Endocytosis of GABA_B receptors modulates membrane excitability in the single-celled organism Paramecium

GABA_B receptors modulate swimming behavior in Paramecium by inhibiting dihydropyridine-sensitive Ca²⁺ channels via G-proteins. Prolonged occupancy of GABA_B receptors by baclofen results in a decrease in GABA_B receptor functions. Since changes in the number of cell-surface GABA_A receptors have been postulated to be of importance in modulating inhibitory synaptic transmission in neurons, we have studied the cell-surface expression and maintenance of GABA_B receptors in P. primaurelia. In this study, we use immunostaining in electron and confocal microscopy to demonstrate that constitutive internalization of GABA_B receptors in P. primaurelia is mediated by clathrin-dependent and -independent endocytosis. Indeed, GABA_B receptors colocalize with the adaptin complex AP2, which is implicated in the selective recruitment of integral membrane proteins to clathrin-coated vesicles, and with caveolin 1, which is associated with uncoated membrane invaginations. Furthermore, when endocytosis is blocked with hypertonic medium, cytosol acidification, filipin or with a peptide that disrupts the association between amphiphysin and dynamin, the effect of baclofen on swimming is increased. These results suggest that GABA_B receptor endocytosis into clathrin-coated and -uncoated vesicles represents an important mechanism in the modulation of swimming behavior in Paramecium.