An electron microscope study of resin production and secretion by the glands of seedlings of Betula pendula Roth.

Transmission and scanning electron microscopy were used to study the ultrastructure and secretory processes of resin glands on shoot stems of Betula pendula seedlings during seasonal growth. The multicellular peltate glands possess a cortex of columnar cells surrounding a parenchymal medulla differing from the stem parenchyma below. Myelin-like deposits comprising concentric layers of membranes and osmiophilic substances accumulate mainly in the cortical cells, while only the medullar cells have chloroplasts. Both of these deposits appear to be synthesized, initially, in the endoplasmic reticulum. The myelin-like material is believed to consist of steroidal triterpenoids in cytoplasmic membranes, and the osmiophilic deposits to represent phenolics. Studies of glands of different ages suggest that the electron-transparent resin ultimately exported is formed by combining the two initial products. In the cortical cells the secretion first accumulates in vesicles that fuse with larger ones and the periplasmatic space. From the latter the secretion diffuses through the cell wall and the resin is finally deposited in the subcuticular space. Secretion vesicles, but no periplasmatic deposits were seen in the medullar cells. At the end of seasonal growth the cortical cells are markedly vacuolate and appear to have lost their organelles. Some of the cells accumulate an electron-opaque material not seen earlier. The medullar cells of the glands are suberized before winter dormancy, but usually later than in the surrounding stem bark.

Interrelationships between glucose metabolism, energy state, and the cytosolic free calcium concentration in cortical synaptosomes from the guinea pig.

The stoichiometries of glycolysis and pyruvate oxidation were determined in cortical synaptosomes under varying rates of ATP consumption. Glycolysis was measured by using D-3-[3H]glucose as a marker and pyruvate oxidation by using D-3,4-[14C]glucose, which has to be metabolized to 1-[14C]pyruvate before being decarboxylated by the pyruvate dehydrogenase complex of intrasynaptosomal mitochondria. Cytosolic free Ca2+ concentration [( Ca2+]c) was determined in parallel and was manipulated by using EGTA in the incubation. The results show that in nonstimulated synaptosomes glycolysis and pyruvate oxidation are tightly coupled and stoichiometric. In the absence of Ca2+, when [Ca2+]c drops from 260 nM to 40 nM, glucose utilization increases, following the increase in energy demand, which has been shown to be due to elevated Na+ cycling. KCl depolarization, veratridine, and a mitochondrial uncoupler, carbonyl cyanide m-chlorophenylhydrazone, all stimulate glycolysis and pyruvate oxidation stoichiometrically, independently of the presence of external Ca2+. A rise in [Ca2+]c, therefore, is not required to regulate mitochondrial pyruvate metabolism. It is concluded that synaptosomes exhibit a high degree of respiratory control, that they rely on glucose oxidation for their energetics, and that stimulation of energy production can be achieved independently of changes in [Ca2+]c.

Ruthenium red inhibits the voltage-dependent increase in cytosolic free calcium in cortical synaptosomes from guinea-pig.

Effects of ruthenium red (RuR) on adenylates, plasma membrane potential (delta psi p) and cytosolic free calcium concentration ([Ca2+]c) in cortical synaptosomes from guinea-pig were investigated. Ten micromoles of RuR did not affect either energy levels as indicated by ATP/ADP ratio or the basal delta psi p. The resting [Ca2+]c in the presence of RuR was unchanged, but above 5 microM it inhibited by more than 50% of the voltage-activated increase in [Ca2+]c by K+-depolarization. In another experiment the potencies of 10 microM RuR and 100 microM verapamil to inhibit high K+-induced increase in [Ca2+]c were compared. It was found that either produced 59% inhibition and this inhibition was not potentiated by the substances together (65% inhibition). The extent of depolarisation of delta psi p by high external K+ was independent of the presence of RuR. RuR blocked only 20% of the increase in [Ca2+]c by veratridine treatment, indicating that Ca2+ accumulation into synaptosomal cytoplasm by veratridine involves some additional mechanisms other than depolarisation of delta psi p. The mechanism of inhibition of evoked release of neurotransmitters by RuR is discussed.

Chloride-dependent uncoupling of oxidative phosphorylation by triethyllead and triethyltin increases cytosolic free calcium in guinea pig cerebral cortical synaptosomes.

Metabolically competent isolated cerebral cortical nerve terminals were used to determine the effects of triethyllead (TEL) and triethyltin (TET) on cytosolic free calcium ([Ca2+]c), on plasma and mitochondrial membrane potentials, and on oxidative metabolism. In the presence of physiological concentrations of extracellular ions, 20 microM TEL and 20 microM TET increase [Ca2+]c from 185 nM to 390 and 340 nM, respectively. A simultaneous depolarization of plasma membrane potential (delta psi p) by only 3-4 mV occurs, a drop which is insufficient to open the voltage-sensitive Ca2+ channels. In contrast, an instant and substantial depolarization of mitochondrial membrane potential (delta psi m) upon addition of TEL and TET is evident, as monitored with safranine O fluorescence. At the same concentration, TEL and TET stimulate basal respiration of synaptosomes by 45%, induce oxidation of endogenous NAD(P)H, and reduce the terminal ATP/ADP ratio by 45%. Thus, TEL and TET inhibit ATP production of intrasynaptosomal mitochondria by a mechanism consistent with uncoupling of oxidative phosphorylation. This bioenergetic effect by TEL and TET can be prevented by omitting external chloride, and a concomitant reduction of the increase in [Ca2+]c by about 60% is observed. Uncoupling of mitochondrial ATP synthesis from oxidation by TEL and TET, [corrected] a process that is dependent on external chloride, is the main mechanism by which they [corrected] increase [Ca2+]c.