Identification of Adenosine Receptor in Rat Pineal Gland: Evidence for A-2 Selectivity

We have examined the binding of the adenosine agonist radioligands [3H]cyclohexyladenosine [( 3H]CHA), R-N6-[3H]phenylisopropyladenosine [( 3H]R-PIA), and 5'-N-ethylcarboxamido[3H]adenosine [( 3H]NECA) to membranes prepared from rat pineal gland. The results showed that the A-1-selective ligands (CHA and R-PIA) had less than or equal to 10% specific binding. By contrast, [3H]NECA, a nonselective A-1/A-2 ligand, gave 72% specific binding of the total binding. This specific binding was insensitive to cyclopentyladenosine (50 nM) or R-PIA (50 microM). To characterize this binding, we used the N-ethylmaleimide pretreatment method. Under these conditions, this binding was of high affinity with a KD of 51 +/- 10 nM and an apparent Bmax of 1,060 +/- 239 fmol/mg of protein. Specific binding was unaffected by the presence of MgCl2 (10 mM) but was sensitive to guanylylimidodiphosphate (100 microM) (-25%), a result suggesting the involvement of an N-protein mechanism in the coupling of the adenosine receptor labeled by [3H]NECA to other components of the receptor complex. The rank of activity of adenosine analogues in displacing specific [3H]NECA binding was NECA greater than 2-chloroadenosine greater than S-adenosyl-L-homocysteine greater than CHA. Binding was also displaced by 3-isobutyl-1-methylxanthine (IC50 = 23.6 microM). These findings are consistent with the selective labeling by [3H]NECA of an A-2-type adenosine receptor in rat pineal membranes.     

Hydroponics versus field lysimeter studies of urea, ammonium and nitrate uptake by oilseed rape(Brassica napus L.)

N-fertilizer use efficiencies are affected by their chemical composition and suffer from potential N-losses by volatilization. In a field lysimeter experiment, (15)N-labelled fertilizers were used to follow N uptake by Brassica napus L. and assess N-losses by volatilization. Use of urea with NBPT (urease inhibitor) showed the best efficiency with the lowest N losses (8% of N applied compared with 25% with urea alone). Plants receiving ammonium sulphate, had similar yield achieved through a better N mobilization from vegetative tissues to the seeds, despite a lower N uptake resulting from a higher volatilization (43% of applied N). Amounts of (15)N in the plant were also higher when plants were fertilized with ammonium nitrate but N-losses reached 23% of applied N. In parallel, hydroponic experiments showed a deleterious effect of ammonium and urea on the growth of oilseed rape. This was alleviated by the nitrate supply, which was preferentially taken up. B. napus was also characterized by a very low potential for urea uptake. BnDUR3 and BnAMT1, encoding urea and ammonium transporters, were up-regulated by urea, suggesting that urea-grown plants suffered from nitrogen deficiency. The results also suggested a role for nitrate as a signal for the expression of BnDUR3, in addition to its role as a major nutrient. Overall, the results of the hydroponic study showed that urea itself does not contribute significantly to the N nutrition of oilseed rape. Moreover, it may contribute indirectly since a better use efficiency for urea fertilizer, which was further increased by the application of a urease inhibitor, was observed in the lysimeter study.     

Inhibition of pancreatic and microbial lipases by proteins

We have compared the effect of several proteins, including melittin, beta-lactoglobulin A, serum albumin, ovalbumin and myoglobin, on the hydrolysis of tributyrin and triolein by lipases from various origins. All proteins tested inactivate pancreatic lipase in absence of colipase and bile salt. Inhibition is not significantly reversed by colipase in absence of bile salt except in systems containing tributyrin and melittin or triolein and beta-lactoglobulin A. In all other cases, activation of pancreatic lipase by colipase in presence of inhibitory protein requires the presence of bile salt. Lipase from Rhizopus delemar is also inhibited by the proteins that inactivate pancreatic lipase. In contrast, the activity of lipase from Rhizopus arrhizus is not affected by the proteins in the same concentration range. Inhibition of lipase activity by amphiphiles such as proteins or detergents appears to be a general phenomenon not directly related to a decrease in tension at the triacylglycerol-water interface. Inhibition could be the result of desorption of lipase from its substrate due to a change in interfacial quality.     

Differential pulse voltammetry: focus on the measurement of 5-hydroxyindole compounds and uric acid in the brain

The electrochemical signal obtained at the +300 mV potential (peak 3) in rats implanted for more than one week could be dependent upon extracellular fraction of 5-hydroxyindolacetic acid (5-HIAA) since a single injection of Pargyline is sufficient to suppress it in caudate and raphe dorsalis nuclei. In contrast, in rats implanted for less than one week, this signal could be dependent upon extracellular fractions of 5-HIAA and uric acid since consecutive injections of Pargyline and Allopurinol are necessary to suppress it.     

Evaluating the contributions of change in investment and change in efficiency to age‐related declines in male and female reproduction

It is commonly observed that reproduction decreases with age, often at a different rate in males and females. This phenomenon is generally interpreted as senescence. Such reproductive declines may stem from at least two sources: a change in resource allocation and a decline in the ability to convert resources into offspring. This distinction is important because a shift in resource allocation may be favoured by selection, while reduced efficiency is purely deleterious. We propose a way to distinguish whether a decline in reproduction is purely deleterious based on estimating reproductive investment, output, and their ratio, efficiency. We apply this approach to the hermaphroditic snail Physa acuta and demonstrate that both male and female functions decline with age. The male decline largely stems from reduced investment into male activity while female decline is due to increased reproductive inefficiency. This shows that age‐related declines in reproduction can occur for a number of different reasons, a distinction that is usually masked by the general term ‘senescence’. This approach could be applied to any species to evaluate age‐related reproductive decline. We advocate that future studies measure age trajectories of reproductive investment and output to explore the potential processes hidden behind the observation that reproduction declines with age.     

Brain protein synthesis in awake rats: equilibrium of L-methionine between plasma and direct precursor pool

After 1 hr. of continuous infusion of L-(35S)methionine the specific activities of L-methionine in plasma and tissue-free and tRNA-bound L-methionine in brain were in the same range. This result indicates that, under steady-state conditions, dilution of the precursor pool for protein synthesis (L-methionyl-tRNA) by L-methionine derived from a source other than plasma can be considered as negligible.     

Inhibition of lipases by proteins: a binding study using dicaprin monolayers

We previously reported that the inhibition of pancreatic and Rhizopus delemar lipases by proteins is due to the protein associated with lipid and is not caused by direct protein-enzyme interaction in the aqueous phase [Gargouri, Y., Piéroni, G., Rivière, C., Sugihara, A., Sarda, L., & Verger, R. (1985) J. Biol. Chem. 260, 2268-2273]. In this study, using radiolabeled lipases, serum albumin, and beta-lactoglobulin A, we investigated their respective binding with respect to lipolysis of dicaprin monolayers. Lipase inhibition was found to be correlated with a lack of lipase binding to mixed protein-dicaprin films or to a desorption of lipase from the interface when inhibitory protein was added later. Since a large proportion of the lipid film remained potentially accessible to the enzyme in the presence of inhibitory protein, it was concluded that the observed decrease in lipase binding to the interface was due to a variation of the physiochemical properties of the lipid-water interface following binding of inhibitory protein. On the basis of the results presented here, it is proposed that mixed protein-glyceride films could be used to characterize the interaction of various lipases with lipid substrates and to classify these enzymes according to their penetration power.