TEMPORAL AND DEPOLARIZATION-INDUCED CHANGES IN THE ABSOLUTE AMOUNTS OF MESSENGER-RNAS ENCODING METABOTROPIC GLUTAMATE RECEPTORS IN CEREBELLAR GRANULE NEURONS IN-VITRO

Cerebellar granule neurons in primary culture express metabotropic glu tamate receptors (mGluRs) coupled to the stimulation of phosphoinositi de hydrolysis and to the inhibition of cyclic AMP (cAMP) formation. To evaluate which mGluR mRNAs are expressed in granule neurons under dif ferent depolarizing conditions, we measured the absolute amounts of se lected receptor mRNAs in neurons cultured for 3-13 days in the presenc e of either 10 or 25 mM KCl. mGluR-specific primer pairs and internal standards, corresponding to unique regions of mGluR1a, mGluR2, mGluR3, mGluR4, and mGluR5, were constructed and used in a competitive PCR-de rived assay to quantify the corresponding mRNA levels. For phosphoinos itide-coupled receptors, the absolute content of mGluR1a mRNA was thre e to 10 times higher than the content of mGluR5 mRNA. The expression o f mGluR5 mRNA increased up to 9 days in vitro and was much higher in 1 0 mM than in 25 mM KCl. For the cAMP-coupled receptors, there was a la rge amount of mGluR4 mRNA and a much smaller content of the mGluR3 and mGluR2 mRNAs. Maintaining the granule neurons in vitro in 10 mM KCl i ncreased the absolute amount of mRNAs encoding mGluR2 and mGluR4 at 9 and 13 days in vitro. In contrast, the content of the mGluR3 mRNA was consistently higher in neurons cultured in 25 mM KCl. These data are c onsistent with the possibility that in primary cultures of cerebellar neurons, phosphoinositide responses may be predominantly mediated by m GluR1a, rather than mGluR5, and that cAMP inhibition involves preferen tially mGluR4 and mGluR3.