Abstract
Metabotropic GABA(B) receptors are abundantly expressed at glutamatergic synapses where they control excitability of the synapse. Here, we tested the hypothesis that glutamatergic neurotransmission may regulate GABA(B) receptors. We found that application of glutamate to cultured cortical neurons led to rapid down-regulation of GABA(B) receptors via lysosomal degradation. This effect was mimicked by selective activation of AMPA receptors and further accelerated by coactivation of group I metabotropic glutamate receptors. Inhibition of NMDA receptors, blockade of L-type Ca(2+) channels, and removal of extracellular Ca(2+) prevented glutamate-induced down-regulation of GABA(B) receptors, indicating that Ca(2+) influx plays a critical role. We further established that glutamate-induced down-regulation depends on the internalization of GABA(B) receptors. Glutamate did not affect the rate of GABA(B) receptor endocytosis but led to reduced recycling of the receptors back to the plasma membrane. Blockade of lysosomal activity rescued receptor recycling, indicating that glutamate redirects GABA(B) receptors from the recycling to the degradation pathway. In conclusion, the data indicate that sustained activation of AMPA receptors down-regulates GABA(B) receptors by sorting endocytosed GABA(B) receptors preferentially to lysosomes for degradation on the expense of recycling. This mechanism may relieve glutamatergic synapses from GABA(B) receptor-mediated inhibition resulting in increased synaptic excitability.
Highlights
31003A_121963. 1 To whom correspondence should be addressed: Institute of Pharmacology and Toxicology, University of Zurich, Winterthurerstrasse 190, 8057 Zurich, Switzerland
Sustained Activation of Glutamate Receptors Triggers Lysosomal Degradation of GABAB Receptors—To investigate whether modulation of neuronal activity regulates degradation of GABAB receptors, cultured cortical neurons grown in 96-well plates were incubated for 90 min with agonists and antagonists of the main neurotransmitter systems and were analyzed for changes in GABAB receptor expression using the in-cell Western assay
Among the drugs tested, only glutamate and AMPA led to a significant reduction of GABAB receptor signals in cultured neurons with glutamate being more effective than AMPA (90 min of glutamate: 46 Ϯ 14%; AMPA: 64 Ϯ 13% of control; Fig. 1A)
Summary
Treatment of cultured cortical neurons with glutamate resulted in the loss of cell-surface GABAB receptors [12], and NMDA-induced excitotoxicity in hippocampal slices led to increased GABAB1 and decreased GABAB2 expression levels [13]. The glutamate-induced down-regulation was triggered by sustained activation of AMPA receptors, depended on Ca2ϩ influx through NMDA receptors as well as L-type Ca2ϩ channels, and resulted in reduced constitutive recycling and increased lysosomal degradation of GABAB receptors.
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