Action of polyvalent cations on sodium transport across skin of larval and adult Rana catesbeiana

The actions of alkaline earth (AE) and transition element (TE) cations on Na+ transport across skin of larval and adult Rana catesbeiana were compared. Bathed on the outside by Ca+2-free Ringer's, both larval and adult skins maintained a stable short-circuit current (3-4 mu Amps cm-2 for larval skin and 20-30 mu Amps cm-2 for adult skin). Addition of Ca+2 to the external bath reduced the SCC; maximal inhibition was about 36% for larval skin and 22% for adult skin. Other AE divalent cations were also inhibitory. The order of effectiveness was: Ba+2 = Ca+2 greater than Sr+2 greater than Mg+2 for larval skin and Ba+2 greater than Ca+2 = Mg+2 for adult skin. Sodium influx was markedly elevated when Ca+2 was removed from the external medium. Current-voltage analysis indicated that Ca+2 increases the resistance of the active pathway without affecting the shunt resistance or the electromotive force of Na+ transport (ENa) in larval and adult skins. The SCC across adult skin was stimulated by TE cations (Co+2, Cd+2, La+3). These ions were inhibitory on larval skin. The transition in the response occurred at stage XXI. The inhibitory effect of TE on larvel skin resembles that seen in response to AE cations and we postulate a common mechanism. Since larval skin lacks the selective Na+ channels found in apical membranes of adult skin, we infer that the mechanism of inhibition by AE cations is not on these channels. A more general phenomenon such as change in surface charge at the apical membrane seems more reasonable.(ABSTRACT TRUNCATED AT 250 WORDS)     

pH-dependent inhibitory effects of tris and lithium ion on intestinal brush-border sucrase

Tris and two of its hydroxylated amine analogs were examined in a metal-free, universal n-butylamine buffer, for their interaction with intestinal brush border sucrase. Our recent three-proton-families model (Vasseur, van Melle, Frangne and Alvarado (1988) Biochem. J., 251, 667-675) has provided the sucrase pK values necessary to interpret the present work. At pH 5.2, 2-amino-2-methyl-l-propanol (PM) causes activation whereas Tris has a concentration-dependent biphasic effect, first causing activation, then fully competitive inhibition. The amine species causing activation is the protonated, cationic form. The difference between the two amines is related to the fact that Tris has a much lower pKa value than PM (respectively, 8.2 and 9.8). Even at pH 5.2, Tris (but not PM) exists as a significant proportion of the free base which, by inhibiting the enzyme fully competitively, overshadows the activating effect of the cationic, protonated amine. Above pH 6.8, both Tris and PM act as fully competitive inhibitors. These inhibitions increase monotonically between pH 6.5 and 8.0 but, above pH 8, inhibition by 2.5 mM Tris tends to diminish whereas inhibition by 40 mM PM increases abruptly to be essentially complete at pH 9.3 and above. As pH increases from 7.6 to 9.0, the apparent affinity of the free amine bases decreases whereas that of the cationic, protonated amines, increases. In this way, the protonated amines replace their corresponding free bases as the most potent inhibitors at high pH. The pH-dependent inhibition by 300 mM Li+ is essentially complete at pH 8, independent of the presence or absence of either 2.5 mM Tris or 40 mM PM. Even at pH 7.6, an excess (300 mM) of Li+ causes significant increases in the apparent Ki value of each Tris, PD (2-amino-2-methyl-1-3-propanediol) and PM, suggesting the possibility of a relation between the effects of Li+ and those of the hydroxylated amines which in fact are mutually exclusive inhibitors. The inhibitory results are interpreted in terms of a mechanistic model in which the free bases bind at two distinct sites in the enzyme's active center. Binding at the glucosyl sub-site occurs through the amine's free hydroxyl groups. This positioning facilitates the interaction between the lone electron pair of the deprotonated amino group with a proton donor in the enzyme's active center, characterized by a pK0 around 8.1. When this same group deprotonates, then the protonated amines acting as proton donors replace the free bases as the species giving fully competitive inhibition of sucrase.     

Similar Effects of Ozone on Four Cultivars of Lettuce in Open Top Chambers During Winter

Ozone is the major phytotoxic air pollutant that reduces the yield of several agricultural crops in the Spanish Mediterranean area. We studied four lettuce cultivars (Lactuca sativa L.) for the effects of different O₃ concentrations during the winter on chlorophyll (Chl) a fluorescence, lipid peroxidation, and root length in outdoor open-top chambers. Under O₃ the photosynthetic quantum conversion declined while heat emissions increased in all cultivars; these results provide more evidence of non-filtered air with additional ozone (NFA+O₃) treatment compared with non-filtered air (NFA) and charcoal filtered ozone-free air (CFA). Changes in the Chl a fluorescence may be associated with an increase in membrane lipid peroxidation as well as with observed reduction of root length under O₃ stress.