Catecholamines produce mitotic inhibition in primary cell cultures of human keratinocytes probably via a block in the G2 part of the cell cycle. Epinephrine produced significant mitotic inhibition (49%) at a concentration as low as 4.5 X 10(-10) M, while its analog, isoproterenol, produced 47% inhibition at 1 X 10(-10) M. Norepinephrine elicited a 49% inhibitory response at 1 X 10(-8) M. One other catecholamine, dopamine, caused a 53% decrease in mitosis at 1 X 10(-6) M. Other structurally related amines to exhibit mitotic inhibition were phenylephrine, 58% at 1 X 10(-7) M; octopamine, 47% at 1 X 10(-5) M; and tyramine, 52% at 1 X 10(-4) M. Serotonin showed no mitotic inhibition at 1 X 10(-4) M. Various alpha and beta adrenergic blocking agents were added to the cell system. The alpha blocking agent, phentolamine, had no effect on mitosis. When added in conjunction with epinephrine or norepinephrine, no reduction of the catecholamine-induced mitotic inhibition was observed. The beta blocking agent, propranolol, by itself showed slight mitotic inhibition at 1 X 10(-6) M. When added along with epinephrine or noreinephrine, propranolol reduced the catecholamine-induced mitotic inhibition approximately 65%. In addition, propranolol blocked mitotic inhibition caused by phenylephrine, an alpha adrenergic agent. However, another beta blocking agent, dichloroisoproterenol, showed strong mitotic inhibition (53%) when added to the cultures at a concentration of 1 X 10(-8) M. The effect was reduced to zero in the presence of propranolol. These data suggest that while beta receptors may be involved in the catecholamine-induced mitotic inhibition of human keratinocytes in vitro, the nature of the receptor-molecule interaction may be complex.

Effects of sodium nitrate were compared with sodium chloride loading on transport of electrolytes by the nephron. Maximal levels of free water clearance/clomerular filtration rate (CH2O/GFR) averaged 8.4% with nitrate loading and 14.4% with saline loading. Since ethacrynic acid and chlorothiazide exert their major natriuretic effect in the distal nephron, the increment in Na ad Cl reabsorbed beyond the proximal tubule. The administration of these agents resulted in an increase in fractional sodium excretion (CNa/GFR) of 21.1%, urinary sodium excretion (UNaV) of 1,126 mueq/min, and urinary chloride excretion (UClV) of 848 mueq/min during nitrate loading compared with an increase in CNa/GFR of 37.6%, UNaV of 2,362 mueq/min, and UClV of 2,397 mueq/min during saline loading. The smaller diuretic-induced increment in Na and Cl excretion in the nitrate studies suggests, as do the hydrated studies, that less Cl and Na are reabsorbed in the distal nephron during nitrate than saline loading. At every level of UNaV, fractional bicarbonate reabsorption was higher, urine pH was lower, and urinary potassium excretion (UKV) was higher in the nitrate studies. Thus, compared with saline loading, sodium nitrate decreases chloride and sodium reabsorption in the distal nephron. The higher hydrogen and potassium secretion in the nitrate studies may be consequent to the decreased ability of the distal nephron to reabsorb chloride.

Choline acetyltransferase (EC 2.3.1.6) catalyzes the biosynthesis of acetylcholine according to the following chemical equation: acetyl-CoA + choline in equilibrium to acetylcholine + CoA. In addition to nervous tissue, primate placenta is the only other animal source which contains appreciable acetylcholine and its biosynthetic enzyme. Human brain caudate nucleus and human placental choline acetyltransferase were purified to electrophoretic homogeneity using ion-exchange and blue dextran-Sepharose affinity chromatography. The molecular weights determined by Sephadex G-150 gel filtration and sodium dodecyl sulfate gel electrophoresis are 67000 plus or minus 3000. N-Ethylmaleimide, p-chloromercuribenzoate, and dithiobis(2-nitrobenzoic acid) inhibit the enzyme. Dithiothreitol reverses the inhibition produced by the latter two reagents. The pKa of the group associated with N-ethylmaleimide inhibition is 8.6 plus or minus 0.3. A chemically competent acetyl-thioenzyme is isolable by Sephadex gel filtration. The enzymes from the brain and placenta are thus far physically and biochemically indistinguishable.

Delta5-3beta HSDH activity has been assayed either by spectrophotometric method or by use of radioactive substrates. The enzymatic activity is equally distributed between mitochondrial and microsomal fractions verified by electronic microscopy. The specific activity is comparable in both fractions, as well as the optimal pH and the Km for NAD and for the substrates. The delta5-3beta Hut optimal pH, specific activity and sensitivity to the inhibitory action of various steroids are different when C19 and C21 steroids are used as substrates. Estrogens and cyclic AMP have also an inhibitory action on the oxidation of C21 steroids. Treatment of microsomal or mitochondrial membranes with phospholipase A releases fatty acids (mainly arachidonic) and decreases the enzymatic activity. "Adsorbtion" of the fatty acids on bovine serum albumin partially reactivates the delta5-3beta HSDH.

During penicillin treatment of an autolysin defective mutant pneumococcus we have observed three novel phenomena: (i) Growth of the mutant cultures is inhibited by the same concentrations of penicillin that induce lysis in the wild type. (ii) Mutant bacteria treated with the minimum growth inhibitory concentration of penicillin will lyse upon the addition of wild-type autolysin to the growth medium. Chloramphenicol and other inhibitors of protein synthesis protect the cells against lysis by exogenous enzyme. Sensitivity of the cells to exogenous autolysin requires treatment with penicillin or other inhibitors of cell wall synthesis (e.g., D-cycloserine or fosfonomycin) since exogenous autolysin alone has no effect on bacterial growth. (iii) Treatment with penicillin (or other inhibitors of cell wall synthesis) causes the escape into the medium of a choline-containing macromolecule that has properties suggesting that it contains pneumococcal lipoteichoic acid (Forssman antigen). Each one of these three phenomena (growth inhibition, sensitization to exogenous autolysin, and leakage of lipoteichoic acid) shows the same dose response as that of the penicillin-induced lysis of wild-type pneumococci. On the basis of these findings we propose a new hypothesis for the mechanism of penicillin-induced lysis of bacteria. It is suggested that inhibition of cell wall synthesis by any means triggers bacterial autolytic enzymes by destabilizing the endogenous complex of an autolysin inhibitor (lipoteichoic acid) and autolytic enzyme. Escape of lipoteichoic acid-like material to the growth medium is a consequence of this labilization. Chloramphenicol protects bacteria against penicillin-induced lysis by interfering with the activity of the autolytic enzyme, rather than by depleting the concentration of the enzyme at the cell surface.

The distributions of vibrational, rotational and translational energies in the products of atom-diatomic molecule reactions are studied on the basis of their ‘‘temperature parametere’. The validity of the vibrational temperature concept, which my characterize the variaous distributions, is examined from different points of view. The analysis is done on the basis of available data from chemiluminescence, chemical laser and molecular beams measurements and from classical trajectory calculations. All the reactions investigated are exothermic with exothermicities ranging between approximate to 17 kcal/mole and approximate to 85 kcal/mole. In most products a high degree of population inversion is found. An attempt is made to predict the complete vibrotational state distribution using the vibrational temperature only. This attempt is equivalent to the assumption that apart from the vibration, the other degrees of freedom have reached a (microcanonical) equilibrium. The agreement with experimental results is good.