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MAP5 in cultured hippocampal neurons: expression diminishes with time and growth cones are not immunostained
1Division of Life Sciences, University of Texas, San Antonio 78285.
Abstract:
A monoclonal antibody was used to determine both the expression of the microtubule-associated protein MAP5 in cultured foetal rat hippocampal neurons as a function of culture age and the cellular distribution of the protein. When cultures at days 2 and 3 were examined by fluorescence microscopy, MAP5 immunostaining was localized intensely in neuronal cell bodies and neurites but not in growth cones. Extensive labelling of axons was seen at days 4 and 5. MAP5 staining was still prominent in neurons after 16 days in culture, and neurites at this time had grown over astrocytes but had completely avoided islands of non-astrocytic cells. MAP5 immunostaining was almost undetectable in cells that had been in culture for 20 days. The decreasing expression of MAP5 in cultured neurons as a function of time parallels that previously shown for MAP5 in intact neonatal rat brain. The effect of elevated temperature on MAP5 expression was also examined. Neurons grown for 9 days at 40 degrees C showed the same cellular distribution of MAP5 as cells grown at 37 degrees C. In particular, growth cones were again negative for MAP5 immunostaining. The absence of MAP5 in growth cones appears consistent with the fact that these structures contain labile microtubules. MAP5 has been shown to be a component of microtubule crossbridges and its absence might thus be expected to contribute to microtubule lability.
Insights
Microtubule-associated protein 5 (MAP5) expression decreases in cultured rat neurons over time, mirroring its decline in neonatal brain. MAP5 is absent in growth cones, consistent with their labile microtubules.
Area of Science:
- Neuroscience
- Cell Biology
- Developmental Biology
Background:
- Microtubule-associated proteins (MAPs) are crucial for microtubule stability and neuronal development.
- Microtubule-associated protein 5 (MAP5) plays a significant role in neurite outgrowth and microtubule organization.
Purpose of the Study:
- To investigate the expression and cellular distribution of MAP5 in cultured fetal rat hippocampal neurons over time.
- To examine the effect of elevated temperature on MAP5 expression and localization.
Main Methods:
- Primary neuronal cultures from fetal rat hippocampi were established.
- Immunofluorescence microscopy was used to detect MAP5 expression and localization at different culture ages (days 2-20).
- Neurons were cultured at standard (37°C) and elevated (40°C) temperatures.
Main Results:
- MAP5 immunostaining was intense in neuronal cell bodies and neurites early in culture (days 2-5), with extensive axonal labeling by days 4-5.
- MAP5 expression remained prominent up to 16 days but became nearly undetectable by 20 days.
- Elevated temperature (40°C) did not alter MAP5 cellular distribution, with growth cones consistently negative for MAP5.
- MAP5 was absent in growth cones, which contain labile microtubules, and MAP5 is a component of microtubule crossbridges.
Conclusions:
- The temporal decrease in MAP5 expression in cultured neurons parallels its decrease in the neonatal rat brain.
- The absence of MAP5 in growth cones supports its role in stabilizing microtubules, as growth cones possess dynamic, labile microtubules.