N-ACETYLASPARTYLGLUTAMATE STIMULATES METABOTROPIC GLUTAMATE-RECEPTOR-3 TO REGULATE EXPRESSION OF THE GABA(A) ALPHA-6 SUBUNIT IN CEREBELLAR GRANULE CELLS

We have shown that the vertebrate neuropeptide N-acetylaspartylglutama te (NAAG) meets the criteria for a neurotransmitter, including functio n as a selective metabotropic glutamate receptor (mGluR) 3 agonist. Sh ort-term treatment of cerebellar granule cells with NAAG (30 mu M) res ults in the transient increase in content of GABA(A) alpha 6 subunit m RNA. Using quantitative PCR, this increase was determined to be up to 170% of control values. Similar effects are seen following treatment w ith trans-1-aminocyclopentane-1,3-dicarboxylate and glutamate and are blocked by the mGluR antagonists (2S,3S,4S)-2-methyl-2-(carboxycyclopr opyl) glycine and (2S)-alpha-ethylglutamic acid. The effect is pertuss is toxin-sensitive. The increase in alpha 6 subunit mRNA level can be simulated by activation of other receptors negatively linked to adenyl ate cyclase activity, such as adenosine A1, alpha 2-adrenergic, muscar inic, and GABA(B) receptors. Forskolin stimulation of cyclic AMP (cAMP ) levels abolished the effect of NAAG. The change in alpha 6 levels in duced by 30 mu M NAAG can be inhibited in a dose-dependent manner by s imultaneous application of increasing doses of the beta-adrenergic rec eptor agonist isoproterenol. The increase in alpha 6 mRNA content is f ollowed by a fourfold increase in alpha 6 protein level 6 h posttreatm ent. Under voltage-clamped conditions, NAAG-treated granule cells demo nstrate an increase in the furosemide-induced inhibition of GABA-gated currents in a concentration-dependent manner, indicating an increase in functional alpha 6-containing GABA(A) receptors. These data support the hypothesis that NAAG, acting through mGluR3, regulates expression of the GABA(A) alpha 6 subunit via a cAMP-mediated pathway and that c AMP-coupled receptors for other neurotransmitters may similarly influe nce GABA(A) receptor subunit composition.