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Purified rat brain microvessels exhibit both acid and neutral sphingomyelinase activities
J B Carré1, O Morand, P Homayoun
1Laboratoire de Neurochimie, INSERM Unité 134, Hôpital de la Salpêtrière, Paris, France.
Abstract:
Purified rat brain microvessels have been shown to hydrolyze radiolabeled sphingomyelin by means of two different enzyme systems. Enzymatic activity was detected at pH 7.4 and was strongly stimulated by magnesium or manganese and inhibited by calcium. Activity at pH 5.1 could also be found and was not dependent on any of these cations. At neutral pH and in the presence of magnesium, the rate of sphingomyelin hydrolysis did not exhibit a linear relationship with protein concentration. In contrast, increasing the protein concentration from 0.05 to 0.5 mg/ml resulted in a constant increase of sphingomyelin hydrolysis at pH 5.1. Kinetic parameters of both neutral and acid activities have been determined and were similar in magnitude to values reported previously for neural sphingomyelinases. This work demonstrates the occurrence of a neutral sphingomyelinase activity in purified rat brain microvessels, an observation raising the question of its role at the level of the blood-brain interface.
Insights
Rat brain microvessels contain neutral sphingomyelinase activity, an enzyme that hydrolyzes sphingomyelin. This finding suggests a potential role for this enzyme at the blood-brain barrier.
Area of Science:
- Biochemistry
- Neuroscience
- Cell Biology
Background:
- Sphingomyelin is a key component of cell membranes.
- Enzymatic hydrolysis of sphingomyelin is crucial for cellular processes.
- The presence and function of sphingomyelinases in brain microvessels are not fully understood.
Purpose of the Study:
- To investigate the presence and characteristics of sphingomyelinase activity in purified rat brain microvessels.
- To determine the kinetic properties of these enzymes.
- To explore the potential role of neutral sphingomyelinase at the blood-brain interface.
Main Methods:
- Purification of rat brain microvessels.
- Assay of sphingomyelin hydrolysis at different pH values (neutral and acidic).
- Investigation of the effects of cations (Mg2+, Mn2+, Ca2+) on enzymatic activity.
- Determination of kinetic parameters (Km, Vmax) for the identified enzymes.
- Analysis of the relationship between protein concentration and enzyme activity.
Main Results:
- Two distinct sphingomyelinase activities were detected in purified rat brain microvessels.
- Neutral activity (pH 7.4) was stimulated by Mg2+ or Mn2+ and inhibited by Ca2+.
- Acidic activity (pH 5.1) was cation-independent.
- Enzyme kinetics at both pH values were comparable to previously reported neural sphingomyelinases.
- A non-linear relationship between protein concentration and neutral sphingomyelinase activity was observed at pH 7.4, while acidic activity showed a linear increase.
Conclusions:
- Purified rat brain microvessels possess both neutral and acidic sphingomyelinase activities.
- The neutral sphingomyelinase activity identified in brain microvessels warrants further investigation into its function at the blood-brain barrier.
- These findings contribute to understanding lipid metabolism and enzyme localization within the neurovasculature.