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     

Glucagon activates mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase in vivo by decreasing the extent of succinylation of the enzyme

1. 3-Hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase (EC 4.1.3.5) in extracts of rat liver mitochondria can be inactivated by succinyl-CoA and activated by incubation in a medium designed to cause desuccinylation ('desuccinylation medium'). 2. The enzyme is less active in extracts of whole liver from control rats than from rats treated with glucagon or mannoheptulose. Incubation in desuccinylation medium raises the activity in extracts from control rats to the same value as treated rats, suggesting that the extent of succinylation in vivo is greater in controls than in hormone-treated animals. 3. This result is also obtained in liver homogenates and in isolated mitochondria. 4. Increasing the succinyl-CoA content of mitochondria to the same high level lowers the enzyme activity to the same value in mitochondria isolated from control or treated rats. In each case subsequent incubation of the lysates in desuccinylation medium raises the enzyme activity by the same extent. 5. Measurement of the incorporation of radiolabel from 2-oxo[5-14C]glutarate into protein is consistent with the proposal that all these changes in activity in isolated mitochondria may be explained by changes in the extent of succinylation of the enzyme. 6. From these data and our earlier work we conclude that, in vivo, mitochondrial HMG-CoA synthase in fed rats is normally substantially succinylated (about 40%) and inactivated, and that glucagon increases the activity of HMG-CoA synthase by lowering the concentration of succinyl-CoA and thus decreasing the extent of succinylation of the enzyme (to less than 10%). This may be an important control mechanism in ketogenesis.     

Sugar transport by the bacterial phosphotransferase system. The intrinsic fluorescence of enzyme I

Enzyme I of the bacterial phosphoenolpyruvate: glycose phosphotransferase system has 2 tryptophan residues/monomer, as determined spectrophotometrically. The tryptophan fluorescence has been investigated with the aid of nanosecond time-resolved techniques. The decay of the fluorescence intensity was analyzed in terms of a biexponential function. The contribution of the emission associated with the shorter decay constant increases from 17-19% at 1 degree C to 43-44% at room temperature. Decay-associated spectra obtained with Enzyme I indicate different spectral distributions associated with the two decay constants. The measurement of tumbling of Enzyme I as a function of temperature revealed a transition of rotational rates between 5 and 15.5 degrees C. Global analysis allowed decomposition of the anisotropy decay into a formulation consistent with monomer and dimer rotational contributions.     

Convenient preparative synthesis of [14C]trehalose from [14C]glucose by intact Escherichia coli cells

At high osmolarity, Escherichia coli synthesizes trehalose intracellularly, irrespective of the nature of the carbon source. Synthesis proceeds via the transfer of UDP-glucose to glucose 6-phosphate, yielding trehalose 6-phosphate, followed by its dephosphorylation to trehalose (H.M. Giaeyer, B.O. Styrvold, I. Kaasen, and A.R. Str?m, J. Bacteriol. 170:2841-2849, 1988). This reaction was exploited to preparatively synthesize [14C]trehalose from exogenous [14C]glucose by using intact bacteria of a mutant (DF214) that could not metabolize glucose. The total yield of radiochemically pure trehalose from glucose was routinely more than 50%.     

Hypothyroidism in rats decreases mitochondrial inner membrane cation permeability

We investigated the cation permeability of liver mitochondria isolated from hypothyroid or euthyroid rats by measuring the rate of swelling of respiring mitochondria in acetate salts as a function of membrane potential. Mitochondria from hypothyroid rats have a decreased permeability of roughly 3-fold in the presence of monovalent cations K and tetramethylammonium at any (measured) membrane potential. Since the monovalent cation leak and the proton leak are known to respond similarly to membrane potential our results support the theory that the difference in non-phosphorylating respiration rate between mitochondria from hypothyroid and euthyroid rats is due to a difference in proton leak.     

A quantitative assessment of the use of 36Cl- distribution to measure plasma membrane potential in isolated hepatocytes

The plasma membrane potential of isolated rat hepatocytes was clamped at different values between 0 and -68 mV by addition of valinomycin in the presence of different extracellular concentrations of K+, and measured by the distribution of 86Rb+ between cells and medium. 36Cl- distribution came to steady state in 10-15 min. This steady-state distribution was compared to the plasma membrane potential over a range of values. 36Cl- distribution provided an accurate measurement of plasma membrane potential between -4 and -40 mV. At higher potentials intracellular chloride concentration is less than 20% of the extracellular concentration and errors due to uncertainties in the measurement of intracellular volume and of the contamination of cell pellets by extracellular medium precluded accurate determination of membrane potential: thus in our experiments 36Cl- underestimated the plasma membrane potential at -68 mV by 8 mV.     

Odorant response of isolated olfactory receptor cells is blocked by amiloride

Summary Olfactory receptor cells were isolated from the nasal mucosa ofRana esculenta and patch clamped. Best results were obtained with free-floating cells showing ciliary movement. 1)On-cell mode: Current records were obtained for up to 50 min. Under control conditions they showed only occasional action potentials. The odorants cineole, amyl acetate and isobutyl methoxypyrazine were applied in saline by prolonged superfusion. At 500 nanomolar they elicited periodic bursts of current transients arising from cellular action potentials. The response was rapidly, fully and reversibly blocked by 50 m amiloride added to the odorant solution. With 10 m amiloride, the response to odorants was only partially abolished. 2)Whole-cell mode: Following breakage of the patch, the odorant response was lost within 5 to 15 min. Prior to this, odorants evoked a series of slow transient depolarizations (0.1/sec, 45 mV peak to peak) which reached threshold and thus elicited the periodic discharge of action potentials. These slow depolarizing waves were reversibly blocked by amiloride, which stabilized the membrane voltage between –80 and –90 mV. We conclude that amiloride inhibits chemosensory transduction of olfactory receptor cells, probably by blocking inward current pathways which open in response to odorants.     

A morphometric comparison of dissimilar early development in sibling species of Platynereis (Annelida,Polychaeta)

Summary Early development of Platynereis massiliensis was studied in serial sections of fixed embryos and in living or fixed embryos whose nuclei had been made visible with a fluorescent label. The unfertilized egg is an ellipsoid with three axes of differing length. The longest axis corresponds to the dorsoventral axis of the developing embryo. Egg volume is ten times that in the sibling species, P. dumerilii, mainly due to increased yolk content. The timing and spatial pattern of cleavage were observed from first cleavage to the 62-cell stage. Volumes of the blastomeres, their nuclei, their yolk-free cytoplasm and their yolk were determined from serial sections up to the 29-cell stage. In the P. massiliensis embryo, cell cycles are on average 3.7 times longer than in P. dumerilii; volume proportions among the blastomeres also differ and the macromeres containing the bulk of yolk are particularly large, but otherwise the cleavage patterns, differential segregation of yolk and yolk-free cytoplasm, and the histogenetic fates of the blastomeres are the same as in P. dumerilii. This equivalence of cell lineage and of cytoplasmic segregation mechanisms in both species, maintained in spite of the different appearance of the embryos, suggests functional importance of and selective constraint on these developmental features. The relatively accelerated divisions of the 2d cell line in P. massiliensis may be interpreted as the precocious development of cell lines which give rise to adult structures. Several structures, obviously functional in developing P. dumerilii, have lost their function in P. massiliensis: the egg contains few cortical granules, giving rise to only a moderate egg jelly layer in the zygote; prototroch cells develop cilia, but the heavy embryo is unable to swim; the larva develops three pairs of parapodia but, unlike the corresponding stage in P. dumerilii, is not capable of coordinate locomotion. This loss of motility is related to the brooding habit of the species developing inside the parental tube and is explained as the result of a switch from pelagic to benthic, protected reproduction in P. massiliensis. Offprint requests to: A.W.C. Dorresteijn