previously found to increase tissue content of cyclic AMP requires the co-presence of adenosine. This alpha adrenergic receptor complex was characterized with blocking agents and contrasted with other activities by examining the effect of other biogenic amines on cyclic AMP content in the presence of adenosine. Phentolamine (but not propranolol) reduced the potentiated response to norepinephrine (NE) (or epinephrine) plus adenosine to the level seen with adenosine alone. Theophylline, an adenosine antagonist, blocked the entire effect of NE plus adenosine. The failure of a high Mg++/Ca++ ratio to block the effect of NE plus adenosine argues against indirect mediation of the alpha receptor effect via the release of K+ or via an unknown neurohumoral agent. The complex variety of potentiative interactions between biogenic amines and adenosine is unique to brain. These interactions may be explained by the proposed existence of both independent and dependent receptors. The dependent receptors respond only to the co-presence of two or more neurohumoral agents. An alternative explanation would involve a compartmentally selective impairment of cyclic AMP degradation.


The effect of liver disease in man on the disposition of phenobarbital.

The disposition of phenobarbital (PB) was studied in normal individuals and in patients with cirrhosis or acute viral hepatitis to determine 1) if there is significant impairment of PB metabolism in hepatic disease and 2) to what extent such abnormal disposition of the drug affects its disappearance from blood. The diagnosis of liver disease was based on characteristic clinical findings, biochemical liver "function" tests and liver biopsy when necessary. All individuals had normal renal function and were free of other drug and alcohol intake for at least 3 weeks. With radiotracer methodology, PB and its principal metabolites, p-hydroxyphenobarbital (PBOH) and conjugated PBOH (PBOC), were monitored in blood and urine for 5 days after a single dose of 14-C-PBadministered intraduodenally. PB blood half-life (T1/2) in the control group was 86 plus or minus 3 hours (S.E.). In cirrhotics the T1/2 was prolonged to 130 plus or minus 15 hours (P less than .001) and this was accompanied by a 50% reduction in urinary PBOC excretion (P less than .05). Urinary excretion of PB and PBOH was unaltered by cirrhosis. In patients with acute viral hepatitis, PB T1/2 was not significantly prolonged and urinary excretion of PB and its metabolites was in the normal range (P greater than .05). No PBOH and only traces of PBOC were detected in the blood of either control individuals or patients with liver disease. Urinary excretion of unchanged PB was an important elimination pathway of the drug in all groups. As a result of this, PB T1/2 in cirrhosis was only moderately prolonged.


Effect of prolactin on androgen metabolism by the guinea-pig sex accessory organs.

The in vitro actions of prolactin on the metabolism of testosterone by the guinea-pig liver and sex accessory organs have been investigated under a variety of conditions including various concentrations of ovine or bovine prolactin, addition of reduced nicotinamide adenine dinucleotide phosphatate (NADPH) or nicotinamide adenine dinucleotide (NAD) to incubation media and various lengths of incubations. Treatment of the epithelium and muscle of the guinea-pig seminal vesicle with NAD elicited a shift from normal reductive metabolism of testosterone toward oxidative metabolism. This shift toward oxidative metabolism of testosterone was particularly marked in the muscle of the guinea-pig seminal vesicle. Ovine prolactin was highly effective in inhibiting the reductive metabolism of testosterone by the sex accessory organs. This effect appeared primarily as a decrease in dihydrotestosterone formation and was noted in both the presence and absence of NADPH or NAD. Oxidized metabolites, androstenedione and androstandione, were apparently insensitive to prolactin treatment in vitro. Variations induced by prolactin in the concentrations of intracellular androgens may be important in either the normal or abnormal growth and function of male sex accessory organs.


Increased running response to morphine in morphine-pretreated mice.

The running response of B6AF1/J mice to 25 mg/kg of morphine sulfate was increased up to 3-fold when this dose was administered either twice daily for 5 days or once a week for 2 or 3 weeks. The effect of weekly pretreatment was proportional to the dose of morphine and lasted as long as 1 month after pretreatment was stopped. There was no sensitization when the mice were less than 15 days old at the time of pretreatment. Of the parental strains, untreated C57Bl/6J mice showed a good running response to morphine, while A/J mice showed little response. Pretreatment of either of these strains produced only slight sensitization. Pretreatment of the hybrids with levorphanol increased the response to morphine. Dextrorphan and naloxone were ineffective. Sensitization by morphine was blocked by naloxone. Increased morphine running was not associated with analgesic tolerance as measured by the tail-flick assay. Morphine pretreatment produced some increase in the running response to amphetamine and to cocaine. Pretreatment with amphetamine or cocaine did not increase the response to morphine.


Further properties of stereospecific opiate binding sites in rat brain: on the nature of the sodium effect.

Addition of sodium salts has been reported to enhance stereospecific binding of opiate antagonists while reducing binding of agonists to rat brain homogenate. We have tested, in addition to sodium and potassium, a number of organic cations. Our results support the suggestion that the ability to enhace antagonist binding is not a general characteristic of cations or high ionic strength, but a property of sodium ions. We have shown that the increase in antagonist binding results from an enhancement of binding affinity and not from unmasking of new binding sites. The reduction in etorphine binding in the presence of sodium is due to a decrease in binding affinity. This decrease is largely accounted for by an acceleration in the dissociation rate, while the greater affinity of naltrexone binding appears to be due to an increase in rate of association. Our results are consistent with the hypothesis of a conformational change in opiate binding sites in the presence of sodium, transformed sites exhibiting greater affinity for antagonists and reduced affinity for agonists. Preheating of rat brain P2 fraction at 50 degrees C results in gradual inactivation of stereospecific binding, but an increase in naltrexone binding is consistently observed after heating at 50 degrees C for 2 to 3 minutes, even at optimal concentrations of sodium.


Effects of dopamine on carotid chemo- and baroreceptors in vitro.

1. The effects of dopamine on the sensory discharges originating from arterial chemo- and baroreceptors were studied in vitro using carotid bodies or sinuses excised from anaesthetized cats and superfused with Locke's solution. 2. Intrastream injections of dopamine 10-200 mug produced a transient depression of the frequency of chemoreceptor discharges. This effect was observed in response to the first injection in eighteen out of twenty preparations. 3. The inhibitory effect of dopamine can counteract partially or totally the excitation of chemoreceptors evoked by simultaneous application of acetylcholine or cyanide. 4. This inhibitory effect of dopamine is reduced or abolished by pretreatment with dopaminergic (Spiroperidol) or alpha-adrenergic (Dibenamine) blockers. 5. In response to repeated injections of dopamine applied at short intervals, the inhibitory effect is replaced by a biphasic effect (early inhibition followed by late excitation), a late and long-lasting excitation or no changes in chemoreceptor activity. The late excitatory effects of dopamine are not blocked by dopaminergic or alpha-adrenergic blockers. 6. Noradrenaline does not affect the chemoreceptor activity of the superfused carotid body. DL-DOPA induces only a late and long-lasting excitatory effect. 7. In carotid sinus preparations, dopamine induces a weak but long-lasting increase in the frequency of baroreceptor discharges. 8. It is concluded that dopamine may play a modulatory role in the generation of chemoreceptor activity through local regulatory processes.


The isolation and characterization of lactoferrin from sow milk and boar seminal plasma.

Lactoferrin isolated from sow milk (about 0-6 mg/ml) was shown to be chromatographically homogeneous, an observation supported by electrophoresis and by reaction against monospecific anti-lactoferrin antiserum. Isoelectric focusing showed multiple forms of the protein (i.e.p., 9-3 to 10-0) converted by neuraminidase to one form (i.e.p., 9-65). Boar seminal plasma contains immunologically identical lactoferrin (0-1 to 0-5 mg/ml) which binds strongly to boar spermatozoa.


Cardioselective beta-adrenergic blocking agents. 1. 1-((3,4-Dimethoxyphenethyl)amino)-3-aryloxy-2-propanols.

A series of 1-amino-3-aryloxy-2-propanols has been synthesized and examined for cardioselective beta-blockade. The introduction of the (3,4-dimethoxyphenethyl)amino group lead to the most cardioselective agents. Structure-activity relationships are discussed. Of the compounds tested 1-[(3,4-dimethoxyphenethyl)amino]-3-(m-tolyloxy)-2-propanol was selected for clinical trial because of optimal potency and selectivity.


Substituted thiadiazolines as inhibitors of central nervous system carbonic anhydrase.

A series (24-30) of substituted thiadiazolines was synthesized and tested for in vitro carbonic anhydrase inhibition and for protective ability against pentylenetetrazole-induced convulsions. ED50 (pentylenetetrazole protection), TD50, and LD50 values are reported for each compound. With the exception of 30, all compounds approximated the model compound methazolamide as in vitro carbonic anhydrase inhibitors. Several of the compounds produced extended protection against pentylenetetrazole-induced convulsions. Ring methoxy substitution in the ortho position appeared to produce maximum activity.


Clinical and ultrastructural observations in a kindred with normo-hyperkalaemic periodic paralysis.

Electron microscopic studies of muscle biopsies from clinically unaffected sibs in a family with normo-hyperkalaemic periodic paralysis with variable myotonia have revealed dilatation of the sarcoplasmic reticulum similar to that observed in affected members. This supports the view that such dilatation is not only a significant and likely primary ultrastructural change but that it may precede clinical manifestations and represent an anatomical marker of the genetic trait. Identical dilatation of the sarcoplasmic reticulum was found in the clinically unaffected father of the affected and unaffected grandchildren of the propositus. This raises the possibility that this non-consanguineous member contributed to the genetic trait or its manifestations in the grandchildren of the index patient since similar dilatation of the sarcoplasmic reticulum was not observed in the muscles of healthy control subjects.


Effect of electric fields on light-scattering and fluorescence of chromaffin granules.

Electric field pulses (in the 5 to 25 kV/cm range) were found to cause a transient increase in the intensity of scattered light (wavelength equals 369 nm, scattering angle equals 90 degrees) from aqueous sucrose suspensions of chromaffin granules. Similar observations were made with the membranes of osmotically lysed chromaffin granules. Under the same experimental conditions the degree of polarization of the scattered light changed only very slightly. The fluorescence of the hydrophobic probe diphenyl-hexatriene, incorporated into the membrane of intact chromaffin granules, showed similar transient changes in the intensity. The calculated relaxation times for these changes in optical properties were approximately 150 musec for the rising phase, and approximately 1 msec for the early stage of the decay. A further relaxation time of about 30 msec was also observed by using this probe. Essentially, all of these signals originated from the granule membrane, and could be attributed to rather small changes in particle size, membrane thickness or refractive index. Moreover, these signals were found to be completely reversible. Catecholamine release from intact granules, pulsed at voltages of 25 kV/cm, occurs already during the first few milliseconds of the transient membrane change.


The reaction of chemical probes with the erythrocyte membrane.

Trinitrobenzenesulfonate (TNBS), fluorodinitrobenzene (FDNB) and suberimidate have been reacted with intact human erythrocytes. TNBS does not penetrate the cell membrane significantly at 23 degrees C in bicarbonate-NaCl buffer, pH 8.6, as estimated by the labeling of the N-terminal valine of hemoglobin. Hence, under these conditions it can be used as a vectorial probe. However, at 37 degrees C, especially in phosphate buffer, at pH 8.6, TNBS does penetrate the cell membrane. FDNB and suberimidate both penetrate the erythrocyte membrane. The time course reaction of TNBS with intact erythrocytes over a 24-hr period at 23 degrees C is complex and shows transition zones for both membrane phosphatidylethanolamine (PE) and membrane proteins. No significant cell lysis occurs up to 10 hr. The fraction of total PE or phosphatidylserine (PS) which reacts with TNBS by this time period can be considered to be located on the outer surface of the cell membrane. Under these conditions it can be located on the outer surface of the cell membrane. Under these conditions it can be shown that 10 to 20% of the total PE and no PS is located on the outer surface of the membrane and hence these amino phospholipids are asymmetrically arranged. The pH gradient between the inside and outside of the cell in our system is 0.4 pH units. Nigericin has no effect on the extent of labeling of PE or PS by TNBS. Isotonic sucrose gives a slight enhancement of the labeling of PE by TNBS. Hence, the inability of PE and PS to react with the TNBS is considered not due to the inside of the cell having a lower pH. The extent of reaction of TNBS with PE is not influenced by changing the osmolarity of the medium or by treatment of cells with pronase, trypsin, phospholipase A or phospholipase D. However, bovine serum albumin (BSA) does protect some of the PE molecules from reacting with TNBS. Cels treated with suberimidate were suspended in either isotonic NaCl or in distilled water. In both cases the suberimidate-treated cells became refractory to hypotonic lysis. Pretreatment of cells with TNBS did not prevent them from interacting with suberimidate and becoming refractory to lysis. However, pretreatment of cells with the penetrating probe FDNB abolished the suberimidate effect. Electron-microscopic analysis of the cells showed a continuous membrane in the case of cells suspended in isotonic saline. The cells suspended in water did not lyse but their membranes had many large holes, sufficient to let the hemoglobin leak out. Since the hemoglobin did not leak out we know that the hemoglobin is cross-linked into a large supramolecular aggregate.


Cardiac lesions in collagen disease.

Clinical and pathological abnormalities of the heart that may have major functional significance in those conditions were studied in collagen disease (total 242 cases). 1. atypical verrucous endocarditis (Libmann-Sacks) were found in 2 cases with loupus erythematosus (25 cases) and in 1 case with scleroderma (3 cases). 2. Rheumatoid nodules were not seen in the heart of rheumatoid arthritis patients (9 cases). 3. The most frequent findings in the heart with collagen diseases were pericarditis, fibrous scarring of the heart muscle, focal inflammation and vasculitis is similar to periarteritis nodosa or endoarteritis obliterans. 4. Morphological changes of the A-V node were found in patients who had A-V dissociation or A-V block before death. Heart muscle infarctions were found in 2 cases.


[Reaction of the neurosecretory system of dogs with reno-vascular hypertension in the postnatal ontogenesis].

In dogs of different age with simulated renovascular hypertension neurosecretion of the hypothalamo-neurohypophysial system was studied. Experiments were conducted with puppies 18-22 days, 11/2-3 months and dogs 3-5 years old. It was established that in the ontogenesis of dogs the differentiation of neurons in the supraoptic nucleus occurred earlier than that of cells in the paraventricular nucleus. The simulation of hypertension accelerates the differentiation of tbe neurosecretory cells. The development of arterial hypertension leads to prevalence of precesses of the neurosecretory material outflow over the latter's synthesis, irrespective of the maturity of the neurosecretory system. With the arterial pressure stabilized at a high level a concordance between an elevated synthesis of the neurosecretion and its increased outflow is noted. Visual evaluation of the activity is supported by the results of cellular micrometry.


[Problem of increasing the viability of the ischemic myocardium in the light of experimental studies].

With the author's own observations and literature sources as a background the key issues concerned with increasing the viability of the myocardium in acute ischemia are considered. The possibility of a material enlargement of the collateral coronary circulation through administration of mesatonemonoaminoxidase inhibitors, diprazine, preparations of the metabolites type and of other agents is shown. Under consideration are data on the membranes-stabilizing effect in acute ischemia of the myocardium of dimedrol, diprazine and prednisolone, as well as possible ways of increasing the survival of the myocardium by activating the redox-processes and through an adequate supply of energy to ensure the vital functions of the myocardial cell at rest by using pertinent pharmacological agents (cytochrome C, NADP, ubiquinone, hexose-phosphate, monoaminodicarboxylic amino acids).


Effect of diphosphonate on crystallization of calcium oxalate in vitro.

Reliable techniques for the calculation of activity product (state of saturation), formation product (limit of metastability) and crystal growth of calcium oxalate were devised.  The activity product at saturation was 2.53 times 10 minus 9 M2, and was independent of duration of incubation, solid-to-solution ratio, pH, calcium concentration or ionic strength. These tehcniques were utilized to assess the effect of disodium ethane-1-hydroxy-1, 1-diphosphonate (EHDP) on crystallization of calcium oxalate in an aqueous salt solution in vitro. EHDP increased the formation product of calcium oxalate, indicating an inhibition of nucleation of calcium oxalate. Further, it inhibited the crystal growth of calcium oxalate.


Dailysate calcium and plasma calcium fractions during and after haemodialysis.

Dialysate calcium and plasma calcium fractions during and after haemodialysis: The effect of differenct dialysate Ca concentrations on the plasma Ca fractions was examined in 28 patients. In 10 patients dialysed with a dialysate Ca concentration of 3.0 mEq/l the Ca fractions were determined at the start and end of dialysis. 8 patients were dialysed with dialysate with dialysate Ca of 3.5 mEq/l. In this group the Ca fractions were also estimated in the dialysis-free interval. The third group was dialysed with a dialysate Ca of 4.5 mEq/l. Total calcium and protein-bound calcium rose significantly in all groups. Ionised calcium in the first group was significantly reduced, in the second group it remained constant and in the third group it was significantly raised. Since parathyroid function  depends on the plasma ionised calcium it is concluded that a dialysed concentration of 3.0 mEq/l is partly responsible for the pathogenesis of secondary hyperparathyroidism and of renal osteodystophy. In normocalcaemic patients a dialysate Ca concentration of 3.5 to 4.0 mEq/l is optimal. In patients entering long-term haemodialysis treatment with pronounced calcium deficiency symptoms a dialysate Ca of up to 4.5 mEq/l may be indicated for a short period after having normalized the inorganic phosphate levels in order to prevent extraosseous calcification.


Angiotensin stimulated AVP-release in humans.

Release of arginine vasopressin (AVP) from rat neurohypophysis in in vitro studies is significantly augmented by the addition of angiotensin (A-II), and in in vivo studies in dogs renin and A-II were found to stimulate secretion of AVP. Both these results suggest the existence of a direct relationship between the salt regulating renin-angiotensin-aldosterone system and the water controlling AVP system. To evaluate whether such observations apply also in man a sensitive double antibody radioimmunoassay for AVP was developed [17, 18]. Basal plasma levels of AVP in recumbent humans without salt and fluid restriction at room temperature were 3.4 plus or minus 2.2 pg/ml, and 30 min after the onset of an A-II infusion at a concentration of 3-30 ng/min-kg, a significant increase of AVP was found. Maximum  measurements were 2-5 times basal levels which returned to normal within 90 min. During the A-II infusion one also noted a 20 mm Hg rise in blood pressure, accompanied by a significant decrease in plasma renin activity. During the same period serum osmolality and serum sodium concentration did not change. Elevation of blood pressure by norepinephrine was not followed by any detectable change of plasma AVP levels, thus excluding a nonspecific blood pressure effect.


[Determination of the ammonia content in the atmosphere of biosatellite mock-ups and approaches to its standardization].

The concentrations of ammonia having an irritating and acute effect on intact rats and hypokinetic rats were determined. The thresholds of the chronic effect were calculated for each group of animals. The 30-day hypokinesia increased the animal sensitivity to ammonia: the threshold of the irritating effect decreased twice. The atmosphere in an enclosure equipped with an automatic life support system was examined during 30-day integrated animal experiments. Ammonia proved to be the major contaminant. Its concentration increased gradually, remaining for the longest part of the experiments (28 days) below the threshold of the chronic effect (40 mg/m3).


[Toxicological evaluation of ethyl acetate in conditions of sealed cabin atmospheres].

Ethyl acetate was studied as an atmosphere contaminant in an enclosed environment. A 90-day continuous experiment on 160 white rats and 120 white mice with three concentrations of the compound (43, 10 and 2 mg/m3) was carried out. Ethyl acetate used in the first two concentrations caused functional disturbances in the state of animals and morphological changes in their viscera. Ethyl acetate at a concentration of 2 mg/m3 was ineffective. The data obtained allow recommendations of permissible concentrations of ethyl acetate vapours in an enclosed atmosphere in relation to a different time of man's exposure.


Production and consumption of alcoholic beverages in the USSR. A statistical study.

The Soviet government has published few official statistics on alcoholic beverages. Using Soviet statistical, commerical and technical sources, however, estimates can be derived on the production and consumption of alcoholic beverages during the 1957-71 period, including urban and rural consumption, expenditures, tax revenues, imports and exports and use of agricultural products.


Drinking attitudes and practices among Wind River Reservation Indian youth.

The attitudes toward drinking of a cohort of 7th- and 8th-grade American Indians, mostly Arapahoe and Shoshone, living on Wyoming's Wing River Reservation were studied. The data suggest that these adolescents and their peers both approve of drinking and do drink even though they can get into trouble for illegal alcohol use.


Attitudes toward alcohol and alcoholism among professionals and nonprofessionals.

Analysis of the responses of graduate students, professional social service agency workers and community residents to scales measuring attitudes and beliefs about alcohol and alcoholism suggests that the professionals tend to reflect the opinions and biases of the community within which they work. The results are discussed in terms of the usefulness of the multidimensional model of attitude structure for comparing attitudes among various groups and the implications for professional and community education programs.


Relationship between treatment facilities and prevalence of alcoholism and drug abuse.

Data are presented which suggest that, in the Commonwealth of Pennsylvania, the availability of treatment of drug abuse and alcoholism is inversely related to the prevalence of the problems. It is hypothesized that as the treatment response to one increases, the treatment response to the other declines. Some of the possible causes and consequences are outlined.


Glucose intolerance in alcoholism.

Intravenous glucose tolerance tests were given to 31 nondiabetic alcoholics and 11 healthy nonalcoholic controls. In almost half of the alcoholics peak glucose concentration was higher and glucose elimination from the plasma was slower than in the controls.


[Preservation of food wastes by a mixture of quinosol and salicyclic acid].

A 90-day experiment has shown that a number of toxic gaseous contaminants are released into the atmosphere as a result of storage of mixed food wastes. The microorganisms dwelling upon decaving food wastes include different microbial species from the environment and food products. A preserving agent consisting of a mixture of quinosole and salicylic acid (1:1) has been used. The agent inhibited microbial activity and terminated processes of decay. This was indicated by a decline in the concentration of ammonia and amine compounds. In the presence of the agent fermentation processes, on the contrary, continued and increased.


Treatment outcome in a drinking-decisions program.

Of 98 alcoholics who were free to decide whether or not to drink during a 6-week inpatient treatment program, 55 chose to drink. The nondrinkers tended to show more improvement at a 6-month follow-up than did the drinkers.


Intellectual competence of alcoholics.

Alcoholic and psychiatric inpatients matched in age and education performed comparably on a battery of cognitive tests. The absence of impairment in the alcoholics is attributed to the relatively short duration of their alcoholism (mean, 7.2 years) and the nature of the tasks involved.


Field dependence and self-actualization in alcoholics.

Field independence and self-actualization were significantly increased in alcoholic patients following a 6-week treatment program. There were no differences, however, between patients treated by drug-assisted individual therapy, individual therapy, or milieu therapy. Some causal factors are hypothesized, and implications for treatment are discussed.


Personality and temperament differences between alcoholics with high and low records of traffic accidents and violations.

Significant differences on scales of the Guilford-Zimmerman Temperament Survey and the Minnesota Multiphasic Personality Inventory were found between alcoholics with high and low records of traffic accidents and violations.


Spouse response to a self-administered alcoholism screening test.

A self-administered, expanded version of the Michigan Alcoholism Screening Test was completed by a group of hospitalized alcoholics and by their spouses and counselors, describing the patients. The data obtained from spouse responses was accurate in 90% of the cases; the spouse may be a more reliable source of information about drinking problems than the alcoholic.


The impact of specialized training in alcoholism on management-level professionals.

Administrative and management-level social workers and nurses significantly increased their alcohol-related work activities during the year after they attended an 8-week alcoholism training program. Measures of attitude change, however, showed no appreciable differences.


A descriptive analysis of alcohol education materials.

Analysis of 832 alcohol education materials (books, pamphlets and leaflets) indicates that the quantity and quality of the materials have increased in recent years. Alcoholism and the effects of alcohol are the major themes for all audience levels. There are few good quality resources for the junior-high and elementary school levels.


Defense mechanisms in men and women alcoholics.

The responses of men and women alcoholics to the Defense Mechanism Inventory showed that the women scored significantly higher in "turning against self" and the men in "turning against others." Although such differences may underlie reported sex differences in the incidence of depression and sociopathic personality disorder, they have also been found in studies of nonalcoholics and psychiatric outpatients.


A report of a 1972 survey of physician's assistant training programs.

This article presents select results of a 1972 survey of all physician's assistant training programs in the United States and its territories. The survey gathered a broad range of information useful to federal health manpower policymakers, physician assistant training program administrators, guidance counselors, and prospective students. Responses were received from 75 operational programs and 13 programs still in planning for an estimated response rate of 81 per cent from the estimated 108 programs in existence.


Peanut alkaline lipase.

Peanut alkaline lipase, (glycerol ester hydrolase EC 3.1.1.3), pH optimum 8.5, was isolated from acetone powders prepared from developing and germinated peanut seed (arachis hypogaea L. var. NC-2). Enzyme activity/seed increased in successive developmental stages. The course of the hydrolytic reaction was linear with regard to enzyme concentration and all times tested up to periods exceeding 60 min. Km for the reaction was determined to be 2.6 times 10-4M. Molecular weight of peanut lipase, as estimated by Sephadex gel filtration and sodium dodecyl sulfate gel electrophoresis, was ca. 55,000.


The clinical pharmacology of hypotensive vasodilator drugs.

Drugs which lower arterial blood pressure by direct vasodilator action usually have to be combined with beta adrenergic blocking drugs and diuretics for maximum clinical effectiveness. Appreciation of the pharmacodynamic interactions is necessary to individualize drug programmes rationally. Vasodilator drug therapy is remarkable for the lack of side effects such as central nervous system effects, postural hypotension and sexual difficulties which usually characterize most "antiadrenergic" drug regiments.


Vasodilators in the treatment of hypertension.

Vasodilator drugs play an important role in the treatment of hypertension in 1974. A powerful vasodilator such as diazoxide may be necessary in the control of the most severely affected patients. It should not be forgotten that drugs of the benzothiadiazine group probably act mainly as vasodilators and that their effect is not dependent upon natriuresis and diuresis. New vasodilators such as minoxidil may have an important role in the treatment of hypertension, but these have not yet been adequately assessed in Australia. Both hydrallazine and prazosin produce a significant fall in the blood pressure when added to a combination of a beta blocker and a diuretic, and double-blind studies have shown that the side effects are more predominant with hydrallazine than with prazosin.


The non-enzymic reduction of nitrite by benzyl viologen (free-radical) in the presence and absence of ammonium sulphate.

Some similarity is inferred between the reaction or reduced benzyl viologen with undissociated nitrous acid, which is significant at pH values below 7 and that with the undissociated product of nitrite ion and ammonium sulphate; presumably ammonium nitrite. This would explain why the presence of ammonium sulphate appreciably offsets the effects of decreasing pH and also the exponential relationship between rate of nitrite loss and ammonium sulphate concentration. There are other features of the reaction which cannot be explained at present, especially with regard to the degree of reduction of benzyl viologen. It is nevertheless apparent that a complex non-enzymic reaction yielding several products occurs when ammonium sulphate is present and that the presence of likely residual quantities after its use in enzyme purification may cause serious errors in enzyme assay.


On the involvement of a glucose 6-phosphate transport system in the function of microsomal glucose 6-phosphatase.

A model for microsomal glucose 6-phosphatase (EC 3.1.3.9) is presented. Glucose 6-phosphatase is postulated to be resultant of the coupling of two components of the microsomal membrane: 1) a glucose 6-phosphate - specific transport system which functions to shuttle the sugar phosphate from the cytoplasm to the lumen of the endoplasmic reticulum; and 2) a catalytic component, glucose-6-P phosphohydrolase, bound to the luminal surface of the membrane. A large body of existing data was shown to be consistent with this hypothesis. In particular, the model reconciles well-documented differences in the kinetic properties of the enzyme of untreated and modified microsomal preparations. Characteristic responses of the enzyme to changes in nutritional and hormonal states may be attributed to adaptations which alter the relative capacities of the transport and catalytic components.