The differential effects of the carcinogen dimethylnitrosamine on isocitrate and 6-phosphogluconate metabolism in single intact cells

A microspectrofluorimetric study is made of the influence of dimethylnitrosamine on NADP reduction, following sequential microinjections into the same L cell, of two substrates: (1) isocitrate, with activity of isocitrate dehydrogenase both in the extramitochondrial and intramitochondrial compartments, (2) 6-phosphogluconate, with activity of the dehydrogenase in the extramitochondrial compartment. In control L cells a two-step reduction of NAD(P) is obtained followed by relatively slow reoxidation. In the minutes which follow addition of carcinogen, e.g., dimethylnitrosamine, to the cell medium the isocitrate and 6-phosphogluconate-induced transient NADP reoxidation is decreased in magnitude compared to control, while the rate constant of NADPH reoxidation is considerably accelerated, possibly due to requirements at the level of the microsomal metabolizing system. Observations within the first hour of carcinogen addition suggest an interesting system for evaluating the immediate actions of carcinogens at extranuclear sites: i.e., a comparative study of NADP reduction-reoxidation rate constants via injection of substrates for extra- vs. intramitochondrial pathways.     

Transient and long-term differential modulations of branched-chain α-keto acid decarboxylase activity in hypophysectomized rats

Hypophysectomy caused a marked but transient increase in branched-chain α-keto acid decarboxylase activities in rat liver mitochondria, peaking at about nine days post-surgery. The magnitude of increase is different for each of the three branched-chain α-keto acids. The activities then fall to a new steady state in three weeks with α-ketoisovalerate decarboxylase activity within the normal range, α-keto-β-methylvalerate decarboxylase activity at twice normal, and α-ketoisocaproate decarboxylase activity decreased to a level too low for accurate measurements.     

Block by phencyclidine of acetylcholine and barium induced adrenal catecholamine secretion

Activation of the sympathetic system by phencyclidine (PCP) should result in catecholamine release from the adrenals. However, adrenalectomy does not reduce PCP-induced hypertension. In an attempt to rectify this inconsistency, the direct effects of PCP on the bovine adrenal medulla were examined. At (3×10^?6M), PCP reduced the acetylcholine-(ACh)-induced catecholamine release by 50%. Surprisingly, barium-induced secretion of catecholamines was also reduced by PCP. ACh-induced catecholamine release was not altered by 10^?3M 4-aminopyridine (4 AP), the potassium channel blocker. Thus, calcium antagonist actions of PCP and consequent block of catecholamine secretion from adrenal medulla may explain the lack of effect of adrenalectomy on PCP-induced hypertension. Possible contributions of calcium and/or potassium channel blockade to other manifestations of PCP overdosage are discussed.     

Taurodontism: A biometric study

This study has been carried out in order to clarify various aspects of the phenomenon known as taurodontism. Samples of both taurodont and non-taurodont teeth were measured and statistical analyses were performed on the resultant data from which various conclusions may be drawn. The most important conclusion is that this phenomenon may be defined metrically. The distributions have shown that the taurodont trait grades into the normal and it is quite difficult to make a subjective judgment at the low end of the taurodont range. It is for this reason that an accurate mathematical means for performing this task is so invaluable. The question of the nature of cuneiform single rooted molars was also investigated, and they were found not to be true taurodonts although the statistical procedure classed them as such. It was demonstrated that through the construction of simple plots these teeth could be eliminated from any group classed by analysis as taurodonts. Various metrical attributes were evaluated with respect to their ability to distinguish taurodonts, and two were found to be satisfactory for this purpose in most cases. Certain changes in molar morphology regarding caries and age were also investigated.     

Solubilization and characterization of cardiac sarcolemmal taurine-binding proteins

Cardiac sarcolemma preparations of both pig and rat ventricles were found to possess two sets of taurine-binding components. The two proteins from pig heart were solubilized with the detergent Ammonyx-Lo. Characterization of these solubilized proteins revealed that both components are glycoproteins and retain the binding properties observed for the membrane isolate. However, the characterization also revealed several differences between the proteins including their binding specificities, their affinities for taurine, their binding isotherms, and their molecular sizes. Possible functions of these two taurine-binding proteins are discussed.     

High resolution scanning electron micrographic study of dissociated mouse taste cells

New techniques for enzymatic dissociation of mammalian tastecells allowed us to study, for the first time, the morphologyof murine taste receptor cells using high resolution scanningelectron microscopy. Cell shape varied from spindle to bipolarto lamellar, similar to shapes previously described in cellsfrom amphibian taste buds. Cell length varied from 19 to 65µm (39 ± 19 µm), with width averaging 6 ±3.4 µm. A rare picture of the apical microvilli of a tastereceptor cell, and a view of microvilli within a taste pore,suggest that at any given time, five to eight taste cells maybe exposed to the oral cavity. Assuming a cell life-span of10 days, and 50 cells per bud, all of which eventually reachthe taste pore, one can calculate that the average cell is exposedto the oral environment for     

Kinetic Basis for the Conjugation of Auxin by a GH3 Family Indole-acetic Acid-Amido Synthetase

The GH3 family of acyl-acid-amido synthetases catalyze the ATP-dependent formation of amino acid conjugates to modulate levels of active plant hormones, including auxins and jasmonates. Initial biochemical studies of various GH3s show that these enzymes group into three families based on sequence relationships and acyl-acid substrate preference (I, jasmonate-conjugating; II, auxin- and salicylic acid-conjugating; III, benzoate-conjugating); however, little is known about the kinetic and chemical mechanisms of these enzymes. Here we use GH3-8 from Oryza sativa (rice; OsGH3-8), which functions as an indole-acetic acid (IAA)-amido synthetase, for detailed mechanistic studies. Steady-state kinetic analysis shows that the OsGH3-8 requires either Mg²⁺ or Mn²⁺ for maximal activity and is specific for aspartate but accepts asparagine as a substrate with a 45-fold decrease in catalytic efficiency and accepts other auxin analogs, including phenyl-acetic acid, indole butyric acid, and naphthalene-acetic acid, as acyl-acid substrates with 1.4–9-fold reductions in k_cat/K_m relative to IAA. Initial velocity and product inhibition studies indicate that the enzyme uses a Bi Uni Uni Bi Ping Pong reaction sequence. In the first half-reaction, ATP binds first followed by IAA. Next, formation of an adenylated IAA intermediate results in release of pyrophosphate. The second half-reaction begins with binding of aspartate, which reacts with the adenylated intermediate to release IAA-Asp and AMP. Formation of a catalytically competent adenylated-IAA reaction intermediate was confirmed by mass spectrometry. These mechanistic studies provide insight on the reaction catalyzed by the GH3 family of enzymes to modulate plant hormone action.