Evidence for the presence of the glyoxylate cycle in Chloroflexus

The key enzymes of the glyoxylate cycle, isocitrate lyase and malate synthase, were present in cell-free extracts of the phototrophic, green, thermophilic bacterium Chloroflexus aurantiacus grown with acetate as the sole organic carbon source.The optimum temperature of these enzymes was 40° C, and their specific activities were high enough to account for the observed growth rate. Lower levels of the enzymes were found in extracts from cells grown on a complete medium.Itaconate was shown to inhibit isocitrate lyase from C. aurantiacus 96% at a concentration of 0.25 mM and also had a profound effect on the growth of the organism on acetate, 0.25 mM inhibiting completely. Itaconate also inhibited the growth when added to the complex medium, but in this case much higher concentrations were required.     

Stimulation of plasmalemma adenosine triphosphatase from oat roots by inorganic and organic cations: concentration-dependence, selectivity, and sites

K⁺-stimulated ATPase activity of a plasmalemma-enriched fraction from excised roots of oat was triphasic in the range 5 to 80 millimolar KCl. The phases obeyed Michaelis-Menten kinetics and were separated from each other by jumps or sharp breaks at about 10 and 20 millimolar. Stimulation by alkali cations was in the order K⁺ > Rb⁺ > Na⁺ > Cs⁺ > Li⁺ or in a closely related sequence. The specificity reflected differences in V_max, not in affinity (K_m⁻¹). Stimulation by the organic cations ethanolamine and choline in the interval 11 to 80 millimolar appeared monophasic rather than biphasic. Substitution on the quaternary nitrogen of the amino alcohols decreased their effectiveness, as did extension and branching of the chain. Stimulation was maximal at about pH 7 both for K⁺ and choline.     

Total Thyroxine and Free Thyroxine Index in Plasma of Dairy Cows in Relation To Strength of Heat

Total thyroxine (TT4) and free thyroxine index (FT4I) were measured in peripheral plasma of cows. The samples were collected at the time of insemination from 66 cows showing pronounced signs of the heat and from 56 cows showing weak or silent heat. Neither TT4 or FT4I in plasma differed significantly between the two categories of oestrous cows.     

In vitro enhancement of natural and antibody-dependent lymphocyte-mediated cytotoxicity against tumor target cells by interferon.

Interferon derived from virus-infected human leukocytes or fibroblasts was found to enhance spontaneous and antibody-dependent lymphocyte cytotoxicity against human target cell lines in vitro. The greater enhancement occurred with spontaneous lymphocyte cytotoxicity. Interferon exerted its effect directly on lymphocytes; no effect on target cells was seen. The mechanism of enhancement was unclear: It did not reflect antibody production or lymphocyte proliferation. Enhancement appeared to be immunologically nonspecific, but clarification of this effect awaits further study.     

Effects in vitro of alloxan on the glucose metabolism of mouse pancreatic B-cells

To facilitate detailed studies of the B-cytotoxic action of alloxan we developed a model using isolated pancreatic islets of normal mice. An essential feature of this model is the low temperature employed during exposure to alloxan, which minimizes the degradation of the drug. The islets were incubated with alloxan for 30min at 4 degrees C and subsequently various aspects of their metabolism were studied. The O(2) consumption was measured by the Cartesian-diver technique. Islets exposed to 2mm-alloxan and control islets had the same endogenous respiration, whereas the O(2) uptake of the alloxan-treated islets was inhibited and that of the control islets stimulated when they were incubated with 28mm-glucose as an exogenous substrate. The islet glucose oxidation was estimated by measurement of the formation of (14)CO(2) from [U-(14)C]glucose at 37 degrees C. Compared with the controls, alloxan-treated islets showed a decrease in the glucose-oxidation rate in a dose-dependent manner. Pretreatment of the islets with 28mm-glucose for 30min at 37 degrees C completely protected against this effect, whereas preincubations at glucose concentrations below 16.7mm failed to exert any protective effect. The glucose utilization was estimated as the formation of (3)H(2)O from [5-(3)H]glucose. Alloxan (2mm) failed to affect islet glucoseutilization rate in the presence of either 2.8 or 28mm-glucose. In contrast, islets exposed to 5 or 10mm-alloxan exhibited lowered glucose utilization. It is concluded that in vitro alloxan has an acute inhibitory effect on the islet glucose metabolism, and that this action can be prevented by previous exposure to a high glucose concentration. The results are consistent with the idea that the B-cytotoxicity of alloxan reflects an interaction with intracellular sites involved in the oxidative metabolism of the B-cell.     

High-affinity uptake of γ-aminobutyric acid in cultured glial and neuronal cells

Both glial and neuronal cells maintained in primary culture were found to accumulate [³H]GABA by an efficient high-affinity uptake system (apparentK _m=9 M,V _max=0.018 and 0.584 nmol/mg/min, respectively) which required sodium ions and was inhibited by 1 mM ouabain. Strychnine and parachloromercuriphenylsulfonate (pCS) (both at 1 mM) also strongly inhibited uptake of [³H]GABA, but metabolic inhibitors (2,4-dinitrophenol, potassium cyanide, and malonate) were without effect. Only three structural analogs of GABA (nipecotate, -alanine, and 2,4-diaminobutyrate) inhibited uptake of [³H]GABA, while several other compounds with structural similarities to GABA (e.g. glycine,l-proline, and taurine) did not interact with the system. The kinetic studies indicated presence of a second uptake (K _m=92 M,V _max=0.124 nmol/mg/min) in the primary cultures containing predominantly glioblasts. On the other hand, only one of the neuronal cell lines transformed by simian virus SV40 appeared to accumulate [³H]GABA against a concentration gradient. ApparentK _m of this uptake was relatively high (819 M), and it was only weakly inhibited by 1 mM ouabain and 1 mM pCS. The structural specificity also differed from that of the uptake observed in the primary cultures. Significantly, none of the nontransformed continuous cell lines of either tumoral (glioma, C₆; neuroblastoma, Ml; MINN) or normal (NN; I₆) origin actively accumulated [³H]GABA. It is suggested that for the neurochemical studies related to GABA and requiring homogeneous cell populations, the primary cultures offer a better experimental model than the continuous cell lines.     

Genetic effects on male mouse kidney glucuronidase activity

Kidney β-glucuronidase activity in C57BL/Kl and DBA/2/Kl male mice differs about tenfold, C57 giving low and DBA high values. Another C57 subline, C57BL/6J, has slightly higher activity than C57BL/Kl. There is an association between the kidney glucuronidase activity and coat color determined by the buff locus, which indicates that part of the variation is due to differences at the Gur locus. The bf allele per se raises the activity of the enzyme. The backcross distributions give evidence that at least one more locus is involved.     

Adenosine triphosphate hydrolysis in rat dental tissues

Summary The purpose of this study was to try to differentiate histochemically between the various enzymes which may catalyze the hydrolysis of ATP in developing rat dental tissues. Freeze cut and freeze dried sections of molar and incisor teeth were incubated in lead capture-based media at pH 5.0, 7.2 or 9.4 with one of the following substrates: -glycerophosphate, AMP, ADP, ATP, AMP-PNP and tetrasodium pyrophosphate. To establish the enzymatic nature of the hydrolysis parallel sections were incubated after prior fixation in either formaldehyde or glutaraldehyde.By comparing the enzymatic stainings obtained with the various substrates and at the different pH: s, it was concluded that ATP can be visibly hydrolyzed in rat dental tissues by alkaline phosphatase (stratum intermedium, apical part of maturation ameloblasts, basal part of all ameloblasts, odontoblasts and subodontoblastic layer), specific ATPase (apical and basal parts of secretory ameloblasts) and ATP pyrophosphatase and/or adenylate cyclase (stratum intermedium, odontoblasts). Acid phosphatase, specific ADPase, 5-nucleotidase, inorganic pyrophosphatase, 3:5-cyclic-AMP-phosphodiesterase and adenylate kinase on the other hand, seem not to be engaged in the ATP hydrolysis to such a degree as to complicate the interpretation of the histochemical staining. The alkaline phosphatase part of the ATP hydrolysis appeared to be rather insensitive to aldehyde fixation, while the hydrolysis effected by specific ATPase and ATP pyrophosphatase and/or adenylate cyclase was extinguished after fixation with formaldehyde for 4 h or glutaraldehyde for 10 min.