Fluorescence polarization studies of rat intestinal microvillus membranes.

Summary. The macrolide, FK-506, is a potent and effective inhibitor of lymphocyte activation. We studied the effects of FK-506 on human neutrophil activation induced by chemoattractants and by various substances which circumvent receptor stimulation. After preincubation for 5 min at 37 °C, FK-506(fMet-LeuPhe)-or platelet-activating factor-induced superoxide production in neutrophils by about 30°7o. At therapeutic concentrations (0.1-1nM) FK-506 was ineffective. FK-506 did not inhibit exocytosis and rises in cytosolic Ca 2÷ concentration [Ca2+] i mediated by these stimuli, and it did not at all inhibit neutrophil activation induced by C5a, leukotriene B 4 and 4fl-phorbol 12-myristate 13-acetate. FK-506 (1 gM) inhibited A23187-induced exocytosis by about 35%, but A23187-induced superoxide production was unaffected. After preincubation for 5 min at 37 °C, FK-506 inhibited fMet-Leu-Phe-induced superoxide production in dibutyryl cAMP-differentiated HL-60 cells by about 20%; preincubation with the drug for 24 h did not result in inhibition of superoxide production. FK-506 did not inhibit agonist-binding to formyl peptide receptors and fMet-Leu-Phe-stimulated GTP hydrolysis of heterotrimeric regulatory guanine nucleotidebinding proteins (G-proteins) in membranes from dibutyryl cAMP-differentiated HL-60 cells. FK-506 did not change steady-state and differential polarized phase fluorescence in Hb60 membranes using 1,6-diphenylhexa-1,3,5-triene and 12-(9-anthroyloxy)-stearate as probes. Our results show that FK-506 at supratherapeutic concentrations partially inhibits neutrophil activation. Inhibition by FK-506 of fMet-Leu-Phe-induced superoxide production is rapid in onset and is not due to inhibition of agonist-binding to receptors, interference with G-proteins or protein kinase C, reduction of rises in [Ca2+]i or alteration in physical membrane state.

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“…1979). R 0 values of 0.390 (DPH) (Lakowicz et al 1979) and 0.285 (12-AS) (Schachter and Shinitzky 1977) were used for calculations.…”

Section: Steady-state and Differential Polarized Phase Fluorometry Dmentioning

confidence: 99%

Partial inhibition of human neutrophil activation by FK-506 at supratherapeutic concentrations

Wenzel-Seifert

Seifert

1993

Naunyn-Schmiedeberg's Arch Pharmacol

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“…Total fluorescence intensity F =fvv + 2(/vH)· Relative microviscosity is proportional to rofr_i. where r 0 is the maximal limiting anisotropy of the probe; 0.362 for DPH (33] and 0.39 for TMA-DPH (31). A correction factor (G) for instrument asymmetry was also made using G = !Hv!IHH where /Hv is fluorescence intensity with horizontal excitation light and emitted light read vertically, and /HH is the corresponding value with the entire light path horizontally aligned.…”

Section: Determinations Of Membrane Fluiditymentioning

confidence: 99%

Subacute exposure to low concentrations of toluene affects dopamine-mediated locomotor activity in the rat

et al. 1991

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SummaryThe effects of low concentrations of toluene (4~80 ppm. 3 days. 6 h/day) were investigated on spontaneous and on apomorphine-induced locomotor activity in the rat. and were correlated to effects on S(-) [N-propyl-3 H(N)]-propylnorapomorphine ([ 3 H]NPA) binding in rat neostriatal membranes. on membrane fluidity. membrane leakage. and calcium levels in synaptosomes from the frontoparietal cortex, the neostriatum and the subcortical limbic area. and on serum hormone levels. Toluene exposure (80 ppm, post-exposure delay 18 h) alone did not affect locomotor activity, but attenuated apomorphineinduced (0.05 mg/kg, s.c.) suppression of rearing. and potentiated apomorphine-induced (I mg/kg. s.c.) increases in locomotion and rearing. Toluene exposure increased the K 0 value of [3H]NPA binding without affecting the Bmax· All these effects were absent at 40 ppm of toluene or at a post-exposure delay of 42 h. Toluene exposure (80 ppm. post-exposure delay of 18 h) did not affect the serum levels of prolactin. TSH, corticosterone. or aldosterone, or synaptosomal membrane fluidity and calcium levels. whereas membrane leakage was increased in the neostriatum. The present study indicates that the reduction of D-2 receptor affinity by short-term. low-dose toluene exposure is accompanied by a reduced D-2 autoreceptor function and an enhanced postsynaptic D-2 receptor function.

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“…Interpretation of the data derived utilizes the Perrin equation, which is written: ro/r = 1 + 3 TIP, where ro is the maximal limiting anisotropy (0.362) for DPH (23), r is the anisotropy, T is the excited state fluorophobe lifetime, and P is the rotational relaxation time. P is usually expressed as the anisotropy parameter [(ro/r)-11-I, where r = (Il1 -I l l ) + (Il1 + 2 1 ,).…”

Section: Methodsmentioning

confidence: 99%

“…The probe TMA-DPH may be less sensitive in rat renal brush border membranes (31) and, in general, is a less sensitive probe (32). Furthermore, the probe DPH is less influenced by changes in ionic composition (23). Finally since we found no change in transition temperature, P, and in apparent energy of activation in relation to dietary manipulation using DPH as a probe, we purposely chose not to examine other probes.…”

Section: Lt Adultmentioning

confidence: 99%

Membrane Fluidity and Phospholipid Composition in Relation to Sulfur Amino Acid Intake in Brush Border Membranes of Rat Kidney

1986

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ABSTRAn. The transport of ions and solutes across PS, phosphatidylserine biological membranes may relate to changes in the lipid PE, phosphatidylethanolamine microenvironment of the membrane which could alter the PG, phosphatidylglycerol activity or configuration of transport sites. Changes in the ANOVA, analysis of variance sulfur amino acid content of diets fed to young rats results in an increase in Na+-taurine symport in brush border membranes isolated from animals fed a low sulfur amino acid diet and a reduction in symport after a high taurine diet in comparison to uptake by membranes from normally fed animals ("the renal adaptive response"). We explored the possibility that diet-induced changes in brush border membrane symport relate to altered membrane fluidity and phospholipid composition in response to diet. An Arrhenius plot of initial rate (15 s) taurine uptake in breakpoint at 22" C, but no change in relation to dietary alteration.Fluorescence polarization data employing the probe 1-6-diphenyl-1,3,5-hexatriene best fits a two-phase linear model employing a computer model fitting program. Dietary manipulations did not change the breakpoint upper segment slope or lower segment slope after incubating fresh membranes with DPH over the temperature range 4-56" C. No change was evident in membrane phospholipid composition in relation to diet. This study indicates that the changes in initial rate Na+-taurine symport in relation to diet are less likely to be due to changes in the configuration of the transporter from an alteration of the lipid microenvironment of the membrane. (Pediatr Res 20: 1305-1309, 1986 Abbreviations LTD, low sulfur amino acid diet NTD, normal sulfur amino acid diet HTD, high taurine diet DPH, 1,6-diphenyl-1,3,5-hexatriene tPnA, transparanic acid TMA-DPH, 1,-(4-trimethylammoniumphenyl)-6-phenyl- Renal tubular epithelium is capable of adapting to changes in the dietary intake of various nutrients. This renal adaptive response has been demonstrated for the @-amino acid, taurine, which is conserved following diminished sulfur amino acid intake and hyperexcreted in the urine after dietary taurine excess (1-3). Studies employing isolated luminal brush border membrane vesicles are consistent with these changes in urinary excretion of taurine occurring as the consequence of a membrane-related event (3, 4), since the LTD appears to increase the Vmax of initial Naf-taurine symport and the HTD decrease the Vmax of uptake into vesicles. We conducted these studies to determine if these diets influence the microviscosity and phospholipid composition of these vesicles.Changes in the physical status or in the lipid composition of biological membranes can alter the movement of ions and solutes across these membranes (5). The technique of fluorescence polarization has been employed to define the "fluidity" of hepatic, erythrocyte, bacterial cell, gastrointestinal, and renal plasma membranes (6-1 1). The importance of the lipid microenvironment of biological membranes is also of interest since the activ...

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Fluorescence polarization studies of rat intestinal microvillus membranes.

Cited by 188 publications

References 52 publications

Partial inhibition of human neutrophil activation by FK-506 at supratherapeutic concentrations

Partial inhibition of human neutrophil activation by FK-506 at supratherapeutic concentrations

Subacute exposure to low concentrations of toluene affects dopamine-mediated locomotor activity in the rat

Membrane Fluidity and Phospholipid Composition in Relation to Sulfur Amino Acid Intake in Brush Border Membranes of Rat Kidney

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