We have studied the developmental changes of glutamate-induced calcium (Ca2+) response in primary cultured hippocampal neurons at three different stages of cultures, 3, 7-8, and 14-16 days in vitro (DIV), using fura-2 single-cell digital micro-fluorimetry. We found that glutamate-induced Ca2+ signaling was altered during development, and that two different ionotropic glutamate receptors, α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors (AMPARs) and N-methyl-D-aspartate receptors (NMDARs), were differently involved in the modulation of calcium response at different stages of neuronal culture. In the stages of culture at 3 and 8 DIV, glutamate-induced Ca2+ influx was mostly because of AMPAR activation and subsequent opening of voltage-dependent calcium channels, as Ca2+ response can be largely reduced by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and by nifedipine. In the advanced culture (14-17 DIV), glutamate-induced Ca 2+ response was less sensitive to 6-cyano-7-nitroquinoxaline-2,3- dione and nifedipine. Furthermore, AMPA-induced Ca2+ response increased in a time-dependent manner during the cultures of 3-8 DIV and then reduced in the advanced culture of 14-17 DIV. NMDA-induced Ca2+ influx increased in a time-dependent manner, with a marked increase in the advanced culture (14-17 DIV). These results suggest that glutamate-induced Ca2+ signaling switched from AMPA-voltage-dependent calcium channel to NMDA-calcium signaling during development.
- α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptor
- Cell culture
- N-methyl-D-aspartate receptor
- Voltage-dependent calcium channels
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