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Decrease in the fluidity of brush-border membrane vesicles induced by gentamicin. A spin-labeling study
T Moriyama1, H Nakahama, Y Fukuhara
1First Department of Medicine, Osaka University Medical School, Japan.
Abstract:
In our previous paper (Horio et al., Biochim Biophys Acta 858: 153-160, 1986), we reported that the addition of gentamicin in vitro to rabbit renal brush-border membrane vesicles decreases the apparent Vmax of Na+-dependent D-glucose transport without affecting the apparent Km. In the present study, we investigated the effects of gentamicin on the physical state of spin-labeled rabbit renal brush-border membranes, using electron spin resonance spectrometry. Brush-border membrane vesicles were prepared from outer cortex (mainly contains early proximal tubule) and outer medulla (containing primarily late proximal tubule), and the gentamicin toxicities in both preparations were compared. Significant decreases were observed in the membrane fluidity of 5 mM gentamicin-treated brush-border membranes. The fluidity of outer cortical brush-border membranes was affected at both 25 degrees and 35 degrees, whereas that of outer medullary membranes was affected only at 35 degrees. Two different stearic acid spin labels revealed that gentamicin affected the fluidity only in the superficial region of the membranes. We also demonstrated that the gentamicin-induced decreases in Na+-dependent D-glucose transport and in the membrane fluidity were recovered by washing gentamicin-treated brush-border membranes. We suggest that gentamicin binds to the superficial region of brush-border membranes and inhibits Na+-dependent D-glucose transport across brush-border membranes through the decrease in the membrane fluidity.
Insights
Gentamicin reduces kidney membrane fluidity, inhibiting glucose transport. This effect is reversible and localized to the membrane surface, suggesting a specific binding interaction.
Area of Science:
- Nephrology
- Biochemistry
- Membrane Biophysics
Background:
- Previous work showed gentamicin decreases Na+-dependent D-glucose transport Vmax in rabbit renal brush-border membranes.
- Gentamicin is an aminoglycoside antibiotic known for nephrotoxicity.
Purpose of the Study:
- To investigate gentamicin's effects on the physical state of renal brush-border membranes.
- To compare gentamicin's toxicity in outer cortex and outer medulla brush-border membranes.
- To elucidate the mechanism of gentamicin-induced inhibition of D-glucose transport.
Main Methods:
- Electron spin resonance (ESR) spectrometry using spin-labeled brush-border membrane vesicles.
- Preparation of vesicles from outer cortex and outer medulla.
- Assessment of membrane fluidity and D-glucose transport.
- Recovery experiments involving washing treated membranes.
Main Results:
- Gentamicin (5 mM) significantly decreased brush-border membrane fluidity.
- Membrane fluidity was affected in outer cortical membranes at 25°C and 35°C, and outer medullary membranes only at 35°C.
- Gentamicin's effect on fluidity was localized to the superficial membrane region.
- Inhibition of D-glucose transport and reduced membrane fluidity were reversible upon washing.
Conclusions:
- Gentamicin impairs renal brush-border membrane fluidity, particularly in the superficial region.
- This biophysical change is linked to the inhibition of Na+-dependent D-glucose transport.
- The observed effects are reversible, suggesting a binding interaction rather than irreversible damage.